The effect of C-X-C motif chemokine ligand 13 in cutaneous squamous cell carcinoma treated with aminolevulinic acid-photodynamic therapy.
Zhu Lude,Zhang Guolong,Wang Peiru,Zhang Linglin,Ji Jie,Liu Xiaojing,Zhou Zhongxia,Zhao Jingjun,Wang Xiuli
Photodiagnosis and photodynamic therapy
BACKGROUND:Active recruitment of inflammatory cells into tumors may be vital for antitumor immunity in cutaneous squamous cell carcinoma (cSCC) after photodynamic therapy. Chemokines play important roles in inflammatory cell recruitment. Moreover, C-X-C motif chemokine ligand 13 (CXCL13) is thought to be a pivotal chemokine involved in inflammatory response and antitumor effect. Here, we examined the roles of CXCL13 in the response of cSCC to ALA-PDT. METHODS:Microarray analysis was used to select the chemokines involved in cSCC treated with ALA-PDT. The expression and transcriptional activity of CXCL13 were assessed by immunohistochemistry and quantitative real-time polymerase chain reaction. Western blotting was used to detect C-X-C motif chemokine receptor 5 (CXCR5) expression. The role of CXCL13 in ALA-PDT efficacy was assessed in vivo. RESULTS:Microarray analysis of total 63 chemokines and their receptors showed that the expression of 21 chemokines and 13 receptors were up-regulated in cSCC after ALA-PDT; in particular, CXCL13 was significantly upregulated. Immunohistochemistry showed that cancer-associated fibroblasts (CAFs) may be the main source of CXCL13 upregulation in the cSCC microenvironment after ALA-PDT. The efficacy of ALA-PDT in the treatment of cSCC was significantly reduced after CXCL13 inhibition. CONCLUSION:CXCL13 plays important roles in the antitumor effect of ALA-PDT for cSCC and may originate mainly from CAFs in the cSCC microenvironment.
The Tumor Microenvironment in Follicular Lymphoma: Its Pro-Malignancy Role with Therapeutic Potential.
International journal of molecular sciences
In the follicular lymphoma (FL) microenvironment, CXCR5ICOSPD1BCL6 follicular helper T (Tfh) cells, which closely correlate with FL B cells in neoplastic follicles, play a major role in supporting FL. Interleukin-4 secreted by Tfh cells triggers the upregulation of the lymphocyte chemoattractant CXCL12 in stromal cell precursors, in particular by fibroblastic reticular cells (FRCs). In turn, mesenchymal stem cells (MSCs) can be committed to FRC differentiation in the bone marrow and lymph nodes involved by FL. Noteworthy, MSCs can promote the differentiation of Tfh cells into highly immunosuppressive T-follicular regulatory cells. The tumor suppressor HVEM is highly mutated in FL cells, and its deficiency increases Tfh cell frequency. In contrast, PI3Kδ inhibition impedes the recruitment of Tfh/regulatory T cells and impairs the proliferation of follicular dendritic cells (FDCs) and FDC-induced angiogenesis. Since TIGIT ligands are expressed by FDCs, the immune checkpoint receptor TIGIT plays an important role in tumor-infiltrating T cells. Thus, TIGIT blockade might invigorate cytotoxic T cells in the FL microenvironment. Given their potential to simultaneously reduce the neoplastic B cells, Tfh, and TFR cells could also reinforce the effects of the cytotoxic T cells. This combinatory strategy should be explored as a treatment option to tackle FL.
Prognostic value and immune infiltration of a novel stromal/immune score-related P2RY12 in lung adenocarcinoma microenvironment.
Yu Lian,Cao Shuhui,Li Jingwen,Han Baohui,Zhong Hua,Zhong Runbo
OBJECTIVE:Increasing evidence highlights the clinical implications of P2RY12 that belongs to the family of G-protein coupled receptors in carcinogenesis. Here, this study was designed to explore the associations between P2RY12 and tumor immune microenvironment of lung adenocarcinoma (LUAD). METHODS:Based on 352 LUAD samples from The Cancer Genome Atlas (TCGA), stromal and immune scores of each sample were estimated by ESTIMATE algorithm. Differential expression analysis was presented between stromal/immune high- and low-score groups. Protein-protein interaction (PPI) was then constructed by STRING database. Univariate and multivariate Cox regression analysis was utilized to screen prognosis-related factors. Co-expressed genes of P2RY12 were analyzed, followed by functional enrichment analysis. Furthermore, the correlation between P2RY12 and immune cell infiltrations was estimated using the TIMER database. P2RY12 expression was validated between 37 pairs of LUAD and normal tissues using RT-qPCR and immunohistochemistry. After overexpressing P2RY12, the proliferation and migration of A549 cells was detected by CCK-8 and scratch test. RESULTS:145 up- and 102 down-regulated stromal- and immune-related mRNAs were identified for LUAD. Based on 145 up-regulated mRNAs, a PPI network was conducted, consisting of 95 nodes and 210 relationship pairs. Ten hub genes were then identified. Among them, CCR8, CNR2, CXCR5, GPR18, GPR31 and P2RY12 were in association with overall survival of patients with LUAD. After adjusting other variables, P2RY12 expression was an independent prognostic factor for LUAD. 288 co-expressed genes of P2RY12 were determined and these co-expressed genes were primarily involved in immune-related biological processes or pathways. Moreover, P2RY12 was significantly correlated with M2 macrophage and dendritic cell infiltration. After validation, P2RY12 expression was significantly decreased in LUAD than normal tissues. Its overexpression distinctly suppressed proliferation and migration of A549 cells. CONCLUSION:Our findings suggest that P2RY12 is an immune infiltration-related prognostic marker for LUAD.
Abnormalities of Bone Marrow B Cells and Plasma Cells in Primary Immune Thrombocytopenia.
Yu Tianshu,Wang Haoyi,Zhao Yajing,Yu Yafei,Hou Yu,Liu Shuang,Han Panpan,Ni Xiaofei,Ji Xuebin,Peng Jun,Liu Xinguang,Hou Ming
Primary immune thrombocytopenia (ITP) is an autoantibody-mediated hemorrhagic disorder where B cells play an essential role. Previous studies have focused on peripheral blood (PB), but B cells in bone marrow (BM) have not been well characterized. We aimed to explore the profile of B cell subsets and their cytokine environments in BM of ITP patients to further clarify the pathogenesis of the disease. B cell subpopulations and their cytokine/chemokine receptors were detected by flow cytometry. Plasma concentrations of cytokines/chemokines were measured by ELISA. mRNA levels of B cell-related transcription factors were determined by qPCR. Regulatory B cell (Breg) function was assessed by quantifying their inhibitory effects on monocytes and T cells in vitro. Decreased proportions of total B cells, naïve B cells and defective Bregs were observed in ITP patients compared with healthy controls (HCs), whereas elevated frequency of long-lived plasma cells was found in BM of autoantibody-positive patients. No statistical difference was observed in plasmablasts or in short-lived plasma cells between ITP patients and HCs. The immunosuppressive capacity of BM Bregs from ITP patients was considerably weaker than that from HCs. In vivo study using an active ITP murine model revealed that Breg transfusion could significantly alleviate thrombocytopenia. Moreover, over-activation of CXCL13-CXCR5 and BAFF/APRIL systems were found in ITP patient BM. Taken together, B cell subsets in BM were skewed toward a proinflammatory profile in ITP patients, suggesting the involvement of dysregulated BM B cells in the development of the disease.
Human Lymph Nodes Maintain TCF-1 Memory T Cells with High Functional Potential and Clonal Diversity throughout Life.
Miron Michelle,Kumar Brahma V,Meng Wenzhao,Granot Tomer,Carpenter Dustin J,Senda Takashi,Chen Dora,Rosenfeld Aaron M,Zhang Bochao,Lerner Harvey,Friedman Amy L,Hershberg Uri,Shen Yufeng,Rahman Adeeb,Luning Prak Eline T,Farber Donna L
Journal of immunology (Baltimore, Md. : 1950)
Translating studies on T cell function and modulation from mouse models to humans requires extrapolating in vivo results on mouse T cell responses in lymphoid organs (spleen and lymph nodes [LN]) to human peripheral blood T cells. However, our understanding of T cell responses in human lymphoid sites and their relation to peripheral blood remains sparse. In this study, we used a unique human tissue resource to study human T cells in different anatomical compartments within individual donors and identify a subset of memory CD8 T cells in LN, which maintain a distinct differentiation and functional profile compared with memory CD8 T cells in blood, spleen, bone marrow, and lungs. Whole-transcriptome and high-dimensional cytometry by time-of-flight profiling reveals that LN memory CD8 T cells express signatures of quiescence and self-renewal compared with corresponding populations in blood, spleen, bone marrow, and lung. LN memory T cells exhibit a distinct transcriptional signature, including expression of stem cell-associated transcription factors TCF-1 and LEF-1, T follicular helper cell markers CXCR5 and CXCR4, and reduced expression of effector molecules. LN memory T cells display high homology to a subset of mouse CD8 T cells identified in chronic infection models that respond to checkpoint blockade immunotherapy. Functionally, human LN memory T cells exhibit increased proliferation to TCR-mediated stimulation and maintain higher TCR clonal diversity compared with memory T cells from blood and other sites. These findings establish human LN as reservoirs for memory T cells with high capacities for expansion and diverse recognition and important targets for immunotherapies.
A single-cell map of intratumoral changes during anti-PD1 treatment of patients with breast cancer.
Bassez Ayse,Vos Hanne,Van Dyck Laurien,Floris Giuseppe,Arijs Ingrid,Desmedt Christine,Boeckx Bram,Vanden Bempt Marlies,Nevelsteen Ines,Lambein Kathleen,Punie Kevin,Neven Patrick,Garg Abhishek D,Wildiers Hans,Qian Junbin,Smeets Ann,Lambrechts Diether
Immune-checkpoint blockade (ICB) combined with neoadjuvant chemotherapy improves pathological complete response in breast cancer. To understand why only a subset of tumors respond to ICB, patients with hormone receptor-positive or triple-negative breast cancer were treated with anti-PD1 before surgery. Paired pre- versus on-treatment biopsies from treatment-naive patients receiving anti-PD1 (n = 29) or patients receiving neoadjuvant chemotherapy before anti-PD1 (n = 11) were subjected to single-cell transcriptome, T cell receptor and proteome profiling. One-third of tumors contained PD1-expressing T cells, which clonally expanded upon anti-PD1 treatment, irrespective of tumor subtype. Expansion mainly involved CD8 T cells with pronounced expression of cytotoxic-activity (PRF1, GZMB), immune-cell homing (CXCL13) and exhaustion markers (HAVCR2, LAG3), and CD4 T cells characterized by expression of T-helper-1 (IFNG) and follicular-helper (BCL6, CXCR5) markers. In pre-treatment biopsies, the relative frequency of immunoregulatory dendritic cells (PD-L1), specific macrophage phenotypes (CCR2 or MMP9) and cancer cells exhibiting major histocompatibility complex class I/II expression correlated positively with T cell expansion. Conversely, undifferentiated pre-effector/memory T cells (TCF7, GZMK) or inhibitory macrophages (CX3CR1, C3) were inversely correlated with T cell expansion. Collectively, our data identify various immunophenotypes and associated gene sets that are positively or negatively correlated with T cell expansion following anti-PD1 treatment. We shed light on the heterogeneity in treatment response to anti-PD1 in breast cancer.
The splenic marginal zone shapes the phenotype of leukemia B cells and facilitates their niche-specific retention and survival.
Stache Vanessa,Verlaat Lydia,Gätjen Marcel,Heinig Kristina,Westermann Jörg,Rehm Armin,Höpken Uta E
Microenvironmental regulation in lymphoid tissues is essential for the development of chronic lymphocytic leukemia. We identified cellular and molecular factors provided by the splenic marginal zone (MZ), which alter the migratory and adhesive behavior of leukemic cells. We used the leukemia mouse model, in which tumor cells are excluded from B cell follicles and instead accumulate within the MZ. Genes involved in MZ B cell development and genes encoding for adhesion molecules were upregulated in MZ-localized cells. Likewise, surface expression of the adhesion and homing molecules, CD49d/VLA-4 and CXCR7, and of NOTCH2 was increased. , exposing cells or human CLL cells to niche-specific stimuli, like B cell receptor- or Toll-like receptor ligands, caused surface expression of these molecules characteristic for a follicular or MZ-like microenvironment, respectively. , inhibition of VLA-4-mediated adhesion and CXCL13-mediated follicular homing displaced leukemic cells not only from the follicle, but also from the MZ and reduced leukemia progression. We conclude that MZ-specific factors shape the phenotype of leukemic cells and facilitate their niche-specific retention. This strong microenvironmental influence gains pathogenic significance independent from tumor-specific genetic aberrations.
Colorectal cancer-infiltrating T lymphocytes display a distinct chemokine receptor expression profile.
Löfroos Ann-Britt,Kadivar Mohammad,Resic Lindehammer Sabina,Marsal Jan
European journal of medical research
BACKGROUND:T lymphocytes exert important homeostatic functions in the healthy intestinal mucosa, whereas in case of colorectal cancer (CRC), infiltration of T lymphocytes into the tumor is crucial for an effective anti-tumor immune response. In both situations, the recruitment mechanisms of T lymphocytes into the tissues are essential for the immunological functions deciding the outcome. The recruitment of T lymphocytes is largely dependent on their expression of various chemokine receptors. The aim of this study was to identify potential chemokine receptors involved in the recruitment of T lymphocytes to normal human colonic mucosa and to CRC tissue, respectively, by examining the expression of 16 different chemokine receptors on T lymphocytes isolated from these tissues. METHODS:Tissues were collected from patients undergoing bowel resection for CRC. Lymphocytes were isolated through enzymatic tissue degradation of CRC tissue and nearby located unaffected mucosa, respectively. The expression of a broad panel of chemokine receptors on the freshly isolated T lymphocytes was examined by flow cytometry. RESULTS:In the normal colonic mucosa, the frequencies of cells expressing CCR2, CCR4, CXCR3, and CXCR6 differed significantly between CD4 and CD8 T lymphocytes, suggesting that the molecular mechanisms mediating T lymphocyte recruitment to the gut differ between CD4 and CD8 T lymphocytes. In CRC, the frequencies of cells expressing CCR2 and CXCR5 were significantly lower in both the CD4 and CD8 T lymphocyte populations compared to unaffected colonic mucosa, and the frequency of CCR9 cytotoxic T lymphocytes was significantly decreased in CRC tissue. CONCLUSIONS:With regard to the normal gut mucosa, the results suggest that the molecular mechanisms mediating T lymphocyte recruitment differ between CD4 and CD8 T lymphocytes, which are important for understanding gut homeostasis. Importantly, T lymphocytes from CRC compared to normal colonic tissue displayed a distinct chemokine receptor expression profile, suggesting that mechanisms for recruitment of T lymphocytes to CRC tissue are skewed compared to normal colonic mucosa. Understanding these mechanisms could help in developing new strategies in cancer immunotherapy and to optimize already available alternatives such as immune checkpoint inhibitors.
CXC family of chemokines as prognostic or predictive biomarkers and possible drug targets in colorectal cancer.
Cabrero-de Las Heras Sara,Martínez-Balibrea Eva
World journal of gastroenterology
Colorectal cancer (CRC) is the third most common cancer in men and the second most common cancer in women, worldwide. In the early stages of the disease, biomarkers predicting early relapse would improve survival rates. In metastatic patients, the use of predictive biomarkers could potentially result in more personalized treatments and better outcomes. The CXC family of chemokines (CXCL1 to 17) are small (8 to 10 kDa) secreted proteins that attract neutrophils and lymphocytes. These chemokines signal through chemokine receptors (CXCR) 1 to 8. Several studies have reported that these chemokines and receptors have a role in either the promotion or inhibition of cancer, depending on their capacity to suppress or stimulate the action of the immune system, respectively. In general terms, activation of the CXCR1/CXCR2 pathway or the CXCR4/CXCR7 pathway is associated with tumor aggressiveness and poor prognosis; therefore, the specific inhibition of these receptors is a possible therapeutic strategy. On the other hand, the lesser known CXCR3 and CXCR5 axes are generally considered to be tumor suppressor signaling pathways, and their stimulation has been suggested as a way to fight cancer. These pathways have been studied in tumor tissues (using immunohistochemistry or measuring mRNA levels) or serum [using enzyme-linked immuno sorbent assay (ELISA) or multiplexing techniques], among other sample types. Common variants in genes encoding for the CXC chemokines have also been investigated as possible biomarkers of the disease. This review summarizes the most recent findings on the role of CXC chemokines and their receptors in CRC and discusses their possible value as prognostic or predictive biomarkers as well as the possibility of targeting them as a therapeutic strategy.
Characterization of Follicular Helper CD4 T Cells Using Multiplexed Imaging.
Ioannidou Kalliopi,Ndiaye Daba-Rokhya,Noto Alessandra,Fenwick Craig,Fortis Sotirios P,Pantaleo Giuseppe,Petrovas Constantinos,de Leval Laurence
Frontiers in immunology
Follicular helper CD4 T (Tfh) cells play an essential role in the formation of germinal centers (GCs), where mature B cells proliferate, differentiate, and provide long-term protective humoral responses. Despite the extensive phenotypic characterization and identification of human Tfh cell subsets, their spatial positioning at tissue level is not well understood. Here, we describe a quantitative multiplexed immunofluorescence approach allowing for the comprehensive characterization of Tfh cells in human tonsils and lymph nodes (LNs) from individuals with angioimmunoblastic T-cell lymphoma (AITL). We have developed eight multiplexed panels comprising a spectrum of Tfh cell markers, like PD-1, CXCR5, and ICOS, along with transcription factors (Bcl6, Tbet, GATA3), to assess their expression, frequencies, spatial distribution and co-localization in a quantitative manner. Combined analysis of relevant markers revealed the presence of several Tfh cell subsets at tissue level based on the differential expression of surface receptors, nuclear factors as well as their distinct localization within the follicular areas. Interestingly, we found a considerable amount of tonsillar Tfh cells expressing high levels of the Th2 regulator GATA3. The co-expression of GATA3, CXCR5, and BCL6, points to an important role of GATA3 for the generation of effector human Tfh cells. Furthermore, our data revealed significantly different Tfh cell profile signatures between health and disease. Therefore, our imaging platform generates meaningful information for the characterization of human Tfh cells and could provide the base for future studies aiming to a comprehensive understanding of Tfh cell tissue heterogeneity.
The impacts of zanubrutinib on immune cells in patients with chronic lymphocytic leukemia/small lymphocytic lymphoma.
Zou Yi-Xin,Zhu Hua-Yuan,Li Xiao-Tong,Xia Yi,Miao Kou-Rong,Zhao Si-Shu,Wu Yu-Jie,Wang Li,Xu Wei,Li Jian-Yong
Ibrutinib, a first-generation Bruton's tyrosine kinase (BTK) inhibitor, could improve immunity of relapsed or refractory (R/R) chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) patients. Whether zanubrutinib, a second-generation selective BTK inhibitor, has similar effects as ibrutinib remains to be determined. Dynamics of number and immunophenotype of immune cells during zanubrutinib treatment in 25 R/R CLL/SLL patients were examined by flow cytometry and blood routine tests. The expression intensity of programmed death-1 (PD-1) on total CD4 (P < .01), total CD8 (P < .01), and T helper cells (P < .05) and cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) on total CD4 (P = .010) and regulatory T cells (P < .05) reduced after treatment. There were significant differences in expression intensity of CD19 (P < .01), C-X-C chemokine receptor type 5 (CXCR5) (P < .01), and CD49d (P < .05) on B cells before and after treatment. Downregulation of PD-1 on T cells and CXCR5 and CD19 on B cells were observed in nearly all patients after zanubrutinib treatment. Programmed death-ligand 1 expression downregulated, especially in the female, CLL, normal spleen, normal β2-macroglobulin (β2-MG) and abnormal lactate dehydrogenase (LDH) subgroups, and CTLA-4 expression on CD4+ T cells tended to decrease in the male, old, CLL, splenomegaly, abnormal β2-MG, normal LDH, IGHV-mutated and wild-type tumor protein 53 subgroups after zanubrutinib treatment. These findings suggest that zanubrutinib can regulate immunity primarily by improving T cell exhaustion, inhibiting suppressor cells and disrupting CLL cells migration through downregulation of adhesion/homing receptors. Furthermore, favorable changes in cell number and immunophenotype were preferably observed in patients without adverse prognostic factors.
Characterization of the prognostic values of CXCR family in gastric cancer.
Yu Chaoran,Zhang Yujie
INTRODUCTION:The role of CXC chemokine receptors (CXCRs) in gastric cancer (GC) has been an increasing focus. However, comprehensive prognostic values of CXCR members in GC are yet to be clearly defined. METHODS:Multiple public available datasets, including Kaplan-Meier (KM) plotter, oncomine, the cancer genome atlas (TCGA), SurvExpress platform and the tumor immune estimation resource (TIMER), were used for mRNA expression and prognostic characterization. Nomogram method was used for clinical model prediction. RESULTS:CXCR3, CXCR4 and CXCR5 displayed significantly up-regulated expression in tumor compared to normal. High mRNA expression of CXCR2 (HR = 0.77, 95%CI: 0.62-0.95, p = 0.014), CXCR3 (HR = 0.74, 95%CI: 0.61-0.90, p = 0.0024), CXCR4 (HR = 0.7, 95%CI: 0.58-0.86, p = 0.00048), CXCR5 (HR = 0.72, 95%CI: 0.59-0.87, p = 0.00093) and CXCR6 (HR = 0.66, 95%CI: 0.54-0.81, p = 4.9e-05) was significantly associated with favorable overall survival (OS). The prognostic values of CXCR members were also explored in subtypes, including HER2 status, Lauren classification, pathological stages. The low risk group of CXCR signature displayed a significantly favorable OS compared to the high risk group (HR = 3.22, 95% CI = 2.21-4.69, p = 1.057e-09). Nomogram clinical models were established for both OS (C-index: 0.692; 95%CI: 0.648-0.736) and recurrence free survival (C-index: 0.731; 95%CI: 0.675-0.786). In addition, CXCR6 and CD8+T cells featured the highest correlation (partial-cor = 0.781, p = 4.17e-77). CONCLUSION:This study identified distinct expression and prognostic values of CXCR members in GC using public databases.
Immunohistochemical analysis indicates that the anatomical location of B-cell non-Hodgkin's lymphoma is determined by differentially expressed chemokine receptors, sphingosine-1-phosphate receptors and integrins.
Middle Stephen,Coupland Sarah E,Taktak Azzam,Kidgell Victoria,Slupsky Joseph R,Pettitt Andrew R,Till Kathleen J
Experimental hematology & oncology
BACKGROUND:The aim of this study was to elucidate the mechanisms responsible for the location of B-cell non-Hodgkin's lymphoma (B-NHL) at different anatomical sites. We speculated that the malignant B cells in these disorders have the potential for trafficking between blood and secondary lymphoid organs (SLO) or extranodal sites and that their preferential accumulation at different locations is governed by the expression of key molecules that regulate the trafficking of normal lymphocytes. METHODS:Biopsy or blood samples from 91 cases of B-NHL affecting SLO (n = 27), ocular adnexae (n = 51) or blood (n = 13) were analysed by immunohistochemistry or flow cytometry for the expression of the following molecules: CCR7, CCL21 and αL (required for the entry of normal lymphocytes into SLO); CXCR4, CXCL12 and α4 (required for entry into extranodal sites); CXCR5, CXCL13 and S1PR2 (required for tissue retention); S1PR1 and S1PR3 (required for egress into the blood). The expression of each of these molecules was then related to anatomical location and histological subtype. RESULTS:The expression of motility/adhesion molecules varied widely between individual patient samples and correlated much more strongly with anatomical location than with histological subtype. SLO lymphomas [comprising 10 follicular lymphoma (FL), 8 diffuse large B-cell lymphoma (DLBCL), 4 mantle-cell lymphoma (MCL) and 5 marginal-zone lymphoma (MZL)] were characterised by pronounced over-expression of S1PR2, suggesting that the malignant cells in these lymphomas are actively retained at the site of clonal expansion. In contrast, the malignant B cells in ocular adnexal lymphomas (10 FL, 9 DLBCL, 4 MCL and 28 MZL) expressed a profile of molecules suggesting a dynamic process of trafficking involving not only tissue retention but also egress via S1PR3 and homing back to extranodal sites via CXCR4/CXCL12 and α4. Finally, leukaemic lymphomas (6 FL, 5 MCL and 2 MZL) were characterised by aberrant expression of the egress receptor S1PR1 and low expression of molecules required for tissue entry/retention. CONCLUSIONS:In summary, our study strongly suggests that anatomical location in B-NHL is governed by the differential expression of specific adhesion/motility molecules. This novel observation has important implications for therapeutic strategies that aim to disrupt protective micro-environmental interactions.
Programmed death ligand-1 and CD8 tumor-infiltrating lymphocytes (TILs) as prognostic predictors in ovarian high-grade serous carcinoma (HGSC).
Farrag Mayada Saad,Abdelwahab Khaled,Farrag Nesrine Saad,Elrefaie Waleed Elsayed,Emarah Ziad
Journal of the Egyptian National Cancer Institute
BACKGROUND:P D-L1 is expressed in tumor cells and plays a crucial role in tumor immune escape. Tumor-infiltrating lymphocytes (TILs) as CD8 T cells contribute to reduced tumor growth. Few studies investigated the prognostic effect of PD-L1 and CD8 TILs in ovarian high-grade serous carcinoma (HGSC). In the present study, we analyzed the expression of PD-L1 and CD8 TILs in HGSC by immunohistochemistry, and results were correlated to prognosis. It was carried on 54 cases of ovarian HGSC who attended the Oncology Centre, Mansoura University, Egypt, from 2012 till 2019. RESULTS:Nearly 60% of cases showed positive PD-L1 expression in tumor cells. Regarding the clinicopathological characteristics, higher PD-L1 expression was found among patients with residual tumor (82.4%) compared to patients with no residual tumor (54.5%), with marginal statistical significance (p 0.07). PD-L1 was significantly associated with CD8 TILs expression. Higher PD-L1 expression was found among tumors with low expression of CD8 TILs with statistically significant difference (p≤0.001). Disease-free survival (DFS) was significantly lower among the group with positive expression of PD-L1 compared to the group with negative expression of PD-L1 (p 0.01), while overall survival (OS) was not associated with PD-L1 expression. On the other hand, the overall survival (OS) in patients with high CD8 expression was significantly higher than patients with low CD8 expression (p 0.043), while DFS was not significantly different among both CD8 TILS groups. CONCLUSIONS:PD-L1 and CD8 TILs may become a promising therapeutic target for patients with ovarian HGSC. More studies are needed to further validate their prognostic effect. Precise identification of patients who will benefit from PD-L1 checkpoint blockade and TILs adaptive immunotherapy is mandatory.
PD-1CXCR5CD4 Th-CXCL13 cell subset drives B cells into tertiary lymphoid structures of nasopharyngeal carcinoma.
Li Jiang-Ping,Wu Chang-You,Chen Ming-Yuan,Liu Shang-Xin,Yan Shu-Mei,Kang Yin-Feng,Sun Cong,Grandis Jennifer R,Zeng Mu-Sheng,Zhong Qian
Journal for immunotherapy of cancer
BACKGROUND:A major current challenge is to exploit tertiary lymphoid structures (TLSs) to promote the lymphocyte infiltration, activation and differentiation by tumor antigens to increase antitumor immune responses. The mechanisms that underlie the role of TLS formation in the adaptive immune responses against nasopharyngeal carcinoma (NPC) remain largely unknown. METHODS:Cell populations and the corresponding markers were identified by single-cell RNA sequencing and fluorescence-activated cell sorting analysis. In vitro differentiation experiments were used to simulate the generation, regulation and function of the Th-CXCL13 cell subset in the tumor microenvironment of NPC. These were followed by histological evaluation of the colocalization of tumor-associated B cells (TABs) and Th-CXCL13 cells within TLSs, and statistical analysis of the relationship between the cells in TLSs and overall survival. RESULTS:A PD-1CXCR5CD4 Th-CXCL13 cell subset was identified in NPC. This subset was a major source of CXCL13, representing the majority of the CD4 T cells at levels comparable with Th1 and Tfh cells present in the TLSs. Monocytes activated by toll-like receptor 4 agonists served as the antigen-presenting cells that most efficiently triggered the expansion of Th-CXCL13 cells. Transforming growth factor beta 1 (TGF-β1) stimulation and activation of were critical for the induction and polarization of Th-CXCL13 cells in this process. The potential functional contributions of TABs recruited by Th-CXCL13 cells which induced plasma cell differentiation and immunoglobulin production via interleukin-21 and CD84 interactions in the TLSs demonstrated improved survival. CONCLUSIONS:Induction of Th-CXCL13 cells links innate inflammation to immune privilege in tumor-associated TLSs and might predict better survival.
CDK4/6 inhibition promotes immune infiltration in ovarian cancer and synergizes with PD-1 blockade in a B cell-dependent manner.
Zhang Qian-Feng,Li Jia,Jiang Kuo,Wang Rui,Ge Jun-Li,Yang Hong,Liu Shu-Juan,Jia Lin-Tao,Wang Lei,Chen Bi-Liang
Great progress has been made in the field of tumor immunotherapy in the past decade. However, the therapeutic effects of immune checkpoint blockade (ICB) against ovarian cancer are still limited. Recently, an inhibitor of cyclin-dependent kinases 4 and 6 (CDK4/6i) has been reported to enhance antitumor immunity in preclinical models. The combined use of CDK4/6i and ICB may be beneficial, but the effects of CDK4/6is on the tumor immune microenvironment and whether they can synergize with ICB in treating ovarian cancer remain unknown. In this study, we first assessed the antitumor efficacy of abemaciclib, an FDA-approved CDK4/6i, in a syngeneic murine ovarian cancer model. Then, immunohistochemistry, immunofluorescence and flow cytometry were performed to evaluate the number, proportion, and activity of tumor-infiltrating lymphocytes. Cytokine and chemokine production was detected both and by PCR array analysis and cytokine antibody arrays. The treatment efficacy of combined abemaciclib and anti-PD-1 therapy was evaluated , and CD8+ and CD4+ T cell activities were analyzed using flow cytometry. Lastly, the requirement for both CD8+ T cells and B cells in combination treatment was evaluated and potential cellular mechanisms were further analyzed by flow cytometry. We observed that abemaciclib monotherapy could enhance immune infiltration, especially CD8+ T cell and B cell infiltration, in the ID8 murine ovarian cancer model. Immunophenotyping analysis showed that abemaciclib induced a proinflammatory immune response in the tumor microenvironment. PCR array analysis suggested the presence of a Th1-polarized cytokine profile in abemaciclib-treated ID8 tumors. studies showed that abemaciclib-treated ID8 cells secreted more CXCL10 and CXCL13, thus recruiting more lymphocytes than control groups. Combination treatment achieved better tumor control than monotherapy, and the activities of CD8+ and CD4+ T cells were further enhanced when compared with monotherapy. The synergistic antitumor effects of combined abemaciclib and anti-PD-1 therapy depended on both CD8+ T cells and B cells. These findings suggest that combined treatment with CDK4/6i and anti-PD-1 antibody could improve the efficacy of anti-PD-1 therapy and hold great promise for the treatment of poorly immune-infiltrated ovarian cancer.
CXCL13 shapes immunoactive tumor microenvironment and enhances the efficacy of PD-1 checkpoint blockade in high-grade serous ovarian cancer.
Yang Moran,Lu Jiaqi,Zhang Guodong,Wang Yiying,He Mengdi,Xu Qing,Xu Congjian,Liu Haiou
Journal for immunotherapy of cancer
BACKGROUND:Most patients with high-grade serous ovarian cancer (HGSC) lack an effective response to immune checkpoint blockade, highlighting the need for more knowledge about what is required for successful treatment. As follicular cytotoxic CXCR5CD8 T cells are maintained by reinvigoration by immune checkpoint blockade in tumors, we attempted to reveal the relationship between CXCR5CD8 T cells and the tumor microenvironment to predict immunotherapy responses in HGSC. METHODS:264 patients with HGSC from two cohorts and 340 HGSC cases from The Cancer Genome Atlas cohort were enrolled. Ex vivo and in vivo studies were conducted with human HGSC tumors and murine tumor models. The spatial correlation between CXC-chemokine ligand 13 (CXCL13), CXCR5, CD8, and CD20 was evaluated by immunohistochemistry and immunofluorescence. Survival was compared between different subsets of patients using Kaplan-Meier analysis. The therapeutic effect of CXCL13 and programmed cell death-1 (PD-1) blockade was validated using human HGSC tumors and murine models. RESULTS:High CXCL13 expression was associated with prolonged survival. Tumors with high CXCL13 expression exhibited increased infiltration of activated and CXCR5-expressing CD8 T cells. Incubation with CXCL13 facilitated expansion and activation of CXCR5CD8 T cells ex vivo. CXCR5CD8 T cells appeared in closer proximity to CXCL13 in tumors and chemotaxis towards CXCL13 in vitro. The combination of CXCL13, CXCR5, and CD8 T cells was an independent predictor for survival. In addition, CXCL13 was associated with clusters of CD20 B cells. CD20 B cells predicted better patient survival in the presence of CXCL13. Histological evaluation highlighted colocalization of CXCL13 with tertiary lymphoid structures (TLSs). TLSs carried prognostic benefit only in the presence of CXCL13. CXCL13 in combination with anti-PD-1 therapy retarded tumor growth in a CD8 T-cell-dependent manner, resulting in increased infiltration of cytotoxic CD8 T cells and CXCR5CD8 T cells. CONCLUSIONS:These data define a critical role of CXCL13 in shaping antitumor microenvironment by facilitating the maintenance of CXCR5CD8 T cells in TLSs and support a clinical investigation for a combination of CXCL13 and PD-1 blockade therapy in HGSC.
Erratum to: CXCL13-CXCR5 co-expression regulates epithelial to mesenchymal transition of breast cancer cells during lymph node metastasis.
Biswas Subir,Sengupta Suman,Roy Chowdhury Sougata,Jana Samir,Mandal Gunjan,Mandal Palash Kumar,Saha Nipun,Malhotra Vivek,Gupta Arnab,Kuprash Dmitry V,Bhattacharyya Arindam
Breast cancer research and treatment
CXC Chemokine Receptor Type 5 Gene Polymorphisms in a Cohort of Egyptian Patients with Diffuse Large B-Cell Lymphoma.
Makhlouf Manal Mohamed,Radwan Eman Roshdy,Khorshed Ola Mohamed,Fathi Lamees Mohamed,Elmasry Manal Mohamed
Pathobiology : journal of immunopathology, molecular and cellular biology
BACKGROUND:The chemokine receptor CXCR5 is selectively expressed on B cells; it is involved in lymphocyte homing and the development of normal lymphoid tissue. Its principle ligand is CXCL13 or B lymphocyte chemoattractant. Three polymorphisms in the CXCR5 gene, rs148351692 C/G, rs6421571 C/T, and rs78440425 G/A, have been identified. OBJECTIVE:To assess the genetic polymorphisms of CXCR5 and evaluate their possible contribution to the susceptibility and response to therapy of diffuse large B-cell lymphoma (DLBCL). PATIENTS AND METHODS:Fifty DLBCL (not otherwise specified) patients and 50 control subjects were included in this study. CXCR5 genotypes were determined by PCR-RFLP. RESULTS:Our study revealed that the CXCR5 rs148351692 C/G and rs6421571 C/T gene polymorphisms are associated with an increased risk of developing DLBCL (OR 28.57 [95% CI 8.96-96.56] and 3.45 [1.67-11.83] respectively), while CXCR5 rs78440425 G/A showed no association with the risk of lymphoma. Moreover, the double and triple combined gene polymorphisms are associated with an increased risk of developing DLBCL of approximately 120-fold and 105-fold, respectively. CXCR5 gene polymorphisms had no significant impact on disease outcome or response to therapy. CONCLUSIONS:CXCR5 gene polymorphisms could be considered a potential risk factor for the development of DLBCL.
CXCR5-mediated shaping of the lymphoid follicle in chronic lymphocytic leukemia.
López-Guerra Mònica,Xargay-Torrent Sílvia,Colomer Dolors
Heinig and colleagues, using the Eμ-Tcl1 mouse model of chronic lymphocytic leukemia (CLL), shed light on the trafficking routes of CLL cells into the protective microenvironmental niches in secondary lymphoid organs. The authors propose a crucial role of the resident follicular dendritic cells for leukemia pathogenesis that is essentially orchestrated by the chemokine receptor CXCR5.
High CXCR3 on Leukemic Cells Distinguishes from in Chronic Lymphocytic Leukemia: Evidence from CD5 and CD5 Clones.
Manukyan Gayane,Papajik Tomas,Mikulkova Zuzana,Urbanova Renata,Kraiczova Veronika Smotkova,Savara Jakub,Kudelka Milos,Turcsanyi Peter,Kriegova Eva
Journal of immunology research
Despite the shared pattern of surface antigens, neoplastic cells in chronic lymphocytic leukemia (CLL) are highly heterogeneous in CD5 expression, a marker linked to a proliferative pool of neoplastic cells. To further characterize CD5 and CD5 neoplastic cells, we assessed the chemokine receptors (CCR5, CCR7, CCR10, CXCR3, CXCR4, CXCR5) and adhesion molecules (CD54, CD62L, CD49d) on the CD5 and CD5 subpopulations, defined by CD5/CD19 coexpression, in peripheral blood of CLL patients ( = 60) subgrouped according to the mutational status ( , = 24; , = 36). CD5 subpopulation showed a high percentage of CXCR3 ( < 0.001), CCR10 ( = 0.001), and CD62L ( = 0.031) and high levels of CXCR5 ( = 0.005), CCR7 ( = 0.013) compared to CD5 cells expressing high CXCR4 ( < 0.001). Comparing and patients, high levels of CXCR3 on CD5 and CD5 subpopulations were detected in the patients, with better discrimination in CD5 subpopulation. Levels of CXCR3 on CD5 subpopulation were associated with time to the next treatment, thus further confirming its prognostic value. Taken together, our analysis revealed higher CXCR3 expression on both CD5 and CD5 neoplastic cells in with a better prognosis compared to patients. Contribution of CXCR3 to CLL pathophysiology and its suitability for prognostication and therapeutic exploitation deserves future investigations.
CXCL13 (BCA-1) is produced by follicular lymphoma cells: role in the accumulation of malignant B cells.
Husson Hervé,Freedman Arnold S,Cardoso Angelo A,Schultze Joachim,Munoz Olivier,Strola Giuliana,Kutok Jeffery,Carideo Elizabeth G,De Beaumont Rosalie,Caligaris-Cappio Federico,Ghia Paolo
British journal of haematology
Follicular lymphomas (FLs) localize in lymphoid tissues and recapitulate the structure of normal secondary follicles. The chemokine/chemokine receptor pair CXCL13/CXCR5 is required for the architectural organization of B cells within lymphoid follicles. In this study, we showed that CXCL13 was secreted by FL cells. FL cells expressed CXCR5 and migrated in response to CXCL13. Furthermore, we observed a synergistic effect between CXCL13 and CXCL12 (SDF-1), a chemokine produced by stromal cells in lymphoid tissues. The production of CXCL13 by FL cells and CXCL12 by stromal cells probably directs and participates in the accumulation of FL cells within specific anatomic sites.
CXCR3 from chemokine receptor family correlates with immune infiltration and predicts poor survival in osteosarcoma.
Tang Yin,Gu Zhiqian,Fu Youwei,Wang Junjie
BACKGROUND:Chemokine receptors have a crucial role in regulating tumor mediating immunity and are also implicated in the prognosis of some cancers. Here, the association between CXC chemokine receptors (CXCR2-5) and prognosis in osteosarcoma was studied. METHODS:Differences between CXCR2, CXCR3, CXCR4, and CXCR5 expression and overall survival (OS) and event-free survival (EFS) were compared using Kaplan-Meier analyses. The associations of CXCR3 expression with clinical features and the prognosis were also analyzed. The signaling pathways modulated by CXCR3 were investigated. The correlations between CXCR3 and immune infiltrates were investigated. RESULTS:The expression of CXCR2, CXCR4, and CXCR5 was not associated with the prognosis, but CXCR3 low expression was correlated with worse OS and EFS of osteosarcoma, especially for female, patients aged less than 15.1 years, or patients without metastasis. Low CXCR3 expression was related to tumor site and histologic response (P<0.05), but not associated with other clinical characteristics. Multivariate Cox analysis revealed that CXCR3 remained independently associated with the prognosis, especially for OS (hazard ratio (HR) = 3.26, 95% CI = 1.15-9.24, P=0.026). The cell adhesion, apoptosis, metabolism, KRAS, P53, NOTCH, reactive oxygen species (ROS), PI3K/Akt/mTOR, vascular endothelial growth factor (VEGF), inflammation, and immune-related pathways such as IL-6/JAK/STAT3, TNF-α via NF-κB, Toll/NOD-like receptor, and complement were modulated by CXCR3. CXCR3 expression showed an especially positive correlation with immune infiltration of T cells CD8, macrophages M1, plasma cells, and NK cells activated. CONCLUSIONS:CXCR3 may be an independent risk factor for the prognosis and is most likely to benefit from immunotherapy in osteosarcoma.
Protein kinase D-dependent CXCR4 down-regulation upon BCR triggering is linked to lymphadenopathy in chronic lymphocytic leukaemia.
Saint-Georges Stéphane,Quettier Maude,Bouyaba Marouane,Le Coquil Stéphanie,Laurienté Vanessa,Guittat Lionel,Lévy Vincent,Ajchenbaum-Cymbalista Florence,Varin-Blank Nadine,Le Roy Christine,Ledoux Dominique
In Chronic Lymphocytic Leukemia (CLL), infiltration of lymph nodes by leukemic cells is observed in patients with progressive disease and adverse outcome. We have previously demonstrated that B-cell receptor (BCR) engagement resulted in CXCR4 down-regulation in CLL cells, correlating with a shorter progression-free survival in patients. In this study, we show a simultaneous down-regulation of CXCR4, CXCR5 and CD62L upon BCR triggering. While concomitant CXCR4 and CXCR5 down-regulation involves PKDs, CD62L release relies on PKC activation. BCR engagement induces PI3K-δ-dependent phosphorylation of PKD2 and 3, which in turn phosphorylate CXCR4 Ser324/325. Moreover, upon BCR triggering, PKD phosphorylation levels correlate with the extent of membrane CXCR4 decrease. Inhibition of PKD activity restores membrane expression of CXCR4 and migration towards CXCL12 in BCR-responsive cells in vitro. In terms of pathophysiology, BCR-dependent CXCR4 down-regulation is observed in leukemic cells from patients with enlarged lymph nodes, irrespective of their IGHV mutational status. Taken together, our results demonstrate that PKD-mediated CXCR4 internalization induced by BCR engagement in B-CLL is associated with lymph node enlargement and suggest PKD as a potential druggable target for CLL therapeutics.
Regulators of Tfh Cell Differentiation.
Jogdand Gajendra M,Mohanty Suchitra,Devadas Satish
Frontiers in immunology
The follicular helper T (Tfh) cells help is critical for activation of B cells, antibody class switching, and germinal center (GC) formation. The Tfh cells are characterized by the expression of CXC chemokine receptor 5 (CXCR5), ICOS, programed death 1 (PD-1), B cell lymphoma 6 (BCL-6), and IL-21. They are involved in clearing infections and are adversely linked with autoimmune diseases and also have a role in viral replication as well as clearance. On the one hand, Tfh cells are generated from naive CD4 T cells with sequential steps involving cytokine signaling (IL-21, IL-6, IL-12, activin A), migration, and positioning in the GC by CXCR5, surface receptors (ICOS/ICOSL, signaling lymphocyte activation molecule-associated protein/signaling lymphocyte activation molecule) as well as transcription factor (BCL-6, c-Maf, and signal transducer and activator of transcription 3) signaling and repressor miR155. On the other hand, Tfh generation is negatively regulated at specific steps of Tfh generation by specific cytokine (IL-2, IL-7), surface receptor (PD-1, CTLA-4), transcription factors B lymphocyte maturation protein 1, signal transducer and activator of transcription 5, T-bet, KLF-2 signaling, and repressor miR 146a. Interestingly, miR-17-92 and FOXO1 act as a positive as well as a negative regulator of Tfh differentiation depending on the time of expression and disease specificity. Tfh cells are also generated from the conversion of other effector T cells as exemplified by Th1 cells converting into Tfh during viral infection. The mechanistic details of effector T cells conversion into Tfh are yet to be clear. To manipulate Tfh cells for therapeutic implication and or for effective vaccination strategies, it is important to know positive and negative regulators of Tfh generation. Hence, in this review, we have highlighted and interlinked molecular signaling from cytokines, surface receptors, transcription factors, ubiquitin ligase, and microRNA as positive and negative regulators for Tfh differentiation.
Chemokines and their receptors promoting the recruitment of myeloid-derived suppressor cells into the tumor.
Li Bao-Hua,Garstka Malgorzata A,Li Zong-Fang
Myeloid-derived suppressor cells (MDSCs) expand in tumor-bearing host. They suppress anti-tumor immune response and promote tumor growth. Chemokines play a vital role in recruiting MDSCs into tumor tissue. They can also induce the generation of MDSCs in the bone marrow, maintain their suppressive activity, and promote their proliferation and differentiation. Here, we review CCL2/CCL12-CCR2, CCL3/4/5-CCR5, CCL15-CCR1, CX3CL1/CCL26-CX3CR1, CXCL5/2/1-CXCR2, CXCL8-CXCR1/2, CCL21-CCR7, CXCL13-CXCR5 signaling pathways, their role in MDSCs recruitment to tumor tissue, and their correlation with tumor development, metastasis and prognosis. Targeting chemokines and their receptors may serve as a promising strategy in immunotherapy, especially combined with other strategies such as chemotherapy, cyclin-dependent kinase or immune checkpoints inhibitors.
The Role of CXC Chemokine Receptors 1-4 on Immune Cells in the Tumor Microenvironment.
Susek Katharina Helene,Karvouni Maria,Alici Evren,Lundqvist Andreas
Frontiers in immunology
Chemokines govern leukocyte migration by attracting cells that express their cognate ligands. Many cancer types show altered chemokine secretion profiles, favoring the recruitment of pro-tumorigenic immune cells and preventing the accumulation of anti-tumorigenic effector cells. This can ultimately result in cancer immune evasion. The manipulation of chemokine and chemokine-receptor signaling can reshape the immunological phenotypes within the tumor microenvironment in order to increase the therapeutic efficacy of cancer immunotherapy. Here we discuss the three chemokine-chemokine receptor axes, CXCR1/2-CXCL1-3/5-8, CXCR3-CXCL9/10/11, and CXCR4-CXCL12 and their role on pro-tumorigenic immune cells and anti-tumorigenic effector cells in solid tumors. In particular, we summarize current strategies to target these axes and discuss their potential use in treatment approaches.
Exploration of prognostic biomarkers and therapeutic targets in the microenvironment of bladder cancer based on CXC chemokines.
Sun Xiaoqi,Chen Qunxi,Zhang Lihong,Chen Jiewei,Zhang Xinke
Mathematical biosciences and engineering : MBE
BACKGROUND:Bladder cancer (BLCA) has a high rate of morbidity and mortality, and is considered as one of the most malignant tumors of the urinary system. Tumor cells interact with surrounding interstitial cells, playing a key role in carcinogenesis and progression, which is partly mediated by chemokines. CXC chemokines exert anti-tumor biological roles in the tumor microenvironment and affect patient prognosis. Nevertheless, their expression and prognostic values patients with BLCA remain unclear. METHODS:We used online tools, including Oncomine, UALCAN, GEPIA, GEO databases, cBioPortal, GeneMANIA, DAVID 6.8, Metascape, TRUST (version 2.0), LinkedOmics, TCGA, and TIMER2.0 to perform the relevant analysis. RESULTS:The mRNA levels of C-X-C motif chemokine ligand (CXCL)1, CXCL5, CXCL6, CXCL7, CXCL9, CXCL10, CXCL11, CXCL13, CXCL16, and CXCL17 were increased significantly increased, and those of CXCL2, CXCL3, and CXCL12 were decreased significantly in BLCA tissues as assessed using the Oncomine, TCGA, and GEO databases. GEO showed that high levels of CXCL1, CXCL6, CXCL10, CXCL11, and CXCL13 mRNA expression are associated significantly with the poor overall survival (all p < 0.05), and similarly, those of CXCL2 and CXCL12 in the TCGA database (p < 0.05). The predominant signaling pathways involving the differentially expressed CXC chemokines are cell cycle, chemokine, and cytokine-cytokine receptor interaction. Moreover, transcription factors such as Sp1 transcription factor (SP1), nuclear factor kappa B subunit 1 (NFKB1), and RELA proto-oncogene, NF-KB subunit (RELA) were likely play critical roles in regulating CXC chemokine expression. LYN proto-oncogene, src family tyrosine kinase (LYN) and LCK proto-oncogene, src family tyrosine kinase (LCK) were identified as the key targets of these CXC chemokines. MicroRNAs miR200 and miR30 were identified as the main microRNAs that interact with several CXC chemokines through an miRNA-target network. The expression of these chemokines is closely associated with the infiltration of six categories of immune cells. CONCLUSION:We explored the CXC chemokines superfamily-based biomarkers associated with BLCA prognosis using public databases, and provided possible chemokine targets for patients with BLCA.
The role of CXC chemokines and their receptors in cancer.
Vandercappellen Jo,Van Damme Jo,Struyf Sofie
Chemokines, or chemotactic cytokines, and their receptors have been discovered as essential and selective mediators in leukocyte migration to inflammatory sites and to secondary lymphoid organs. Besides their functions in the immune system, they also play a critical role in tumor initiation, promotion and progression. There are four subgroups of chemokines: CXC, CC, CX(3)C, and C chemokine ligands. The CXC or alpha subgroup is further subdivided in the ELR(+) and ELR(-) chemokines. Members that contain the ELR motif bind to CXC chemokine receptor 2 (CXCR2) and are angiogenic. In contrast, most of the CXC chemokines without ELR motif bind to CXCR3 and are angiostatic. An exception is the angiogenic ELR(-)CXC chemokine stromal cell-derived factor-1 (CXCL12/SDF-1), which binds to CXCR4 and CXCR7 and is implicated in tumor metastasis. This review is focusing on the role of CXC chemokines and their receptors in tumorigenesis, including angiogenesis, attraction of leukocytes to tumor sites and induction of tumor cell migration and homing in metastatic sites. Finally, their therapeutic use in cancer treatment is discussed.
The Effect of Hypoxia on the Expression of CXC Chemokines and CXC Chemokine Receptors-A Review of Literature.
Korbecki Jan,Kojder Klaudyna,Kapczuk Patrycja,Kupnicka Patrycja,Gawrońska-Szklarz Barbara,Gutowska Izabela,Chlubek Dariusz,Baranowska-Bosiacka Irena
International journal of molecular sciences
Hypoxia is an integral component of the tumor microenvironment. Either as chronic or cycling hypoxia, it exerts a similar effect on cancer processes by activating hypoxia-inducible factor-1 (HIF-1) and nuclear factor (NF-κB), with cycling hypoxia showing a stronger proinflammatory influence. One of the systems affected by hypoxia is the CXC chemokine system. This paper reviews all available information on hypoxia-induced changes in the expression of all CXC chemokines (CXCL1, CXCL2, CXCL3, CXCL4, CXCL5, CXCL6, CXCL7, CXCL8 (IL-8), CXCL9, CXCL10, CXCL11, CXCL12 (SDF-1), CXCL13, CXCL14, CXCL15, CXCL16, CXCL17) as well as CXC chemokine receptors-CXCR1, CXCR2, CXCR3, CXCR4, CXCR5, CXCR6, CXCR7 and CXCR8. First, we present basic information on the effect of these chemoattractant cytokines on cancer processes. We then discuss the effect of hypoxia-induced changes on CXC chemokine expression on the angiogenesis, lymphangiogenesis and recruitment of various cells to the tumor niche, including myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), tumor-associated neutrophils (TANs), regulatory T cells (T) and tumor-infiltrating lymphocytes (TILs). Finally, the review summarizes data on the use of drugs targeting the CXC chemokine system in cancer therapies.
G-protein coupled receptors and ligands that organize humoral immune responses.
Lu Erick,Cyster Jason G
B-cell responses are dynamic processes that depend on multiple types of interactions. Rare antigen-specific B cells must encounter antigen and specialized systems are needed-unique to each lymphoid tissue type-to ensure this happens efficiently. Lymphoid tissue barrier cells act to ensure that pathogens, while being permitted entry for B-cell recognition, are blocked from replication or dissemination. T follicular helper (Tfh) cells often need to be primed by dendritic cells before supporting B-cell responses. For most responses, antigen-specific helper T cells and B cells need to interact, first to initiate clonal expansion and the plasmablast response, and later to support the germinal center (GC) response. Newly formed plasma cells need to travel to supportive niches. GC B cells must become confined to the follicle center, organize into dark and light zones, and interact with Tfh cells. Memory B cells need to be positioned for rapid responses following reinfection. Each of these events requires the actions of multiple G-protein coupled receptors (GPCRs) and their ligands, including chemokines and lipid mediators. This review will focus on the guidance cue code underlying B-cell immunity, with an emphasis on findings from our laboratory and on newer advances in related areas. We will discuss our recent identification of geranylgeranyl-glutathione as a ligand for P2RY8. Our goal is to provide the reader with a focused knowledge about the GPCRs guiding B-cell responses and how they might be therapeutic targets, while also providing examples of how multiple types of GPCRs can cooperate or act iteratively to control cell behavior.
The characteristics and novel clinical implications of CD4+CXCR5+Foxp3+ follicular regulatory T cells in breast cancer.
Miao Xianyuan,Guo Qiusheng,Pan Zhiwen,Xu Xiaohong,Shao Xiying,Wang Xiaojia
Annals of translational medicine
Background:Follicular regulatory T cells (Tfr) are a subset of regulatory T cells (Tregs) that suppress the humoral immune response in the germinal center. They are associated with increased rates of disease stabilization and decreased autoantibody levels in a variety of tumor and autoimmune diseases. The binding of T-cell immunoglobulin mucin 3 (TIM-3) and its ligand on the surface of Tfr cells could result in the depletion of T lymphocytes and the termination of the immune response mediated by helper T cell 1. However, the role of Tfr cells in breast cancer (BC) remains unclear. Methods:In this study, we detected the expression of CD4+CXCR5+Foxp3+Tfr cells in the peripheral blood of 35 BC patients and 30 healthy control patients by flow cytometry, and analyzed the relationship between Tfr cells and the clinical characteristics of patients. In addition, the expression of TIM-3 on the surface of Tfr cells in 6 triple-negative BC (TNBC) patients was further investigated using mass spectrometry. Results:We found a significant increase in Tfr cells in BC patients compared to healthy control patients (23.47%±9.70% . 10.99%±4.68%; P=0.001). Notably, the increase was more significant in early stage than advanced stage TNBC patients (28.52%±10.75% . 18.69%±5.19%; P=0.006), and there was a negative correlation between Tfr cells and serum lactate dehydrogenase (LDH) in early stage TNBC patients (r=-0.585; P=0.008). Additionally, we found that the expression of Tfr cells was higher in TNBC patients than luminal BC patients (28.25%±10.11% . 18.5%±8.15%; P=0.028); however, there was no significant difference in expression in hormone receptor positive (HR+) BC and hormone receptor negative (HR-) BC (P=0.141) patients. Notably, the surface of Tfr cells of TNBC patients had higher levels of TIM-3 expression than those of healthy control patients (3.93±0.92 . 2.65±0.15, respectively; t=-3.02; P<0.05), which the mass spectrometry showed were positively correlated with the intracellular Foxp3 expression of Tfr cells (r=0.82; P=0.036). Conclusions:Our results suggest that circulating Tfr cells and the expression of TIM-3 were significantly increased in BC patients, which were related to stage and histological type, and may be involved in the pathogenesis of BC.
Determination of Potential Therapeutic Targets and Prognostic Markers of Ovarian Cancer by Bioinformatics Analysis.
Zhang Jing,Huang Shouguo,Quan Lini,Meng Qiu,Wang Haiyan,Wang Jie,Chen Jin
BioMed research international
This study is to study the expression of CXCRs in ovarian cancer tissues and their value in prognosis. The expressions of CXCR1-CXCR7 mRNA between ovarian tumor tissues and normal tissues and in different pathological types of ovarian tumor tissues were compared by ONCOMINE online tool. The relationship between the expression of CXCRs and clinical pathological staging was studied by GEPIA. Kaplan-Meier plotter online tool was used to analyze prognosis. Finally, GO and KEGG analyses and protein interaction network analysis were performed for CXCRs by the DAVID software to predict their function, and cBioPortal was used to identify the key functional genes. The expression of CXCR3/4/7 mRNA in ovarian cancer tissues was higher than that in normal ovarian tissues, and the expression of CXCR4 was the highest (fold change = 306.413, < 0.05). The expression of CXCR1/2/3/4/7 mRNA in different pathological types of ovarian tumors was significantly different ( < 0.05). Only CXCR5 expression level was associated with tumor staging. Survival analysis showed that high CXCR7 mRNA expression and low CXCR5/6 expression were associated with the shortening of overall survival. High CXCR4/7 expression and low CXCR5/6 expression were associated with the shortening of progression-free survival. High CXCR2/4 expression and low CXCR5/6 expression were closely related to the shortening of postprogressing survival. Protein interaction network analysis showed that GNB1, PTK2, MAPK1, PIK3CA, GNB4, GNA11, KNG1, and ARNT proteins were closely related to the CXC receptor family. CXCR3/4/7 are potential therapeutic targets, and CXCR2/4/5/6/7 are new markers for the prognosis of ovarian cancer.
Enhanced Phenotype Definition for Precision Isolation of Precursor Exhausted Tumor-Infiltrating CD8 T Cells.
Martinez-Usatorre Amaia,Carmona Santiago J,Godfroid Céline,Yacoub Maroun Céline,Labiano Sara,Romero Pedro
Frontiers in immunology
In the context of adoptive T cell transfer (ACT) for cancer treatment, it is crucial to generate large amounts of tumor-specific CD8 T cells with high potential to persist . PD-1, Tim3, and CD39 have been proposed as markers of tumor-specific tumor-infiltrating CD8 T lymphocytes (CD8 TILs). However, these molecules are highly expressed by terminally differentiated exhausted CD8 T cells (Tex) that lack proliferation potential. Therefore, optimized strategies to isolate tumor-specific TILs with high proliferative potential, such as Tcf1+ precursor exhausted T cells (Tpe) are needed to improve persistence of ACT. Here we aimed at defining cell surface markers that would unequivocally identify Types for precision cell sorting increasing the purity of tumor-specific PD-1+ Tcf1+ Tpe from total TILs. Transcriptomic analysis of Tpe vs. Tex CD8 TIL subsets from B16 tumors and primary human melanoma tumors revealed that Tpes are enriched in and lack and expression, which encode Slamf6, CD39, and Tim3 cell surface proteins, respectively. Indeed, we observed by flow cytometry that CD39- Tim3- Slamf6+ PD-1+ cells yielded maximum enrichment for tumor specific PD-1+ Tcf1+ OT1 TILs in B16.OVA tumors. Moreover, this population showed higher re-expansion capacity upon an acute infection recall response compared to the CD39+ counterparts or bulk PD-1+ TILs. Hence, we report an enhanced sorting strategy (CD39- Tim3- Slamf6+ PD-1+) of Tpes. In conclusion, we show that optimization of CD8 TIL cell sorting strategy is a viable approach to improve recall capacity and persistence of transferred cells in the context of ACT.
Signaling via the CXCR5/ERK pathway is mediated by CXCL13 in mice with breast cancer.
Xu Licheng,Liang Zhi,Li Shuyan,Ma Jianjun
Breast cancer is the most common cause of cancer-associated mortality and the most frequently diagnosed type of cancer in women worldwide. It has been revealed that the chemokine C-X-C motif chemokine ligand 13 (CXCL13) serves a pivotal role in breast cancer growth and is associated with lymph node metastasis. However, to the best of our knowledge, the mechanism by which CXCL13 mediates breast cancer growth remains uncharacterized. Female BALB/c mice were used in this study. Tumor volume was calculated and changes of gross tumor morphology were observed by hematoxylin and eosin staining. The expression of CXCL13, C-X-C motif chemokine receptor 5 (CXCR5) and extracellular signaling-related kinase (ERK) mRNA and protein expression were detected by reverse transcriptase quantitative-polymerase chain reaction and western blot analysis. Simultaneously, the production of cytokines [interleukin-1β (IL-1β), tumor necrosis factor (TNF) and tumor growth factor β1 (TGF-β1)] was detected by an ELISA. The CXCL13 inhibitor reduced tumor volume and growth, and reduced the mRNA and protein expression levels of key members of the CXCR5/ERK signaling pathway: CXCL13, CXCR5 and ERK. Furthermore, the detectable concentration of the cytokines IL-1β and TNF decreased following CXCL13 inhibition, whereas the concentration of TGF-β1 was increased. The attenuation of tumor growth resulting from CXCL13 inhibition may be associated with the CXCR5/ERK signaling pathway. This study provides a theoretical basis for treating breast cancer through CXCL13 inhibition in clinical trials.
Astragalus-mediated stimulation on antigen-presenting cells could result in higher IL-21 production from CXCR5 Tfh-like cells and better IL-21-mediated effector functions.
Dong Qin,Pu Jin,Du Tingting,Xu Shenqian,Liu Wuxia,Liu Ling,Wang Zhenlong,Cai Chen
T cells in renal cell carcinoma (RCC) patients display multiple features of impairment and exhaustion. Here, we hypothesize that Astragalus membranaceus, a herbal medicine commonly used to accompany chemotherapy, might have adjuvating effects on T cells from RCC patients. To investigate this, circulating T cells from healthy individuals and RCC patients were cocultured ex vivo with aqueous extract from Astragalus. Functional characteristics of T cells in the absence and presence of Astragalus extract were then compared. We first identified a downregulation of IL-21 expression in RCC patients in association with a functional dysregulation of CXCR5 Tfh-like cells. Astragalus extract could significantly increase IL-21 expression in a dose-dependent manner. This Astragalus-mediated effect depended on the presence of antigen-presenting cells (APCs), as purified CXCR5 Tfh-like cells presented little IL-21 upregulation following Astragalus stimulation. APCs primed by Astragalus extract also promoted IL-21 expression from Tfh-like cells. Interestingly, Astragalus-stimulated Tfh-like cells presented enhanced helper function and resulted in higher humoral responses and better CD8 T cell survival. This effect was dependent on the presence of IL-21. Overall, these data indicated that Astragalus could enhance IL-21 production and effector function from CXCR5 Tfh-like cells in a manner that depended on the presence of APCs.
T follicular helper cell subsets: a potential key player in autoimmunity.
Kurata Izumi,Matsumoto Isao,Sumida Takayuki
Follicular helper T (Tfh) cells are one of CD4+ helper T subsets which promote B cell maturation, activation and antigen-specific antibody production. Autoantibodies are hallmarks of autoimmune diseases, and crucial contributions of Tfh cells in development of these diseases are now evident. Deregulation of Tfh activities can contribute to a pathogenic autoantibody production and can play an important role in the promotion of autoimmune diseases. These days multiple researchers reported three subpopulations which has distinct effector functions in Tfh cells: Tfh1, Tfh2 and Tfh17 cells. In this review, we summarize the observed alterations in whole Tfh cells and subset distribution during autoimmune diseases.
γδTFH cells promote B cell maturation and antibody production in neuroblastoma.
Mou Wenjun,Han Wei,Ma Xiaoli,Wang Xiaolin,Qin Hong,Zhao Wen,Ren Xiaoya,Chen Xi,Yang Wei,Cheng Haiyan,Wang Xisi,Zhang Hui,Ni Xin,Wang Huanmin,Gui Jingang
BACKGROUND:Previous studies have shown that γδ TFH cells are capable of modulating antibody production in immunized and infected mouse model. In recent studies, human γδ TFH cells are shown to contribute to the activation of humoral immunity and promote the maturation of B cells. However, little information is available on their involvement in neuroblastoma (NB) pathogenesis. RESULTS:In the present study, the frequency of γδ TFH cells in 74 NB patients was significantly higher compared with that in 60 healthy controls. Moreover, most γδ TFH cells in NB patients had a naive phenotype with up-regulation of CD25, CD69, HLA-DR and CD40L and down-regulation of ICOS. Importantly, γδ TFH cells in NB patients produced more IL-4 and IL-10 than those in healthy controls. Furthermore, serum total IgG level was significantly increased in NB patients compared with healthy controls. The expression of CD23 on B cells was up-regulated while CD80 expression was significantly down-regulated in NB patients. Further analysis of B cell compartment showed that the frequency of CD19CD27 plasma cells was enhanced in NB patients. Spearman's correlation analysis revealed that the frequency of γδ TFH cells was positively correlated to serum total IgG level and CD19CD27 plasma cells in NB patients, but negatively correlated to CD19 B cells. CONCLUSIONS:We concluded that γδ TFH cells might promote B cell maturation and antibody production in NB patients.
Follicular cytotoxic CD8 T cells present high cytokine expression, and are more susceptible to Breg-mediated suppression in non-small cell lung cancer.
Ma Qin-Yun,Chen Jun,Zhao Jun
Tumor-infiltrating CD8 T cells are instrumental to antitumor immunity. In this study, we found that a subset of CXCR5-expressing CD8 T cells, termed follicular cytotoxic T (Tfc) cells, potently infiltrated the untreated tumors from non-small cell lung cancer (NSCLC) patients. On average, Tfc cells represented 14% of total tumor-infiltrating CD8 T cells and 6.6% of total tumor-infiltrating lymphocytes. Upon antigenic stimulation, Tfc cells presented significantly higher degranulation and stronger release of proinflammatory cytokines, including IFNg, IL2, and TNF, and the pleiotropic cytokine IL10 than non-Tfc cells. However, the expression of granzyme B and perforin was significantly lower in Tfc cells than in non-Tfc CD8 T cells. B regulatory (Breg) cells could significantly suppress proinflammatory cytokine production in both Tfc cells and non-Tfc CD8 T cells, but in Tfc cells, a lower concentration was required. Moreover, Breg cells could significantly elevate IL10 expression by Tfc cells but could not affect IL-10 expression by non-Tfc CD8 T cells. The neutralization of IL10 significantly reduced the extent of Breg-mediated regulation. Together, this study demonstrated that Tfc cells represented a significant proportion of tumor-infiltrating CD8 T cells in lung carcinoma. These Tfc cells were different from non-Tfc CD8 T cells in terms of cytokine expression and granzyme and perforin release and were more susceptible to Breg-mediated suppression in an IL-10-dependent manner.
The transcription factor Runx3 guards cytotoxic CD8 effector T cells against deviation towards follicular helper T cell lineage.
Shan Qiang,Zeng Zhouhao,Xing Shaojun,Li Fengyin,Hartwig Stacey M,Gullicksrud Jodi A,Kurup Samarchith P,Van Braeckel-Budimir Natalija,Su Yao,Martin Matthew D,Varga Steven M,Taniuchi Ichiro,Harty John T,Peng Weiqun,Badovinac Vladimir P,Xue Hai-Hui
Activated CD8 T cells differentiate into cytotoxic effector (T) cells that eliminate target cells. How T cell identity is established and maintained is not fully understood. We found that Runx3 deficiency limited clonal expansion and impaired upregulation of cytotoxic molecules in T cells. Runx3-deficient CD8 T cells aberrantly upregulated genes characteristic of follicular helper T (T) cell lineage, including Bcl6, Tcf7 and Cxcr5. Mechanistically, the Runx3-CBFβ transcription factor complex deployed H3K27me3 to Bcl6 and Tcf7 genes to suppress the T program. Ablating Tcf7 in Runx3-deficient CD8 T cells prevented the upregulation of T genes and ameliorated their defective induction of cytotoxic genes. As such, Runx3-mediated Tcf7 repression coordinately enforced acquisition of cytotoxic functions and protected the cytotoxic lineage integrity by preventing T-lineage deviation.
Characteristics of B lymphocyte infiltration in HPV head and neck squamous cell carcinoma.
Zhang Siwei,Wang Bozhi,Ma Fen,Tong Fangjia,Yan Bingqing,Liu Tianyang,Xie Huanhuan,Song Lianhao,Yu Siyang,Wei Lanlan
Human papillomavirus (HPV) is an important etiological factor of head and neck squamous cell carcinoma (HNSCC). HPV HNSCC patients usually have a better prognosis, which probably results from the higher infiltration of B lymphocytes. This study was purposed to detect the infiltration of B lymphocyte subsets and the correlation between B lymphocyte subsets and the prognosis in HPV-related HNSCC. In this study, 124 HPV and 513 HPV HNSCC samples were obtained from the Gene Expression Omnibus (GEO) database and The Cancer Genome Atlas (TCGA) database for transcriptomic analysis. Infiltration of B lymphocytes subsets was detected with 7 HPV HNSCC and 13 HPV HNSCC tissues through immunohistochemistry and immunofluorescence. One HPV HNSCC sample was detected with single-cell sequencing for chemokine analysis. In the results, the infiltration of plasma cells (CD19 CD38 ) and memory B cells (MS4A1 CD27 ) was higher in HPV HNSCC samples. High infiltration of plasma cells and memory B cells was related to a better prognosis. High density of B lymphocytes was positively correlated with high CXCL13 production mainly from CD4 T lymphocytes in HNSCC. These results indicated that a high density of plasma cells and memory B cells could predict excellent prognosis. CD4 T lymphocytes might affect B lymphocytes and their subsets through the CXCL13/CXCR5 axis in HNSCC.
PD-L1hi B cells are critical regulators of humoral immunity.
Khan Adnan R,Hams Emily,Floudas Achilleas,Sparwasser Tim,Weaver Casey T,Fallon Padraic G
Specific B-cell subsets can regulate T-cell immune responses, and are termed regulatory B cells (Breg). The majority of Breg cells described in mouse and man have been identified by IL-10 production and are known to suppress allergy and autoimmunity. However, Breg cell mediated immune suppression, independent of IL-10, also occurs. Here we show that Breg cells play a critical role in regulating humoral immunity mediated by CD4(+)CXCR5(+)PD-1(+) follicular helper T cells, and can suppress inflammation in autoimmune disease through elevated expression of PD-L1. We have also identified that these B cells are resistant to αCD20 B-cell depletion. This work describes how Breg cells are critical in humoral homoeostasis and may have implications for the regulation of autoimmune diseases.
Increased levels of CCR7(lo)PD-1(hi) CXCR5 CD4 T cells, and associated factors Bcl-6, CXCR5, IL-21 and IL-6 contribute to repeated implantation failure.
Gong Qiaoqiao,Zhu Yuejie,Pang Nannan,Ai Haiquan,Gong Xiaoyun,La Xiaolin,Ding Jianbing
Experimental and therapeutic medicine
fertilization-embryo transfer (IVF-ET) can be used by infertile couples to assist with reproduction; however, failure of the embryo to implant into the endometrial lining results in failure of the IVF treatment. The present study investigated the expression of chemokine receptor 7 (CCR7)(lo) programmed death-1(PD-1)(hi) chemokine receptor type 5 (CXCR5) cluster of differentiation 4 (CD4) T cells and associated factors in patients with repeated implantation failure (RIF). A total of 30 females with RIF and 30 healthy females were enrolled in the current study. Flow cytometry was used to detect the proportion of CCR7(lo)PD-1(hi) CXCR5 CD4 T cells in the peripheral blood. Cytokine bead arrays were performed to detect the levels of interleukin (IL)-6, -4 and -2 in the serum. ELISAs were used to detect the level of IL-21 in the serum. Quantitative real time polymerase chain reaction analysis and immunohistochemistry were used to investigate the expression of B-cell lymphoma 6 (Bcl-6), chemokine receptor type 5 (CXCR5) and IL-21 in the endometrium. The results revealed that the percentage of CCR7(lo)PD-1(hi) CXCR5 CD4 T cells was increased in the RIF group compared with the control group during the mid luteal phase. The mRNA and protein levels of Bcl-6, IL-21 and CXCR5 in the endometrium and the concentrations of IL-21 and IL-6 in the serum were significantly increased in the RIF group; however, no significant difference was observed between the two groups in regards to the expression of IL-4 and IL-2. Furthermore, a significant positive correlation was identified between the percentage of CCR7(lo)PD-1(hi) CXCR5 CD4 T cells and IL-21 and IL-6 levels. The expression of IL-21 also had a positive correlation with Bcl-6 and CXCR5 expression in the RIF group. These results suggest that increased levels of CCR7(lo)PD-1(hi) CXCR5 CD4 T cells and associated factors contribute to RIF and could therefore be a potential therapeutic target.
Interleukin-30/IL27p28 Shapes Prostate Cancer Stem-like Cell Behavior and Is Critical for Tumor Onset and Metastasization.
Sorrentino Carlo,Ciummo Stefania L,Cipollone Giuseppe,Caputo Sara,Bellone Matteo,Di Carlo Emma
Prostate cancer stem-like cells (PCSLC) are believed to be responsible for prostate cancer onset and metastasis. Autocrine and microenvironmental signals dictate PCSLC behavior and patient outcome. In prostate cancer patients, IL30/IL27p28 has been linked with tumor progression, but the mechanisms underlying this link remain mostly elusive. Here, we asked whether IL30 may favor prostate cancer progression by conditioning PCSLCs and assessed the value of blocking IL30 to suppress tumor growth. IL30 was produced by PCSLCs in human and murine prostatic intraepithelial neoplasia and displayed significant autocrine and paracrine effects. PCSLC-derived IL30 supported PCSLC viability, self-renewal and tumorigenicity, expression of inflammatory mediators and growth factors, tumor immune evasion, and regulated chemokine and chemokine receptor genes, primarily via STAT1/STAT3 signaling. IL30 overproduction by PCSLCs promoted tumor onset and development associated with increased proliferation, vascularization, and myeloid cell recruitment. Furthermore, it promoted PCSLC dissemination to lymph nodes and bone marrow by upregulating the CXCR5/CXCL13 axis, and drove metastasis to lungs through the CXCR4/CXCL12 axis. These mechanisms were drastically hindered by IL30 knockdown or knockout in PCSLCs. Collectively, these results mark IL30 as a key driver of PCSLC behavior. Targeting IL30 signaling may be a potential therapeutic strategy against prostate cancer progression and recurrence. IL30 plays an important role in regulating prostate cancer stem-like cell behavior and metastatic potential, therefore targeting this cytokine could hamper prostate cancer progression or recurrence. .
PD-1 regulates CXCR5 CD4 T cell-mediated proinflammatory functions in non-small cell lung cancer patients.
Shi Weiwei,Yang Bo,Sun Qiong,Meng Jing,Zhao Xiao,Du Sicheng,Li Xiaoyan,Jiao Shunchang
PD-1 inhibitors have been used to revive exhausted T cell responses in non-small cell lung cancer (NSCLC) and other malignancies. CXCR5 T follicular helper (Tfh) cells are characterized by constitutive high PD-1 expression and have been associated with the formation of tertiary lymphoid structures and implicated in antitumor immunity. In this study, we investigated the effect of PD-1 and PD-1 inhibition on CXCR5 CD4 T cells. Data showed that CXCR5 CD4 T cells in both healthy subjects and NSCLC patients presented markedly higher PD-1 expression than CXCR5 CD4 T cells. Both CXCR5 and CXCR5 CD4 T cells from NSCLC patients presented higher PD-1 expression than their counterparts in healthy subjects. PD-1 CXCR5 CD4 T cells were functional, could express IL-21, IL-10, and CXCL13 upon stimulation, demonstrated auxiliary effects toward CD8 T cell-mediated IFN-γ production and proliferation, and promoted IgM and IgG production. However, the potency of PD-1 CXCR5 CD4 T cells was lower than the potency of PD-1 CXCR5 CD4 T cells. PD-1 blocking could significantly enhance the effector functions of PD-1 CXCR5 CD4 T cells. Overall, this study demonstrated that PD-1 CXCR5 CD4 T cells could promote CD8 T cell and B cell inflammation and could be modulated by PD-1 inhibition.
Tumor-Infiltrating Lymphocytes and Their Prognostic Value in Cutaneous Melanoma.
Maibach Fabienne,Sadozai Hassan,Seyed Jafari S Morteza,Hunger Robert E,Schenk Mirjam
Frontiers in immunology
Recent breakthroughs in tumor immunotherapy such as immune checkpoint blockade (ICB) antibodies, have demonstrated the capacity of the immune system to fight cancer in a number of malignancies such as melanoma and lung cancer. The numbers, localization and phenotypes of tumor-infiltrating lymphocytes (TIL) are not only predictive of response to immunotherapy but also key modulators of disease progression. In this review, we focus on TIL profiling in cutaneous melanoma using histopathological approaches and highlight the observed prognostic value of the primary TIL subsets. The quantification of TIL in formalin-fixed tumor samples ranges from visual scoring of lymphocytic infiltrates in H&E to multiplex immunohistochemistry and immunofluorescence followed by enumeration using image analysis software. Nevertheless, TIL enumeration in the current literature primarily relies upon single marker immunohistochemistry analyses of major lymphocyte subsets such as conventional T cells (CD3, CD4, CD8), regulatory T cells (FOXP3) and B cells (CD20). We review key studies in the literature on associations between TIL subsets and patient survival. We also cover recent findings with respect to the existence of ectopic lymphoid aggregates found in the TME which are termed tertiary lymphoid structures (TLS) and are generally a positive prognostic feature. In addition to their prognostic significance, the existence of various TIL sub-populations has also been reported to predict a patient's response to ICB. Thus, the literature on the predictive potential of TIL subsets in melanoma patients receiving ICB has also been discussed. Finally, we describe recently developed state-of-the-art profiling approaches for tumor infiltrating immune cells such as digital pathology scoring algorithms (e.g., Immunoscore) and multiplex proteomics-based immunophenotyping platforms (e.g., imaging mass cytometry). Translating these novel technologies have the potential to revolutionize tumor immunopathology leading to altering our current understanding of cancer immunology and dramatically improving outcomes for patients.
B cells are associated with survival and immunotherapy response in sarcoma.
Petitprez Florent,de Reyniès Aurélien,Keung Emily Z,Chen Tom Wei-Wu,Sun Cheng-Ming,Calderaro Julien,Jeng Yung-Ming,Hsiao Li-Ping,Lacroix Laetitia,Bougoüin Antoine,Moreira Marco,Lacroix Guillaume,Natario Ivo,Adam Julien,Lucchesi Carlo,Laizet Yec Han,Toulmonde Maud,Burgess Melissa A,Bolejack Vanessa,Reinke Denise,Wani Khalid M,Wang Wei-Lien,Lazar Alexander J,Roland Christina L,Wargo Jennifer A,Italiano Antoine,Sautès-Fridman Catherine,Tawbi Hussein A,Fridman Wolf H
Soft-tissue sarcomas represent a heterogeneous group of cancer, with more than 50 histological subtypes. The clinical presentation of patients with different subtypes is often atypical, and responses to therapies such as immune checkpoint blockade vary widely. To explain this clinical variability, here we study gene expression profiles in 608 tumours across subtypes of soft-tissue sarcoma. We establish an immune-based classification on the basis of the composition of the tumour microenvironment and identify five distinct phenotypes: immune-low (A and B), immune-high (D and E), and highly vascularized (C) groups. In situ analysis of an independent validation cohort shows that class E was characterized by the presence of tertiary lymphoid structures that contain T cells and follicular dendritic cells and are particularly rich in B cells. B cells are the strongest prognostic factor even in the context of high or low CD8 T cells and cytotoxic contents. The class-E group demonstrated improved survival and a high response rate to PD1 blockade with pembrolizumab in a phase 2 clinical trial. Together, this work confirms the immune subtypes in patients with soft-tissue sarcoma, and unravels the potential of B-cell-rich tertiary lymphoid structures to guide clinical decision-making and treatments, which could have broader applications in other diseases.
Tertiary lymphoid structures improve immunotherapy and survival in melanoma.
Cabrita Rita,Lauss Martin,Sanna Adriana,Donia Marco,Skaarup Larsen Mathilde,Mitra Shamik,Johansson Iva,Phung Bengt,Harbst Katja,Vallon-Christersson Johan,van Schoiack Alison,Lövgren Kristina,Warren Sarah,Jirström Karin,Olsson Håkan,Pietras Kristian,Ingvar Christian,Isaksson Karolin,Schadendorf Dirk,Schmidt Henrik,Bastholt Lars,Carneiro Ana,Wargo Jennifer A,Svane Inge Marie,Jönsson Göran
Checkpoint blockade therapies that reactivate tumour-associated T cells can induce durable tumour control and result in the long-term survival of patients with advanced cancers. Current predictive biomarkers for therapy response include high levels of intratumour immunological activity, a high tumour mutational burden and specific characteristics of the gut microbiota. Although the role of T cells in antitumour responses has thoroughly been studied, other immune cells remain insufficiently explored. Here we use clinical samples of metastatic melanomas to investigate the role of B cells in antitumour responses, and find that the co-occurrence of tumour-associated CD8 T cells and CD20 B cells is associated with improved survival, independently of other clinical variables. Immunofluorescence staining of CXCR5 and CXCL13 in combination with CD20 reveals the formation of tertiary lymphoid structures in these CD8CD20 tumours. We derived a gene signature associated with tertiary lymphoid structures, which predicted clinical outcomes in cohorts of patients treated with immune checkpoint blockade. Furthermore, B-cell-rich tumours were accompanied by increased levels of TCF7 naive and/or memory T cells. This was corroborated by digital spatial-profiling data, in which T cells in tumours without tertiary lymphoid structures had a dysfunctional molecular phenotype. Our results indicate that tertiary lymphoid structures have a key role in the immune microenvironment in melanoma, by conferring distinct T cell phenotypes. Therapeutic strategies to induce the formation of tertiary lymphoid structures should be explored to improve responses to cancer immunotherapy.
C-X-C chemokine receptor type 5 gene polymorphisms are associated with non-Hodgkin lymphoma.
Song Haihan,Tong Danian,Cha Zhanshan,Bai Jianwen
Molecular biology reports
The C-X-C chemokine receptor type 5 (CXCR5) is one of the principal regulators for targeting T cells, B cells and dendritic cells into secondary lymphoid organs. Polymorphism studies of CXCR5 gene remain extremely scarce. The aim of this study was to examine the effect of polymorphisms in the CXCR5 gene on the development of non-Hodgkin lymphoma (NHL) in the Chinese population. Four polymorphisms in CXCR5 gene, rs148351692C/G, rs6421571C/T, rs80202369G/A and rs78440425G/A, were tested by polymerase chain reaction-restriction fragment length polymorphism in 404 NHL cases and 456 age-matched healthy controls. Data were analyzed using the χ(2) test. Results showed that individuals with the rs6421571 CT, rs6421571 TT and rs80202369 AA genotype had significantly increased susceptibility to NHL [Odd ratio (OR) = 1.41, 95 % confidence interval (CI): 1.04-1.92, p = 0.028; OR = 2.30, 95 % CI: 1.44-3.65, p < 0.001; and OR = 3.24, 95 % CI: 1.26-8.32, p = 0.010, respectively]. When analyzing the haplotypes of these polymorphisms, the prevalence of the TGG (rs6421571, rs80202369, and rs78440425) haplotype was significantly higher in NHL cases than in controls (OR = 1.59, 95 % CI: 1.25-2.03, p < 0.001). In addition, numbers of rs6421571 TT genotype and T allele were significantly increased in NHL patients with high Ann Arbor stages (p < 0.03) or NHL with B cell subtype (p < 0.02). These data indicate that CXCR5 gene polymorphisms may be new risk factors for NHL. The finding that the adjacent SNPs, rs6421571C/T and rs80202369G/A, are both associated with NHL suggests that the 87 bp region carrying these 2 polymorphisms may have important functional significance.
CD8 cytotoxic T lymphocytes in cancer immunotherapy: A review.
Farhood Bagher,Najafi Masoud,Mortezaee Keywan
Journal of cellular physiology
CD8 cytotoxic T lymphocytes (CTLs) are preferred immune cells for targeting cancer. During cancer progression, CTLs encounter dysfunction and exhaustion due to immunerelated tolerance and immunosuppression within the tumor microenvironment (TME), with all favor adaptive immune-resistance. Cancer-associated fibroblasts (CAFs), macrophage type 2 (M2) cells, and regulatory T cells (Tregs) could make immunologic barriers against CD8 T cell-mediated antitumor immune responses. Thus, CD8 T cells are needed to be primed and activated toward effector CTLs in a process called tumor immunity cycle for making durable and efficient antitumor immune responses. The CD8 T cell priming is directed essentially as a corroboration work between cells of innate immunity including dendritic cells (DCs) and natural killer (NK) cells with CD4 T cells in adoptive immunity. Upon activation, effector CTLs infiltrate to the core or invading site of the tumor (so-called infiltrated-inflamed [I-I] TME) and take essential roles for killing cancer cells. Exogenous reactivation and/or priming of CD8 T cells can be possible using rational immunotherapy strategies. The increase of the ratio for costimulatory to coinhibitory mediators using immune checkpoint blockade (ICB) approach. Programmed death-1 receptor (PD-1)-ligand (PD-L1) and CTL-associated antigen 4 (CTLA-4) are checkpoint receptors that can be targeted for relieving exhaustion of CD8 T cells and renewing their priming, respectively, and thereby eliminating antigen-expressing cancer cells. Due to a diverse relation between CTLs with Tregs, the Treg activity could be dampened for increasing the number and rescuing the functional potential of CTLs to induce immunosensitivity of cancer cells.
Application of PD-1 Blockade in Cancer Immunotherapy.
Wu Xiaomo,Gu Zhongkai,Chen Yang,Chen Borui,Chen Wei,Weng Liqiang,Liu Xiaolong
Computational and structural biotechnology journal
The programmed cell death protein 1 (PD-1) pathway has received considerable attention due to its role in eliciting the immune checkpoint response of T cells, resulting in tumor cells capable of evading immune surveillance and being highly refractory to conventional chemotherapy. Application of anti-PD-1/PD-L1 antibodies as checkpoint inhibitors is rapidly becoming a promising therapeutic approach in treating tumors, and some of them have successfully been commercialized in the past few years. However, not all patients show complete responses and adverse events have been noted, suggesting a better understanding of PD-1 pathway mediated immunosuppression is needed to predict patient response and improve treatment efficacy. Here, we review the progresses on the studies of the mechanistic role of PD-1 pathway in the tumor immune evasion, recent clinical development and commercialization of PD-1 pathway inhibitors, the toxicities associated with PD-1 blockade observed in clinical trials as well as how to improve therapeutic efficacy and safety of cancer immunotherapy.
Poor clinical outcomes of intratumoral dendritic cell-specific intercellular adhesion molecule 3-grabbing non-integrin-positive macrophages associated with immune evasion in gastric cancer.
Liu Xin,Cao Yifan,Li Ruochen,Gu Yong,Chen Yifan,Qi Yangyang,Lv Kunpeng,Wang Jieti,Yu Kuan,Lin Chao,Liu Hao,Zhang Heng,He Hongyong,Chen Lingli,Zhang Peipei,Shen Zhenbin,Qin Jing,Sun Yihong,Li He,Huang Hua,Zhang Weijuan,Xu Jiejie
European journal of cancer (Oxford, England : 1990)
AIM:Tumour-associated macrophages (TAMs) are prominent immune cells infiltrating in solid tumours with phenotypic and functional heterogeneity. However, the clinical significance of heterogeneous subtypes of TAMs in gastric cancer still remains obscure. Here, we aimed to explore the clinical significance of TAMs expressing dendritic cell-specific intercellular adhesion molecule 3-grabbing non-integrin (DC-SIGN) and its relevance with immune contexture in gastric cancer. METHODS:We selected 453 formalin-fixed and paraffin-embedded samples and 51 fresh tissue specimens of patients with gastric cancer from Zhongshan Hospital. The association of DC-SIGN macrophages with clinicopathological parameters, overall survival (OS) and responsiveness to fluorouracil-based adjuvant chemotherapy (ACT) was inspected. Immunohistochemistry (IHC) and flow cytometry (FCM) were applied to characterize immune cells in gastric cancer. RESULTS:We demonstrated that high intratumoral DC-SIGN macrophages infiltration predicted poor OS and inferior therapeutic responsiveness to fluorouracil-based ACT in patients with gastric cancer. Furthermore, higher infiltration of DC-SIGN macrophages indicated an increased number of Foxp3 regulatory T cells (Tregs), CD8 T cells and a higher ratio of Foxp3/CD8 within the tumour microenvironment (TME). In addition, CD8 T cells in DC-SIGN macrophages high subgroup were functionally impaired, showing decreased interferon-γ (IFN-γ), granzyme B (GZMB) and perforin production yet elevated programmed cell death protein 1 (PD-1) and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) expression. CONCLUSIONS:DC-SIGN macrophages were associated with immunoinvasive TME and indicated poor prognosis and inferior therapeutic responsiveness to fluorouracil-based ACT. DC-SIGN macrophages might be an independent prognosticator and a potential immunotherapeutic target for gastric cancer.
Intratumoral TIGIT CD8 T-cell infiltration determines poor prognosis and immune evasion in patients with muscle-invasive bladder cancer.
Liu Zhaopei,Zhou Quan,Wang Zewei,Zhang Hongyu,Zeng Han,Huang Qiuren,Chen Yifan,Jiang Wenbin,Lin Zhiyuan,Qu Yang,Xiong Ying,Bai Qi,Xia Yu,Wang Yiwei,Liu Li,Zhu Yu,Xu Le,Dai Bo,Guo Jianming,Wang Jiajun,Chang Yuan,Zhang Weijuan
Journal for immunotherapy of cancer
BACKGROUND:T-cell immunoglobulin and ITIM domain (TIGIT) is identified as a novel checkpoint receptor that can facilitate immune escape via mediating T-cell exhaustion in tumors. However, the clinical significance and immune contexture correlation of intratumoral TIGIT CD8 T-cells remain to be further explored in muscle-invasive bladder cancer (MIBC). METHODS:259 patients with MIBC from two clinical centers (Zhongshan Hospital, n=141; Shanghai Cancer Center, n=118) were analyzed to evaluate the prognostic value and immune contexture association of TIGIT CD8 T-cells through immunohistochemistry. Fresh tumor tissue samples from 26 patients with MIBC were examined to discover the phenotype of this CD8 subpopulation by flow cytometry. RESULTS:High infiltration of intratumoral TIGIT CD8 T-cells predicted poor overall survival (OS) and recurrence-free survival (RFS) in MIBC. For patients with stage II MIBC with low infiltration of TIGIT CD8 cells, adjuvant chemotherapy (ACT) could significantly prolong their OS and RFS. Intratumoral TIGIT CD8 T-cell abundance was correlated with impaired CD8 T-cell cytotoxicity and exhibited production of immunosuppressive cytokine IL-10. Further analysis of tumor-infiltrating immune cell landscape revealed TIGIT CD8 T-cells were associated with suppressive immune contexture, including Th2 cells, regulatory T-cells, mast cells and neutrophils. CONCLUSION:Intratumoral TIGIT CD8 T-cell abundance could serve as an independent prognosticator for clinical outcome and a predictive biomarker for inferior ACT responsiveness. Intratumoral TIGIT CD8 T-cell abundance correlated with dampened CD8 T-cell antitumor immunity and immunosuppressive contexture abundance, highlighting a tumor-promoting role of TIGIT CD8 T-cells.
Poor clinical outcomes and immunoevasive contexture in CXCL13+CD8+ T cells enriched gastric cancer patients.
Jin Kaifeng,Cao Yifan,Gu Yun,Fang Hanji,Fei Yuchao,Wang Jieti,Liu Xin,Lv Kunpeng,He Xudong,Lin Chao,Liu Hao,Li He,He Hongyong,Li Ruochen,Zhang Heng,Xu Jiejie
As an adverse survival prognosticator, chemokine (C-X-C motif) ligand 13 (CXCL13) has been studied in several types of malignancies. The secretion and physiological roles of CXCL13 in follicular helper T cells (T) cells have been well described, while the clinical significance of CD8 tumor-infiltrating lymphocytes (TILs)-associated CXCL13 remains unknown. This study aims to investigate the clinical significance of CXCL13CD8 T cells in survival and chemotherapeutic responsiveness prediction in gastric cancer. In this study, 440 patients enrolled from Zhongshan Hospital with tumor microarray (TMA) specimens were randomly divided into testing set (n = 220) and validation set (n = 220) for analysis. CXCL13CD8 T cells were detected by multicolor immunohistochemistry. Fresh tumor tissue samples from another 60 gastric cancer patients were collected to detect CXCL13CD8 T cells functional status by flow cytometry (FCM). We found that high intratumoral CXCL13CD8 T cells infiltration predicted poor overall survival and inferior chemotherapeutic responsiveness in gastric cancer. CXCL13CD8T cells were associated with immunoevasive contexture with increased regulatory T (T) cells and dysfunctional cytotoxic T lymphocytes (CTLs). Moreover, the combinational analysis of CXCL13CD8 T cells and CD8 T cells infiltration stratified patients into distinct risk groups with different clinical outcomes and chemotherapeutic responsiveness. Conclusively, intratumoral CXCL13CD8 T cells infiltration could be an independent prognostic and predictive marker for gastric cancer patients. CXCL13CD8 T cells represented an exhausted CD8 T cell subset, and might be a potential immunotherapeutic target in gastric cancer.
Intratumoral CXCL13CD8T cell infiltration determines poor clinical outcomes and immunoevasive contexture in patients with clear cell renal cell carcinoma.
Dai Siyuan,Zeng Han,Liu Zhaopei,Jin Kaifeng,Jiang Wenbin,Wang Zewei,Lin Zhiyuan,Xiong Ying,Wang Jiajun,Chang Yuan,Bai Qi,Xia Yu,Liu Li,Zhu Yu,Xu Le,Qu Yang,Guo Jianming,Xu Jiejie
Journal for immunotherapy of cancer
BACKGROUND:Chemokine (C-X-C motif) ligand 13 (CXCL13) was known as a selective chemotaxis for B cells, a product of follicular helper CD4T cells (T) and a contributor to tertiary lymphoid structures (TLS). Although secretion and function of CXCL13 produced by T have been deeply explored, the immune function and prognostic significance of CXCL13 secreted by CD8T cells still remain unrevealed. This study aims to investigate the clinical merit of CXCL13CD8T cells in clear cell renal cell carcinoma (ccRCC). METHODS:We analyzed prognostic value and immune contexture that associated with CXCL13CD8T cells infiltration level in a total of 755 patients from Zhongshan Hospital cohort (n=223) and The Cancer Genome Atlas cohort (n=532). In vitro analyses were conducted on 42 samples of resected tumor tissue from Zhongshan Hospital in order to detect the immune status of CXCL13CD8T cells and total CD8T cells. Immunohistochemistry (IHC) and flow cytometry were applied to characterize immune cells and portray the tumor microenvironment (TME) in ccRCC. RESULTS:Intratumoral CXCL13CD8T cells abundance was associated with inferior overall survival and disease-free survival. CXCL13CD8T cells possessed higher level of immune checkpoints like programmed cell-death protein 1 (PD-1), T-cell immunoglobulin mucin 3 (Tim-3), T cell immunoreceptor with Ig and ITIM domains (TIGIT) and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), higher Ki-67 expression and lower tumor necrosis factor α (TNF-α), interferon γ (IFN-γ) expression. Total CD8T cells in high-level CXCL13CD8T cells infiltration subgroup exhibited elevated exhausted markers (PD-1, Tim-3, TIGIT) and descended activated markers (TNF-α, IFN-γ) without quantity variance. Furthermore, the abundance of intratumoral CXCL13CD8T cell was correlated with immunoevasive TME accompanied by increased T helper 2 cells, tumor-associated macrophages, Foxp3 regulatory T cells, TLS and decreased natural killer cells, GZMB cells. CONCLUSIONS:Intratumoral CXCL13CD8T cells infiltration indicated inferior clinical outcome in patients with ccRCC. CXCL13CD8T cells possessed increased exhausted markers, decreased effector molecules and better proliferation ability. CXCL13CD8T cells abundance impaired total CD8T cells' immune function. Intratumoral CXCL13CD8T cells abundance was associated with immunoevasive contexture. The abundance of CXCL13CD8T cells was an independent prognosticator and a potential immunotherapeutic target marker for ccRCC treatment.
Effect of CXCR5-Positive Cell Infiltration on the Immune Contexture and Patient Prognosis in Head and Neck Squamous Cell Carcinoma.
Chen Jun,Meng Xiangchao,Zhou Qinyi,Feng Jialin,Zheng Wenjie,Wang Zhuoying,Wang Jiadong,Wang You
OncoTargets and therapy
Purpose:CXCR5-positive (CXCR5) tumor cell infiltration has different prognostic values in different types of cancer. The objective was to evaluate the effect of CXCR5 cell infiltration in head and neck squamous cell carcinoma (HNSCC). Patients and Methods:The study included two patient cohorts: The Cancer Genome Atlas cohort (TCGA, n = 472) and the Renji Hospital cohort (RJHC, n = 201). The TCGA and RJHC cohorts were analyzed for CXCR5-related mRNAs and CXCR5+ cell infiltration, respectively. We then evaluated the correlation between CXCR5 mRNA and CXCR5 cell infiltration in terms of overall survival and the immune contexture. Results:The 5-year overall survival rate was significantly correlated with high CXCR5 mRNA expression and CXCR5 cell infiltration in the TCGA and RJHC cohorts, respectively (p < 0.01), even after adjusting for confounders. Moreover, high CXCR5 mRNA expression was associated with more CD4 T cells, CD8 T cells, plasma cells, and less dendritic cells. A high CXCR5 mRNA expression was also correlated with increased expression of cytotoxic IFNG, TNFSF11 (RANKL), GZMA, GZMB, GZMK, GZMM, and PRF1 and increased expression of the immunosuppressive gene PDCD1 (PD-1), CD274 (PD-L1), CTLA4, LAG3, HAVCR2 (TIM-3), BTLA, and TIGIT. Conclusion:HNSCC patients with a high intratumoral CXCR5 expression had a better prognosis than those with low intratumoral CXCR5 expression. Moreover, CXCR5 cell infiltration could be used as an independent prognostic biomarker or as a potential therapeutic target. The presence of CXCR5 cells affects the infiltration of immunocytes in head and neck cancer, differently from what was reported in other cancer types. Further randomized controlled trials or studies with more patients are needed to validate our results.
ETV4 promotes pancreatic ductal adenocarcinoma metastasis through activation of the CXCL13/CXCR5 signaling axis.
Gao Xiaoliang,Jiang Mingzuo,Chu Yi,Han Yuying,Jin Yirong,Zhang Wenyao,Wang Weijie,Yang Suzhen,Li Wenjiao,Fan Ahui,Cao Jiayi,Wang Jiayao,Liu Hao,Fu Xin,Chen Di,Nie Yongzhan,Fan Daiming
Pancreatic ductal adenocarcinoma (PDAC) has the highest fatality rate of any solid tumor, with a five-year survival rate of only 10% in the USA. PDAC is characterized by early metastasis. More than 50% of patients present with distant metastases at the time of diagnosis, and the majority of patients will develop metastasis within 4 years after tumor resection. Despite extensive studies, the molecular mechanisms underlying PDAC metastasis remain unclear. The polyoma enhancer activator protein (PEA3) subfamily was reported to play a vital role in the initiation and progression of multiple tumors. Herein, we found that ETS variant 4 (ETV4) was highly expressed in PDAC tissues and associated with poor survival. Univariate and multivariate analyses revealed that ETV4 expression was an independent prognostic factor for patient survival. Further experiments showed that ETV4 overexpression promoted PDAC invasion and metastasis both in vitro and in vivo. For the first time, we demonstrated that, mechanistically, ETV4 increased CXCR5 expression by directly binding to the CXCR5 promoter region. Knockdown of CXCR5 significantly reversed ETV4-mediated PDAC migration and invasion, while CXCR5 overexpression exerted the opposite effects. Intriguingly, we found that CXCL13, a specific ligand of CXCR5, increased ETV4 expression and promoted PDAC invasion and metastasis by activating the ERK1/2 pathway. ETV4 knockdown significantly abrogated the enhanced migratory and invasive abilities induced by the CXCL13/CXCR5 axis. In addition, a CXCR5 neutralizing antibody disrupted the CXCL13/ETV4/CXCR5 positive feedback loop and inhibited cell migration and invasion. Overall, in this study, we demonstrated that ETV4 plays a vital role in PDAC metastasis and defined a novel CXCL13/ETV4/CXCR5 positive feedback loop. Targeting this pathway has implications for potential therapeutic strategies for PDAC treatment.
METTL3-dependent mA modification programs T follicular helper cell differentiation.
Yao Yingpeng,Yang Ying,Guo Wenhui,Xu Lifan,You Menghao,Zhang Yi-Chang,Sun Zhen,Cui Xiao,Yu Guotao,Qi Zhihong,Liu Jingjing,Wang Fang,Liu Juanjuan,Zhao Tianyan,Ye Lilin,Yang Yun-Gui,Yu Shuyang
T follicular helper (T) cells are specialized effector CD4 T cells critical to humoral immunity. Whether post-transcriptional regulation has a function in T cells is unknown. Here, we show conditional deletion of METTL3 (a methyltransferase catalyzing mRNA N-methyladenosine (mA) modification) in CD4 T cells impairs T differentiation and germinal center responses in a cell-intrinsic manner in mice. METTL3 is necessary for expression of important T signature genes, including Tcf7, Bcl6, Icos and Cxcr5 and these effects depend on intact methyltransferase activity. mA-miCLIP-seq shows the 3' UTR of Tcf7 mRNA is subjected to METTL3-dependent mA modification. Loss of METTL3 or mutation of the Tcf7 3' UTR mA site results in accelerated decay of Tcf7 transcripts. Importantly, ectopic expression of TCF-1 (encoded by Tcf7) rectifies T defects owing to METTL3 deficiency. Our findings indicate that METTL3 stabilizes Tcf7 transcripts via mA modification to ensure activation of a T transcriptional program, indicating a pivotal function of post-transcriptional regulation in promoting T cell differentiation.
GPA33 is expressed on multiple human blood cell types and distinguishes CD4 central memory T cells with and without effector function.
Opstelten Rianne,Suwandi Jessica S,Slot Manon C,Morgana Florencia,Scott Andrew M,Laban Sandra,Nikolic Tatjana,Turksma Annelies W,Kroeze Anna,Voermans Carlijn,Zwaginga Jaap-Jan,Roep Bart O,Amsen Derk
European journal of immunology
The Ig superfamily protein glycoprotein A33 (GPA33) has been implicated in immune dysregulation, but little is known about its expression in the immune compartment. Here, we comprehensively determined GPA33 expression patterns on human blood leukocyte subsets, using mass and flow cytometry. We found that GPA33 was expressed on fractions of B, dendritic, natural killer and innate lymphoid cells. Most prominent expression was found in the CD4 T cell compartment. Naïve and CXCR5 regulatory T cells were GPA33 , and naïve conventional CD4 T cells expressed intermediate GPA33 levels. The expression pattern of GPA33 identified functional heterogeneity within the CD4 central memory T cell (Tcm) population. GPA33 CD4 Tcm cells were fully undifferentiated, bona fide Tcm cells that lack immediate effector function, whereas GPA33 Tcm cells exhibited rapid effector functions and may represent an early stage of differentiation into effector/effector memory T cells before loss of CD62L. Expression of GPA33 in conventional CD4 T cells suggests a role in localization and/or preservation of an undifferentiated state. These results form a basis to study the function of GPA33 and show it to be a useful marker to discriminate between different cellular subsets, especially in the CD4 T cell lineage.
Elevated levels of follicular T helper cells and their association with therapeutic effects in patients with chronic lymphocytic leukaemia.
Qiu Liannv,Zhou Yonglie,Yu Qinhua,Zheng Sujie,Wang Zhenni,Huang Qiang
Chronic lymphocytic leukaemia (CLL) is characterized by an abnormal expansion of mature B cells with variable progression. Follicular T helper (Tfh) cells help B cells differentiate into plasma cells or long-lived memory B cells in germinal centres (GCs). However, the role of Tfh cells in CLL is poorly understand, and whether it plays a critical role in disease progression in vivo is lacking. In this study, we investigate the dynamic change of circulating Tfh cells in peripheral blood from patients with CLL during the treatment periods to evaluate their utility to predict disease progression. Our findings revealed the expansion of circulating CD4CXCR5, CD4ICOS, CD4PD-1 and CD4CXCR5ICOSPD-1 (Tfh) cells but lower serum IL-21 levels and CD4 T cell polarization not only to Tfh2 subtypes but also to Tfh17 subtypes in patients with CLL at pretreatment compared to patients with monoclonal B cell lymphocytosis (MBL) and healthy individuals, especially in those with advanced stage, which indicate these Tfh cells could be employed as a novel indicator for disease progression. Moreover, we observed significant correlations of Tfh17 and immunoglobulin heavy chain variable region (IGHV) mutation. Importantly, significantly decreased CD4ICOS, CD4PD-1 and Tfh cells were found after effective treatments, whereas a significantly high CD4ICOS, CD4PD-1 and Tfh cells were still found in those with progressive disease after treatments, suggesting that circulating CD4ICOS, CD4PD-1, Tfh cells could predict therapeutic effects.
γδ T cells control humoral immune response by inducing T follicular helper cell differentiation.
Rezende Rafael M,Lanser Amanda J,Rubino Stephen,Kuhn Chantal,Skillin Nathaniel,Moreira Thais G,Liu Shirong,Gabriely Galina,David Bruna A,Menezes Gustavo B,Weiner Howard L
γδ T cells have many known functions, including the regulation of antibody responses. However, how γδ T cells control humoral immunity remains elusive. Here we show that complete Freund's adjuvant (CFA), but not alum, immunization induces a subpopulation of CXCR5-expressing γδ T cells in the draining lymph nodes. TCRγδCXCR5 cells present antigens to, and induce CXCR5 on, CD4 T cells by releasing Wnt ligands to initiate the T follicular helper (Tfh) cell program. Accordingly, TCRδ mice have impaired germinal center formation, inefficient Tfh cell differentiation, and reduced serum levels of chicken ovalbumin (OVA)-specific antibodies after CFA/OVA immunization. In a mouse model of lupus, TCRδ mice develop milder glomerulonephritis, consistent with decreased serum levels of lupus-related autoantibodies, when compared with wild type mice. Thus, modulation of the γδ T cell-dependent humoral immune response may provide a novel therapy approach for the treatment of antibody-mediated autoimmunity.
Apoptosis Susceptibility Prolongs the Lack of Memory B Cells in Acute Leukemic Patients After Allogeneic Hematopoietic Stem Cell Transplantation.
Mensen Angela,Oh Youngseong,Becker Sonya C,Hemmati Philipp G,Jehn Christian,Westermann Jörg,Szyska Martin,Göldner Henning,Dörken Bernd,Scheibenbogen Carmen,Arnold Renate,Na Il-Kang
Biology of blood and marrow transplantation : journal of the American Society for Blood and Marrow Transplantation
Long-term survival after allogeneic hematopoietic stem cell transplantation requires intact immunosurveillance, which is hampered by lymphoid organ damage associated with conditioning therapy, graft-versus-host disease, and immunosuppression. Our study aimed to identify the mechanisms contributing to sustained low memory B cell numbers after transplantation. Peripheral B and T cell subset recovery and functional marker expression were investigated in 35 acute leukemic patients up to 1 year after transplantation. Apoptosis of B cells after CD40/TLR-9, CD40/BCR, and CD40/BCR/TLR-9-dependent stimulation and drug efflux capacity were analyzed. One half of the patients suffered from infections after day 180. All patients had strongly diminished CD27(+) memory B cells despite already normalized total B cell numbers and fully recovered CD27(-)IgD(-) memory B cells, putatively of extra-follicular origin. Circulating memory follicular helper T cells were reduced in the majority of patients as well. Naïve B cells exhibited a decreased expression of CXCR5, which mediates follicular B cell entry. Additionally, a lower HLA-DR expression was found on naïve B cells, impairing antigen presentation. Upon CD40/TLR-9-dependent activation, B cells underwent significantly increased apoptosis paralleled by an aberrant up-regulation of Fas-L on activated T cells and Fas on resting B cells. Significantly increased B cell apoptosis was also observed after CD40/BCR and CD40/BCR/TLR-9-dependent activation. Drug efflux capacity of naïve B cells was diminished in cyclosporin A-treated patients, additionally contributing to an apoptosis-prone phenotype. We conclude that B cell survival and migration and T cell communication defects are contributing candidates for an impaired germinal center formation of memory B cells after allogeneic hematopoietic stem cell transplantation. Follow-up studies should evaluate effectiveness of revaccinations on the cellular level and should address the long-term sequelae of B cell defects after transplantation.
Implication of TIGIT human memory B cells in immune regulation.
Hasan Md Mahmudul,Nair Sumi Sukumaran,O'Leary Jacqueline G,Thompson-Snipes LuAnn,Nyarige Verah,Wang Junwen,Park Walter,Stegall Mark,Heilman Raymond,Klintmalm Goran B,Joo HyeMee,Oh SangKon
Regulatory B cells (Bregs) contribute to immune regulation. However, the mechanisms of action of Bregs remain elusive. Here, we report that T cell immunoreceptor with Ig and ITIM domains (TIGIT) expressed on human memory B cells especially CD19CD24CD27CD39IgDIgMCD1c B cells is essential for effective immune regulation. Mechanistically, TIGIT on memory B cells controls immune response by directly acting on T cells and by arresting proinflammatory function of dendritic cells, resulting in the suppression of Th1, Th2, Th17, and CXCR5ICOS T cell response while promoting immune regulatory function of T cells. TIGIT memory B cells are also superior to other B cells at expressing additional inhibitory molecules, including IL-10, TGFβ1, granzyme B, PD-L1, CD39/CD73, and TIM-1. Lack or decrease of TIGIT memory B cells is associated with increased donor-specific antibody and TFH response, and decreased Treg response in renal and liver allograft patients. Therefore, TIGIT human memory B cells play critical roles in immune regulation.
Ageing promotes early T follicular helper cell differentiation by modulating expression of RBPJ.
Webb Louise M C,Fra-Bido Sigrid,Innocentin Silvia,Matheson Louise S,Attaf Noudjoud,Bignon Alexandre,Novarino Julien,Fazilleau Nicolas,Linterman Michelle A
Ageing profoundly changes our immune system and is thought to be a driving factor in the morbidity and mortality associated with infectious disease in older people. We have previously shown that the impaired immunity to vaccination that occurs in aged individuals is partly attributed to the effect of age on T follicular helper (Tfh) cell formation. In this study, we examined how age intrinsically affects Tfh cell formation in both mice and humans. We show increased formation of Tfh precursors (pre-Tfh) but no associated increase in germinal centre (GC)-Tfh cells in aged mice, suggesting age-driven promotion of only early Tfh cell differentiation. Mechanistically, we show that ageing alters TCR signalling which drives expression of the Notch-associated transcription factor, RBPJ. Genetic or chemical modulation of RBPJ or Notch rescues this age-associated early Tfh cell differentiation, and increased intrinsic Notch activity recapitulates this phenomenon in younger mice. Our data offer mechanistic insight into the age-induced changes in T-cell activation that affects the differentiation and ultimately the function of effector T cells.
PD-1CXCR5CD4 T Cells Play Defense in Cancer and Offense in Arthritis.
Gu-Trantien Chunyan,Willard-Gallo Karen
Trends in immunology
T follicular helper (TFH) cells are characteristically defined by their CXCR5 positivity and homing to B cell follicles in secondary lymphoid organs (SLO). An expanded subpopulation of functionally comparable and phenotypically similar PD-1CXCR5CD4 T cells were recently identified in breast cancer (BC) and rheumatoid arthritis (RA) to have beneficial or detrimental roles, respectively, but are they inflammatory tissue effector TFH cells?
Gastric cancer patients display a distinctive population of IFNgIL10 double positive CD8 T cells, which persists longer during prolonged activation.
Zhong Chen,Song Zongchang,Li Min
Experimental cell research
IL10 is generally regarded as a broad-spectrum regulatory cytokine. However, the role of IL10 in CD8 T cells remains controversial. In this study, we investigated the characteristics of endogenous IL10 by CD8 T cells in gastric cancer (GC) patients. Using intracellular staining, we found that in both GC patients and healthy controls, the majority of IL10-expressing CD8 T cells also presented concurrent IFNg expression. Interestingly, the frequency of IFNgIL10 CD8 T cells was significantly higher in GC patients than in healthy controls, while the frequency of IFNgIL10 CD8 T cells was significantly lower in GC patients than in healthy controls. Compared to the IFNgIL10 CD8 T cells, both IFNgIL10 and IFNgIL10 CD8 T cells presented significantly higher expression of activation/inhibitory markers. Interestingly, the IFNgIL10 cells presented lower PD1 and TIM3 and higher KLRG1 than the IFNgIL10 CD8 T cells. Remarkably, the IFNgIL10 CD8 T cells, but not the IFNgIL10 CD8 T cells, were highly enriched in the CD45ROCXCR5 subset. Prolonged activation resulted in significant enrichment of IFNgIL10 CD8 T cells over time. Interestingly, compared to the CD45ROCXCR5 CD8 T cells, the CD45ROCXCR5 CD8 T cells presented stronger proliferation capacity at later stages of stimulation, and higher viability throughout the stimulation process. Overall, our investigation demonstrated that GC patients were enriched with a distinctive population of IFNgIL10 double positive CD8 T cells, which resembled T follicular cytotoxic cells and could persist longer during prolonged activation.
PD-1 and TIGIT coexpression identifies a circulating CD8 T cell subset predictive of response to anti-PD-1 therapy.
Simon Sylvain,Voillet Valentin,Vignard Virginie,Wu Zhong,Dabrowski Camille,Jouand Nicolas,Beauvais Tiffany,Khammari Amir,Braudeau Cécile,Josien Régis,Adotevi Olivier,Laheurte Caroline,Aubin François,Nardin Charles,Rulli Samuel,Gottardo Raphael,Ramchurren Nirasha,Cheever Martin,Fling Steven P,Church Candice D,Nghiem Paul,Dreno Brigitte,Riddell Stanley R,Labarriere Nathalie
Journal for immunotherapy of cancer
BACKGROUND:Clinical benefit from programmed cell death 1 receptor (PD-1) inhibitors relies on reinvigoration of endogenous antitumor immunity. Nonetheless, robust immunological markers, based on circulating immune cell subsets associated with therapeutic efficacy are yet to be validated. METHODS:We isolated peripheral blood mononuclear cell from three independent cohorts of melanoma and Merkel cell carcinoma patients treated with PD-1 inhibitor, at baseline and longitudinally after therapy. Using multiparameter flow cytometry and cell sorting, we isolated four subsets of CD8 T cells, based on PD-1 and TIGIT expression profiles. We performed phenotypic characterization, T cell receptor sequencing, targeted transcriptomic analysis and antitumor reactivity assays to thoroughly characterize each of these subsets. RESULTS:We documented that the frequency of circulating PD-1TIGIT (DPOS) CD8 T-cells after 1 month of anti-PD-1 therapy was associated with clinical response and overall survival. This DPOS T-cell population was enriched in highly activated T-cells, tumor-specific and emerging T-cell clonotypes and T lymphocytes overexpressing CXCR5, a key marker of the CD8 cytotoxic follicular T cell population. Additionally, transcriptomic profiling defined a specific gene signature for this population as well as the overexpression of specific pathways associated with the therapeutic response. CONCLUSIONS:Our results provide a convincing rationale for monitoring this PD-1TIGIT circulating population as an early cellular-based marker of therapeutic response to anti-PD-1 therapy.
CXCR4 inhibition in human pancreatic and colorectal cancers induces an integrated immune response.
Biasci Daniele,Smoragiewicz Martin,Connell Claire M,Wang Zhikai,Gao Ya,Thaventhiran James E D,Basu Bristi,Magiera Lukasz,Johnson T Isaac,Bax Lisa,Gopinathan Aarthi,Isherwood Christopher,Gallagher Ferdia A,Pawula Maria,Hudecova Irena,Gale Davina,Rosenfeld Nitzan,Barmpounakis Petros,Popa Elizabeta Cristina,Brais Rebecca,Godfrey Edmund,Mir Fraz,Richards Frances M,Fearon Douglas T,Janowitz Tobias,Jodrell Duncan I
Proceedings of the National Academy of Sciences of the United States of America
Inhibition of the chemokine receptor CXCR4 in combination with blockade of the PD-1/PD-L1 T cell checkpoint induces T cell infiltration and anticancer responses in murine and human pancreatic cancer. Here we elucidate the mechanism by which CXCR4 inhibition affects the tumor immune microenvironment. In human immune cell-based chemotaxis assays, we find that CXCL12-stimulated CXCR4 inhibits the directed migration mediated by CXCR1, CXCR3, CXCR5, CXCR6, and CCR2, respectively, chemokine receptors expressed by all of the immune cell types that participate in an integrated immune response. Inhibiting CXCR4 in an experimental cancer medicine study by 1-wk continuous infusion of the small-molecule inhibitor AMD3100 (plerixafor) induces an integrated immune response that is detected by transcriptional analysis of paired biopsies of metastases from patients with microsatellite stable colorectal and pancreatic cancer. This integrated immune response occurs in three other examples of immune-mediated damage to noninfected tissues: Rejecting renal allografts, melanomas clinically responding to anti-PD1 antibody therapy, and microsatellite instable colorectal cancers. Thus, signaling by CXCR4 causes immune suppression in human pancreatic ductal adenocarcinoma and colorectal cancer by impairing the function of the chemokine receptors that mediate the intratumoral accumulation of immune cells.
Migratory cues controlling B-lymphocyte trafficking in human lymph nodes.
Park Saem Mul,Brooks Anna Es,Chen Chun-Jen J,Sheppard Hilary M,Loef Evert Jan,McIntosh Julie D,Angel Catherine E,Mansell Claudia J,Bartlett Adam,Cebon Jonathan,Birch Nigel P,Dunbar P Rod
Immunology and cell biology
B-cell migration within lymph nodes (LNs) is crucial to adaptive immune responses. Chemotactic gradients are proposed to drive migration of B cells into follicles, followed by their relocation to specific zones of the follicle during activation, and ultimately egress. However, the molecular drivers of these processes and the cells generating chemotactic signals that affect B cells in human LNs are not well understood. We used immunofluorescence microscopy, flow cytometry and functional assays to study molecular mechanisms of B-cell migration within human LNs, and found subtle but important differences to previous murine models. In human LNs we find CXCL13 is prominently expressed at the follicular edge, often associated with fibroblastic reticular cells located in these areas, whereas follicular dendritic cells show minimal contribution to CXCL13 expression. Human B cells rapidly downregulate CXCR5 on encountering CXCL13, but recover CXCR5 expression in the CXCL13-low environment. These data suggest that the CXCL13 gradient in human LNs is likely to be different from that proposed in mice. We also identify CD68 CD11c PU.1 tingible body macrophages within both primary and secondary follicles as likely drivers of the sphingosine-1-phosphate (S1P) gradient that mediates B-cell egress from LNs, through their expression of the S1P-degrading enzyme, S1P lyase. Based on our findings, we present a model of B-cell migration within human LNs, which has both similarities and interesting differences to that proposed for mice.
C-X-C chemokine receptor type 5 gene polymorphism affects gene expression in CD4+ T cells and is associated with increased risk of colorectal cancer.
Xing Junjie,Li Xu,Sui Jinke,Cao Guangwen,Fu Chuangang
Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine
Dysregulation of the immune system may play important roles in the development of colorectal cancer (CRC). The C-X-C chemokine receptor type 5 (CXCR5) is one of the principal regulators for targeting T cells, B cells, and dendritic cells into secondary lymphoid organs. The current study investigated the association between CXCR5 gene polymorphisms and the risk of CRC, and the potential effect of these polymorphisms on different immune cells. Two polymorphisms in CXCR5 gene, rs6421571C/T and rs80202369G/A, were examined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in 302 cases and 316 controls. Results showed that individuals with the rs6421571CT and TT genotypes had a strong correlation with the incidence of CRC (odds ratio (OR) = 1.46; 95 % confidence interval (CI), 1.02-2.09; p = 0.041 and OR = 2.62; 95 % CI, 1.50-4.95; p < 0.001, respectively). Also, rs80202369AA genotype revealed significantly higher distribution in CRC patients than in controls (p = 0.002). We further investigated the possible effects of the polymorphisms by assessing messenger RNA (mRNA) and protein levels of CXCR5 in peripheral blood mononuclear cells (PBMCs), CD4+ T cells, CD8+ T cells, and B cells. Data presented that healthy controls with rs6421571CT and TT genotypes had higher mRNA and protein levels of CXCR5 than those with wild-type CC genotype specifically in CD4+ T cells. These findings suggest novel risk factors of CRC and indicate a potential mechanism of CXCR5 gene polymorphism.
Cyclosporine a directly affects human and mouse b cell migration in vitro by disrupting a hIF-1 αdependent, o sensing, molecular switch.
Hilchey Shannon P,Palshikar Mukta G,Emo Jason A,Li Dongmei,Garigen Jessica,Wang Jiong,Mendelson Eric S,Cipolla Valentina,Thakar Juilee,Zand Martin S
BACKGROUND:Hypoxia is a potent molecular signal for cellular metabolism, mitochondrial function, and migration. Conditions of low oxygen tension trigger regulatory cascades mediated via the highly conserved HIF-1 α post-translational modification system. In the adaptive immune response, B cells (Bc) are activated and differentiate under hypoxic conditions within lymph node germinal centers, and subsequently migrate to other compartments. During migration, they traverse through changing oxygen levels, ranging from 1-5% in the lymph node to 5-13% in the peripheral blood. Interestingly, the calcineurin inhibitor cyclosporine A is known to stimulate prolyl hydroxylase activity, resulting in HIF-1 α destabilization and may alter Bc responses directly. Over 60% of patients taking calcineurin immunosuppressant medications have hypo-gammaglobulinemia and poor vaccine responses, putting them at high risk of infection with significantly increased morbidity and mortality. RESULTS:We demonstrate that O tension is a previously unrecognized Bc regulatory switch, altering CXCR4 and CXCR5 chemokine receptor signaling in activated Bc through HIF-1 α expression, and controlling critical aspects of Bc migration. Our data demonstrate that calcineurin inhibition hinders this O regulatory switch in primary human Bc. CONCLUSION:This previously unrecognized effect of calcineurin inhibition directly on human Bc has significant and direct clinical implications.
Impaired function of CD4+ T follicular helper (Tfh) cells associated with hepatocellular carcinoma progression.
Jia Yiqiong,Zeng Zhen,Li Yuanyuan,Li Zhiwei,Jin Lei,Zhang Zheng,Wang Lifeng,Wang Fu-Sheng
BACKGROUND AND AIMS:CD4+ T follicular helper (Tfh) cells, a new subset of immune cells, have been demonstrated to be involved in the development and prognosis of tumors. However, their functional role in human hepatocellular carcinoma (HCC) is relatively unknown, and the detailed mechanisms in HCC development remain to be described. METHODS:A total of 85 HCC patients with hepatitis B virus (HBV) infection, 25 HBV-relative liver cirrhosis (LC) patients, and 20 healthy controls (HC) were randomly enrolled. Flow cytometric analysis, immunohistochemical staining, and relative function (i.e., cytokine secretion, B cell maturation) assays were used to analyze the properties of CXCR5+CD4+ T cells. In addition, the relationship between the frequency of CXCR5+CD4+ T cells and overall survival rates or disease-free survival rates was also analyzed by the Kaplan-Meier method. RESULTS:The frequency of circulating CXCR5+CD4+ T cells was significantly decreased in HCC patients compared with HBV-relative liver cirrhosis (LC) patients and healthy controls, and the decrease in circulating CXCR5+CD4+ T cells correlated with disease progression. The proportion of infiltrated CXCR5+CD4+ T cells was significantly decreased in tumor regions compared with nontumor regions. Furthermore, compared with healthy controls, the function of circulating CXCR5+CD4+ T cells in HCC was impaired, with reduced IL-21 secretion and dysfunction in promoting B cell maturation. Importantly, follow-up data indicated that a decreased frequency of circulating CXCR5+CD4+ T cells was also associated with reduced disease-free survival time in HCC patients. CONCLUSIONS:Impairment of CD4+ T follicular helper cells may influence the development of HBV-associated HCC. Decreased CD4+ T follicular helper cells may represent a potential prognostic marker and serve as a novel therapeutic target for HCC patients.
T follicular helper cells in human efferent lymph retain lymphoid characteristics.
Vella Laura A,Buggert Marcus,Manne Sasikanth,Herati Ramin S,Sayin Ismail,Kuri-Cervantes Leticia,Bukh Brody Irene,O'Boyle Kaitlin C,Kaprielian Hagop,Giles Josephine R,Nguyen Son,Muselman Alexander,Antel Jack P,Bar-Or Amit,Johnson Matthew E,Canaday David H,Naji Ali,Ganusov Vitaly V,Laufer Terri M,Wells Andrew D,Dori Yoav,Itkin Maxim G,Betts Michael R,Wherry E John
The Journal of clinical investigation
T follicular helper cells (Tfh), a subset of CD4+ T cells, provide requisite help to B cells in the germinal centers (GC) of lymphoid tissue. GC Tfh are identified by high expression of the chemokine receptor CXCR5 and the inhibitory molecule PD-1. Although more accessible, blood contains lower frequencies of CXCR5+ and PD-1+ cells that have been termed circulating Tfh (cTfh). However, it remains unclear whether GC Tfh exit lymphoid tissues and populate this cTfh pool. To examine exiting cells, we assessed the phenotype of Tfh present within the major conduit of efferent lymph from lymphoid tissues into blood, the human thoracic duct. Unlike what was found in blood, we consistently identified a CXCR5-bright PD-1-bright (CXCR5BrPD-1Br) Tfh population in thoracic duct lymph (TDL). These CXCR5BrPD-1Br TDL Tfh shared phenotypic and transcriptional similarities with GC Tfh. Moreover, components of the epigenetic profile of GC Tfh could be detected in CXCR5BrPD-1Br TDL Tfh and the transcriptional imprint of this epigenetic signature was enriched in an activated cTfh subset known to contain vaccine-responding cells. Together with data showing shared TCR sequences between the CXCR5BrPD-1Br TDL Tfh and cTfh, these studies identify a population in TDL as a circulatory intermediate connecting the biology of Tfh in blood to Tfh in lymphoid tissue.
Phenotypically defined subpopulations of circulating follicular helper T cells in common variable immunodeficiency.
Yesillik Sait,Gupta Sudhir
Immunity, inflammation and disease
BACKGROUND:Common variable immunodeficiency (CVID) is characterized by low immunoglobulin G and IgA/IgM, decreased switched memory B cells, impaired response to vaccine, and an increased susceptibility to infections and autoimmunity. T cells play an important role in germinal center reaction where it supports isotype switching, somatic hypermutation, generation of memory B cells, and differentiation of B cells to plasma cells. The objective was to study the distribution of three subsets of T cells and their relationship with autoimmune diseases associated with CVID. METHODS:T cells have been divided into T 1 (interleukin 21 [IL-21] and interferon γ), T 2 (IL-21 and IL-4), and T 17 (IL-21 and IL-17) cells. Mononuclear cells from 25 patients with CVID and age and gender-matched controls were stained with various monoclonal antibodies (anti-CD4 APC, anti-CXCR5 FITC, anti-CCR6 PerCP, and anti-CXCR3 PE) and isotype controls and analyzed for T 1 (CD4 CXCR5 CXCR3 CCR6 ), T 2 (CD4 CXCR5 CXCR3 CCR6 ), and T 17 (CD4 CXCR5 CXCR3 CCR6 ) cells by multicolor flow cytometry. Twenty thousand cells were acquired and analyzed by FlowJo software. Statistical analysis of comparison of patients and healthy controls was performed by paired t test using PRISM 7 software. RESULTS:T 2 and T 17 cells subpopulations of T cells were significantly decreased (P < .003 and P < .006, respectively) in CVID as compared with controls. No significant difference was observed in any of T cell subpopulations between CVID with and those without autoimmunity group. CONCLUSION:Alterations in T cell subpopulation may play a role in defects in B cell compartment in CVID.
Control of Germinal Center Localization and Lineage Stability of Follicular Regulatory T Cells by the Blimp1 Transcription Factor.
Shen Erxia,Rabe Hardis,Luo Lin,Wang Lei,Wang Qin,Yin Jie,Yang Xueying,Liu Wenquan,Sido Jessica M,Nakagawa Hidetoshi,Ao Lin,Kim Hye-Jung,Cantor Harvey,Leavenworth Jianmei W
Follicular regulatory T (T) cells are a specialized suppressive subset that controls the germinal center (GC) response and maintains humoral self-tolerance. The mechanisms that maintain T lineage identity and suppressive activity remain largely unknown. Here, we show that expression of Blimp1 by FoxP3 T cells is essential for T lineage stability, entry into the GC, and expression of regulatory activity. Deletion of Blimp1 in T cells reduced FoxP3 and CTLA-4 expression and increased pro-inflammatory cytokines and spontaneous production of autoantibodies, including elevated IgE. Maintenance of T stability reflected Blimp1-dependent repression of the IL-23R-STAT3 axis and activation of the CD25-STAT5 pathway, while silenced IL-23R-STAT3 or increased STAT5 activation rescued the Blimp1-deficient T phenotype. Blimp1-dependent control of CXCR5/CCR7 expression also regulated T homing into the GC. These findings uncover a Blimp1-dependent T checkpoint that enforces suppressive activity and acts as a gatekeeper of GC entry.
Sustained delivery and molecular targeting of a therapeutic monoclonal antibody to metastases in the central nervous system of mice.
Wen Jing,Wu Di,Qin Meng,Liu Chaoyong,Wang Lan,Xu Duo,Vinters Harry V,Liu Yang,Kranz Emiko,Guan Xin,Sun Guibo,Sun Xiaobo,Lee YooJin,Martinez-Maza Otoniel,Widney Daniel,Lu Yunfeng,Chen Irvin S Y,Kamata Masakazu
Nature biomedical engineering
Approximately 15-40% of all cancers develop metastases in the central nervous system (CNS), yet few therapeutic options exist to treat them. Cancer therapies based on monoclonal antibodies are widely successful, yet have limited efficacy against CNS metastases, owing to the low levels of the drug reaching the tumour site. Here, we show that the encapsulation of rituximab within a crosslinked zwitterionic polymer layer leads to the sustained release of rituximab as the crosslinkers are gradually hydrolysed, enhancing the CNS levels of the antibody by approximately tenfold with respect to the administration of naked rituximab. When the nanocapsules were functionalized with CXCL13-the ligand for the chemokine receptor CXCR5, which is frequently found on B-cell lymphoma-a single dose led to improved control of CXCR5-expressing metastases in a murine xenograft model of non-Hodgkin lymphoma, and eliminated lymphoma in a xenografted humanized bone marrow-liver-thymus mouse model. Encapsulation and molecular targeting of therapeutic antibodies could become an option for the treatment of cancers with CNS metastases.
Primary Intra-Axial Diffuse Large B-Cell Lymphoma in Immunocompetent Patients: Clinical Impact of Molecular Analysis and Histogenetic Evaluation.
Primary central nervous system (CNS) diffuse large B-cell lymphoma (DLBCL) represents less than 1% of non-Hodgkin lymphomas and 2%-3% of brain tumors. Primary CNS DLBCL occurs sporadically in healthy patients. Tumor development and progression have been associated with reduced/absent expression of human leukocyte antigen class I and II proteins; increased expression of CXCR4, CXCL12, CXCR5, and CCR7; mutations of VH4/34, BCL6, MYC, and PAX5 genes; and rearrangement of immunoglobulin heavy and light chain genes. Generally, DLBCL is a single supratentorial lesion (60%-70%), and stereotactic biopsy and intraoperative examination are the main diagnostic methods. Distinctive histologic features are a diffuse growth pattern and angioinvasiveness. Most neoplastic cells resemble centroblasts and exhibit positive CD20, CD22, PAX5, CD79a, and MUM1 expression. The prognosis of primary CNS DLBCL is less favorable than that of nodal DLBCL, and DLBCL subtype, strong FOXP1 immunoreactivity, MYC and BCL2 overexpression, and BCL6 translocations are associated with poor prognosis.
Expression of the chemokine receptors CXCR4, CXCR5, and CCR7 in primary central nervous system lymphoma.
Jahnke Kristoph,Coupland Sarah E,Na Il-Kang,Loddenkemper Christoph,Keilholz Ulrich,Korfel Agnieszka,Stein Harald,Thiel Eckhard,Scheibenbogen Carmen
Pathologically expanded peripheral T helper cell subset drives B cells in rheumatoid arthritis.
Rao Deepak A,Gurish Michael F,Marshall Jennifer L,Slowikowski Kamil,Fonseka Chamith Y,Liu Yanyan,Donlin Laura T,Henderson Lauren A,Wei Kevin,Mizoguchi Fumitaka,Teslovich Nikola C,Weinblatt Michael E,Massarotti Elena M,Coblyn Jonathan S,Helfgott Simon M,Lee Yvonne C,Todd Derrick J,Bykerk Vivian P,Goodman Susan M,Pernis Alessandra B,Ivashkiv Lionel B,Karlson Elizabeth W,Nigrovic Peter A,Filer Andrew,Buckley Christopher D,Lederer James A,Raychaudhuri Soumya,Brenner Michael B
CD4 T cells are central mediators of autoimmune pathology; however, defining their key effector functions in specific autoimmune diseases remains challenging. Pathogenic CD4 T cells within affected tissues may be identified by expression of markers of recent activation. Here we use mass cytometry to analyse activated T cells in joint tissue from patients with rheumatoid arthritis, a chronic immune-mediated arthritis that affects up to 1% of the population. This approach revealed a markedly expanded population of PD-1CXCR5CD4 T cells in synovium of patients with rheumatoid arthritis. However, these cells are not exhausted, despite high PD-1 expression. Rather, using multidimensional cytometry, transcriptomics, and functional assays, we define a population of PD-1CXCR5 'peripheral helper' T (T) cells that express factors enabling B-cell help, including IL-21, CXCL13, ICOS, and MAF. Like PD-1CXCR5 T follicular helper cells, T cells induce plasma cell differentiation in vitro through IL-21 secretion and SLAMF5 interaction (refs 3, 4). However, global transcriptomics highlight differences between T cells and T follicular helper cells, including altered expression of BCL6 and BLIMP1 and unique expression of chemokine receptors that direct migration to inflamed sites, such as CCR2, CX3CR1, and CCR5, in T cells. T cells appear to be uniquely poised to promote B-cell responses and antibody production within pathologically inflamed non-lymphoid tissues.
Development of an Antigen-Antibody Co-Display System for Detecting Interaction of G-Protein-Coupled Receptors and Single-Chain Variable Fragments.
Zhang Yinjie,Wu Boyang Jason,Yu Xiaolan,Luo Ping,Ye Hao,Yu Yan,Han Wei,Li Jingjing
International journal of molecular sciences
G-protein-coupled receptors (GPCRs), especially chemokine receptors, are ideal targets for monoclonal antibody drugs. Considering the special multi-pass transmembrane structure of GPCR, it is often a laborious job to obtain antibody information about off-targets and epitopes on antigens. To accelerate the process, a rapid and simple method needs to be developed. The split-ubiquitin-based yeast two hybrid system (YTH) was used as a blue script for a new method. By fusing with transmembrane peptides, scFv antibodies were designed to be anchored on the cytomembrane, where the GPCR was co-displayed as well. The coupled split-ubiquitin system transformed the scFv-GPCR interaction signal into the expression of reporter genes. By optimizing the topological structure of scFv fusion protein and key elements, including signal peptides, transmembrane peptides, and flexible linkers, a system named Antigen-Antibody Co-Display (AACD) was established, which rapidly detected the interactions between antibodies and their target GPCRs, CXCR4 and CXCR5, while also determining the off-target antibodies and antibody-associated epitopes. The AACD system can rapidly determine the association between GPCRs and their candidate antibodies and shorten the research period for off-target detection and epitope identification. This system should improve the process of GPCR antibody development and provide a new strategy for GPCRs antibody screening.
Long non‑coding RNA PLK1S1 was associated with renal cell carcinoma progression by interacting with microRNA‑653 and altering C‑X‑C chemokine receptor 5 expression.
Li Weiyuan,Yang Dengke,Zhang Yu,Zhao Shutian,Li Dong,Liu Min
Renal cell carcinoma (RCC) is the most common type of renal cancer. Long non‑coding RNA (lncRNA) has been reported to play a vital role in the development and progression of various types of cancer type. However, the underlying molecular mechanisms of PLK1S1 in regulating RCC progression remain unclear. In the present study, PLK1S1 was upregulated in RCC tissues and cells, and PLK1S1 expression was also significantly elevated in stage IV RCC tissues. Kaplan‑Meier analysis showed that patients with high PLK1S1 expression had a shorter overall survival time compared with those with low PLK1S1 expression. Moreover, bioinformatics analysis and luciferase reporter assay demonstrated that PLK1S1 inhibited microRNA (miR)‑653 expression by direct interaction. Functional analyses demonstrated that a miR‑653 inhibitor promoted short hairpin PLK1S1‑attenuated cell proliferation, invasion and sorafenib resistance of RCC cells. In addition, C‑X‑C motif chemokine receptors 5 (CXCR5) was identified as an effector of PLK1S1/miR‑653‑mediated tumorigenesis and drug resistance in RCC cells. Lastly, xenograft experiments demonstrated that PLK1S1 knockdown inhibited tumor growth in vivo. Reverse transcription‑quantitative PCR and western blot analysis revealed that PLK1S1 knockdown upregulated the expression level of miR‑653, whilst downregulating the expression level of CXCR5. In conclusion, the present study revealed that PLK1S1 promoted tumor progression and sorafenib resistance in RCC through regulation of the miR‑653/CXCR5 axis, which may offer a novel treatment strategy for patients with RCC.
[Relation of Circulating Follicular Helper T Cell Changes with B Cell Dysfunction in MDS Patients].
Liu Hui,Cao Hui-Qin,Zhao Jian-Qiang
Zhongguo shi yan xue ye xue za zhi
OBJECTIVE:To explore the relation of circulating follicular helper T cell (c Tfh) changes with B cell dysfunction in MDS patients. METHODS:20 patients diagnosed as MDS from Auguct 2015 to October 2017 were enrolled in MDS group, and 20 healthy valuntears matching in age and sex were enrolled in healthy control (HC) group. The perepheral blood in 2 groups were collected, the mononuclear cells (PBMC) from which were isolated by densily gradient contrifugation, at the same time, the serum left in isolation process was reserved for further study. The flow cytometry was used to detect the ratio of cTfh such as CD4CXCR5 T cells and its subset CD4CXCR5ICOS T cells, CD4CXCR5PD-1 T cells in PBMC, as well as the ratio of plasmablast CD19CD20CD38 B cells. The ELISA was used to detect the concentration of IgA, IgM and IgG. The differences in ratio of cTfh cells and plasmablast B cells, as well as the concentration of IgA, IgM and IgG between MDS and HC groups were compared, at the same time, the correlation of cTfh cell ratio with the plasmablast B cell ratio and the concentration of IgA, IgM and IgG in MDS patient was analyzed. RESULTS:The ratio of CD4CXCR5T, CD4CXCR5ICOST cells and CD19CD20CD38B cells and the concentration of IgA, IgM and IgG decreased in MDS patients, while the ratio of CD4CXCR5PD-1T cells increased in MDS patients. The ratio of CD4CXCR5T cells, CD4CXCR5ICOST cells positively correlated with the ratio of CD19CD20CD38B cells, as well as with the concentration of IgA, IgM and IgG in MDS patients. However, the ratio of CD4CXCR5PD-1T cells negatively correlated with the ratio of CD19CD20CD38B cells, as well as with the concentration of IgA, IgM and IgG. CONCLUSION:The ratio of circulating Tfh cells and their subsets showed significant changes, that correlate with B cell dysfunction in MDS patients.
Circulating T Follicular Helper Cell Abnormalities Associated to Different Clinical Forms of Chronic Chagas Disease.
Quebrada Palacio Luz P,Fernández Esteban R,Hernández-Vásquez Yolanda,Petray Patricia B,Postan Miriam
Frontiers in cellular and infection microbiology
Multiple perturbations of the immune response affecting a range of cells have been reported in -infected individuals and associated to clinical manifestations of chronic Chagas disease. There is a paucity of knowledge about the role of T follicular helper (Tfh) cells in this infection. Here, we sought to characterize circulating Tfh (cTfh) cells in chronic Chagas disease patients and to identify potential associations with disease severity in humans. cTfh cells were characterized by flow cytometry in freshly isolated PBMCs from 7 -infected asymptomatic patients (ASYMP), 5 patients with chronic chagasic dilated cardiomyopathy (CCC) and 8 healthy controls, using antibodies against chemokine receptors CXCR5, CXCR3, CCR6, and CCR7. Our results showed significant expansion of CD4+CD45RO+CXCR5+CCR6+ cells in ASYMP and CCC patients, along with a contraction of CD4+CD45RO+CXCR5+CXCR3-CCR6- (cTfh2) cells. ASYMP patients further exhibited decreased CD4+CD45RO+CXCR5+CXCR3+CCR6- (cTfh1) cells and expanded CD4+CD45RO+CXCR5+CXCR3-CCR6+ (cTfh17) cells while CCC patients exhibited significantly increased frequencies of CD4+CD45RO+CXCR5+CCR7+ cells. Linear regression analysis revealed a positive trend of CD4+CD45RO+CXCR5+CXCR3+CCR6+ (cTfh1/17) cells and negative trends of cTfh1 and cTfh2 cells as disease was more severe. There was no correlation between the frequencies of cTfh cells and circulating CD19+IgD-IgG+ cells or serum levels of -specific IgG. These results demonstrate that the cTfh compartment of humans chronically infected with comprises expanded CCR6-expressing cells and reduced cTfh2 cells. The association of discrete phenotypic changes in cTfh subsets with different clinical forms suggests the potential contribution of T follicular helper cells to Chagas heart disease progression.
Hypermethylation of Single CpG Dinucleotides at the Promoter of CXCL13 Gene Promoting Cell Migration in Cervical Cancer.
Ma Dong,Fan Shao-Bei,Hua Na,Li Guo-Hua,Chang Quan,Liu Xiao
Current cancer drug targets
BACKGROUND:Chemokine 13 (CXCL13) and its chemokine receptor 5 (CXCR5) are involved in the onset of various types of cancer. However, their role in cervical cancer (CC) remains unknown. OBJECTIVE:To investigate the role of chemokine 13 (CXCL13) and its receptor in CC. METHODS:The expression of CXCL13/CXCR5 and the infiltration of CXCR5+CD8+ T cells in CC, cervical intraepithelial neoplasia (CIN), normal cervical epithelial (NCE) tissues, and in CC cell lines were analysed and the associated clinical significance was determined. In vitro, CXCL13 overexpression and DNA methyltransferase inhibition (through S110) were used to investigate the biological function and the underlying mechanism that regulates CXCL13 expression. Tumor growth and liver metastasis were also evaluated in the xenogenous subcutaneously implant model. RESULTS:CXCL13/CXCR5 expression levels and the infiltration of CXCR5+CD8+ T cells were significantly decreased in CC tissues compared with CIN and NCE tissues. CXCL13 downregulation was significantly correlated with the FIGO stages, lymph node metastasis, interstitial infiltration depth, and pathological grade. The overexpression of CXCL13 suppressed CC cell migration. CXCL13 downregulation was associated with hypermethylation in CC cell lines, and primary tumor biopsies. Furthermore, a CpG dinucleotide at the HIF-1a transcription factor motifs in the promoter element of CXCL13 was consistently methylated in CC cells and associated with HIF-1a. CXCL13 overexpression and S110 treatment dramatically repressed tumor growth and liver metastasis in the xenograft model; whereas it's low expression increased the risk of death in CC patients. CONCLUSION:DNA methylation-dependent CXCL13 downregulation may promote cervical carcinogenesis and progression.
Human Lymphoid Stromal Cells Contribute to Polarization of Follicular T Cells Into IL-4 Secreting Cells.
Misiak Jan,Jean Rachel,Rodriguez Stéphane,Deleurme Laurent,Lamy Thierry,Tarte Karin,Amé-Thomas Patricia
Frontiers in immunology
Fibroblastic reticular cells (FRCs) are the specialized lymphoid stromal cells initially identified as triggering T-cell recruitment and dynamic motion in secondary lymphoid organs. Interestingly, FRCs also display antigen presentation capacities and support lymphocyte survival. CXCR5CD4 follicular T cells are important players of B-cell maturation and antibody response. Our study reported that -differentiated FRC-like cells enhanced the growth of the whole CXCR5CD4 T-cell compartment, while enhancing IL-4 secretion specifically by the PD1CXCR5CD4 cell subset, in a Notch- and ICAM1/LFA1-dependent manner. In addition, we revealed that in follicular lymphoma (FL) tissues, previously identified as enriched for PD1CXCR5CD4 mature follicular helper T cells, PD1CXCR5CD4 T cells displayed an enrichment for Notch and integrin gene signatures, and a Notch and ICAM-1-dependent overexpression of IL-4 compared to their non-malignant counterparts. These findings suggest that the crosstalk between FRCs and CXCR5PD1CD4 T cells may contribute to the FL IL-4 rich environment, thus providing new insights in FL lymphomagenesis.
Follicular helper T cells promote the effector functions of CD8 T cells via the provision of IL-21, which is downregulated due to PD-1/PD-L1-mediated suppression in colorectal cancer.
Shi Weiwei,Dong Lin,Sun Qiong,Ding Hong,Meng Jing,Dai Guanghai
Experimental cell research
Recent studies have demonstrated that higher expression of follicular helper T cell (Tfh)-associated genes is associated with better prognosis in breast cancer and colorectal cancer (CRC), but the underlying mechanisms remain unclear. In this study, we compared the function of Tfh cells in non-cancer (NC) controls and in CRC patients between stages II and IV. Data showed that the level of CD4CXCR5 Tfh cells were significantly upregulated in stage II CRC patients but was progressively reduced in stage III and stage IV patients. The frequency of PD-1 cells in CD4CXCR5 Tfh cells, on the other hand, was progressively increased from NC patients to CRC patients with increasing severity. Interestingly, the CD4CXCR5PD-1 Tfh cells, but not the CD4CXCR5PD-1 Tfh cells, promoted the CD107a expression and IFN-γ expression by CD8 T cells. This CD8 T cell-promoting capacity was dependent on the expression of IL-21, as this capacity was significantly impaired by IL-21 neutralization. Tfh cells from CRC patients with advanced stages presented gradual reduction in the capacity to stimulate CD8 T cells and the capacity to produce IL-21. In addition, treatment with autologous PD-L1-expressing tumor cells further suppressed the IL-21 expression by PD-1 Tfh cells. Together, these data demonstrated that Tfh cells potently enhanced the effector functions of CD8 T cells in an IL-21-dependent pathway; however, this role of Tfh cells was limited in CRC patients due to PD-1/PD-L1-mediated suppression.
The N-terminal length and side-chain composition of CXCL13 affect crystallization, structure and functional activity.
Rosenberg Eric M,Herrington James,Rajasekaran Deepa,Murphy James W,Pantouris Georgios,Lolis Elias J
Acta crystallographica. Section D, Structural biology
CXCL13 is the cognate chemokine agonist of CXCR5, a class A G-protein-coupled receptor (GPCR) that is essential for proper humoral immune responses. Using a `methionine scanning' mutagenesis method on the N-terminus of CXCL13, which is the chemokine signaling region, it was shown that minor length alterations and side-chain substitutions still result in CXCR5 activation. This observation indicates that the orthosteric pocket of CXCR5 can tolerate these changes without severely affecting the activity. The introduction of bulk on the ligand was well tolerated by the receptor, whereas a loss of contacts was less tolerated. Furthermore, two crystal structures of CXCL13 mutants were solved, both of which represent the first uncomplexed structures of the human protein. These structures were stabilized by unique interactions formed by the N-termini of the ligands, indicating that CXCL13 exhibits substantial N-terminal flexibility while the chemokine core domain remains largely unchanged. Additionally, it was observed that CXCL13 harbors a large degree of flexibility in the C-terminal extension of the ligand. Comparisons with other published structures of human and murine CXCL13 validate the relative rigidity of the core domain as well as the N- and C-terminal mobilities. Collectively, these mutants and their structures provide the field with additional insights into how CXCL13 interacts with CXCR5.
CXCL14 Preferentially Synergizes With Homeostatic Chemokine Receptor Systems.
Kouzeli Ariadni,Collins Paul J,Metzemaekers Mieke,Meyrath Max,Szpakowska Martyna,Artinger Marc,Struyf Sofie,Proost Paul,Chevigne Andy,Legler Daniel F,Eberl Matthias,Moser Bernhard
Frontiers in immunology
Reflecting their importance in immunity, the activity of chemokines is regulated on several levels, including tissue and context-specific expression and availability of their cognate receptor on target cells. Chemokine synergism, affecting both chemokine and chemokine receptor function, has emerged as an additional control mechanism. We previously demonstrated that CXCL14 is a positive allosteric modulator of CXCR4 in its ability to synergize with CXCL12 in diverse cellular responses. Here, we have extended our study to additional homeostatic, as well as a selection of inflammatory chemokine systems. We report that CXCL14 strongly synergizes with low (sub-active) concentrations of CXCL13 and CCL19/CCL21 in chemotaxis with immune cells expressing the corresponding receptors CXCR5 and CCR7, respectively. CXCL14 by itself was inactive, not only on cells expressing CXCR5 or CCR7 but also on cells expressing any other known conventional or atypical chemokine receptor, as assessed by chemotaxis and/or β-arrestin recruitment assays. Furthermore, synergistic migration responses between CXCL14 and inflammatory chemokines CXCL10/CXCL11 and CCL5, targeting CXCR3 and CCR5, respectively, were marginal and occasional synergistic Ca flux responses were observed. CXCL14 bound to 300-19 cells and interfered with CCL19 binding to CCR7-expressing cells, suggesting that these cellular interactions contributed to the reported CXCL14-mediated synergistic activities. We propose a model whereby tissue-expressed CXCL14 contributes to cell localization under steady-state conditions at sites with prominent expression of homeostatic chemokines.
Simulating CXCR5 Dynamics in Complex Tissue Microenvironments.
Cosgrove Jason,Alden Kieran,Stein Jens V,Coles Mark C,Timmis Jon
Frontiers in immunology
To effectively navigate complex tissue microenvironments, immune cells sense molecular concentration gradients using G-protein coupled receptors. However, due to the complexity of receptor activity, and the multimodal nature of chemokine gradients , chemokine receptor activity is poorly understood. To address this issue, we apply a modelling and simulation approach that permits analysis of the spatiotemporal dynamics of CXCR5 expression within an B-follicle with single-cell resolution. Using this approach, we show that that B-cell scanning is robust to changes in receptor numbers and changes in individual kinetic rates of receptor activity, but sensitive to global perturbations where multiple parameters are altered simultaneously. Through multi-objective optimization analysis we find that the rapid modulation of CXCR5 activity through receptor binding, desensitization and recycling is required for optimal antigen scanning rates. From these analyses we predict that chemokine receptor signaling dynamics regulate migration in complex tissue microenvironments to a greater extent than the total numbers of receptors on the cell surface.
CXCL13/CXCR5 Axis Predicts Poor Prognosis and Promotes Progression Through PI3K/AKT/mTOR Pathway in Clear Cell Renal Cell Carcinoma.
Zheng Zaosong,Cai Yuhong,Chen Haicheng,Chen Zhiliang,Zhu Dingjun,Zhong Qiyu,Xie Wenlian
Frontiers in oncology
The chemokine ligands and their receptors play critical roles in cancer progression and patients outcomes. We found that CXCL13 was significantly upregulated in ccRCC tissues compared with normal tissues in both The Cancer Genome Atlas (TCGA) cohort and a validated cohort of 90 pairs ccRCC tissues. Statistical analysis showed that high CXCL13 expression related to advanced disease stage and poor prognosis in ccRCC. We also revealed that serum CXCL13 levels in ccRCC patients ( = 50) were significantly higher than in healthy controls ( = 40). Receiver operating characteristic (ROC) curve revealed that tissue and serum CXCL13 expression might be a diagnostic biomarker for ccRCC with an area under curve (AUC) of 0.809 and 0.704, respectively. CXCL13 was significantly associated with its receptor, CXCR5, in ccRCC tissues, and ccRCC patients in high CXCL13 high CXCR5 expression group have a worst prognosis. Functional and mechanistic study revealed that CXCL13 promoted the proliferation and migration of ccRCC cells by binding to CXCR5 and activated PI3K/AKT/mTOR signaling pathway. These results suggested that CXCL13/CXCR5 axis played a significant role in ccRCC and might be a therapeutic target and prognostic biomarker.
Serum levels of the chemokine CXCL13, genetic variation in CXCL13 and its receptor CXCR5, and HIV-associated non-hodgkin B-cell lymphoma risk.
Hussain Shehnaz K,Zhu Weiming,Chang Shen-Chih,Breen Elizabeth Crabb,Vendrame Elena,Magpantay Larry,Widney Dan,Conn Daniel,Sehl Mary,Jacobson Lisa P,Bream Jay H,Wolinsky Steven,Rinaldo Charles R,Ambinder Richard F,Detels Roger,Zhang Zuo-Feng,Martínez-Maza Otoniel
Cancer epidemiology, biomarkers & prevention : a publication of the American Association for Cancer Research, cosponsored by the American Society of Preventive Oncology
BACKGROUND:CXCL13 and CXCR5 are a chemokine and receptor pair whose interaction is critical for naïve B-cell trafficking and activation within germinal centers. We sought to determine whether CXCL13 levels are elevated before HIV-associated non-Hodgkin B-cell lymphoma (AIDS-NHL), and whether polymorphisms in CXCL13 or CXCR5 are associated with AIDS-NHL risk and CXCL13 levels in a large cohort of HIV-infected men. METHODS:CXCL13 levels were measured in sera from 179 AIDS-NHL cases and 179 controls at three time-points. TagSNPs in CXCL13 (n = 16) and CXCR5 (n = 11) were genotyped in 183 AIDS-NHL cases and 533 controls. OR and 95% confidence intervals (CI) for the associations between one unit increase in log CXCL13 levels and AIDS-NHL, as well as tagSNP genotypes and AIDS-NHL, were computed using logistic regression. Mixed linear regression was used to estimate mean ratios (MR) for the association between tagSNPs and CXCL13 levels. RESULTS:CXCL13 levels were elevated for more than 3 years (OR = 3.24; 95% CI = 1.90-5.54), 1 to 3 years (OR = 3.39; 95% CI = 1.94-5.94), and 0 to 1 year (OR = 3.94; 95% CI = 1.98-7.81) before an AIDS-NHL diagnosis. The minor allele of CXCL13 rs355689 was associated with reduced AIDS-NHL risk (OR(TCvsTT) = 0.65; 95% CI = 0.45-0.96) and reduced CXCL13 levels (MR(CCvsTT) = 0.82; 95% CI = 0.68-0.99). The minor allele of CXCR5 rs630923 was associated with increased CXCL13 levels (MR(AAvsTT) = 2.40; 95% CI = 1.43-4.50). CONCLUSIONS:CXCL13 levels were elevated preceding an AIDS-NHL diagnosis, genetic variation in CXCL13 may contribute to AIDS-NHL risk, and CXCL13 levels may be associated with genetic variation in CXCL13 and CXCR5. IMPACT:CXCL13 may serve as a biomarker for early AIDS-NHL detection.
Therapeutic regulatory T-cell adoptive transfer ameliorates established murine chronic GVHD in a CXCR5-dependent manner.
McDonald-Hyman Cameron,Flynn Ryan,Panoskaltsis-Mortari Angela,Peterson Nicholas,MacDonald Kelli P A,Hill Geoffrey R,Luznik Leo,Serody Jonathan S,Murphy William J,Maillard Ivan,Munn David H,Turka Laurence A,Koreth John,Cutler Corey S,Soiffer Robert J,Antin Joseph H,Ritz Jerome,Blazar Bruce R
Chronic graft-versus-host disease (cGVHD) is a major complication of allogeneic hematopoietic stem cell transplantation. In cGVHD, alloreactive T cells and germinal center (GC) B cells often participate in GC reactions to produce pathogenic antibodies. Although regulatory T cells (Tregs) can inhibit GC reactions, Treg numbers are reduced in cGVHD, contributing to cGVHD pathogenesis. Here, we explored 2 means to increase Tregs in cGVHD: interleukin-2/monoclonal antibody (IL-2/mAb) complexes and donor Treg infusions. IL-2/mAb complexes given over 1 month were efficacious in expanding Tregs and treating established cGVHD in a multi-organ-system disease mouse model characterized by GC reactions, antibody deposition, and lung dysfunction. In an acute GVHD (aGVHD) model, IL-2/mAb complexes given for only 4 days resulted in rapid mortality, indicating IL-2/mAb complexes can drive conventional T-cell (Tcon)-mediated injury. In contrast, Treg infusions, which uniformly suppress aGVHD, increased Treg frequency and were effective in preventing the onset of, and treating, established cGVHD. Efficacy was dependent upon CXCR5-sufficient Tregs homing to, and inhibiting, GC reactions. These studies indicate that the infusion of Tregs, especially ones enriched for GC homing, may be desirable for cGVHD therapy. Although IL-2/mAb complexes can be efficacious in cGVHD, a cautious approach needs to be taken in settings in which aGVHD elements, and associated Tcon, are present.
A Portrait of CXCR5 Follicular Cytotoxic CD8 T cells.
Yu Di,Ye Lilin
Trends in immunology
CD8 T cells differentiate into multiple effector and memory subsets to carry out immune clearance of infected and cancerous cells and provide long-term protection. Recent research identified a CXCR5Tcf1Tim-3 subset that localizes in, or proximal to, B cell follicles in secondary lymphoid organs of mice, non-human primates, and humans, hereby termed follicular cytotoxic T (T) cells. With remarkable similarity to follicular helper T (T) cells, T differentiation is dependent on transcription factors E2A, Bcl6, and Tcf1, but inhibited by other regulators, including Blimp1, Id2, and Id3. This review summarizes the phenotype, function, and differentiation of this new subset. Owing to its follicular location and self-renewal capability, we propose immunotherapeutic strategies to target T cells to potentially treat certain cancers and chronic infections such as HIV-1.
CXCR5 polymorphisms in non-Hodgkin lymphoma risk and prognosis.
Charbonneau Bridget,Wang Alice H,Maurer Matthew J,Asmann Yan W,Zent Clive S,Link Brian K,Ansell Stephen M,Weiner George J,Ozsan Nazan,Feldman Andrew L,Witzig Thomas E,Cunningham Julie M,Dogan Ahmet,Habermann Thomas M,Slager Susan L,Novak Anne J,Cerhan James R
Cancer immunology, immunotherapy : CII
CXCR5 [chemokine (C-X-C motif) receptor 5; also known as Burkitt lymphoma receptor 1 (BCR1)] is expressed on mature B-cells, subsets of CD4+ and CD8+ T-cells, and skin-derived migratory dendritic cells. Together with its ligand, CXCL13, CXCR5 is involved in guiding B-cells into the B-cell zones of secondary lymphoid organs as well as T-cell migration. This study evaluated the role of common germline genetic variation in CXCR5 in the risk and prognosis of non-Hodgkin lymphoma (NHL) using a clinic-based study of 1,521 controls and 2,694 NHL cases including 710 chronic lymphocytic leukemia/small lymphocytic lymphoma, 586 diffuse large B-cell lymphoma (DLBCL), 588 follicular lymphoma (FL), 137 mantle cell lymphoma (MCL), 230 marginal zone lymphoma (MZL), and 158 peripheral T-cell lymphoma (PTCL). Of the ten CXCR5 tag SNPs in our study, five were associated with risk of NHL, with rs1790192 having the strongest association (OR 1.19, 95% CI 1.08-1.30; p = 0.0003). This SNP was most strongly associated with the risk of FL (OR 1.44, 95 % CI 1.25-1.66; p = 3.1 × 10(-7)), with a lower degree of association with DLBCL (OR 1.16, 95% CI 1.01-1.33; p = 0.04) and PTCL (OR 1.29, 95 % CI 1.02-1.64; p = 0.04) but no association with the risk of MCL or MZL. For FL patients that were observed as initial disease management, the number of minor alleles of rs1790192 was associated with better event-free survival (HR 0.64; 95% CI 0.47-0.87; p = 0.004). These results provide additional evidence for a role of host genetic variation in CXCR5 in lymphomagenesis, particularly for FL.
Differential G protein subunit expression by prostate cancer cells and their interaction with CXCR5.
El-Haibi Christelle P,Sharma Praveen,Singh Rajesh,Gupta Pranav,Taub Dennis D,Singh Shailesh,Lillard James W
BACKGROUND:Prostate cancer (PCa) cell lines and tissues differentially express CXCR5, which positively correlate with PCa progression, and mediate PCa cell migration and invasion following interaction with CXCL13. However, the differential expression of G protein α, β, and γ subunits by PCa cell lines and the precise combination of these proteins with CXCR5 has not been elucidated. METHODS:We examined differences in G protein expression of normal prostate (RWPE-1) and PCa cell lines (LNCaP, C4-2B, and PC3) by western blot analysis. Further, we immunoprecipitated CXCR5 with different G protein subunits, and CXCR4, following CXCL13 stimulation. To investigate constitutive coupling of CXCR5 with CXCR4 and PAR-1 we performed invasion assay in PCa cells transfected with Gαq/i2 or Gα13 siRNA, following CXCL13 treatment. We also investigated Rac and RhoA activity by G-LISA activation assay in PCa cells following CXCL13/thrombin stimulation. RESULT:Of the 22 G proteins studied, Gαi1-3, Gβ1-4, Gγ5, Gγ7, and Gγ10 were expressed by both normal and PCa cell lines. Gαs was moderately expressed in C4-2B and PC3 cell lines, Gαq/11 was only present in RWPE-1 and LNCaP cell lines, while Gα12 and Gα13 were expressed in C4-2B and PC3 cell lines. Gγ9 was expressed only in PCa cell lines. Gα16, Gβ5, Gγ1-4, and Gγ13 were not detected in any of the cell lines studied. Surprisingly, CXCR4 co-immunoprecipitated with CXCR5 in PCa cell lines irrespective of CXCL13 treatment. We also identified specific G protein isoforms coupled to CXCR5 in its resting and active states. Gαq/11/Gβ3/Gγ9 in LNCaP and Gαi2/Gβ3/Gγ9 in C4-2B and PC3 cell lines, were coupled to CXCR5 and disassociated following CXCL13 stimulation. Interestingly, Gα13 co-immunoprecipitated with CXCR5 in CXCL13-treated, but not in untreated PCa cell lines. Inhibition of Gαq/i2 significantly decreased the ability of cells to invade, whereas silencing Gα13 did not affect CXCL13-dependent cell invasion. Finally, CXCL13 treatment significantly increased Rac activity in Gαq/i2 dependent manner, but not RhoA activity, in PCa cell lines. CONCLUSIONS:These findings offer insight into molecular mechanisms of PCa progression and can help to design some therapeutic strategies involving CXCR5 and/or CXCL13 blockade and specific G protein inhibition to abrogate PCa metastasis.
Association of peripheral CD4+ CXCR5+ T cells with chronic lymphocytic leukemia.
Cha Zhanshan,Zang Yan,Guo Huijun,Rechlic James R,Olasnova Lindsay M,Gu Haihui,Tu Xiaohua,Song Haihan,Qian Baohua
Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine
Accumulating evidences indicate that immune dysregulation plays a key role in both lymphomagenesis and patient outcome of chronic lymphocytic leukemia (CLL). Peripheral blood CD4+ CXCR5+ T cells, known as circulating follicular helper T cells (Tfh), can induce B cell activation and production of specific antibody responses. The aim of the study was to investigate changes of circulating Tfh in CLL. Tfh and it subtypes were tested by measuring CD4, CXCR5, CXCR3, and CCR6 in 72 CLL cases and 86 healthy controls using flow cytometry. Data showed that the percentage of Tfh in the peripheral CD4+ T cells was significantly increased in CLL (25.1%) than in controls (8.4%) (p < 0.001). Further analysis revealed that the upregulation of Tfh was contributed by Tfh-th2 subtype and Tfh-th17 subtype. Investigating staging of the cases demonstrated that the prevalence of Tfh was significantly elevated in cases with Binet stage C (37.3%) than those with stage A (20.1 %) or stage B (23.9 %). In addition, we analyzed Tfh in patients with immunoglobulin variable heavy chain (IGHV) gene mutational status. Results presented that Tfh-th17 subtype had clearly higher frequency in patients with IGHV mutation compared to the unmutated cases (p = 0.035). This study suggested the involvement of Tfh in the pathogenesis and progression of CLL, and provided a potential target for treating this disease.
CXCR5 and ICOS expression identifies a CD8 T-cell subset with T features in Hodgkin lymphomas.
Le Kieu-Suong,Amé-Thomas Patricia,Tarte Karin,Gondois-Rey Françoise,Granjeaud Samuel,Orlanducci Florence,Foucher Etienne D,Broussais Florence,Bouabdallah Reda,Fest Thierry,Leroux Dominique,Yadavilli Sapna,Mayes Patrick A,Xerri Luc,Olive Daniel
A better characterization of T-cell subsets in the microenvironment of classical Hodgkin lymphoma (cHL) would help to develop immunotherapies. Using multicolor flow cytometry, we identified in 6 of 43 cHL tissue samples a previously unrecognized subset of CD8 T cells coexpressing CXCR5 and inducible T-cell costimulator (ICOS) molecules (CD8). These cells shared phenotypic features with follicular helper T (T) cells including low CCR7 expression together with high expression of B-cell lymphoma-6, programmed cell death 1, B and T lymphocyte attenuator, CD200, and OX40. They had deficient cytotoxicity, low interferon-γ secretion, and common functional properties with intratumoral CD4 T cells, such as production of interleukin-4 (IL-4), IL-21, CXCL13, and capacity to sustain B cells. Gene profiling analysis showed a significant similarity between the signatures of CD8 T cells and CD4 T cells. Benign lymphadenitis tissues (n = 8) were devoid of CD8 cells. Among the 35 B-cell lymphoma tissues analyzed, including follicular lymphomas (n = 13), diffuse large cell lymphomas (n = 12), marginal zone lymphomas (MZLs; n = 3), mantle cell lymphomas (n = 3), and chronic lymphocytic leukemias (n = 4), only 1 MZL sample contained CD8 cells. Lymphoma tumors with CD8 cells shared common histopathological features including residual germinal centers, and contained high amounts of activated CD8 cells. These data demonstrate a CD8 T-cell differentiation pathway leading to the acquisition of some T similarities. They suggest a particular immunoediting process with global CD8 activation acting mainly, but not exclusively, in HL tumors.
Expression features of CXCR5 and its ligand, CXCL13 associated with poor prognosis of advanced colorectal cancer.
Qi X-W,Xia S-H,Yin Y,Jin L-F,Pu Y,Hua D,Wu H-R
European review for medical and pharmacological sciences
OBJECTIVE:CXCL13 plays a unique role in the trafficking and homing of B1 cells associated with its cognate receptor, CXCR5. The CXCR5-CXCL13 axis has been previously demonstrated to be a poor prognosis factor in malignancies. However, the clinical significance of the CXCR5-CXCL13 expression in colorectal cancer carcinoma (CRC) remains unclear. The aim of this study was to investigate the CXCR5-CXCL13 expression in CRC and determine its correlation with the progression and prognosis of the tumor. PATIENTS AND METHODS:A total of 144 paraffin-embedded specimens with advanced colon cancer were assessed for CXCR5 and CXCL13 by immunohistochemistry. Patients' long-term survival was also monitored. There were significant differences in lymph node metastasis (p = 0.0066), neural invasion (p = 0.0061) and neural invasion (p = 0.0001) between high and low expression of CXCR5. RESULTS:There were significant differences in distant metastasis (p = 0.0261), TNM stage (p = 0.0409), differentiation (p < 0.0001) and neural invasion of the CXCL13. Both CXCR5 and CXCL13 was associated with poor correlation with the overall survival (OS) and relapse-free survival (RFS). CONCLUSIONS:Our data suggest that the CXCR5 and CXCL13 may play a crucial role in the development, metastasis and relapse of advanced colon cancer. They can be used as prognostic markers of colon cancer in clinical practice.
p53 Deletion promotes myeloma cells invasion by upregulating miR19a/CXCR5.
Yue Zhijie,Zhou Yongxia,Zhao Pan,Chen Yafang,Yuan Ying,Jing Yaoyao,Wang Xiaofang
P53 deletion has been identified as one of the few factors that defined high risk and poor prognosis in MM. It has been reported p53 deletion is associated with resistance to chemotherapy and organ infiltrations of MM. However, p53 deletion in the migration and dissemination of MM cells has not been totally elucidated. In this research, first, we investigated whether p53 is associated with migration of MM cells. We found that p53 regulates the migration of NCI-H929 cells with wild-type p53 but not U266 cells with mutated-type p53. Next, we investigated the related mechanism by which p53 regulates the migration. We found that down-regulation of p53 reduced adhesion of NCI-H929 cells to the BM stroma via decreased expression of E-cadherin and increased EMT-regulating proteins. Further study have identified the miR-19a/CXCR5 pathway as a candidate p53-induced migration mechanism. In conclusion, we have demonstrated for the first time the critical value of p53 deletion in MM cell migration and dissemination, as well as the acquisition of an EMT-like phenotype. Our research provides new insights into the function of p53 in migration of MM and suggests p53/miRNA19a/CXCR5 may provide potentially therapeutic targets for the treatment of myeloma with p53 deletion.
Upregulation of bacterial-specific Th1 and Th17 responses that are enriched in CXCR5CD4 T cells in non-small cell lung cancer.
Ma Qin-Yun,Huang Da-Yu,Zhang Hui-Jun,Wang Shaohua,Chen Xiao-Feng
The microbial community in the mucosal surfaces is involved in the development of human cancers, including gastric cancer and colorectal cancer. The respiratory tract in the lung also hosts a distinctive microbial community, but the correlation between this community and lung cancer is largely unknown. Here, we examined the Th1 and Th17 responses toward several bacterial antigens, in CD4 T cells sourced from the peripheral blood (PB), the lung cancer (LC) tissue, and the gastrointestinal (GI) tract of non-small cell lung cancer (NSCLC) patients. Compared to healthy controls, the NSCLC patients presented significantly higher frequencies of Th1 and Th17 cells reacting to Streptococcus salivarius and S. agalactiae, in the PB, LC, and GI tract. Further investigation showed that the upregulation in anti-bacteria response was likely antigen-specific for two reasons. Firstly, the frequencies of Th1 and Th17 cells reacting to Escherichia coli, a typical GI bacterium, were not upregulated in the PB and the LC of NSCLC patients. Secondly, the S. salivarius and S. agalactiae responses could be partially blocked by Tü39, a MHC class II blocking antibody, suggesting that antigen-specific interaction between CD4 T cells and antigen-presenting cells was required. We also found that S. salivarius and S. agalactiae could potently activate the monocytes to secrete higher levels of interleukin (IL)-6, IL-12, and tumor necrosis factor, which were Th1- and Th17-skewing cytokines. Interestingly, whereas CXCR5CD4 T cells represented <20% of total CD4 T cells, they represented 17%-82% of bacteria-specific Th1 or Th17 cells. Together, these data demonstrated that NSCLC patients presented a significant upregulation of bacterial-specific Th1 and Th17 responses that were enriched in CXCR5CD4 T cells.
Upregulation of peripheral CD4+CXCR5+ T cells in osteosarcoma.
Xiao Hong,Luo Gang,Son Haihang,Zhou Yue,Zheng Wenjie
Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine
Immune dysregulation plays a key role in the development of osteosarcoma (OS). Peripheral blood CD4+CXCR5+ T cells can induce B-cell activation and produce various cytokines and therefore may play critical roles in tumorigenesis. The purpose of the study was to investigate changes of peripheral CD4+CXCR5+ T cells in OS. Peripheral CD4+CXCR5+ T cells and its subtypes were determined by measuring CD3, CD4, CXCR5, CXCR3, and CCR6 in 38 OS patients and 42 healthy controls using flow cytometry. Data demonstrated that percentage of peripheral CD4+CXCR5+ T cells was significantly increased in OS patients (13.9 %) than in controls (8.6 %, p<0.001). Further analysis identified a profound skewing of peripheral CD4+CXCR5+ T cell subsets toward Th2 and Th17 cells in OS patients. Investigating clinical status of the patients showed that prevalence of peripheral CD4+CXCR5+ T cells was significantly elevated in cases with metastasis (17.4 %) than those without metastasis (12.7 %). Similarly, patients with high tumor grade revealed increased percentage of CD4+CXCR5+ T cells compared to those with low tumor grade (15.3 versus 11.0 %). Interestingly, the upregulation of peripheral CD4+CXCR5+ T cells in patients with metastasis or high tumor grade was contributed by Th1 and Th17 subtypes. This study suggests the involvement of peripheral CD4+CXCR5+ T cells in the pathogenesis and progression of OS and provides novel knowledge for understanding this disease.
Expression and clinical significance of CXCR5/CXCL13 in human non‑small cell lung carcinoma.
Singh Rajesh,Gupta Pranav,Kloecker Goetz H,Singh Shailesh,Lillard James W
International journal of oncology
CXCR5 and/or CXCL13 expression is elevated in certain carcinomas and lymphomas. To determine if these factors are involved in progression of non-small cell lung cancer (LuCa), we evaluated their expression in patients with various forms of this disease. Lung biopsies from patients with non-neoplastic cells (n=8), squamous cell carcinoma (SCC; n=24), or adenocarcinoma (AC; n=54) were stained for CXCR5. Histopathological analysis of these samples showed significantly higher expression of CXCR5 (p<0.001) in carcinomas (i.e., SCCs and ACs) relative to non‑neoplastic lung tissue. Nuclear and membrane CXCR5 intensities were highest in ACs, with median values of 185 and 130, respectively, followed by SCCs with median values of 170 and 110, respectively. The lowest nuclear and membrane expressions of CXCR5 were found in non-neoplastic tissues, having median values of 142 and 90, respectively. Sera from SCC patients (n=17), AC patients (n=14), and healthy controls (n=9) were tested for the presence of CXCL13. Serum CXCL13 levels in LuCa patients were higher than in healthy controls. CXCR5 expression in cell lines of human non-small cell lung carcinoma (NCI-H1915) and small cell lung carcinoma (SW-1271) were evaluated by flow cytometry. CXCR5 expression was higher in NCI-H1915 cells relative to SW-1271 cells. The functional significance of CXCR5 expression was tested in a migration assay. In response to CXCL13, more NCI-H1915 cells migrated than SW-1271 cells. These findings suggest that the CXCR5‑CXCL13 axis influences LuCa progression. After validation in larger patient groups, CXCR5 and CXCL13 may prove useful as biomarkers for LuCa. Correspondingly, blockade of this axis could serve as an effective therapy for LuCa.
CXCR5 guides migration and tumor eradication of anti-EGFR chimeric antigen receptor T cells.
Li Guangchao,Guo Jintao,Zheng Yanfang,Ding Wen,Han Zheping,Qin Lingyu,Mo Wenjun,Luo Min
Molecular therapy oncolytics
The efficacy of chimeric antigen receptor (CAR) T is still not optimal for solid tumors, partly due to the lack of T cell infiltration to the tumor site. One promising strategy is to guide T cells through tumor-specific chemokines, provided that the matching chemokine receptors are expressed on T cells. Previous reports showed that, for non-small cell lung cancer (NSCLC) patients, the tumor sites express high levels of chemokine CXCL13, whereas CXCR5, the only receptor for CXCL13, is mainly expressed on B cells and follicle helper T cells. Therefore, we engineered an epidermal growth factor receptor (EGFR) CAR-T cell to express a second receptor CXCR5, to facilitate migration of CAR-T cells to the CXCL13-expressing NSCLC tumors, and to minimize EGFR-CAR-T possible off-tumor, on-target toxicity. We first confirmed CXCL13 expression in NSCLC patient blood and cancer tissues and the absence of CXCR5 expression in normal CD3 T cells. Next, we demonstrated that EGFR-CXCR5-CAR-T cells have similar killing activity as EGFR-CAR-T with a cytotoxicity assay . Furthermore, the Transwell assay and xenograft tumor mouse model were used to confirm that EGFR-CXCR5-CAR-T exhibits a significant increase in T cell infiltration to CXCL13-expressing tumors and eradicates the CXCL13-expressing tumors more efficiently.
Proteomic analysis reveals a FANCA-modulated neddylation pathway involved in CXCR5 membrane targeting and cell mobility.
Renaudin Xavier,Guervilly Jean-Hugues,Aoufouchi Said,Rosselli Filippo
Journal of cell science
The aim of this study was to identify novel substrates of the FANCcore complex, the inactivation of which leads to the genetic disorder Fanconi anemia, which is associated with bone marrow failure, developmental abnormalities and a predisposition to cancer. Eight FANC proteins participate in the nuclear FANCcore complex, which functions as an E3 ubiquitin-ligase that monoubiquitylates FANCD2 and FANCI in response to replicative stress. Here, we use mass spectrometry to compare proteins from FANCcore-complex-deficient cells to those of rescued control cells after treatment with hydroxyurea, an inducer of FANCD2 monoubiquitylation. FANCD2 and FANCI appear to be the only targets of the FANCcore complex. We identify other proteins that are post-translationally modified in a FANCA- or FANCC-dependent manner. The majority of these potential targets localize to the cell membrane. Finally, we demonstrate that (a) the chemokine receptor CXCR5 is neddylated; (b) FANCA but not FANCC appears to modulate CXCR5 neddylation through an unknown mechanism; (c) CXCR5 neddylation is involved in targeting the receptor to the cell membrane; and (d) CXCR5 neddylation stimulates cell migration and motility. Our work has uncovered a pathway involving FANCA in neddylation and cell motility.
Contribution of FoxP3 Tfr cells to overall human blood CXCR5 T cells.
Fonseca V R,Graca L
Clinical and experimental immunology
The identification that T follicular helper (Tfh) cells is critical for the emergence of germinal centre responses prompted the study of CXCR5-expressing CD4 T cell subsets in autoimmunity. However, circulating CXCR5-expressing T cells are heterogeneous by containing Forkhead box protein 3 (FoxP3) T follicular regulatory (Tfr) cells in addition to bona fide Tfh cells. Such heterogeneity may hamper the analysis of the contribution of specific follicular T cell subsets for autoimmune pathogenesis. Therefore, separate assessment of Tfh and Tfr populations offer greater opportunities for stratification of autoimmune patients, such as Sjögren's syndrome patients.
Interferon-γ suppresses the proliferation and migration of human placenta-derived mesenchmal stromal cells and enhances their ability to induce the generation of CD4CXCR5Foxp3Treg subset.
Yi Jun-Zhu,Chen Zheng-Hua,Xu Feng-Huang,Wang Zhuo-Ya,Zhang Hong-Qin,Jiang Guo-Sheng,Luan Xi-Ying
We investigate the effects of interferon (IFN)-γ on human placenta-derived mesenchymal stromal cells (hPMSCs), in particular, their adhesion, proliferation and migration and modulatory effects on the CD4CXCR5Foxp3Treg subset. And we compared hPMSCs ability to induce the generation of different Treg subsets in response to treatment with IFN-γ. We found that IFN-γ suppressed the proliferation and migration for hPMSCs. The ability of hPMSCs to induce the generation of CD4CXCR5Foxp3Treg subset was enhanced by IFN-γ. And maximal effectiveness of IFN-γ treated hPMSCs upon inducing the generation of Treg subsets was for CD4CXCR5Foxp3Treg subset as compared with that of CD4CD25Foxp3, CD8CD25Foxp3, CD4IL-10 and CD8IL-10Treg subsets. These results have important implications for the development and application of hPMSCs in clinical use.
Immunotherapy of B-cell non-Hodgkin lymphoma by targeting the chemokine receptor CXCR5 in a preclinical mouse model.
Panjideh Hossein,Müller Gerd,Koch Mathias,Wilde Frank,Scheu Susanne,Moldenhauer Gerhard,Lipp Martin
International journal of cancer
Bispecific antibodies are promising agents for immunotherapy. Here, we describe a quadroma-based trifunctional bispecific antibody binding the chemokine receptor CXCR5 and the T-cell antigen CD3 that efficiently prevents tumor growth in a mouse B-cell lymphoma model. CXCR5 regulates the tissue homeostasis of mature B cells and is highly expressed on B-cell non-Hodgkin and lymphocyte-predominant Hodgkin lymphoma, as well as on a subset of CD4(+) T cells known as follicular T-helper cells. In vitro, the bispecific CXCR5::CD3 antibody efficiently recruited effector T cells to CXCR5 expressing B cells and induced a co-stimulation-independent activation of CD8(+) and CD4(+) T cells as demonstrated by the de novo expression of CD25 and CD69, and secretion of the cytokines IFN-γ, TNF-α, IL-6 and IL-10 by peripheral blood mononuclear cells. Notably, at low antibody concentrations, CXCR5::CD3 displayed a significantly higher cytotoxic activity against autologous B cells than its parental antibodies or rituximab. In vivo imaging revealed that CXCR5::CD3 and its parental CXCR5 antibody efficiently prevent tumor growth in a xenograft model of B-cell lymphoma in mice and prolong their survival. Taken together, our results identify CXCR5 as a promising target for antibody-based therapies in the treatment of B-cell malignancies.
CXCR5-CXCL13 axis markers in full-term and preterm human neonates in the first weeks of life.
Pietrasanta Carlo,De Leo Pasqualina,Jofra Tatiana,Ronchi Andrea,Pugni Lorenza,Mosca Fabio,Aiuti Alessandro,Cicalese Maria Pia,Fousteri Georgia
European journal of immunology
Term and preterm neonates have very few circulating Tfh-like cells (cTfh), and no circulating Tfr-like cells. Neonatal cTfh are CXCR5 PD-1 CD45RA , suggestive of a naive, possibly recently activated phenotype. CXCL13 is high at birth, but decreases rapidly in the first weeks of life. Overall, signs of GC activity in human neonates are weak, even in those born prematurely or after sepsis.
IRF5 is a novel regulator of CXCL13 expression in breast cancer that regulates CXCR5(+) B- and T-cell trafficking to tumor-conditioned media.
Pimenta Erica Maria,De Saurav,Weiss Ryan,Feng Di,Hall Kelly,Kilic Sarah,Bhanot Gyan,Ganesan Shridar,Ran Sophia,Barnes Betsy J
Immunology and cell biology
Clinical studies using prognostic and predictive signatures have shown that an immune signal emanating from whole tumors reflects the level of immune cell infiltration--a high immune signal linked to improved outcome. Factors regulating immune cell trafficking to the tumor, however, are not known. Previous work has shown that expression of interferon regulatory factor 5 (IRF5), a critical immune regulator, is lost in ~80% of invasive ductal carcinomas examined. We postulated that IRF5-positive and -negative breast tumors would differentially regulate immune cell trafficking to the tumor. Using a focused tumor inflammatory array, differences in cytokine and chemokine expression were examined between IRF5-positive and -negative MDA-MB-231 cells grown in three-dimensional culture. A number of cytokines/chemokines were found to be dysregulated between cultures. CXCL13 was identified as a direct target of IRF5 resulting in the enhanced recruitment of B and T cells to IRF5-positive tumor-conditioned media. The ability of IRF5 to regulate mediators of cell migration was confirmed by enzyme-linked immunosorbent assay, chromatin immunoprecipitation assay, small interfering RNA knockdown and immunofluorescence staining of human breast tumor tissues. Analysis of primary immune cell subsets revealed that IRF5 specifically recruits CXCR5(+) B and T cells to the tumor; CXCR5 is the receptor for CXCL13. Analysis of primary breast tumor tissues revealed a significant correlation between IRF5 and CXCL13 expression providing clinical relevance to the study. Together, these data support that IRF5 directly regulates a network of genes that shapes a tumor immune response and may, in combination with CXCL13, serve as a novel prognostic marker for antitumor immunity.
Identification of CXCL13/CXCR5 axis's crucial and complex effect in human lung adenocarcinoma.
Tian Chen,Li Chang,Zeng Yulan,Liang Jinyan,Yang Qifan,Gu Feifei,Hu Yue,Liu Li
Immune escape and low response to immunotherapy are crucial challenges in present lung cancer treatment. In this study, we constructed a new immune-related classifier based on CXCL13/CXCR5, an important tumor microenvironment component and strongly related with the formation of tertiary lymphoid structures (TLSs) in tumor microenvironment. With the classifier, we divided patients into two main clusters and each cluster was further divided into subcluster (A1, A2, B1, B2, B3). In the later analysis, we noticed that patients in subcluster B3 had a distinct advantage over patients in A1 in survival time and immune infiltration, suggesting a more favorable response to immunotherapy. Moreover, we demonstrated the genetic and epigenetic regulation related to the subclusters and recovered four key differentially expressed genes (ERBB4, GRIN2A, IL2RA, CCND2). With several experiments, we verified the unique role of CCND2 in tumor metastasis and T cell apoptosis. Overexpressing CCND2 could significantly impair cancer cell abilities of migration and invasion and downregulate PD-1/PD-L1 signaling, which may be the cause of T cell apoptosis reduction. In the end, we constructed a regression risk model that could successfully predict ICI response. To sum up, our study established new stratification models that can successfully predict patient survival and response to ICI. And using integrative analysis of multi-omics data, four key DEGs were noticed, and CCND2, one of the four genes, was identified as a potential treatment target because of its effect in tumor metastasis and T cell apoptosis.
Circulating CXCR5+CD4+ T cells assist in the survival and growth of primary diffuse large B cell lymphoma cells through interleukin 10 pathway.
Cha Zhanshan,Qian Guangfang,Zang Yan,Gu Haihui,Huang Yanyan,Zhu Lishuang,Li Jinqi,Liu Yang,Tu Xiaohua,Song Haihan,Qian Baohua
Experimental cell research
Diffuse large B cell lymphoma (DLBCL) is a common and aggressive cancer caused by the malignant transformation of B cells. Although it has been established that the follicular helper T (Tfh) cells play a central role in B cell development, little information is available on their involvement in DLBCL pathogenesis. We studied the role of the peripheral Tfh equivalent, the CXCR5 CD4 T cells, in DLBCL. Data showed that compared to CXCR5 CD4 T cells, CXCR5 CD4 T cells were significantly more effective at promoting the proliferation as well as inhibiting the apoptosis of primary autologous DLBCL tumor cells. Surprisingly, we found that at equal cell numbers, CXCR5 CD4 T cells in DLBCL patients secreted significantly less interleukin (IL)-21 than CXCR5 CD4 T cells, while the level of IL-10 secretion was significant elevated in the CXCR5 compartment compared to the CXCR5 compartment. Neutralization of IL-10 in the primary DLBCL-CXCR5 CD4 T cell coculture compromised the CXCR5 CD4 T cell-mediated pro-tumor effects, in a manner that was dependent on the concentration of anti-IL-10 antibodies. The CXCR5 compartment also contained significantly lower frequencies of cytotoxic CD4 T cells than the CXCR5 compartment. In conclusion, our investigations discovered a previously unknown pro-tumor role of CXCR5-expressing circulating CD4 T cells, which assisted the survival and proliferation of primary DLBCL cells through IL-10.
The Expression and Significance of CXCR5 and MMP-13 in Colorectal Cancer.
Yan Qin,Yuan Yin,Yankui Liu,Jingjie Feng,Linfang Jin,Yong Pu,Dong Hua,Xiaowei Qi
Cell biochemistry and biophysics
The objective of the study is to investigate the CXCR5 and MMP-13 expression in colorectal cancer and explore its correlation between the clinicopathological characteristics and prognosis. The expressions of CXCR5 and MMP-13 proteins in 236 paired specimens of colorectal cancer and incisal edge normal tissues as well as 62 samples of colorectal adenoma tissues were analyzed by immunohistochemistry. The CXCR5 and MMP-13 positive expression rate in colorectal cancer tissues was 43.6 and 80.5 %, respectively. Both rates were higher than those in the incisal edge healthy intestinal mucosal tissues (4.2 and 13.1 %) and colorectal adenoma tissues (24.2 and 64.5 %), P < 0. 05 in both cases. The expressions of the CXCR5 and MMP-13 proteins were positively related to the lymph node and distal metastasis, tumor stage and relapse, P < 0. 05. The expression of the CXCR5 protein was positively related to MMP-13, P < 0. 05. The median and overall survival in the patients with positive CXCR5 and MMP-13 expression were significantly shorter than those with negative expression: median survival, 20.5 months (CXCR5 +) versus 30.8 months (CXCR5 -), 20.3 months (MMP-13 +) versus 24.6 months; overall survival, 26.5 months (CXCR5 +) versus 47.5 months (CXCR5 -), 22.7 months (MMP-13 +) versus 29.3 months. The expression of CXCR5 and MMP-13 could promote the pathogenesis, development, metastasis, and relapse of colorectal cancer. It could also serve as a valuable indicator for the prediction of metastasis and relapse of colorectal cancer.
CD40 controls CXCR5-induced recruitment of myeloid-derived suppressor cells to gastric cancer.
Ding Yixin,Shen Jin,Zhang Guangbo,Chen Xiaojuan,Wu JiaMing,Chen Weichang
To explore the mechanisms of MDSC trafficking and accumulation during tumor progression. In this study, we report significant CD40 upregulation in tumor-infiltrating MDSC when compared with splenic MDSC. Microarray analyses comparing CD40(high) and CD40l(ow) MDSC revealed 1872 differentially expressed genes, including CD83, CXCR5, BTLA, CXCL9, TLR1, FLT3, NOD2 and CXCL10. In vivo experiments comparing wild-type (WT) and CD40 knockout (KO) mice demonstrated that CD40 critically regulates CXCR5 expression. Consistently, the transwell analysis confirmed the essential role of CXCR5-CXCL13 crosstalk in the migration of CD40+ MDSC toward gastric cancer. Furthermore, more MDSC accumulated in the gastric cancers of WT mice when compared with KO mice, and the WT tumors mostly contained CD40+ cells. Functionally, tumors grew faster in WT than KO mice. In conclusion, we demonstrate that CD40 expression upregulates the chemokine receptor CXCR5 and promotes MDSC migration toward and accumulation within cancer. Therefore, this study provides preliminary evidence that CD40 may stimulate tumor growth by enabling immune evasion via MDSC recruitment and inhibition of T cell expansion.
The prevalence and function of CD4CXCR5Foxp3 follicular regulatory T cells in diffuse large B cell lymphoma.
Cha Zhanshan,Gu Haihui,Zang Yan,Wang Zi,Li Jinqi,Huang Weihua,Qin Aihua,Zhu Lishuang,Tu Xiaohua,Cheng Ning,Song Haihan,Qian Baohua
CD4CXCR5Foxp3 follicular regulatory T (Tfr) cells possess critical roles in suppressing the germinal center reaction, B cell activation, and follicular helper T cell (Tfh) cytokine secretion. Since diffuse large B cell lymphoma (DLBCL) can arise from B cells undergoing germinal center reaction and/or differentiation, we hypothesized that Tfr cells might be involved in DLBCL. In the present study, we recruited thirty-five DLBCL patients and twenty-five healthy controls. Data showed that DLBCL patients presented an enrichment of circulating CD4CXCR5Foxp3 Tfr cells compared to controls. In the primary tumor isolated from enlarged lymph nodes, Tfr cells made up of roughly 3% to 16% of infiltrating T cells. Higher levels of tumor-infiltrating Tfr cells were observed in patients with less advanced DLBCL stages, and in patients that stayed in remission 24 months after the initial R-CHOP treatment. High BCL6 and high FOXP3 expression was observed in Tfr cells ex vivo. After anti-CD3/CD28 and IL-2 stimulation, the Tfr cells more closely resembled Treg cells and presented high IL10 and TGFB1 expression. CD4CD25CXCR5 Tfr cells and CD4CD25CXCR5 non-Tfr Treg cells could suppress CD4CD25 Tconv cell and CD8 T cell proliferation with similar capacity. However, Tfr cells were less capable of suppressing IFNG expression than Treg cells, and although both cell types supported CD19 tumor cell proliferation, Tfr cells were less supportive than the non-Tfr Treg cells. Overall, this study suggested that Tfr cells were involved in intratumoral immunity, were likely beneficial to DLBCL patients, and were functionally distinctive from non-Tfr Treg cells. The distribution pattern and the prognostic value of Tfr cells in DLBCL should be examined in further studies.
Exosomal miR-214-5p Released from Glioblastoma Cells Modulates Inflammatory Response of Microglia after Lipopolysaccharide Stimulation through Targeting CXCR5.
Yang Jian-Kai,Liu Hong-Jiang,Wang Yuanyu,Li Chen,Yang Ji-Peng,Yang Liang,Qi Xue-Jiao,Zhao Yin-Long,Shi Xue-Fang,Li Jing-Chen,Sun Guo-Zhu,Jiao Bao-Hua
CNS & neurological disorders drug targets
BACKGROUND AND OBJECTIVE:Exosomes communicate inter-cellularly and miRNAs play critical roles in this scenario. MiR-214-5p was implicated in multiple tumors with diverse functions uncovered. However, whether miR-214-5p is mechanistically involved in glioblastoma, especially via exosomal pathway, is still elusive. Here we sought to comprehensively address the critical role of exosomal miR-214-5p in glioblastoma (GBM) microenvironment. METHODS:The relative expression of miR-214-5p was determined by real-time PCR. Cell viability and migration were measured by MTT and transwell chamber assays, respectively. The secretory cytokines were measured with ELISA kits. The regulatory effect of miR-214-5p on CXCR5 expression was interrogated by luciferase reporter assay. Protein level was analyzed by Western blot. RESULTS:We demonstrated that miR-214-5p was aberrantly overexpressed in GBM and associated with poorer clinical prognosis. High level of miR-214-5p significantly contributed to cell proliferation and migration. GBM-derived exosomal miR-214-5p promoted inflammatory response in primary microglia upon lipopolysaccharide challenge. We further identified CXCR5 as the direct target of miR-214- 5p in this setting. CONCLUSION:Overexpression of miR-214-5p in GBM modulated the inflammatory response in microglia via exosomal transfer.
CXCL13 inhibition induce the apoptosis of MDA-MB-231 breast cancer cells through blocking CXCR5/ERK signaling pathway.
Ma J-J,Jiang L,Tong D-Y,Ren Y-N,Sheng M-F,Liu H-C
European review for medical and pharmacological sciences
OBJECTIVE:Treatment of the high-risk triple negative breast cancer (TNBC) is a critical clinical challenge. Here we aimed to explore a novel strategy for TNBC treatment by blocking the tumor-associated chemokine CXCL13 in the MDA-MB-231 TNBC cells. MATERIALS AND METHODS:MDA-MB-231 cells were treated with anti-CXCL13 antibodies (inhibition group), or phosphate-buffered saline (PBS) (control group), followed by determining the levels of interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF-α) and transforming growth factor beta-1 (TGF-β1) with enzyme-linked immunosorbent assay (ELISA). The effects of CXCL13 inhibition on cell proliferation and apoptosis were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and flow cytometry, respectively. Quantitative Real Time-PCR (qRT-PCR) and Western blot were used to compare the levels of CXCL13, CXCR5, extracellular signal-regulated kinase (ERK). The levels of cyclin D1 and cleaved caspase-9 were detected by Western blot. RESULTS:The levels of IL-1, TNF-α and TGF-β1 in MDA-MB-231 cells treated with anti-CXCL13 antibodies were significantly downregulated (p<0.05). Meanwhile, CXCL13 blockade decreased the cell proliferation and increased the apoptosis rate of MDA-MB-231 cells. The inhibition of CXCL13 led to marked reduction in CXCL13 and CXCR5 mRNA and an increase in ERK mRNA. The inhibition of CXCL13 resulted in the downregulation of CXCL13, CXCR5, p-ERK/ERK, cyclin D1 and upregulation of cleaved caspase-9 proteins. CONCLUSIONS:CXCL13 blockade effectively suppressed the proliferation of MDA-MB-231 cells by promoting cell apoptosis. This effect is presumably associated with the downregulation of CXCL13 and suppression of the CXCR5/ERK signaling pathway.
CXCR5CD8 T cells present elevated capacity in mediating cytotoxicity toward autologous tumor cells through interleukin 10 in diffuse large B-cell lymphoma.
Tang Jiahong,Zha Jie,Guo Xutao,Shi Pengcheng,Xu Bing
Diffuse large B-cell lymphoma (DLBCL) is a common and aggressive subtype of non-Hodgkin's lymphomas, with limited treatment options in refractory and relapsed patients. Growing evidence supports the notion that CD8 T cell immunity could be utilized to eliminate B cell lymphomas. CXCR5CD8 T cell is a novel cell subtype and share CXCR5 expression with CD19 tumor cells. In this study, we investigated the frequency and function of existing CXCR5CD8 T cells in DLBCL patients. We found that DLBCL patients as a group demonstrated significantly higher level of CXCR5CD8 T cells than healthy individuals, with huge variability in each patient. Using anti-CD3/CD28-stimulated CD8 T cells as effector (E) cells and autologous CD19 tumor cells as target (T) cells, at high E:T ratio, no difference between the intensities of CXCR5CD8 T cell- and CXCR5CD8 T cell-mediated cytotoxicity were observed. However, at intermediate and low E:T ratios, the CXCR5CD8 T cells presented stronger cytotoxicity than CXCR5CD8 T cells. The expressions of granzyme A, granzyme B, and perforin were significantly higher in CXCR5CD8 T cells than in CXCR5CD8 T cells, with no significant difference in the level of degranulation. Tumor cells in DLBCL were known to secrete high level of interleukin 10 (IL-10). We therefore blocked the IL-10/IL-10R pathway, and found that the expressions of granzyme A, granzyme B, and perforin by CXCR5CD8 T cells were significantly elevated. Together, these results suggest that CXCR5CD8 T cells are potential candidates of CD8 T cell-based immunotherapies, could mediate elimination of autologous tumor cells in DLBCL patients, but are also susceptible to IL-10-mediated suppression.
Follicular Regulatory T Cells Can Access the Germinal Center Independently of CXCR5.
Vanderleyden Ine,Fra-Bido Sigrid C,Innocentin Silvia,Stebegg Marisa,Okkenhaug Hanneke,Evans-Bailey Nicola,Pierson Wim,Denton Alice E,Linterman Michelle A
The germinal center (GC) response is critical for generating high-affinity humoral immunity and immunological memory, which forms the basis of successful immunization. Control of the GC response is thought to require follicular regulatory T (Tfr) cells, a subset of suppressive Foxp3 regulatory T cells located within GCs. Relatively little is known about the exact role of Tfr cells within the GC and how they exert their suppressive function. A unique feature of Tfr cells is their reported CXCR5-dependent localization to the GC. Here, we show that the lack of CXCR5 on Foxp3 regulatory T cells results in a reduced frequency, but not an absence, of GC-localized Tfr cells. This reduction in Tfr cells is not sufficient to alter the magnitude or output of the GC response. This demonstrates that additional, CXCR5-independent mechanisms facilitate Treg cell homing to the GC.
The Differential Expression and Function of the Inflammatory Chemokine Receptor CXCR5 in Benign Prostatic Hyperplasia and Prostate Cancer.
Yang Lu,Gao Liang,Chen Yongji,Tang Zhuang,Zhu Yuchun,Han Ping,Li Xiang,Wei Qiang
International journal of medical sciences
BACKGROUND:Chemokine and chemokine receptors could have played an important role in tumor angiogenesis and distant metastasis. The mechanism of inflammation, expression and function of chemokines and chemokine receptors in benign prostatic hyperplasia (BPH) and prostate cancer (PCa) remain unclear. The purpose of present study is to detect differential expression and function of chemokines and chemokine receptors (CCRs) in BPH and PCa. METHODS:BPH-1 and peripheral blood mononuclear cells (PBMCs) were co-cultured in Transwell chambers, and human normal prostate (NP) tissues, BPH tissues and PCa tissues were collected. CCR gene-chips were used to analyze and compare the differential expression of CCRs in BPH-1 cells, BPH-1 cells co-cultured with PBMCs, and LNCaP cells. The differential expression of CCRs was detected and validated using real-time PCR, western blotting and immunofluorescence (IF). The proliferation of LNCaP cells was also investigated after the knockdown CXCR5. RESULTS:RESULTS of gene-chips indicated that there was low or no expression of CCR10, CXCR1, CXCR3 and CXCR5 in BPH-1 cells, whereas the expression of these receptors in BPH-1 cells was increased by PBMCs, and the expression was high in LNCaP cells. Furthermore, real-time PCR and western blotting confirmed the above mentioned results. IF verified no or low expression of CXCR1, CXCR3 and CXCR5 in NP tissues, low or moderate expression in BPH and high expression in PCa. However, CCR10 was not expressed at detectable levels in the three groups. The growth and proliferation of LNCaP cells was markedly inhibited after down-regulation of CXCR5. CONCLUSIONS:PCa cells expressed high levels of CCR10, CXCR1, CXCR3 and CXCR5. Although BPH cells did not express these factors, their expression was up-regulated when BPH-1 cells were incubated with inflammatory cells. Finally, down-regulation of CXCR5 inhibited the growth and proliferation of LNCaP cells.
p63 and p73 repress CXCR5 chemokine receptor gene expression in p53-deficient MCF-7 breast cancer cells during genotoxic stress.
Mitkin Nikita A,Muratova Alisa M,Sharonov George V,Korneev Kirill V,Sviriaeva Ekaterina N,Mazurov Dmitriy,Schwartz Anton M,Kuprash Dmitry V
Biochimica et biophysica acta. Gene regulatory mechanisms
Many types of chemotherapeutic agents induce of DNA-damage that is accompanied by activation of p53 tumor suppressor, a key regulator of tumor development and progression. In our previous study we demonstrated that p53 could repress CXCR5 chemokine receptor gene in MCF-7 breast cancer cells via attenuation of NFkB activity. In this work we aimed to determine individual roles of p53 family members in the regulation of CXCR5 gene expression under genotoxic stress. DNA-alkylating agent methyl methanesulfonate caused a reduction in CXCR5 expression not only in parental MCF-7 cells but also in MCF-7-p53off cells with CRISPR/Cas9-mediated inactivation of the p53 gene. Since p53 knockout was associated with elevated expression of its p63 and p73 homologues, we knocked out p63 using CRISPR/Cas9 system and knocked down p73 using specific siRNA. The CXCR5 promoter activity, CXCR5 expression and CXCL13-directed migration in MCF-7 cells with inactivation of all three p53 family genes were completely insensitive to genotoxic stress, while pairwise p53+p63 or p53+p73 inactivation resulted in partial effects. Using deletion analysis and site-directed mutagenesis, we demonstrated that effects of NFkB on the CXCR5 promoter inversely correlated with p63 and p73 levels. Thus, all three p53 family members mediate the effects of genotoxic stress on the CXCR5 promoter using the same mechanism associated with attenuation of NFkB activity. Understanding of this mechanism could facilitate prognosis of tumor responses to chemotherapy.
Matrine reduces the secretion of exosomal circSLC7A6 from cancer-associated fibroblast to inhibit tumorigenesis of colorectal cancer by regulating CXCR5.
Gu Chen,Lu Huigang,Qian Zhenghai
Biochemical and biophysical research communications
Tumor microenvironment (e.g., stromal cells) has been suggested to be implicated in colorectal cancer (CRC) progression. Of which, cancer-associated fibroblasts (CAFs) are believed as one of the key stromal cells in tumors. Traditionally, matrine was used to treat cancers, including CRC. Unfortunately, little is known about whether matrine inhibited CRC progression via CAFs. In this research, we analyzed cell proliferation, invasion and apoptosis by cell colony formation assay, transwell assay and Annexin V staining, respectively. circSLC7A6 and CXCR5 expression levels were examined by RT-qPCR. Exosomes were analyzed by NanoSight Tracking Analysis and exosome markers were probed by westernblot. In the results, we found that matrine significantly led to inhibited cell proliferation and invasion, and increased apoptosis. Moreover, matrine blocked circSLC7A6 exosome secretion from CAFs. circSLC7A6 acted as promoter for CRC cell proliferation and invasion, whereas as inhibitor for apoptosis. Clinically, circSLC7A6 was upregulated in CRC tumor tissues compared to adjacent normal tissues. In addition, circSLC7A6 was associated with higher overall survival. Eventually, we confirmed that chemokine receptor CXCR5 was a crucial effector for circSLC7A6-modulated tumorigenesis. Here, our data suggest matrine inhibits CRC tumorigenesis through blocking exosomal circSLC7A6 release from CAFs. This finding will provide strong evidence for treating CRC using matrine.
Increased susceptibility to oral Trichuris muris infection in the specific absence of CXCR5 CD11c cells.
Bradford Barry M,Donaldson David S,Forman Ruth,Else Kathryn J,Mabbott Neil A
Trichuris muris is a natural mouse helminth pathogen which establishes infection specifically in the caecum and proximal colon. The rapid expulsion of T. muris in resistant mouse strains is associated with the induction of a protective T helper cell type 2 (Th2)-polarized immune response. Susceptible mouse strains, in contrast, mount an inappropriate Th1 response to T. muris infection. Expression of the chemokine CXCL13 by stromal follicular dendritic cells attracts CXCR5-expressing cells towards the B-cell follicles. Previous studies using a complex in vivo depletion model have suggested that CXCR5-expressing conventional dendritic cells (cDC) help regulate the induction of Th2-polarized responses. Here, transgenic mice with CXCR5 deficiency specifically restricted to CD11c cells were used to determine whether the specific absence CXCR5 on CD11c cells such as cDC would influence susceptibility to oral T. muris infection by affecting the Th1/Th2 balance. We show that in contrast to control mice, those which lacked CXCR5 expression on CD11c cells failed to clear T. muris infection and developed cytokine and antibody responses that suggested a disturbed Th1/Th2 balance with enhanced IFN-γ expression. These data suggest an important role of CXCR5-expressing CD11c cells such as cDC in immunity to oral T. muris infection.
CXCR5PD-1 follicular helper CD8 T cells control B cell tolerance.
Chen Yuhong,Yu Mei,Zheng Yongwei,Fu Guoping,Xin Gang,Zhu Wen,Luo Lan,Burns Robert,Li Quan-Zhen,Dent Alexander L,Zhu Nan,Cui Weiguo,Malherbe Laurent,Wen Renren,Wang Demin
Many autoimmune diseases are characterized by the production of autoantibodies. The current view is that CD4 T follicular helper (Tfh) cells are the main subset regulating autoreactive B cells. Here we report a CXCR5PD1 Tfh subset of CD8 T cells whose development and function are negatively modulated by Stat5. These CD8 Tfh cells regulate the germinal center B cell response and control autoantibody production, as deficiency of Stat5 in CD8 T cells leads to an increase of CD8 Tfh cells, resulting in the breakdown of B cell tolerance and concomitant autoantibody production. CD8 Tfh cells share similar gene signatures with CD4 Tfh, and require CD40L/CD40 and TCR/MHCI interactions to deliver help to B cells. Our study thus highlights the diversity of follicular T cell subsets that contribute to the breakdown of B-cell tolerance.
Loss of Cxcr5 alters neuroblast proliferation and migration in the aged brain.
Fritze Jonas,Ginisty Aurélie,McDonald Rebecca,Quist Ella,Stamp Eleanor,Monni Emanuela,Dhapola Parashar,Lang Stefan,Ahlenius Henrik
Stem cells (Dayton, Ohio)
Neurogenesis, the production of new neurons from neural stem cells, dramatically decreases during aging concomitantly with increased inflammation both systemically and in the brain. However, the precise role of inflammation and whether local or systemic factors drive the neurogenic decline during aging is poorly understood. Here, we identify CXCR5/5/CXCL13 signaling as a novel regulator of neurogenesis in the aged brain. The chemokine Cxcl13 was found to be upregulated in the brain during aging. Loss of its receptor, Cxcr5, led to increased proliferation and decreased numbers of neuroblasts in the aged subventricular zone (SVZ), together with accumulation of neuroblasts in the rostral migratory stream and olfactory bulb (OB), without increasing the amount of new mature neurons in the OB. The effect on proliferation and migration was specific to neuroblasts and likely mediated through increased levels of systemic IL-6 and local Cxcl12 expression in the SVZ. Our study raises the possibility of a new mechanism by which interplay between systemic and local alterations in inflammation regulates neurogenesis during aging.
CXCR5PD1ICOS Circulating T Follicular Helpers Are Associated With Donor-Specific Antibodies After Renal Transplantation.
Danger Richard,Chesneau Mélanie,Delbos Florent,Le Bot Sabine,Kerleau Clarisse,Chenouard Alexis,Ville Simon,Degauque Nicolas,Conchon Sophie,Cesbron Anne,Giral Magali,Brouard Sophie
Frontiers in immunology
Donor-specific anti-HLA antibodies (DSAs) are a major risk factor associated with renal allograft outcomes. As a trigger of B cell antibody production, T follicular helper cells (Tfhs) promote DSA appearance. Herein, we evaluated whether circulating Tfhs (cTfhs) are associated with the genesis of antibody-mediated rejection. We measured cTfh levels on the day of transplantation and 1 year after transplantation in blood from a prospective cohort of 237 renal transplantation patients without DSA during the first year post-transplantation. Total cTfhs were characterized as CD4CD45RACXCR5, and the three following subsets of activated cTfh were analyzed: CXCR5PD1, CXCR5PD1ICOS, an CXCR5PD1CXCR3. Immunizing events (previous blood transfusion and/or pregnancy) and the presence of class II anti-HLA antibodies were associated with increased frequencies of activated CXCR5PD1, CXCR5PD1ICOS, and CXCR5PD1CXCR3 cTfh subsets. In addition, ATG-depleting induction and calcineurin inhibitor treatments were associated with a relative increase of activated cTfh subsets frequencies at 1 year post-transplantation. In multivariate survival analysis, we reported that a decrease in activated CXCR5PD1ICOS at 1 year after transplantation in the blood of DSA-free patients was significantly associated with the risk of developing DSA after the first year ( = 0.018, HR = 0.39), independently of HLA mismatches ( = 0.003, HR = 3.79). These results highlight the importance of monitoring activated Tfhs in patients early after transplantation and show that current treatments cannot provide early, efficient prevention of Tfh activation and migration. These findings indicate the need to develop innovative treatments to specifically target Tfhs to prevent DSA appearance in renal transplantation.
Inverse relationship between CD40L expression and cytolytic molecule expression by CD8CXCR5 T follicular cytotoxic cells in colorectal cancer.
Xing Junjie,Li Xu,E Jifu,Wang Chen,Wang Hao
Experimental cell research
CXCR5 CD8 T cells, sometimes termed T follicular cytotoxic (Tfc) cells, are characterized by high proinflammatory cytokine and cytolytic molecule expression and low exhaustion and checkpoint molecule expression. Additionally, Tfc cells could promote B cell responses and support Ig release. It is yet unclear how Tfc cells could help B cells when they have the potential to mediate cytotoxicity at the same time. In this study, we found that Tfc cells expressed significantly higher levels of CD40L than non-Tfc CD8 T cells. Interestingly, Tfc cells from colorectal cancer (CRC) patients presented significantly higher CD40L than Tfc cells from healthy controls in a manner that was associated with CRC stage. Coincubation of Tfc cells and autologous B cells resulted in higher CD40L expression in a time-dependent manner. Interestingly, activated Tfc cells, when incubated with B cells, presented rapid downregulation of perforin and granzyme B. In general, greater than 50% of tumor-infiltrating Tfc cells expressed CD40L. In addition, the level of CD40L in tumor-infiltrating Tfc cells was higher in stage IV CRC patients than in stage II and stage III CRC patients. Interestingly, the levels of perforin and granzyme B expression by tumor-infiltrating Tfc cells were inversely correlated with the level of CD40L expression by tumor-infiltrating Tfc cells. Overall, we demonstrated that an inverse association existed between CD40L and cytotoxic molecule expression in Tfc cells from CRC patients.
CXCR5 overexpression in HL-60 cells enhances chemotaxis toward CXCL13 without anticipated interaction partners or enhanced MAPK signaling.
MacDonald Robert J,Yen Andrew
In vitro cellular & developmental biology. Animal
CXCR5 is a serpentine receptor implicated in cell migration in lymphocytes and differentiation in leukocytes. It causes MAPK pathway activation and has known membrane partners for signaling. CXCR5 mRNA is reportedly expressed in neutrophils following isolation, but its role in this cellular context is unknown. CXCR5 is also expressed in HL-60 cells, a human acute myeloid leukemia line, following treatment with all-trans retinoic acid, which induces differentiation toward a neutrophil-like state. CXCR5 is necessary for this process; differentiation was crippled in CXCR5 knockout cells and enhanced in cells ectopically expressing it. Since CXCR5 has various membrane protein partners, we investigated whether CXCR5-driven all-trans retinoic acid-induced differentiation depends on its association with such partners. Pursuing this, we generated HL-60 cells overexpressing the protein. We found that CXCR5 drove migration toward its ligand, CXCL13, and probed for interactions with several candidates using flow cytometry-based Förster resonance energy transfer. Surprisingly, we did not detect interactions with any candidates, including three reported in other cellular contexts. Additionally, we observed no significant changes in all-trans retinoic acid-induced differentiation; this may be due to the stoichiometry of CXCR5 and partner receptors or CXCL13. The anticipated membrane partnerings were surprisingly apparently unnecessary for downstream CXCR5 signaling and all-trans retinoic acid-induced differentiation.
Protective role of follicular CXCR5CD8 T cells against dengue virus 2 infection.
Qiu Liannv,Wang Hong,Yu Qinhua,Liu JieJing,Chen Sufeng,Zhao Zhao
International journal of infectious diseases : IJID : official publication of the International Society for Infectious Diseases
Follicular CXCR5CD8 T cells have antiviral effects in chronic virus infection, but the roles of these cells during dengue virus 2 (DENV2) infection remain poorly understood. OBJECTIVE:This study was conducted to analyzed in detail the dynamic changes and functional properties of circulating follicular CXCR5CD8 T cells to explore their effects on DENV2 infection. METHODS:Circulating follicular CXCR5CD8 T cells and cytokines were analyzed by flow cytometry in DENV2 patients at difference days after DENV2 infection. CD8 T cells were isolated and purified from DENV2 patients, then were stimulated with NS1 peptides and TCR stimulant. After cultivation, multiple parameters were tested. RESULTS:(1) CXCR5CD8 T cells emerged after DENV2 infection, with high PD-1 expression, and were correlated with the reduction in DENV2 RNA viral loads. (2) PD-1CXCR5CD8 T cells were negatively associated with disease progression. (3) Serum IFN-γ, IL-6 and IL-10 levels were increased late in the course of DENV2 infection. (4) CXCR5CD8 T cells from DENV2 patients exhibited increased cytotoxicity and IFN-γ and IL-10 secretion. CONCLUSION:CXCR5CD8 T cells could play a protective role in dengue pathogenesis and may be a novel strategy for controlling DENV2 infection and vaccine development.
Circulating CXCR3-CCR6-CXCR5CD4 T cells are associated with acute allograft rejection in liver transplantation.
Zhang Ke,Sun Yan-Ling,Zhou Shuang-Nan,Xu Ruo-Nan,Liu Zhen-Wen,Wang Fu-Sheng,Shi Ming
Circulating T follicular helper (cTFH) cells have been demonstrated to be involved in B-cell-mediated alloreactive responses in kidney and liver transplantation; however, whether these cells are involved in acute liver allograft rejection after liver transplantation, and which subsets are involved, remains to be clarified. The present study aimed to investigate the profiles of cTFH cells in acute liver allograft rejection, including the CXC motif receptor 3 (CXCR3) chemokine receptor 6 (CCR6) subset, the CXCR3CCR6 subset, and the CXCR3CCR6 subset. Twelve liver transplant patients with acute rejection (AR) and 20 with no acute rejection (NAR) were enrolled in the study. The results showed that the proportion of CXCR3-CCR6-CXCR5CD4 T cells was significantly increased and the proportion of CXCR3CCR6CXCR5CD4 T cells was significantly decreased in patients with AR compared with patients with NAR. In addition, the proportion of CXCR3-CCR6-CXCR5CD4 T cells was positively correlated with the proportion of B cells in patients with AR. The level of serum interleukin (IL)-21 was higher in the AR group than in the NAR groups. Furthermore, the proportion of CXCR3-CCR6-CXCR5CD4 T cells was positively correlated with alanine amino transferase (ALT), whereas the proportion of CXCR3CCR6 CXCR5CD4 T cells was negatively correlated with ALT. B cells and TFH cells were detected in follicular-like structures in liver allograft tissues from patients with AR. These results suggest that CXCR3-CCR6-CXCR5CD4 T cells may be involved in acute allograft rejection after liver transplantation.
CXCR5CD8 T cells are a distinct functional subset with an antitumor activity.
Chu Fuliang,Li Haiyan S,Liu Xindong,Cao Jingjing,Ma Wencai,Ma Ying,Weng Jinsheng,Zhu Zheng,Cheng Xiaoyun,Wang Zhiqiang,Liu Jingwei,Jiang Zi Yang,Luong Amber U,Peng Weiyi,Wang Jing,Balakrishnan Kumudha,Yee Cassian,Dong Chen,Davis Richard Eric,Watowich Stephanie S,Neelapu Sattva S
CXCR5 mediates homing of both B and follicular helper T (T) cells into follicles of secondary lymphoid organs. We found that CXCR5CD8 T cells are present in human tonsils and follicular lymphoma, inhibit T-mediated B cell differentiation, and exhibit strong cytotoxic activity. Consistent with these findings, adoptive transfer of CXCR5CD8 T cells into an animal model of lymphoma resulted in significantly greater antitumor activity than CXCR5CD8 T cells. Furthermore, RNA-Seq-based transcriptional profiling revealed 77 differentially expressed genes unique to CXCR5CD8 T cells. Among these, a signature comprised of 33 upregulated genes correlated with improved survival in follicular lymphoma patients. We also showed that CXCR5CD8 T cells could be induced and expanded ex vivo using IL-23 plus TGF-β, suggesting a possible strategy to generate these cells for clinical application. In summary, our study identified CXCR5CD8 T cells as a distinct T cell subset with ability to suppress T-mediated B cell differentiation, exert strong antitumor activity, and confer favorable prognosis in follicular lymphoma patients.
PD-1+Tim-3+ CD8+ T Lymphocytes Display Varied Degrees of Functional Exhaustion in Patients with Regionally Metastatic Differentiated Thyroid Cancer.
Severson Jill J,Serracino Hilary S,Mateescu Valerica,Raeburn Christopher D,McIntyre Robert C,Sams Sharon B,Haugen Bryan R,French Jena D
Cancer immunology research
Regional metastatic differentiated thyroid cancer (mDTC) provides a unique model in which to study the tumor-immune interface. These lymph node metastases persist for years, generally without progression to distant metastases. Although the immune system likely impedes disease progression, it is unsuccessful in eliminating disease. Our previous studies revealed that programmed death-1 (PD-1)(+) T cells were enriched in tumor-involved lymph nodes (TILN). Tumor-associated leukocytes and tumor cells were collected from grossly involved lymph nodes from 12 patients to further characterize the phenotype and functional potential of mDTC-associated PD-1(+) T cells. PD-1(+)CD4(+) and PD-1(+)CD8(+) T cells were enriched in 8 of 12 TILN samples. PD-1(+) T cells coexpressed Tim-3 and CD69 and failed to downregulate CD27. CD8(+) T cells, but not CD4(+) T cells, from these samples were variably deficient in their ability to produce effector cytokines when compared with control TILNs that lacked resident PD-1(+) T cells. PD-1(+)CD8(+) T cells were capable of exocytosis but lacked intracellular perforin. Surprisingly, T-cell proliferative capacity was largely maintained in all samples. Thus, although PD-1 expression by mDTC-associated CD8(+) T cells was associated with dysfunction, exhaustion was not complete. Notably, molecular markers of exhaustion did not translate to dysfunction in all samples or in CD4(+) T cells. Regulatory T cells (Treg), PD-L1, and galectin-9 were commonly found in mDTC and likely contributed to the initiation of T-cell exhaustion and disease progression. Therapies that release the effects of PD-1 and Tim-3 and reduce the suppressive effects of Tregs may encourage tumor elimination in patients with mDTC.
CD8CXCR5 T cells in tumor-draining lymph nodes are highly activated and predict better prognosis in colorectal cancer.
E Jifu,Yan Feihu,Kang Zhengchun,Zhu Liangliang,Xing Junjie,Yu Enda
Tumor-draining lymph nodes (TDLNs) are the primary sites of tumor antigen presentation, as well as the origin of metastasis in most cases. Hence, the type and function of immune cells in TDLNs are critical to the microenvironment and potentially affect the clinical outcome of the malignancy. CD8CXCR5 T cells are recently described to present high effector functions in infectious diseases, but their role in colorectal cancer (CRC) remains unclear. In forty-four Stage III CRC patients, we examined the CD8CXCR5 T cells in blood, tumor, and TDLN. CD8CXCR5 T cells represented lass than 2% of CD3 T cells in blood, but a much larger population in tumor. In TDLN, the CD8CXCR5 T cells represented the vast majority of CD8 T cells and between 9.3% and 32.9% of CD3 T cells. The prevalence of CD8CXCR5 T cells in tumor was not associated with their frequency in peripheral blood, but was positively correlated with their frequency in TDLN. The transcription of effector genes, including IFNG, TNF, IL2, PRF1, and GZMB, and exhaustion markers, including PD1, TIM3, 2B4, and LAG3, were examined in CD8CXCR5 T cells and CD8CXCR5 T cells. With a few exceptions, CD8CXCR5 T cell presented significantly higher effector gene expression, and significantly lower exhaustion marker expression than their CXCR5 counterparts. In addition, the prognosis of CRC patients was positively associated with the frequency of TDLN CD8CXCR5 T cells, and with the expression of IFNG, PRF1, and GZMB expression by tumor and TDLN CD8CXCR5 T cells. Together, these results demonstrated that CD8CXCR5 T cells were significant participants of CRC-associated immunity and could potentially serve as therapeutic options.
CXCR5 CD8 T cells potently infiltrate pancreatic tumors and present high functionality.
Bai Minghui,Zheng Youwei,Liu Haichao,Su Baowei,Zhan Yong,He Hua
Experimental cell research
Despite continued improvement in conventional therapy, pancreatic cancer continues to be one of the deadliest tumors worldwide with abysmal 5-year survival rate. New immunotherapeutic strategies that aim at improving antitumor cytotoxic CD8 T cell responses are being developed in solid tumors. To assist the development of immunotherapies, we investigated the CD8 T cells in pancreatic cancer patients. Compared to healthy individuals, pancreatic cancer patients presented a significant enrichment in the frequency of CD8CXCR5 T cells. In the tumor microenvironment, the frequencies of CD8CXCR5 T cells were further increased. In most cases, over half of tumor-infiltrating CD8 T cells were CD8CXCR5 T cells. Compared to the circulating population, the tumor-infiltrating CD8CXCR5 T cells expressed higher levels of PD-1 and TIM-3. Functional analyses demonstrated that upon CD3/CD28 activation, the percentages of TNF-expressing and IFN-γ-expressing cells in CD8CXCR5 T cells were significantly higher than that in CD8CXCR5 T cells. CD8CXCR5 T cells also presented enhanced cytotoxicity than CD8CXCR5 T cells. Upon PD-1 and TIM-3 blockade, the functions of CD8CXCR5 T cells were further improved. The disease-free survival of pancreatic cancer patients following tumor resection was positively correlated with the frequencies of circulating and tumor-infiltrating CD8CXCR5 T cells. Together, our study identified that CD8CXCR5 T cells were a potent subset of CD8 T cells that were highly enriched in pancreatic cancer patients and could respond to anti-PD-1/anti-TIM-3 blockade by further upregulation in function.
CXCR5CD8 T cells infiltrate the colorectal tumors and nearby lymph nodes, and are associated with enhanced IgG response in B cells.
Xing Junjie,Zhang Chenxin,Yang Xiaohong,Wang Shaoxuan,Wang Zhongchuan,Li Xu,Yu Enda
Experimental cell research
Colorectal cancer is the third most prevalent cancer type worldwide and contributes to a significant percentage of cancer-related mortality. Recent studies have shown that the CXCR5CD8 T cells present more potent proinflammatory function than CXCR5CD8 T cells in chronic virus infections and in follicular lymphoma, but the role of CXCR5CD8 T cells in colorectal cancer is yet unclear. In this study, we demonstrated that CXCR5CD8 T cells were very rare in peripheral blood mononuclear cells from healthy and colorectal cancer individuals, but were significantly enriched in resected tumors and tumor-associated lymph nodes. Compared to CXCR5CD8 T cells, the CXCR5CD8 T cells demonstrated significantly higher Bcl-6 expression and lower Blimp1 expression, suggesting that CXCR5CD8 T cells might represent a memory CD8 T cell subset. CXCR5CD8 T cells also enhanced the IgG expression by autologous B cells. Under ex vivo condition, the CXCR5CD8 T cells demonstrated lower degranulation, TNFα expression and IFNγ expression than CXCR5CD8 T cells. However, after PMA + ionomycin stimulation, the degranulation and TNFα expression by CXCR5CD8 T cells were significantly elevated to a level comparable with CXCR5CD8 T cells, whereas the IFNγ expression by PMA + ionomycin-stimulated CXCR5CD8 T cells were significantly higher than that by CXCR5CD8 T cells. Following long-term TCR-stimulation, CXCR5CD8 T cells demonstrated significantly more potent proliferation capacity and higher IFNγ expression than CXCR5CD8 T cells. TCR-stimulated CXCR5CD8 T cells also showed a gradual downregulation in CXCR5 expression. We further found that TCR-stimulated CXCR5CD8 T cells demonstrated higher granzyme B production and induced more specific lysis of autologous tumor cells than CXCR5CD8 T cells. Together, these data demonstrate that CXCR5CD8 T cells represent a significant CD8 T cell subset in colorectal tumors and have the potential to contribute to antitumor immunity, but their specific roles require further studies in vivo.
CXCR5 CD8 T cells displayed higher activation potential despite high PD-1 expression, in tumor-involved lymph nodes from patients with thyroid cancer.
Zhou Yu,Guo Linqi,Sun Huawei,Xu Jianbo,Ba Tu
Thyroid cancer is one of the malignancies with better clinical outcomes. However, a minority of patients develops an aggressive anaplastic thyroid carcinoma. Development of innovative and multimodal therapeutic strategies is urgently needed. Here, we investigated the role of CXCR5 CD8 T cells in the peripheral blood, tumor-involved lymph nodes (TILN), and tumor mass of thyroid cancer patients. In peripheral blood mononuclear cells, CXCR5 cells represented 1.4% ± 0.84% (mean ± s.d.) of total CD8 T cells, while in TILN and in tumor, the frequencies of CXCR5 CD8 T cells were significantly higher at 27.7% ± 7.8% and 15.5% ± 2.9%, respectively. Compared to CXCR5 CD8 T cells, CXCR5 CD8 T cells presented significantly higher PD-1 expression and lower or comparable TIM-3 and CTLA-4 expression. To compare and contrast the functional characteristics of CXCR5 CD8 T cells and CXCR5 CD8 T cells, these cells were separated from TILNs and were TCR-stimulated via anti-CD3/CD28. Upon stimulation, CXCR5 CD8 T cells presented stronger downregulation of CD27, higher expression of proinflammatory cytokines IL-2, IFN-γ, and TNF-α, and higher proliferation capacity than CXCR5 CD8 T cells. Moreover, CXCR5 CD8 T cells presented higher expression of cytotoxic molecules Gzm-A, Gzm-B, and perforin. Overall, these results demonstrated that in thyroid cancer patients CXCR5 CD8 T cells infiltrated the TILNs and the tumors, and were functionally more potent compared to their CXCR5 counterpart.
CXCR5-Dependent Entry of CD8 T Cells into Rhesus Macaque B-Cell Follicles Achieved through T-Cell Engineering.
Ayala Victor I,Deleage Claire,Trivett Matthew T,Jain Sumiti,Coren Lori V,Breed Matthew W,Kramer Joshua A,Thomas James A,Estes Jacob D,Lifson Jeffrey D,Ott David E
Journal of virology
Follicular helper CD4 T cells, T, residing in B-cell follicles within secondary lymphoid tissues, are readily infected by AIDS viruses and are a major source of persistent virus despite relative control of viral replication. This persistence is due at least in part to a relative exclusion of effective antiviral CD8 T cells from B-cell follicles. To determine whether CD8 T cells could be engineered to enter B-cell follicles, we genetically modified unselected CD8 T cells to express CXC chemokine receptor 5 (CXCR5), the chemokine receptor implicated in cellular entry into B-cell follicles. Engineered CD8 T cells expressing human CXCR5 (CD8) exhibited ligand-specific signaling and chemotaxis Six infected rhesus macaques were infused with differentially fluorescent dye-labeled autologous CD8 and untransduced CD8 T cells and necropsied 48 h later. Flow cytometry of both spleen and lymph node samples revealed higher frequencies of CD8 than untransduced cells, consistent with preferential trafficking to B-cell follicle-containing tissues. Confocal fluorescence microscopy of thin-sectioned lymphoid tissues demonstrated strong preferential localization of CD8 T cells within B-cell follicles with only rare cells in extrafollicular locations. CD8 T cells were present throughout the follicles with some observed near infected T In contrast, untransduced CD8 T cells were found in the extrafollicular T-cell zone. Our ability to direct localization of unselected CD8 T cells into B-cell follicles using CXCR5 expression provides a strategy to place highly effective virus-specific CD8 T cells into these AIDS virus sanctuaries and potentially suppress residual viral replication. AIDS virus persistence in individuals under effective drug therapy or those who spontaneously control viremia remains an obstacle to definitive treatment. Infected follicular helper CD4 T cells, T, present inside B-cell follicles represent a major source of this residual virus. While effective CD8 T-cell responses can control viral replication in conjunction with drug therapy or in rare cases spontaneously, most antiviral CD8 T cells do not enter B-cell follicles, and those that do fail to robustly control viral replication in the T population. Thus, these sites are a sanctuary and a reservoir for replicating AIDS viruses. Here, we demonstrate that engineering unselected CD8 T cells to express CXCR5, a chemokine receptor on T associated with B-cell follicle localization, redirects them into B-cell follicles. These proof of principle results open a pathway for directing engineered antiviral T cells into these viral sanctuaries to help eliminate this source of persistent virus.
Aberrant expansion of circulating CD4 CXCR5 CCR7 PD1 Tfh precursor cells in idiopathic inflammatory myopathy.
Zhang Lu,Li Wenli,Cai Ying,Liu Xia,Peng Qinglin,Liang Lin
International journal of rheumatic diseases
OBJECTIVE:To determine circulating follicular T helper (Tfh) cell precursor and its relationship with clinical characteristics in idiopathic inflammatory myopathy (IIM). METHODS:The study population included 47 patients with IIM and 30 healthy controls. Circulating CD4 CXCR5 CCR7 PD-1 T cells and intracellular interleukin (IL)-21 were assessed by flow cytometry. Serum IL-21 levels were measured by enzyme-linked immunosorbent assay. The disease activity was evaluated using myositis disease activity assessment visual analog scales (VAS) as well as muscle and physician global assessment (PGA). RESULTS:The percentage of the CCR7 PD-1 subset cells within CD4 CXCR5 T cells was significantly increased in patients with IIM compared to that in healthy controls (14.3 ± 6.5 vs 11.4 ± 2.6, P = .009). Patients with higher percentages of CCR7 PD-1 subsets presented with higher PGA VAS (P = .000), muscle VAS (P = .000), as well as serum creatinine kinase (CK) levels (P = .000) than those with lower percentages of CCR7 PD-1 subsets. IL-21 expression significantly increased in CD4 CXCR5 CCR7 PD-1 T cells in patients with IIM compared to that in healthy controls (26.07 ± 7.38 vs 19.25 ± 5.67, P = .001). Meanwhile, both the CCR7 PD-1 subset and intracellular IL-21 expression in IIM patients showed significantly positive correlation with PGA VAS, muscle VAS and serum CK levels. Circulating CD4 CXCR5 CCR7 PD-1 T cells and intracellular IL-21 decreased significantly when disease was improved (P = .018; P = .028). CONCLUSION:The percentage of circulating CCR7 PD-1 subset among total CD4 CXCR5 T cells and intracellular IL-21 expression expanded and showed significant correlation with disease activity in IIM. The circulating follicular helper T cell precursor may be involved in the pathogenesis, especially muscle injury in IIM.
Emerging Role of CAR T Cells in Non-Hodgkin's Lymphoma.
Avanzi Mauro P,Brentjens Renier J
Journal of the National Comprehensive Cancer Network : JNCCN
Adoptive T-cell therapy with chimeric antigen receptor T cells (CAR-Ts) has produced impressive clinical responses among patients with B-cell malignancies, and several groups have published positive results using anti-CD19 CAR-Ts for the treatment of B-cell acute lymphoblastic leukemia. Recently, new data from clinical trials have demonstrated the benefits of CAR-T therapy in the non-Hodgkin's lymphoma (NHL) setting. This review describes some of the most recent and promising advances in engineered T-cell therapy, with particular emphasis on the clinical benefits of NHL treatment.
Clinical development of anti-CD19 chimeric antigen receptor T-cell therapy for B-cell non-Hodgkin lymphoma.
Makita Shinichi,Yoshimura Kiyoshi,Tobinai Kensei
B-cell non-Hodgkin lymphoma (B-NHL) is the most frequent hematological malignancy. Although refined chemotherapy regimens and several new therapeutics including rituximab, a chimeric anti-CD20 monoclonal antibody, have improved its prognosis in recent decades, there are still a substantial number of patients with chemorefractory B-NHL. Anti-CD19 chimeric antigen receptor (CAR) T-cell therapy is expected to be an effective adoptive cell treatment and has the potential to overcome the chemorefractoriness of B-cell leukemia and lymphoma. Recently, several clinical trials have shown remarkable efficacy of anti-CD19 CAR T-cell therapy, not only in B-acute lymphoblastic leukemia but also in B-NHL. Nonetheless, there are several challenges to overcome before introduction into clinical practice, such as: (i) further refinement of the manufacturing process, (ii) further improvement of efficacy, (iii) finding the optimal infusion cell dose, (iv) optimization of lymphocyte-depleting chemotherapy, (v) identification of the best CAR structure, and (vi) optimization of toxicity management including cytokine release syndrome, neurologic toxicity, and on-target off-tumor toxicity. Several ways to solve these problems are currently under study. In this review, we describe the updated clinical data regarding anti-CD19 CAR T-cell therapy, with a focus on B-NHL, and discuss the clinical implications and perspectives of CAR T-cell therapy.
T cell immune response within B-cell follicles.
Huang Qizhao,Xu Lifan,Ye Lilin
Advances in immunology
B-cell follicle represents a functionally dynamic microstructure within second lymphoid tissues, predominantly consisting of B cells, follicular T cells and DCs. Through intimate interactions with cognate B cells, follicular helper T cells (Tfh) initiate and facilitate germinal center (GC) reactions by providing signals required for producing high-affinity antibodies, as well as for the generation of long-lived antibody-secreting plasma cells and memory B cells. Concomitantly, germinal center reaction needs to be fine controlled to avoid autoimmunity or B-cell malignancies. Among immune cells residing in follicles, follicular regulatory T cells (Tfr), converted from naïve Treg cells, are specifically assigned to repress excessive GC responses by suppressing Tfh and GC B cells within GC structure. Hence, through Yin and Yang (positive and negative) regulation of GC reaction, Tfh cells play concert with Tfr cells in maintaining immune homeostasis. Besides CD4 T cells, a small portion of CXCR5 expressing CD8 T cells, regarded as follicular cytotoxic T cells (Tfc), could migrate into B cell follicles during chronic viral infection and several types of cancers, and this population exhibit lower level of exhaustion than its CXCR5 counterparts. Besides, Tfc cells demonstrate a stem-cell like phenotype during chronic infection which could further differentiate into terminally differentiated CXCR5CD8 T cells. Collectively, in this review, we will discuss the recent advances in our understanding of the ontology and differentiation of B-cell follicle resident Tfh, Tfr and Tfc cells.
Circulating precursor CCR7(lo)PD-1(hi) CXCR5⁺ CD4⁺ T cells indicate Tfh cell activity and promote antibody responses upon antigen reexposure.
He Jing,Tsai Louis M,Leong Yew Ann,Hu Xin,Ma Cindy S,Chevalier Nina,Sun Xiaolin,Vandenberg Kirsten,Rockman Steve,Ding Yan,Zhu Lei,Wei Wei,Wang Changqi,Karnowski Alexander,Belz Gabrielle T,Ghali Joanna R,Cook Matthew C,Riminton D Sean,Veillette André,Schwartzberg Pamela L,Mackay Fabienne,Brink Robert,Tangye Stuart G,Vinuesa Carola G,Mackay Charles R,Li Zhanguo,Yu Di
Follicular B helper T (Tfh) cells support high affinity and long-term antibody responses. Here we found that within circulating CXCR5⁺ CD4⁺ T cells in humans and mice, the CCR7(lo)PD-1(hi) subset has a partial Tfh effector phenotype, whereas CCR7(hi)PD-1(lo) cells have a resting phenotype. The circulating CCR7(lo)PD-1(hi) subset was indicative of active Tfh differentiation in lymphoid organs and correlated with clinical indices in autoimmune diseases. Thus the CCR7(lo)PD-1(hi) subset provides a biomarker to monitor protective antibody responses during infection or vaccination and pathogenic antibody responses in autoimmune diseases. Differentiation of both CCR7(hi)PD-1(lo) and CCR7(lo)PD-1(hi) subsets required ICOS and BCL6, but not SAP, suggesting that circulating CXCR5⁺ helper T cells are primarily generated before germinal centers. Upon antigen reencounter, CCR7(lo)PD-1(hi) CXCR5⁺ precursors rapidly differentiate into mature Tfh cells to promote antibody responses. Therefore, circulating CCR7(lo)PD-1(hi) CXCR5⁺ CD4⁺ T cells are generated during active Tfh differentiation and represent a new mechanism of immunological early memory.
Targeting Antigen to Clec9A Primes Follicular Th Cell Memory Responses Capable of Robust Recall.
Kato Yu,Zaid Ali,Davey Gayle M,Mueller Scott N,Nutt Stephen L,Zotos Dimitra,Tarlinton David M,Shortman Ken,Lahoud Mireille H,Heath William R,Caminschi Irina
Journal of immunology (Baltimore, Md. : 1950)
Targeting Ags to dendritic cell (DC) surface receptors can induce a variety of responses depending on the DC type targeted, the receptor targeted, and the adjuvant used. Clec9A (DNGR-1), which is expressed by CD8(+) DCs, has been shown to bind F-actin exposed on damaged cells. Targeting Ag to this receptor in mice and nonhuman primates induces strong humoral immunity even in the absence of adjuvant, a process seen for a few select DC receptors. In contrast with other receptors, however, targeting Clec9A induces long-lived, affinity-matured Ab responses that are associated with efficient CD4(+) T cell responses shown to possess properties of follicular Th cells (TFH). In this article, we provide definitive evidence that Clec9A targeting promotes the development of TFH by showing that responding CD4 T cells express CXCR5, PD1, the TFH transcription factor Bcl6, and the cytokine IL-21, and that these cells localize to germinal centers. Furthermore, we extend studies from the model Ag OVA to the viral Ag glycoprotein D of HSV-1 and examine the capacity of primed TFH to form functional memory. We show that targeting glycoprotein D to Clec9A even in the absence of adjuvant induced long-lived memory CXCR5(+) PD1(hi) CD4(+) T cells that proliferated extensively upon secondary challenge and rapidly developed into effector TFH. This was associated with enhanced germinal center B cell responses and accelerated Ab production. Our study indicates that targeting Ags to Clec9A in the absence of adjuvant routinely generates TFH responses that form long-lived memory capable of robust secondary TFH responses.
A novel mouse model for tracking the fate of CXCR5-expressing T cells.
Takebe Tomo,Sakamoto Kazuki,Higami Yoshikazu,Harada Yohsuke
Biochemical and biophysical research communications
The germinal center (GC) reaction, a critical process in the humoral immune response, requires follicular helper T (Tfh) cells. Tfh cells express the master transcription factor Bcl6 and chemokine receptor CXCR5, which enable them to migrate from the T cell zone to B cell follicles and interact with GC B cells. However, CXCR5 is downregulated when Tfh cells become memory cells. Therefore, it is difficult to track Tfh cells continuously in vivo. In this study, we generated a mouse strain, Cxcr5R26, in which the fluorescent protein tdTomato is inducibly expressed in CXCR5 cells by tamoxifen administration. After the oral administration of tamoxifen, most Tfh cells in Peyer's patches (PP) from Cxcr5R26 mice were tdTomato. To track antigen-specific Tfh cells in vivo, OVA-specific OT-II T cells derived from Cxcr5R26 mice were transferred to wild-type mice, and the recipient mice were immunized with OVA followed by tamoxifen administration. CXCR5 T cells became tdTomato and were mainly located in B cell follicles and GC areas 8 days after immunization. Four weeks after immunization, tdTomato OT-II T cells migrated from B cell follicles to the T-B border area and T cell zone after CXCR5 downregulation and CCR7 upregulation. These results indicate that Cxcr5R26 mice are a useful tool for studying the cell fate of differentiated Tfh cells in vivo and therefore have implications for the development of therapeutic strategies for infectious and autoimmune diseases.
Simian Immunodeficiency Virus (SIV)-Specific Chimeric Antigen Receptor-T Cells Engineered to Target B Cell Follicles and Suppress SIV Replication.
Haran Kumudhini Preethi,Hajduczki Agnes,Pampusch Mary S,Mwakalundwa Gwantwa,Vargas-Inchaustegui Diego A,Rakasz Eva G,Connick Elizabeth,Berger Edward A,Skinner Pamela J
Frontiers in immunology
There is a need to develop improved methods to treat and potentially cure HIV infection. During chronic HIV infection, replication is concentrated within T follicular helper cells (Tfh) located within B cell follicles, where low levels of virus-specific CTL permit ongoing viral replication. We previously showed that elevated levels of simian immunodeficiency virus (SIV)-specific CTL in B cell follicles are linked to both decreased levels of viral replication in follicles and decreased plasma viral loads. These findings provide the rationale to develop a strategy for targeting follicular viral-producing (Tfh) cells using antiviral chimeric antigen receptor (CAR) T cells co-expressing the follicular homing chemokine receptor CXCR5. We hypothesize that antiviral CAR/CXCR5-expressing T cells, when infused into an SIV-infected animal or an HIV-infected individual, will home to B cell follicles, suppress viral replication, and lead to long-term durable remission of SIV and HIV. To begin to test this hypothesis, we engineered gammaretroviral transduction vectors for co-expression of a bispecific anti-SIV CAR and rhesus macaque CXCR5. Viral suppression by CAR/CXCR5-transduced T cells was measured , and CXCR5-mediated migration was evaluated using both an transwell migration assay, as well as a novel tissue migration assay. The functionality of the CAR/CXCR5 T cells was demonstrated through their potent suppression of SIV and SIV replication in and migration to the ligand CXCL13 , and concentration in B cell follicles in tissues . These novel antiviral immunotherapy products have the potential to provide long-term durable remission (functional cure) of HIV and SIV infections.
CXCR5 CAR-T cells simultaneously target B cell non-Hodgkin's lymphoma and tumor-supportive follicular T helper cells.
Bunse Mario,Pfeilschifter Janina,Bluhm Julia,Zschummel Maria,Joedicke Jara J,Wirges Anthea,Stark Helen,Kretschmer Vivien,Chmielewski Markus,Uckert Wolfgang,Abken Hinrich,Westermann Jörg,Rehm Armin,Höpken Uta E
CAR-T cell therapy targeting CD19 demonstrated strong activity against advanced B cell leukemia, however shows less efficacy against lymphoma with nodal dissemination. To target both B cell Non-Hodgkin's lymphoma (B-NHLs) and follicular T helper (Tfh) cells in the tumor microenvironment (TME), we apply here a chimeric antigen receptor (CAR) that recognizes human CXCR5 with high avidity. CXCR5, physiologically expressed on mature B and Tfh cells, is also highly expressed on nodal B-NHLs. Anti-CXCR5 CAR-T cells eradicate B-NHL cells and lymphoma-supportive Tfh cells more potently than CD19 CAR-T cells in vitro, and they efficiently inhibit lymphoma growth in a murine xenograft model. Administration of anti-murine CXCR5 CAR-T cells in syngeneic mice specifically depletes endogenous and malignant B and Tfh cells without unexpected on-target/off-tumor effects. Collectively, anti-CXCR5 CAR-T cells provide a promising treatment strategy for nodal B-NHLs through the simultaneous elimination of lymphoma B cells and Tfh cells of the tumor-supporting TME.
Frequency of CD4+CXCR5+ TFH cells in patients with hepatitis b virus-associated membranous nephropathy.
Liu Yong,Zhao Pingwei,Qu Zhihui,Ayana Desalegn Admassu,Jiang Yanfang
The frequency of different subsets of CD4(+)CXCR5(+) TFH cells and serum cytokine levels were analyzed in a total of 14 patients with newly diagnosed hepatitis B virus-associated membranous nephropathy (HBV-MN), 12 individuals with immune-tolerant HBV infection (HBV-IT) and 12 healthy controls (HC). Serum cytokine levels were measured before and 10-12 weeks after treatment. Significantly higher frequency of CD4(+)CXCR5(+), CD4(+)CXCR5(+)ICOS(+) and CD4(+)CXCR5(+)PD-1(+) TFH cells, and higher serum levels of IL-17A, IFN-γ, IL-2, IL-10, IL-4 and IL-21 were detected in HBV-MN patients compared to the HC. The percentage of CD4(+)CXCR5(+) TFH cells and serum IL-21 level in HBV-MN patients were also higher than the HBV-IT. The percentage of CD4(+)CXCR5(+) TFH cell was negatively correlated with the value of eGFR, and the percentage of CD4(+)CXCR5(+)ICOS(+) TFH cells was positively correlated with the 24-h urinary protein concentration. Notably, the percentage of CD4(+)CXCR5(+)PD-1(+) TFH cells was positively correlated with serum IL-21 level and 24-h urinary protein concentration. Treatment with prednisone or/and immunosuppressive drugs significantly reduced the frequency of CD4(+)CXCR5(+), CD4(+)CXCR5(+)PD-1(+) TFH cells and serum IL-21 level, but increased IL-4 and IL-10 levels in the patients. CD4(+)CXCR5(+) TFH cells, especially CD4(+)CXCR5(+)PD-1(+) TFH cells may participate in the pathogenesis of HBV-MN.
Tumor-infiltrating lymphocytes and hepatocellular carcinoma: pathology and clinical management.
Shirabe Ken,Motomura Takashi,Muto Jun,Toshima Takeo,Matono Rumi,Mano Yohei,Takeishi Kazuki,Ijichi Hideki,Harada Noboru,Uchiyama Hideaki,Yoshizumi Tomoharu,Taketomi Akinobu,Maehara Yoshihiko
International journal of clinical oncology
The presence of tumor-infiltrating lymphocytes (TILs) in hepatocellular carcinoma (HCC) is relatively rare. The prognosis of patients with HCC and marked TILs is better than that of patients with HCC without TILs. TILs in HCC tissues are mainly T cells, and previous reports suggested that TILs might be important antitumor effector cells. TILs have been extensively analyzed, and subpopulations of CD3(+), CD4(+), and CD8(+) T cells are often present in HCC. Some studies have reported that the percentage of CD8(+) T cells, which might have cytotoxic activity, is decreased in tumors with TILs, as compared with noncancerous tissues. Although the antitumor effects of TILs seem to be impaired in HCCs, the underlying mechanism has remained unclear until quite recently. Pathological and in vitro studies have now shown that regulatory T cells play important roles in the deterioration of the antitumor effects of TILs. The aim of this review is to introduce recent pathological findings for TILs in HCC and to evaluate new therapeutic strategies in this field.
Phenotype and function of CXCR5+CD45RA-CD4+ T cells were altered in HBV-related hepatocellular carcinoma and elevated serum CXCL13 predicted better prognosis.
Duan Zhaojun,Gao Jian,Zhang Ling,Liang Hua,Huang Xiangbo,Xu Qiang,Zhang Yu,Shen Tao,Lu Fengmin
The present study reveals an immunological characterization of circulating and tumor-infiltrating T follicular helper cells (Tfh), namely CXCR5+CD45RA-CD4+ T cells, and their related cytokines in hepatitis B virus-related hepatocellular carcinoma (HCC) patients. In HCC patients, circulating Tfh cells showed a CCR7+ and/or ICOS+ phenotype with increased Th2-like cells and decreased Th1-like and Th17-like subsets. Although the bulk frequency of circulating Tfh cells was not altered in HCC patients, the frequency of infiltrated CXCR5+CD45RA-CD4+ CD3+cells was higher in tumor than in para-tumor tissues, and Th1-like cells were the predominant phenotype. Circulating Tfh cells in HCC patients were defective in the production of IL-21 in vitro, which was in accordance with lower IL-21 levels in tumor tissues than in para-tumor tissues. Serum CXCL13 was increased in HCC patients and associated with recurrence-free survival after hepatectomy. This was confirmed in an additional HCC cohort of 111 patients with up to 5 years follow-up. Immunohistochemical staining indicated that the percentage of CXCR5+ or CXCL13+ cells was higher in poorly differentiated than in well-differentiated tumors. In conclusion, patients with HBV-related HCC showed altered phenotypes and impaired function of Tfh cells or subpopulations. CXCL13 could be a potential biomarker for predicting recurrence in HCC patients after hepatectomy.
CXCR5 CD8 T Cells Indirectly Offer B Cell Help and Are Inversely Correlated with Viral Load in Chronic Hepatitis B Infection.
Jiang Hang,Li Linhai,Han Jiang,Sun Zhiwei,Rong Yihui,Jin Yun
DNA and cell biology
Treatment options for chronic hepatitis B (CHB) infection are extremely limited. CXCR5 CD8 T cell is a novel cell subtype and could possess strong cytotoxic properties in HIV infection. In this study, we investigated the role of CXCR5 CD8 T cells in CHB patients. Compared to healthy individuals, both CHB patients and hepatitis B virus (HBV)-infected hepatocellular carcinoma patients presented significant upregulation of CXCR5 CD8 T cells in peripheral blood, in which CXCR5 CD8 T cells were negatively correlated with the frequency of CXCR5 CD4 T cells in CHB patients. After PMA+ionomycin stimulation, CXCR5 CD8 T cells from CHB patients presented significantly higher transcription level of interferon gamma (IFN-γ), interleukin 10 (IL-10), and IL-21, as well as higher IL-10 and IL-21 protein secretion, than CXCR5 CD8 T cells. Unlike CXCR5 CD4 T cells, when incubated with naive CD19CD27 B cells, CXCR5 CD8 T cells alone did not upregulate IgM, IgG, and IgA secretion. However, addition of CXCR5 CD8 T cells in B cell-CXCR5 CD4 T cell coculture significantly increased the levels of secreted IgG and IgA, demonstrating that CXCR5 CD8 T cell could indirectly offer B cell help. Furthermore, high frequencies of CXCR5 CD8 T cells tended to associate with low HBV DNA load, and the frequency of CXCR5 CD8 T cells was negatively correlated with alanine aminotransferase (ALT) level. Together, these results suggested that CXCR5 CD8 T cells were involved in the antiviral immune responses in CHB and could potentially serve as a therapeutic candidate.
CXCR5CD8 T cells could induce the death of tumor cells in HBV-related hepatocellular carcinoma.
Jin Yun,Lang Cuicui,Tang Jianzhong,Geng Jiawei,Song Haihan K,Sun Zhiwei,Wang Jinfeng
The follicular CXCR5CD8 T cells have recently emerged as a critical cell type in mediating peripheral tolerance as well as antiviral immune responses during chronic infections. In this study, we investigated the function of CXCR5CD8 T cells in HBV-related hepatocellular carcinoma patients. Compared to CXCR5CD8 T cells, CXCR5CD8 T cells presented elevated PD-1 expression but reduced Tim-3 and CTLA-4 expression. Upon anti-CD3/CD28 stimulation, CXCR5CD8 T cells demonstrated higher proliferation potency than CXCR5CD8 T cells, especially after PD-1 blockade. CXCR5CD8 T cells also demonstrated significantly higher granzyme B synthesis and release, as well as higher level of degranulation. Tumor cells were more readily eliminated by CXCR5CD8 T cells than by CXCR5CD8 T cells. Interestingly, we found that B cells were more resistant to CXCR5CD8 T cell-mediated killing than tumor cells, possibly through IL-10-mediated protection. In addition, the CXCR5CD8 T cell-mediated cytotoxic effects on tumor cells could be significantly enhanced by PD-L1 blockade. Together, we presented that in patients with in HBV-related hepatocellular carcinoma, CXCR5CD8 T cells could mediate tumor cell death more potently than the CXCR5CD8 T cells in vitro while the autologous B cells were protected.
CD8+CXCR5+T cells infiltrating hepatocellular carcinomas are activated and predictive of a better prognosis.
Ye Linsen,Li Yang,Tang Hui,Liu Wei,Chen Yunhao,Dai Tianxing,Liang Rongpu,Shi Mengchen,Yi Shuhong,Chen Guihua,Yang Yang
CD8+ T cells are thought to be the primary cytotoxic lymphocytes exerting antitumor effects. However, few studies have focused on the antitumor effects of CD8+ T cell-mediated humoral immunity or on interactions between CD8+ T cells and B cells in hepatocellular carcinoma (HCC). We found that the frequency of IL-21-producing CD8+CXCR5+ T cells was higher in HCC tumor tissue than in peritumoral tissue or peripheral blood from the same patients or in blood from healthy donors. Moreover, CD8+CXCR5+ T cells migrated in response to supernatants from primary HCC (HCC-SN) cells, and HCC-SN cells also powerfully induced CXCR5 expression in CD8+ T cells and IL-21 expression in CD8+CXCR5+ T cells. CD8+CXCR5+ T cells from HCC patients, but not those from healthy individuals, stimulated CD19+ B cells to differentiate into IgG-producing plasmablasts. These findings reveal that CD8+CXCR5+ T cells strongly infiltrate HCC tumors, and their infiltration is predictive of a better prognosis. Surprisingly, moreover, CD8+CXCR5+ T cells produced IL-21, which induced B cells to differentiate into IgG-producing plasmablasts and to play a key role in humoral immunity in HCC.
CXCR5-negative natural killer cells ameliorate experimental autoimmune myasthenia gravis by suppressing follicular helper T cells.
Yang Chun-Lin,Zhang Peng,Liu Ru-Tao,Zhang Na,Zhang Min,Li Heng,Du Tong,Li Xiao-Li,Dou Ying-Chun,Duan Rui-Sheng
Journal of neuroinflammation
BACKGROUND:Recent studies have demonstrated that natural killer (NK) cells can modulate other immune components and are involved in the development or progression of several autoimmune diseases. However, the roles and mechanisms of NK cells in regulating experimental autoimmune myasthenia gravis (EAMG) remained to be illustrated. METHODS:To address the function of NK cells in experimental autoimmune myasthenia gravis in vivo, EAMG rats were adoptively transferred with splenic NK cells. The serum antibodies, and splenic follicular helper T (Tfh) cells and germinal center B cells were determined by ELISA and flow cytometry. The roles of NK cells in regulating Tfh cells were further verified in vitro by co-culturing splenocytes or isolated T cells with NK cells. Moreover, the phenotype, localization, and function differences between different NK cell subtypes were determined by flow cytometry, immunofluorescence, and ex vivo co-culturation. RESULTS:In this study, we found that adoptive transfer of NK cells ameliorated EAMG symptoms by suppressing Tfh cells and germinal center B cells. Ex vivo studies indicated NK cells inhibited CD4 T cells and Tfh cells by inducing the apoptosis of T cells. More importantly, NK cells could be divided into CXCR5 and CXCR5 NK subtypes according to the expression of CXCR5 molecular. Compared with CXCR5 NK cells, which were mainly localized outside B cell zone, CXCR5 NK were concentrated in the B cell zone and exhibited higher expression levels of IL-17 and ICOS, and lower expression level of CD27. Ex vivo studies indicated it was CXCR5 NK cells not CXCR5 NK cells that suppressed CD4 T cells and Tfh cells. Further analysis revealed that, compared with CXCR5 NK cells, CXCR5 NK cells enhanced the ICOS expression of Tfh cells. CONCLUSIONS:These findings highlight the different roles of CXCR5 NK cells and CXCR5 NK cells. It was CXCR5 NK cells but not CXCR5 NK cells that suppressed Tfh cells and inhibited the autoimmune response in EAMG models.
Investigating the role of circulating CXCR5-expressing CD8+ T-cells as a biomarker for bacterial infection in subjects with pneumonia.
Shen Yu,Qu Qiu-Xia,Jin Meng-Ni,Chen Cheng
BACKGROUND:Recently, lymphoid follicle-confined and circulating CD8+ T-cells expressing the C-X-C chemokine receptor type 5 (CXCR5) were described, which was involved in anti-virus immune response. However, the dynamics and role of circulating CXCR5-expressing CD8+ T-cells during bacterial infection is unknown. So, we asked whether CXCR5+ CD8+ T cells were also generated during bacterial infections in lower respiratory tract. METHODS:The clinical data of 65 pneumonia patients were analyzed. The patients were divided into groups as tuberculosis, bronchiectasis and community or hospital acquired pneumonia (CAP, HAP). The sputum/bronchial secretion or bronchoalveolar lavage fluid (BALF) samples were taken for microbiological examination. The procalcitonin (PCT) was used to evaluate disease severity of these groups and compared among patients. We characterized the number and phenotype (PD-1 and CD103) of CXCR5 + CD8+ T cells in the peripheral circulation by flow cytometry in all individuals and analyzed their association with the serum PCT level and disease severity. RESULTS:Patients were mainly infected with Escherichia coli, Acinetobacter baumannii, Klebsiella pneumonia (K.p), Pseudomonas aeruginosa, and Staphylococcus aureus. Of note is the finding that PCT was weakly correlated with severity of respiratory infections. Furthermore, it was revealed an increase of CXCR5-expressing CD8+ T cells in peripheral blood of un-controlled CAP and progressive HAP compared controlled CAP and HAP, respectively (P < 0.05). Strikingly, the circulating CXCR5-expressing CD8+ T-cells in K.p-infected group was higher than that non-K.p-infected group (P < 0.05). Meanwhile, the ratio of CXCR5 + CD8+/CD8 was positively correlated with PCT level (P < 0.05). In clinic, the determination of CXCR5-expressing CD8+ T-cells showed better results compared to PCT and can be useful for the prediction of exacerbation of CAP or HAP. Phenotypically, CXCR5+ CD8 + T cell expressed comparable level of inhibitory molecules PD-1 and lower CD103 compared to their CXCR5- counterparts. CONCLUSION:The circulating CXCR5-expressing CD8+ T-cell has diagnostic value for current pneumonia severity, and could act as a biomarker for identifying a bacteria-associated exacerbation. These cells may provide novel insight for the pathogenesis of pneumonia.
Upregulation of miR-542-3p inhibits the growth and invasion of human colon cancer cells through PI3K/AKT/survivin signaling.
Yang Chuang,Wang Ming-Huan,Zhou Jun-De,Chi Qiang
The present study was designed to assess the expression of microRNA-542-3p (miR-542-3p) in human colon cancer and investigate the possible molecular mechanisms underlying the effect of miR-542-3p on the growth and invasion of colon cancer cells. We found that miR-542-3p expression was downregulated in colon cancer patient tissues, compared with that observed in the control group. Silencing of miR‑542-3p expression significantly promoted cell viability and inhibited apoptosis. In addition, overexpression of miR-542-3p significantly reduced cell viability and promoted apoptosis in colon cancer cells. Meanwhile, silencing of miR-542-3p expression significantly suppressed PI3K and p-AKT and survivin protein expression, while overexpression of miR-542-3p significantly induced PI3K and p-AKT and survivin protein expression in colon cancer cells. PI3K inhibitor (LY294002) or survivin inhibitor (YM155) suppressed PI3K/AKT/survivin signaling and increased the anticancer effects of miR-542-3p on the apoptosis in colon cancer. The present study demonstrated that upregulation of miR-542-3p inhibited the growth and invasion of colon cancer cells through PI3K/AKT/survivin signaling, highlighting a novel therapeutic approach for the treatment of colon cancer.
Protein Kinase C Epsilon Cooperates with PTEN Loss for Prostate Tumorigenesis through the CXCL13-CXCR5 Pathway.
Garg Rachana,Blando Jorge M,Perez Carlos J,Abba Martin C,Benavides Fernando,Kazanietz Marcelo G
PKCε, an oncogenic member of the PKC family, is aberrantly overexpressed in epithelial cancers. To date, little is known about functional interactions of PKCε with other genetic alterations, as well as the effectors contributing to its tumorigenic and metastatic phenotype. Here, we demonstrate that PKCε cooperates with the loss of the tumor suppressor Pten for the development of prostate cancer in a mouse model. Mechanistic analysis revealed that PKCε overexpression and Pten loss individually and synergistically upregulate the production of the chemokine CXCL13, which involves the transcriptional activation of the CXCL13 gene via the non-canonical nuclear factor κB (NF-κB) pathway. Notably, targeted disruption of CXCL13 or its receptor, CXCR5, in prostate cancer cells impaired their migratory and tumorigenic properties. In addition to providing evidence for an autonomous vicious cycle driven by PKCε, our studies identified a compelling rationale for targeting the CXCL13-CXCR5 axis for prostate cancer treatment.
p53-dependent expression of CXCR5 chemokine receptor in MCF-7 breast cancer cells.
Mitkin Nikita A,Hook Christina D,Schwartz Anton M,Biswas Subir,Kochetkov Dmitry V,Muratova Alisa M,Afanasyeva Marina A,Kravchenko Julia E,Bhattacharyya Arindam,Kuprash Dmitry V
Elevated expression of chemokine receptors in tumors has been reported in many instances and is related to a number of survival advantages for tumor cells including abnormal activation of prosurvival intracellular pathways. In this work we demonstrated an inverse correlation between expression levels of p53 tumor suppressor and CXCR5 chemokine receptor in MCF-7 human breast cancer cell line. Lentiviral transduction of MCF-7 cells with p53 shRNA led to elevated CXCR5 at both mRNA and protein levels. Functional activity of CXCR5 in p53-knockdown MCF-7 cells was also increased as shown by activation of target gene expression and chemotaxis in response to B-lymphocyte chemoattractant CXCL13. Using deletion analysis and site-directed mutagenesis of the cxcr5 gene promoter and enhancer elements, we demonstrated that p53 appears to act upon cxcr5 promoter indirectly, by repressing the activity of NFκB transcription factors. Using chromatin immunoprecipitation and reporter gene analysis, we further demonstrated that p65/RelA was able to bind the cxcr5 promoter in p53-dependent manner and to directly transactivate it when overexpressed. Through the described mechanism, elevated CXCR5 expression may contribute to abnormal cell survival and migration in breast tumors that lack functional p53.
Improved outcome of high-risk early HER2 positive breast cancer with high CXCL13-CXCR5 messenger RNA expression.
Razis Evangelia,Kalogeras Konstantine T,Kotoula Vassiliki,Eleftheraki Anastasia G,Nikitas Nikitas,Kronenwett Ralf,Timotheadou Eleni,Christodoulou Christos,Pectasides Dimitrios,Gogas Helen,Wirtz Ralph M,Makatsoris Thomas,Bafaloukos Dimitrios,Aravantinos Gerasimos,Televantou Despina,Pavlidis Nicholas,Fountzilas George
Clinical breast cancer
UNLABELLED:The CXCL13-CXCR5 is a chemokine axis that is activated in some breast cancers. A total of 321 tissue blocks from a group of patients who received adjuvant, dose-dense chemotherapy for high-risk early breast cancer were examined. Activation of this axis was found to be associated with determinants of poor prognosis but also with improved outcome in the human epidermal growth factor receptor 2 overexpressing subpopulation. BACKGROUND:Chemokines are important in cell migration and are thought to play a key role in metastasis. We explored the prognostic significance of C-X-C ligand-motif (CXCL) 12, CXCL13, and receptor (CXCR) 5 on disease-free survival (DFS) and overall survival (OS) in early breast cancer. METHODS:A total of 595 patients with high risk, [corrected] early breast cancer were treated in a 2-arm trial (HE10/97) with dose-dense sequential epirubicin followed by cyclophosphamide, methotrexate, and 5-fluorouracil (CMF) with or without paclitaxel. RNA was extracted from 321 formalin-fixed paraffin-embedded primary tumor tissue samples and quantitative reverse-transcriptase polymerase chain reaction was used to assess messenger RNA (mRNA) expression of CXCL12, CXCL13, and CXCR5; estrogen receptor; progesterone receptor (PgR); microtubule-associated protein tau and human epidermal growth factor receptor 2 (HER2). RESULTS:CXCL13 and CXCR5 were found to be negatively associated with estrogen receptor and microtubule-associated protein tau mRNA expression and with dense lymphocytic infiltration, and were positively associated with nuclear grade. Only CXCL13 was positively associated with HER2. Multivariate analysis revealed an association between high CXCL13 mRNA expression and improved DFS (hazard ratio [HR] 0.48 [95% CI, 0.25-0.90]; Wald, P = .023) but not OS; whereas high CXCL12 expression was significantly associated with increased OS (HR 0.53 [95% CI, 0.33-0.85]; Wald, P = .009). In the HER2 mRNA overexpressing subgroup, high CXCL13 mRNA expression was associated with improved DFS (P < .001), whereas high CXCR5 was associated with increased DFS and OS (P = .004 and P = .049, respectively). CONCLUSIONS:The CXCL13-CXCR5 axis is associated with classic determinants of poor prognosis, such as high grade, hormone receptor negativity, and axillary node involvement. Interestingly, this chemokine axis seems to be strongly associated with improved outcome in patients with HER2(+) disease.
CXCL13-CXCR5 axis promotes the growth and invasion of colon cancer cells via PI3K/AKT pathway.
Zhu Zhenyu,Zhang Xukui,Guo Hongliang,Fu Ling,Pan Ganlin,Sun Yinggang
Molecular and cellular biochemistry
CXCL13, an inflammatory factor in the microenvironment, plays a vital role in the progression of inflammatory diseases and tumors. CXCL13 and its receptor CXCR5 have been reported to be associated with poor prognosis of advanced colon cancer. However, the molecular mechanisms of CXCL13-CXCR5 axis in colon cancer remain elusive. The aim of this study was to investigate the role of CXCR5-CXCL13 axis in the growth and invasion of colon cancer cells. Our results showed that CXCL13 promoted the growth, migration, and matrigel invasion of colon cancer cells. Furthermore, CXCL13 increased the expression and secretion of MMP-13, and stimulated the activation of PI3K/AKT pathway. After knockdown of CXCR5 by siRNA, the biological functions of colon cancer cells regulated by CXCL13 were significantly inhibited. In addition, inhibition of PI3K/AKT pathway by specific inhibitor LY294002 suppressed the CXCL13-mediated growth, migration, and invasion of colon cancer cells. Together, our findings suggest that CXCL13-CXCR5 axis promotes the growth, migration, and invasion of colon cancer cells, probably via PI3K/AKT pathway. Thus, CXCL13 may be a useful biomarker for the detection and treatment of colon cancer.
RelA driven co-expression of CXCL13 and CXCR5 is governed by a multifaceted transcriptional program regulating breast cancer progression.
Biswas Subir,Roy Chowdhury Sougata,Mandal Gunjan,Purohit Suman,Gupta Arnab,Bhattacharyya Arindam
Biochimica et biophysica acta. Molecular basis of disease
Lethal metastasis of primary breast tumors to lymph nodes has been found to be associated with the co-expression of chemokine CXCL13 and its receptor CXCR5. To date, however, the precise molecular events regulating the co-expression of CXCL13 and CXCR5 in the context of breast cancer progression have not been identified. Therefore, to extend our understanding of the drivers of breast cancer metastasis, we undertook a line of investigation in this study in which we demonstrate that the transcriptional regulation of CXCL13 is mediated by the reciprocal activity of RelA and Nrf2, while CXCR5 is transcriptionally silenced by CpG island methylation within its promoter. Critically, we show that intra-tumoral CXCL13 and CXCR5 mRNA expression is positively correlated with intra-tumoral RelA expression within the primary tumor of breast cancer (BCa) patients (n = 98). We demonstrate a role for Nrf2 in the negative transcriptional regulation of cxcl13. Furthermore, using a luciferase assay and deletion analysis of the cxcl13 gene promoter, we demonstrate that RelA and Nrf2 directly act upon the cxcl13 promoter to regulate transcription. Chromatin immunoprecipitation PCR, supported by in silico docking analyses, confirmed that RelA and Nrf2 both occupy multiple positions within the cxcl13 promoter. Collectively, in RelA high conditions, low Nrf2 and lack of cxcr5 promoter DNA-methylation govern CXCL13-CXCR5 co-expression within breast tumors, and thus drive disease progression and metastasis.
CXCL13 and Its Receptor CXCR5 in Cancer: Inflammation, Immune Response, and Beyond.
Kazanietz Marcelo G,Durando Michael,Cooke Mariana
Frontiers in endocrinology
It is well-established that the chemokine C-X-C motif ligand 13 (CXCL13) and its receptor, the G-protein coupled receptor (GPCR) CXCR5, play fundamental roles in inflammatory, infectious and immune responses. Originally identified as a B-cell chemoattractant, CXCL13 exerts important functions in lymphoid neogenesis, and has been widely implicated in the pathogenesis of a number of autoimmune diseases and inflammatory conditions, as well as in lymphoproliferative disorders. Current evidence also indicates that the CXCL13:CXCR5 axis orchestrates cell-cell interactions that regulate lymphocyte infiltration within the tumor microenvironment, thereby determining responsiveness to cytotoxic and immune-targeted therapies. In this review, we provide a comprehensive perspective of the involvement of CXCL13 and its receptor in cancer progression. Studies in recent years postulated novel roles for this chemokine in controlling the cancer cell phenotype, and suggest important functions in the growth and metastatic dissemination of solid tumors. Carcinogens have been found to induce CXCL13 production, and production of this chemokine within the tumor milieu has been shown to impact the proliferation, migration, and invasive properties of cancer cells. Thus, the complex networks of cellular interactions involving tumoral CXCL13 and CXCR5 integrate to promote cancer cell autonomous and non-autonomous responses, highlighting the relevance of autocrine and paracrine interactions in dictating the cancer phenotype. Dissecting the molecular and signaling events regulated by CXCL13 and how this chemokine dynamically controls the interaction between the cancer cell and the tumor microenvironment is key to identify novel effectors and therapeutic targets for cancer treatment.
CXCL13 Signaling in the Tumor Microenvironment.
Hussain Muzammal,Liu Jinsong,Wang Gui-Zhen,Zhou Guang-Biao
Advances in experimental medicine and biology
Chemokines have emerged as important players in tumorigenic process. An extensive body of literature generated over the last two or three decades strongly implicate abnormally activated or functionally disrupted chemokine signaling in liaising most-if not all-hallmark processes of cancer. It is well-known that chemokine signaling networks within the tumor microenvironment are highly versatile and context-dependent: exert both pro-tumoral and antitumoral activities. The C-X-C motif chemokine ligand 13 (CXCL13), and its cognate receptor CXCR5, represents an emerging example of chemokine signaling axes, which express the ability to modulate tumor growth and progression in either way. Collateral evidence indicate that CXCL13-CXCR5 axis may directly modulate tumor growth by inducing proliferation of cancer cells, as well as promoting invasive phenotypes and preventing their apoptosis. In addition, CXCL13-CXCR5 axis may also indirectly modulate tumor growth by regulating noncancerous cells, particularly the immune cells, within the tumor microenvironment. Here, we review the role of CXCL13, together with CXCR5, in the human tumor microenvironment. We first elaborate their patterns of expression, regulation, and biological functions in normal physiology. We then consider how their aberrant activity, as a result of differential overexpression or co-expression, may directly or indirectly modulate the growth of tumors through effects on both cancerous and noncancerous cells.
CXCL13-CXCR5 co-expression regulates epithelial to mesenchymal transition of breast cancer cells during lymph node metastasis.
Biswas Subir,Sengupta Suman,Roy Chowdhury Sougata,Jana Samir,Mandal Gunjan,Mandal Palash Kumar,Saha Nipun,Malhotra Vivek,Gupta Arnab,Kuprash Dmitry V,Bhattacharyya Arindam
Breast cancer research and treatment
We investigated the expression of -CXC chemokine ligand 13 (CXCL13) and its receptor -CXC chemokine receptor 5 (CXCR5) in 98 breast cancer (BC) patients with infiltrating duct carcinoma, out of which 56 were found lymph node metastasis (LNM) positive. Interestingly, co-expression of CXCL13 and CXCR5 showed a significant correlation with LNM. Since, epithelial to mesenchymal transition (EMT) is highly associated with metastasis we investigated EMT-inducing potential of CXCL13 in BC cell lines. In CXCL13-stimulated BC cells, expression of various mesenchymal markers (Vimentin, N-cadherin), EMT regulators (Snail, Slug), and matrix metalloproteinase-9 (MMP9) was increased, whereas the expression of epithelial marker E-cadherin was found to be decreased. In addition, expression of receptor activator of nuclear factor kappa-B ligand (RANKL), which is known to regulate MMP9 expression via Src activation, was also significantly increased after CXCL13 stimulation. Using specific protein kinase inhibitors, we confirmed that CXCL13 stimulated EMT and MMP9 expression via RANKL-Src axis in BC cell lines. To further validate this observation, we examined gene expression patterns in primary breast tumors and detected significantly higher expression of various mesenchymal markers and regulators in CXCL13-CXCR5 co-expressing patients. Therefore, this study showed the EMT-inducing potential of CXCL13 as well as demonstrated the prognostic value of CXCL13-CXCR5 co-expression in primary BC. Moreover, CXCL13-CXCR5-RANKL-Src axis may present a therapeutic target in LNM positive BC patients.
Transcriptome Network Analysis Identifies CXCL13-CXCR5 Signaling Modules in the Prostate Tumor Immune Microenvironment.
Ohandjo Adaugo Q,Liu Zongzhi,Dammer Eric B,Dill Courtney D,Griffen Tiara L,Carey Kaylin M,Hinton Denise E,Meller Robert,Lillard James W
The tumor immune microenvironment (TIME) consists of multiple cell types that contribute to the heterogeneity and complexity of prostate cancer (PCa). In this study, we sought to understand the gene-expression signature of patients with primary prostate tumors by investigating the co-expression profiles of patient samples and their corresponding clinical outcomes, in particular "disease-free months" and "disease reoccurrence". We tested the hypothesis that the CXCL13-CXCR5 axis is co-expressed with factors supporting TIME and PCa progression. Gene expression counts, with clinical attributes from PCa patients, were acquired from TCGA. Profiles of PCa patients were used to identify key drivers that influence or regulate CXCL13-CXCR5 signaling. Weighted gene co-expression network analysis (WGCNA) was applied to identify co-expression patterns among CXCL13-CXCR5, associated genes, and key genetic drivers within the CXCL13-CXCR5 signaling pathway. The processing of downloaded data files began with quality checks using NOISeq, followed by WGCNA. Our results confirmed the quality of the TCGA transcriptome data, identified 12 co-expression networks, and demonstrated that CXCL13, CXCR5 and associated genes are members of signaling networks (modules) associated with G protein coupled receptor (GPCR) responsiveness, invasion/migration, immune checkpoint, and innate immunity. We also identified top canonical pathways and upstream regulators associated with CXCL13-CXCR5 expression and function.
CXCL13/CXCR5 signaling axis in cancer.
Hussain Muzammal,Adah Dickson,Tariq Muqddas,Lu Yongzhi,Zhang Jiancun,Liu Jinsong
The tumor microenvironment comprises stromal and tumor cells which interact with each other through complex cross-talks that are mediated by a variety of growth factors, cytokines, and chemokines. The chemokine ligand 13 (CXCL13) and its chemokine receptor 5 (CXCR5) are among the key chemotactic factors which play crucial roles in deriving cancer cell biology. CXCL13/CXCR5 signaling axis makes pivotal contributions to the development and progression of several human cancers. In this review, we discuss how CXCL13/CXCR5 signaling modulates cancer cell ability to grow, proliferate, invade, and metastasize. Furthermore, we also discuss the preliminary evidence on context-dependent functioning of this axis within the tumor-immune microenvironment, thus, highlighting its potential dichotomy with respect to anticancer immunity and cancer immune-evasion mechanisms. At the end, we briefly shed light on the therapeutic potential or implications of targeting CXCL13/CXCR5 axis within the tumor microenvironment.
CXCL13-mediated recruitment of intrahepatic CXCR5CD8 T cells favors viral control in chronic HBV infection.
Li Yongyin,Tang Libo,Guo Ling,Chen Chengcong,Gu Shuqin,Zhou Yang,Ye Guofu,Li Xiaoyi,Wang Weibin,Liao Xinxin,Wang Yu,Peng Xiaohong,Liu Guangze,Zhang Xiaoyong,Sun Jian,Peng Jie,Hou Jinlin
Journal of hepatology
BACKGROUND & AIMS:Although CD8T cell exhaustion hampers viral control during chronic HBV infection, the pool of CD8T cells is phenotypically and functionally heterogeneous. Therefore, a specific subpopulation of CD8T cells should be further investigated. This study aims to dissect a subset of CD8T cells expressing C-X-C motif chemokine receptor 5 (CXCR5) in chronic HBV infection. METHODS:The frequency of CXCR5CD8T cells and the levels of C-X-C motif chemokine ligand 13 (CXCL13), a chemokine of CXCR5, were measured in patients with chronic HBV infection. C57BL/6, interleukin (IL)-21 receptor- or B cell-deficient mice were hydrodynamically injected with pAAV-HBV1.2 plasmids. Phenotype and functions of peripheral and intrahepatic CXCR5 and CXCR5CD8T cells were assessed. RESULTS:CXCR5CD8T cells were partially exhausted but possessed a stronger antiviral ability than the CXCR5 subset in patients with chronic HBV infection; moreover, CXCR5CD8T cells were associated with a favorable treatment response in patients with chronic hepatitis B (CHB). High levels of CXCL13 from patients with CHB facilitated the recruitment of intrahepatic CXCR5CD8T cells, and this subpopulation produced high levels of HBV-specific interferon (IFN)-γ and IL-21. Notably, PD1 (programmed death 1) blockade and exogenous IL-21 enhanced the production of IFN-γ. More strikingly, mice injected with CXCR5CD8T cells showed remarkably decreased expression of HBsAg. Additionally, an impaired production of HBV-specific IFN-γ from intrahepatic CXCR5CD8T cells was observed in IL-21 receptor- or B cell-deficient mice. CONCLUSION:CXCL13 promotes the recruitment of CXCR5CD8T cells to the liver, and this subpopulation improves viral control in chronic HBV infection. The identification of this unique subpopulation may contribute to a better understanding of CD8T cell functions and provide a potential immunotherapeutic target in chronic HBV infection. LAY SUMMARY:Exhaustion of CD8 T cells is an important factor in the development of chronic hepatitis B virus (HBV) infection. CD8 T cells expressing the receptor CXCR5 are partially exhausted, but have potent antiviral activity, as they produce high levels of HBV-specific cytokines in chronic HBV infection. Increased expression of CXCL13 within the liver facilitates the recruitment of CXCR5CD8T cells and establishes effective immune control of HBV infection.
Characterization of CXCR5 CD8 T-cells in humanized NSG mice.
Perdomo-Celis Federico,Medina-Moreno Sandra,Davis Harry,Bryant Joseph,Taborda Natalia A,Rugeles Maria T,Kottilil Shyamasundaram,Zapata Juan C
Humanized NOD/SCID/IL-2 receptor γ-chain (huNSG) mice recapitulate some features of human T-cell populations that can be exploited in basic and pre-clinical research. CXCR5 T CD8 T-cells play an important role in the control of viral infections and tumors. Indeed, they have been associated with low-level HIV replication, making them a possible novel correlate of protection, and potentially useful in the eradication of HIV reservoirs. Here, by flow cytometry, we evaluated the reconstitution of CXCR5 CD8 T-cells in huNSG mice engrafted with CD34 hematopoietic stem cells. This population was readily generated in huNSG mice, and where particularly confined to spleen and lymph nodes. These cells exhibited a follicular-like phenotype, with expression of Programmed Death (PD)-1, Inducible T-cell costimulatory (ICOS), and absence of CCR7. Moreover, CXCR5 CD8 T-cells had a higher expression of interleukin (IL)-21 and a higher cytotoxic potential compared with CXCR5 cells. HIV infection did not affect the frequencies of CXCR5 CD8 T-cells in secondary lymphoid organs. Finally, taking advantage of the high proportion of naïve T-cells in huNSG mice, we evaluated the in vitro response of splenic T-cells to the follicular profile-polarizing cytokines Transforming Growth Factor (TGF)-β1 and IL-23. After in vitro treatment, there was an increase in CXCR5 CD8 T-cells, which exhibited high levels of PD-1, CD40 L and low expression of CCR7. Thus, there is a reconstitution of CXCR5 CD8 T-cells in huNSG mice, supporting the use of this model for exploring the biology and role of this cell population in healthy and diseased conditions.
Alleviating effect of paeoniflorin-6'-O-benzene sulfonate in antigen-induced experimental Sjögren's syndrome by modulating B lymphocyte migration via CXCR5-GRK2-ERK/p38 signaling pathway.
Chen Xiaoyun,Zhang Pengying,Liu Qi,Zhang Qiaolin,Gu Fang,Xu Shixia,Körner Heinrich,Wu Huaxun,Wei Wei
Primary Sjögren's syndrome (pSS) is an autoimmune disease of unresolved aetiology that affects the exocrine glands. Clinical symptoms frequently also involve skin, liver, kidney and neurovascular components. The pathogenesis of pSS is still unclear but B cell hyperactivity has been identified as a hallmark of pSS. Currently, a curative therapeutic agent is lacking. In this study, we explored whether paeoniflorin-6'-O-benzene (CP-25) exerted therapeutic effects through regulating B lymphocyte migration via CXCR5-GRK2-MAPK mediated signaling pathways in a mouse model of antigen-induced, experimental Sjögren's syndrome (ESS). We found that CP-25 increased the salivary flow and alleviated the histopathology of ESS. Furthermore, CP-25 reduced the viability of B lymphocyte and limited the target organs index. In the peripheral blood and salivary gland of ESS mice, CP-25 down-regulated the proportion of total B cells, CXCR5+ B cells and PDCA1 + CD19- and limited the presence of phosphorylated (p-) p38 and ERK (p-ERK). Besides, CP-25 increased the percentage of memory B cells in the peripheral blood and reduced it in salivary gland. Furthermore, in vitro, CP-25 down-regulated p-p38, p-ERK, CXCR5 and membrane GRK2, and increased cytoplasm GRK2 in Maver-1 cells, a mantle cell lymphoma cell line, causing a lower migration ability of Maver-1 cells. Thus, we define CP-25 as a novel compound that is a potent therapeutic agent for pSS which modulates B lymphocyte subsets and impacts the migration of B lymphocytes through regulating the CXCR5-GRK2-ERK/p38 signaling pathway.
CXCR5/NRF2 double knockout mice develop retinal degeneration phenotype at early adult age.
Huang Hu,Lennikov Anton
Experimental eye research
The objective of this study is to characterize the retinal degeneration (RD) phenotype of CXCR5/NRF2 double knockout (DKO) mice at the early adult age. CXCR5 KO mice and NRF2 KO mice were bred to create CXCR5/NRF2 DKO mice. The assessment of RD features included fundus and optical coherence tomography (OCT) imaging, periodic acid-Schiff (PAS), and immunofluorescence staining of retinal pigment epithelium (RPE)-choroid flatmounts. Stained samples were imaged with fluorescent microscopy, and Western blots were used to monitor protein expression changes. The staining of cleaved caspase-3 and PNA-lectin was performed to assess the presence of photoreceptor cell apoptosis. Quantification and statistical analyses were performed with Image J and Graphpad software. The young adult (2-6 months) DKO mice exhibited increased hypopigmented spots on fundus and sub-RPE abnormalities on OCT as compared to the CXCR5-KO mice, and C57BL6 WT controls. PAS-stained sections demonstrated aberrant RPE/sub-RPE depositions. The DKO mice had increased sub-RPE depositions of IgG and AMD-associated proteins (β-amyloid, Apolipoprotein-E, C5b-9, and αB-crystallin). The protein expression of AMD-associated proteins and microglia marker (TMEM119) were upregulated at the RPE/BM/choroid complex of DKO mice. The adult DKO mice underwent photoreceptor cell apoptosis compared to the single CXCR5 and NRF2 KO and the WT mice at an early adult age. Mechanistically increased expression of CXCL13 and N-cadherin was observed as a sign of epithelial-mesenchymal transition. The data suggest that the CXCR5/NRF2-DKO mice develop RD characteristics at an early age and may serve as a valuable animal model of RD.
Transforming growth factor-β promotes the function of HIV-specific CXCR5 CD8 T cells.
Yang Hong-Ge,Jiao Yan-Mei,Huang Hui-Huang,Zhang Chao,Zhang Ji-Yuan,Xu Ruo-Nan,Song Jin-Wen,Fan Xing,Jin Lei,Shi Ming,Wang Fu-Sheng
Microbiology and immunology
HIV replication can be inhibited by CXCR5 CD8 T cells (follicular cytotoxic T cell [TFC]) which transfer into B-cell follicles where latent HIV infection persists. However, how cytokines affect TFC remain unclear. Understanding which cytokines show the ability to affect TFC could be a key strategy toward curing HIV. Similar mechanisms could be used for the growth and transfer of TFCs and follicular helper T (TFH) cells; as a result, we hypothesized that cytokines IL-6, IL-21, and transforming growth factor-β (TGF-β), which are necessary for the differentiation of TFH cells, could also dictate the development of TFCs. In this work, lymph node mononuclear cells and peripheral blood mononuclear cells from HIV-infected individuals were cocultured with IL-6, IL-21, and TGF-β. We then carried out T-cell receptor (TCR) repertoire analysis to compare the differences between CXCR5 and CXCR5 CD8 T cells. Our results showed that the percentage and function of TFC can be enhanced by stimulation with TGF-β. Besides, TGF-β stimulation enhanced the diversity of TCR and complementarity-determining region 3 sequences. HIV DNA showed a negative correlation with TFC. The use of TGF-β to promote the expression of CXCR5 CD8 T cells could become a new treatment approach for curing HIV.
Increased CXCL13 and CXCR5 in Anterior Cingulate Cortex Contributes to Neuropathic Pain-Related Conditioned Place Aversion.
Wu Xiao-Bo,He Li-Na,Jiang Bao-Chun,Wang Xue,Lu Ying,Gao Yong-Jing
Pain consists of sensory-discriminative and emotional-affective components. The anterior cingulate cortex (ACC) is a critical brain area in mediating the affective pain. However, the molecular mechanisms involved remain largely unknown. Our recent study indicated that C-X-C motif chemokine 13 (CXCL13) and its sole receptor CXCR5 are involved in sensory sensitization in the spinal cord after spinal nerve ligation (SNL). Whether CXCL13/CXCR5 signaling in the ACC contributes to the pathogenesis of pain-related aversion remains unknown. Here, we showed that SNL increased the CXCL13 level and CXCR5 expression in the ACC after SNL. Knockdown of CXCR5 by microinjection of Cxcr5 shRNA into the ACC did not affect SNL-induced mechanical allodynia but effectively alleviated neuropathic pain-related place avoidance behavior. Furthermore, electrophysiological recording from layer II-III neurons in the ACC showed that SNL increased the frequency and amplitude of spontaneous excitatory postsynaptic currents (sEPSCs), decreased the EPSC paired-pulse ratio, and increased the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor/N-methyl-D-aspartate receptor ratio, indicating enhanced glutamatergic synaptic transmission. Finally, superfusion of CXCL13 onto ACC slices increased the frequency and amplitude of spontaneous EPSCs. Pre-injection of Cxcr5 shRNA into the ACC reduced the increase in glutamatergic synaptic transmission induced by SNL. Collectively, these results suggest that CXCL13/CXCR5 signaling in the ACC is involved in neuropathic pain-related aversion via synaptic potentiation.
Identification and validation of an excellent prognosis subtype of muscle-invasive bladder cancer patients with intratumoral CXCR5 CD8 T cell abundance.
Huang Qiuren,Zhou Quan,Zhang Hongyu,Liu Zhaopei,Zeng Han,Chen Yifan,Qu Yang,Xiong Ying,Wang Jiajun,Chang Yuan,Xia Yu,Wang Yiwei,Liu Li,Zhu Yu,Xu Le,Dai Bo,Guo Jianming,Wang Zewei,Bai Qi,Zhang Weijuan
Bladder cancer is the ninth most frequent-diagnosed disease worldwide, bearing high morbidity and mortality rates. Studies have shown that a particular population of CXCR5CD8 T cells was associated with superior prognosis in various tumor types, and yet its role in muscle-invasive bladder cancer (MIBC) remains unclear. In this study, 662 MIBC patients from 3 cohorts (Zhongshan Hospital, = 141; Shanghai Cancer Center, = 108; The Cancer Genome Atlas, = 403) were analyzed retrospectively. 11 fresh resected samples of MIBC were examined to characterize the phenotype of CXCR5CD8 T cells and 402 MIBC patients from TCGA were applied for bioinformatics analysis. It was explored that the abundance of intratumoral CXCR5CD8 T cells indicated superior overall survival and disease-free survival. Patients with a higher infiltration of CXCR5CD8 T cells in tumor tissue benefit more from adjuvant chemotherapy (ACT). Intratumoral CXCR5CD8 T cells displayed cytolytic and self-renewal features. Remarkably, CXCR5CD8 T cells were mainly presented in the basal and stromal-rich subtypes of MIBC and tumors with enriched CXCR5CD8 T cells showed limited FGFR3 signaling signature and activated immunotherapeutic and EGFR associated pathway. In conclusion, we identified an excellent prognosis and ACT sensitive subtype of MIBC with intratumoral CXCR5CD8 T cell abundance. Tumors with high density of CXCR5CD8 T cells possessed potential sensitivity to immunotherapy and EGFR-targeted therapy. CXCR5CD8 T cells provide a new potential biomarker as well as a therapeutic target in MIBC.
CD40 Signaling Promotes CXCR5 Expression in B Cells via Noncanonical NF-B Pathway Activation.
Wei Chuan,Chen Ying,Xu Lei,Yu Beibei,Lu Di,Yu Yanxiong,Lei Zhigang,Tang Rui,Zhou Sha,Zhu Jifeng,Chen Xiaojun,Su Chuan
Journal of immunology research
Chemokine receptor CXCR5-mediated control of B cell trafficking in the lymphoid tissues plays a central role in orchestrating the B cell function, which not only guides the colocalization of B cells with follicular helper T cells in the follicular mantle zone but also determines the position of germinal center dark and light zones. However, the mechanisms that regulate the expression of CXCR5 in B cells remain unclear. Here, we show that the expression level of CXCR5 in B cells was substantially reduced in vitro culture conditions, while being maintained in the presence of CD40 signals. Furthermore, CD40 signaling promotes CXCR5 expression in B cells at least partially through noncanonical NF-B signaling pathway activation. However, other non-B cells also contributed to the optimal expression of CXCR5 in B cells through cell-cell contact and cytokine secretion. Our findings suggest that CD40 signaling-mediated activation of the noncanonical NF-B pathway promotes the expression of CXCR5 in a B cell-intrinsic way to orchestrate the trafficking of B cells.
Chemokine CXCL13 acts via CXCR5-ERK signaling in hippocampus to induce perioperative neurocognitive disorders in surgically treated mice.
Shen Yanan,Zhang Yuan,Chen Lihai,Du Jiayue,Bao Hongguang,Xing Yan,Cai Mengmeng,Si Yanna
Journal of neuroinflammation
BACKGROUND:Perioperative neurocognitive disorders (PNDs) occur frequently after surgery and worsen patient outcome. How C-X-C motif chemokine (CXCL) 13 and its sole receptor CXCR5 contribute to PNDs remains poorly understood. METHODS:A PND model was created in adult male C57BL/6J and CXCR5 mice by exploratory laparotomy. Mice were pretreated via intracerebroventricular injection with recombinant CXCL13, short hairpin RNA against CXCL13 or a scrambled control RNA, or ERK inhibitor PD98059. Then surgery was performed to induce PNDs, and animals were assessed in the Barnes maze trial followed by a fear-conditioning test. Expression of CXCL13, CXCR5, and ERK in hippocampus was examined using Western blot, quantitative PCR, and immunohistochemistry. Levels of interleukin-1 beta (IL-1β) and tumor necrosis factor alpha (TNF-α) in hippocampus were assessed by Western blot. RESULTS:Surgery impaired learning and memory, and it increased expression of CXCL13 and CXCR5 in the hippocampus. CXCL13 knockdown partially reversed the effects of surgery on CXCR5 and cognitive dysfunction. CXCR5 knockout led to similar cognitive outcomes as CXCL13 knockdown, and it repressed surgery-induced activation of ERK and production of IL-1β and TNF-α in hippocampus. Recombinant CXCL13 induced cognitive deficits and increased the expression of phospho-ERK as well as IL-1β and TNF-α in hippocampus of wild-type mice, but not CXCR5 mice. PD98059 partially blocked CXCL13-induced cognitive dysfunction as well as production of IL-1β and TNF-α. CONCLUSIONS:CXCL13-induced activation of CXCR5 may contribute to PNDs by triggering ERK-mediated production of pro-inflammatory cytokines in hippocampus.
CXCL13/CXCR5 signalling is pivotal to preserve motor neurons in amyotrophic lateral sclerosis.
Trolese Maria Chiara,Mariani Alessandro,Terao Mineko,de Paola Massimiliano,Fabbrizio Paola,Sironi Francesca,Kurosaki Mami,Bonanno Silvia,Marcuzzo Stefania,Bernasconi Pia,Trojsi Francesca,Aronica Eleonora,Bendotti Caterina,Nardo Giovanni
BACKGROUND:CXCL13 is a B and T lymphocyte chemokine that mediates neuroinflammation through its receptor CXCR5. This chemokine is highly expressed by motoneurons (MNs) in Amyotrophic Lateral Sclerosis (ALS) SOD1G93A (mSOD1) mice during the disease, particularly in fast-progressing mice. Accordingly, in this study, we investigated the role of this chemokine in ALS. METHODS:We used in vitro and in vivo experimental paradigms derived from ALS mice and patients to investigate the expression level and distribution of CXCL13/CXCR5 axis and its role in MN death and disease progression. Moreover, we compared the levels of CXCL13 in the CSF and serum of ALS patients and controls. FINDINGS:CXCL13 and CXCR5 are overexpressed in the spinal MNs and peripheral axons in mSOD1 mice. CXCL13 inhibition in the CNS of ALS mice resulted in the exacerbation of motor impairment (n = 4/group;Mean_Diff.=27.81) and decrease survival (n = 14_Treated:19.2 ± 1.05wks, n = 17_Controls:20.2 ± 0.6wks; 95% CI: 0.4687-1.929). This was corroborated by evidence from primary spinal cultures where the inhibition or activation of CXCL13 exacerbated or prevented the MN loss. Besides, we found that CXCL13/CXCR5 axis is overexpressed in the spinal cord MNs of ALS patients, and CXCL13 levels in the CSF discriminate ALS (n = 30) from Multiple Sclerosis (n = 16) patients with a sensitivity of 97.56%. INTERPRETATION:We hypothesise that MNs activate CXCL13 signalling to attenuate CNS inflammation and prevent the neuromuscular denervation. The low levels of CXCL13 in the CSF of ALS patients might reflect the MN dysfunction, suggesting this chemokine as a potential clinical adjunct to discriminate ALS from other neurological diseases. FUNDING:Vaccinex, Inc.; Regione Lombardia (TRANS-ALS).
The exhausted CD4CXCR5 T cells involve the pathogenesis of human tuberculosis disease.
Jean Bosco Munyemana,Wei Ming,Hou Hongyan,Yu Jing,Lin Qun,Luo Ying,Sun Ziyong,Wang Feng
International journal of infectious diseases : IJID : official publication of the International Society for Infectious Diseases
OBJECTIVES:CD4CXCR5 T cells have previously been established. However, their decreased frequency during tuberculosis (TB) disease is only partially understood. The aim of this study was to explore the depletion of CD4CXCR5 T cells in human TB. METHODS:The frequency and function of CD4CXCR5 T cells were evaluated in active TB (ATB) patients and healthy control subjects. The function of CD4CXCR5 T cells was determined after blockade of inhibitory receptors. RESULTS:The frequency of CD4CXCR5 T cells was decreased in ATB patients. The expression of activation markers (HLA-DR and ICOS) and inhibitory receptors (Tim-3 and PD-1) on CD4CXCR5 T cells was increased in the ATB group. TB-specific antigen stimulation induced higher expression of inhibitory receptors than phytohemagglutinin stimulation in the ATB group. In contrast, TB antigen stimulation did not induce a significantly increased expression of IL-21 and Ki-67 on CD4CXCR5 T cells. However, blockade of inhibitory receptors Tim-3 and PD-1 not only increased the frequency of CD4CXCR5 T cells, but also restored their proliferation and cytokine secretion potential. CONCLUSIONS:The increased expression of inhibitory receptors involves the depletion of CD4CXCR5 T cells, and blockade of inhibitory receptors can restore the function of CD4CXCR5 T cells in ATB patients.
Improved Functionality of Exhausted Intrahepatic CXCR5+ CD8+ T Cells Contributes to Chronic Antigen Clearance Upon Immunomodulation.
Kumashie Kingsley Gideon,Cebula Marcin,Hagedorn Claudia,Kreppel Florian,Pils Marina C,Koch-Nolte Friedrich,Rissiek Björn,Wirth Dagmar
Frontiers in immunology
Chronic hepatotropic viral infections are characterized by exhausted CD8+ T cells in the presence of cognate antigen in the liver. The impairment of T cell response limits the control of chronic hepatotropic viruses. Immune-modulatory strategies are attractive options to re-invigorate exhausted T cells. However, in hepatotropic viral infections, the knowledge about immune-modulatory effects on the in-situ regulation of exhausted intrahepatic CD8+ T cells is limited. In this study, we elucidated the functional heterogeneity in the pool of exhausted CD8+ T cells in the liver of mice expressing the model antigen Ova in a fraction of hepatocytes. We found a subpopulation of intrahepatic CXCR5+ Ova-specific CD8+ T cells, which are profoundly cytotoxic, exhibiting efficient metabolic functions as well as improved memory recall and self-maintenance. The intrahepatic Ova-specific CXCR5+ CD8+ T cells are possibly tissue resident cells, which may rely largely on OXPHOS and glycolysis to fuel their cellular processes. Importantly, host conditioning with CpG oligonucleotide reinvigorates and promotes exhausted T cell expansion, facilitating complete antigen eradication. The CpG oligonucleotide-mediated reinvigoration may support resident memory T cell formation and the maintenance of CXCR5+ Ova-specific CD8+ T cells in the liver. These findings suggest that CpG oligodinucleotide may preferentially target CXCR5+ CD8+ T cells for expansion to facilitate the revival of exhausted T cells. Thus, therapeutic strategies aiming to expand CXCR5+ CD8+ T cells might provide a novel approach against chronic liver infection.
Human CXCR5 PD-1 CD8 T cells in healthy individuals and patients with hematologic malignancies.
Hofland Tom,Martens Anne W J,van Bruggen Jaco A C,de Boer Renate,Schetters Sjoerd,Remmerswaal Ester B M,Bemelman Frederike J,Levin Mark-David,Bins Adriaan D,Eldering Eric,Kater Arnon P,Tonino Sanne H
European journal of immunology
Immune checkpoint blockade (ICB) has revolutionized cancer therapy, but varying response rates illustrate the need for biomarkers of response. Studies in mice have identified a subset of CD8 T cells that is essential for response to PD-1 ICB. These CD8 T cells co-express CXCR5, PD-1 and Tcf1, and provide effector T cells upon PD-1 ICB. It is unknown whether similar T cells play a role in PD-1 ICB in humans. We studied human peripheral blood and lymph nodes (LNs) for the frequency, phenotype, and functionality of CXCR5 PD-1 CD8 T cells. We find that CXCR5 PD-1 CD8 T cells are memory-like cells, express Tcf1, and lack expression of effector molecules. CXCR5 PD-1 CD8 T cells produce cytokines upon stimulation, but have limited proliferative capacity. We studied patients with hematologic malignancies with varying response rates to PD-1 ICB. Specifically in chronic lymphocytic leukemia, in which PD-1 ICB does not induce clinical responses, CXCR5 PD-1 CD8 T cells show loss of the memory phenotype and increased effector differentiation. In conclusion, we identified CXCR5 PD-1 CD8 T cells in human peripheral blood and LN, which could play a similar role during PD-1 ICB. Future studies should analyze CXCR5 PD-1 CD8 T cells during PD-1 ICB and their importance for therapeutic response.
Possible roles of CXCL13/CXCR5 axis in the development of bullous pemphigoid.
Ohuchi Kentaro,Fujimura Taku,Lyu Chunbing,Amagai Ryo,Muto Yusuke,Aiba Setsuya
The Journal of dermatology
CXCL13 recruits CXCR5 follicular helper T (Tfh) cells in inflammatory lesions to develop secondary lymphoid organs. Tfh cells activate B cells to produce antibodies during humoral immune responses. Indeed, as previous reports suggested, CXCR5 cell numbers were increased in the peripheral blood of bullous pemphigoid (BP) patients when compared with healthy donors, and the ratio of CXCR5 cells was positively correlated with the anti-BP180-NC16A titers. From the above findings, in this report, we hypothesized that a chemokine related to CXCR5 cells, namely CXCL13, may play a role in the development of BP. We performed immunohistochemical staining of CXCR5, CXCL13, LL37, CXCL10 and CCL20 for 10 cases of BP and 10 cases of pemphigus vulgaris (PV), and quantitatively analyzed the staining by digital microscopy. Moreover, we investigated the CXCL10 and CXCL13 production in BP and PV patients by enzyme-linked immunosorbent assay. The immunomodulatory effects of LL37 on the production of T-helper 17-related chemokines were evaluated using monocyte-derived M2 macrophages. Immunohistochemical staining and digital microscopic analysis showed that the ratios of CXCR5 , CXCL13 and LL37 cells in the dermis were significantly higher in BP patients than in PV patients. Notably, the ratio of CXCL13 cells was positively correlated with the anti-BP180-NC16A titers. Moreover, the serum levels of CXCL13 were positively correlated with the anti-BP180-NC16A titers. Furthermore, CD163 M2 macrophages stimulated by LL37 in vitro produced CXCL10 and CCL20. In the lesional skin of BP, CD163 macrophages CXCL10 and CCL20 were produced. The serum levels of CXCL10 were negatively correlated with the anti-BP180-NC16A titers. The present study results indicate that the mechanism of the development of BP may involve the CXCL13/CXCR5-mediated migration of Tfh cells.
Autoimmune-Mediated Retinopathy in CXCR5-Deficient Mice as the Result of Age-Related Macular Degeneration Associated Proteins Accumulation.
Lennikov Anton,Saddala Madhu Sudhana,Mukwaya Anthony,Tang Shibo,Huang Hu
Frontiers in immunology
Previous research has shown that CXCR5 mice develop retinal degeneration (RD) with age, a characteristic related to age macular degeneration (AMD). RD in these mice is not well-understood, and in this study, we sought to characterize further the RD phenotype and to gain mechanistic insights into the function of CXCR5 in the retina. CXCR5 and WT control mice were used. Fundus images demonstrated a significant ( < 0.001) increase of hypo-pigmented spots in the retina of aged CXCR5 mice compared with WT control mice. PAS staining indicated localization of deposits in the sub-retinal pigment epithelia (RPE) layer. AMD-associated proteins Cryab, amyloid beta, and C3d were detected within the RPE/sub-RPE tissues by immunofluorescence (IF). In addition, western blot analysis of COX-2, Arg1, and VEGF-a revealed an increase in the signaling of these molecules within the RPE/choroid complex. Transmission electron microscopy (TEM) indicated a drusen-like structure of sub-RPE deposits with an accumulation of vacuolated cellular debris. Loss of photoreceptors was detected by peanut lectin staining and was corroborated by a reduction in MAP2 signaling. Loss of blood-retinal barrier integrity was demonstrated by a reduction of ZO-1 expression. Inflammatory cells were detected in the sub-RPE space, with an increase in IBA-1 positive microglia cells on the surface of the RPE. Mass spectrometry analysis of CXCR5 mouse RPE/choroid proteins extracts, separated by SDS-page and incubated with autologous serum, identified autoantibodies against AMD-associated proteins: Cryaa, Cryab, and Anxa2. evaluations in BV-2 cell culture indicated a significant increase in production of Arg-1 ( < 0.001) and COX-2 ( < 0.01) in the presence of anti-CXCR5 antibody when compared with Igg-treated control BV-2 cells stimulated with IL-4 and TNFα/IFNγ, respectively. Anti-CXCR5 antibody treatment without stimulating agents did not affect Arg-1 and COX-2 expression; this suggests that CXCR5 may have a regulatory role in microglia cells activation. These results indicate that with age, CXCR5 mice develop RD characterized by microglia dysfunction, increased production of CXCL13 in the RPE progressive photoreceptor, neuronal loss, and sub-RPE deposition of cellular debris, resulting in the production of immunogenic proteins and autoimmune-mediated RD.
The elusive identity of CXCR5 CD8 T cells in viral infection and autoimmunity: Cytotoxic, regulatory, or helper cells?
Fousteri Georgia,Kuka Mirela
Our knowledge on the development and functions of CXCR5 CD8 T cells is rudimentary when confronted to other extensively studied CD8 T cell subsets. A decade ago, it became apparent that CD8 T cells possess two additional and rather unexpected functional properties other than cytotoxicity, one involving what is known as B cell helper activity and the other involving suppression of self-reactive responses generally known as T cell regulation. Although these adaptive responses are well-known functions of CD4 T cells, they remain poorly understood in CD8 T cells. Thus far, three subsets of CXCR5 CD8 T cells have been identified. The first subset of CXCR5 CD8 T cells is present in chronic viral infections and is referred to as progenitors of exhausted T cells showing heightened proliferative and cytotoxic properties as compared to CXCR5 CD8 T cells. The second subset of CXCR5 CD8 T cells functions as regulatory T cells that inhibit CD4 T follicular helper (Tfh) humoral responses and the development of autoantibodies. The third subset of CXCR5 CD8 T cells was identified in mice with mutations in immunoregulatory genes (i.e. FOXP3 and IL-2-deficient mice) and involves CD8 T cells with Tfh-like properties that promote humoral autoimmunity through interaction with B cells. This review summarizes the phenotype, function, and differentiation of CXCR5 CD8 T cells.
CXCR5 induces perineural invasion of salivary adenoid cystic carcinoma by inhibiting microRNA-187.
Zhang Mei,Wu Jia-Shun,Xian Hong-Chun,Chen Bing-Jun,Wang Hao-Fan,Yu Xiang-Hua,Pang Xin,Dai Li,Jiang Jian,Liang Xin-Hua,Tang Ya-Ling
CXCR5 played critical roles in tumorigenesis and metastasis. Nevertheless, little was known about the involvement of CXCR5 in perineural invasion (PNI) of salivary adenoid cystic carcinoma (SACC). Here, we confirmed upregulation of CXCR5 in SACC specimens and cells and identified that CXCR5 exhibited a significant positive correlation with PNI. Functionally, knockdown of CXCR5 suppressed SACC cells migration, invasion and PNI ability, whereas CXCR5 overexpression displayed the opposite effects. Moreover, CXCR5 downregulated microRNA (miR)-187, which could competitively sponge S100A4. The PNI-inhibitory effect of CXCR5 knockdown or miR-187 overexpression could be reversed by elevated expression of S100A4. Conjointly, our data revealed that CXCR5 facilitated PNI through downregulating miR-187 to disinhibit S100A4 expression in SACC.
CXCL13/CXCR5 Interaction Facilitates VCAM-1-Dependent Migration in Human Osteosarcoma.
Liu Ju-Fang,Lee Chiang-Wen,Lin Chih-Yang,Chao Chia-Chia,Chang Tsung-Ming,Han Chien-Kuo,Huang Yuan-Li,Fong Yi-Chin,Tang Chih-Hsin
International journal of molecular sciences
Osteosarcoma is the most common primary tumor of the skeletal system and is well-known to have an aggressive clinical outcome and high metastatic potential. The chemokine (C-X-C motif) ligand 13 (CXCL13) plays a vital role in the development of several cancers. However, the effect of CXCL13 in the motility of osteosarcoma cells remains uncertain. Here, we found that CXCL13 increases the migration and invasion potential of three osteosarcoma cell lines. In addition, CXCL13 expression was upregulated in migration-prone MG-63 cells. Vascular cell adhesion molecule 1 (VCAM-1) siRNA and antibody demonstrated that CXCL13 promotes migration via increasing VCAM-1 production. We also show that CXCR5 receptor controls CXCL13-mediated VCAM-1 expression and cell migration. Our study identified that CXCL13/CXCR5 axis facilitate VCAM-1 production and cell migration in human osteosarcoma via the phospholipase C beta (PLCβ), protein kinase C α (PKCα), c-Src, and nuclear factor-κB (NF-κB) signaling pathways. CXCL13 and CXCR5 appear to be a novel therapeutic target in metastatic osteosarcoma.
Intratumoral CXCR5CD8T associates with favorable clinical outcomes and immunogenic contexture in gastric cancer.
Wang Jieti,Li Ruochen,Cao Yifan,Gu Yun,Fang Hanji,Fei Yuchao,Lv Kunpeng,He Xudong,Lin Chao,Liu Hao,Zhang Heng,Li He,He Hongyong,Xu Jiejie,Huang Hua
Studies that examined an association between CD8T and prognosis in gastric cancer are inconsistent, and a distinct population of CXCR5CD8T associated with better overall survival has been reported among various malignancies. Here, we show that the abundance of intratumoral CXCR5CD8T cells is associated with better overall survival in patients with gastric cancer. Patients with TNM II + III gastric cancer with higher intratumoral CXCR5CD8T cell infiltration are more likely to benefit from adjuvant chemotherapy. Microsatellite-unstable and Epstein-Barr virus positive tumors are enriched with CXCR5CD8T cells. Gastric cancer infiltrating CXCR5CD8T cells represent a specific subtype of stem-like CD8T with effector memory feature. Identification of the clinical significance and phenotype of gastric cancer infiltrating CXCR5CD8T provides a roadmap for patient stratification and trials of targeted therapies.
CXCR5+ CD8 T Cells: Protective or Pathogenic?
Valentine Kristen M,Hoyer Katrina K
Frontiers in immunology
CD8 T cells are infrequently considered part of germinal center reactions. Yet, a distinct CXCR5+ CD8 T cell subset identified within the B cell follicle and germinal center in situations of chronic antigen has recently been defined. CXCR5+ CD8 T cells maintain transcriptional and phenotypic features consistent with the CD8 T cell nomenclature of a non-exhausted, effector memory population. CD8 T cell localization to the B cell follicle suggests a functional profile similar to CD4 T follicular helper cells that are licensed to promote B cell responses. The functional mechanisms defined under different immune settings, while largely similar, differentially control disease pathogenesis. CXCR5+ CD8 T cells control viral load during infection, and also promote antibody-mediated autoimmune disease progression. The existence of this novel CXCR5+ CD8 T cell subset in human and murine models of disease may provide a paradigm shift in our understanding of germinal center reactions.
Antibody-suppressor CD8+ T Cells Require CXCR5.
Zimmerer Jason M,Ringwald Bryce A,Elzein Steven M,Avila Christina L,Warren Robert T,Abdel-Rasoul Mahmoud,Bumgardner Ginny L
BACKGROUND:We previously reported the novel activity of alloprimed CD8 T cells that suppress posttransplant alloantibody production. The purpose of the study is to investigate the expression and role of CXCR5 on antibody-suppressor CD8 T-cell function. METHODS:C57BL/6 mice were transplanted with FVB/N hepatocytes. Alloprimed CD8 T cells were retrieved on day 7 from hepatocyte transplant recipients. Unsorted or flow-sorted (CXCR5CXCR3 and CXCR3CXCR5) alloprimed CD8 T-cell subsets were analyzed for in vitro cytotoxicity and capacity to inhibit in vivo alloantibody production following adoptive transfer into C57BL/6 or high alloantibody-producing CD8 knock out (KO) hepatocyte transplant recipients. Alloantibody titer was assessed in CD8 KO mice reconstituted with naive CD8 T cells retrieved from C57BL/6, CXCR5 KO, or CXCR3 KO mice. Antibody suppression by ovalbumin (OVA)-primed monoclonal OVA-specific t-cell receptor transgenic CD8+ T cells (OT-I) CXCR5 or CXCR3 CD8 T-cell subsets was also investigated. RESULTS:Alloprimed CXCR5CXCR3CD8 T cells mediated in vitro cytotoxicity of alloprimed "self" B cells, while CXCR3CXCR5CD8 T cells did not. Only flow-sorted alloprimed CXCR5CXCR3CD8 T cells (not flow-sorted alloprimed CXCR3CXCR5CD8 T cells) suppressed alloantibody production and enhanced graft survival when transferred into transplant recipients. Unlike CD8 T cells from wild-type or CXCR3 KO mice, CD8 T cells from CXCR5 KO mice do not develop alloantibody-suppressor function. Similarly, only flow-sorted CXCR5CXCR3 (and not CXCR3CXCR5) OVA-primed OT-I CD8 T cells mediated in vivo suppression of anti-OVA antibody production. CONCLUSIONS:These data support the conclusion that expression of CXCR5 by antigen-primed CD8 T cells is critical for the function of antibody-suppressor CD8 T cells.
ER-stressed MSC displayed more effective immunomodulation in RA CD4CXCR5ICOS follicular helper-like T cells through higher PGE2 binding with EP2/EP4.
Wei Jing,Ouyang Xunli,Tang Yawei,Li Han,Wang Bing,Ye Yunshan,Jin Minli,Al Azab Mahmoud,Li Weiping,Li Xia
To analyze the further immunomodulatory effects of endoplasmic reticulum (ER)-stressed umbilical cord-derived mesenchymal stem cells MSCs (UC-MSCs) on rheumatoid arthritis (RA) CD4CXCR5ICOS T (follicular helper-like T, Tfh) cells. MSCs were isolated from umbilical cord and surface markers were identified by flow cytometry. CD4 T cells were purified from RA patients' peripheral blood mononuclear cells (PBMCs) using immunomagnetic beads. Thapsigargin (Tg)-stimulated or unstimulated MSCs were co-cultured with RA CD4 T cells. CD4CXCR5ICOS T cells were analyzed with fluorescence activating cell sorter (FACS) and major soluble factors secreted by MSCs were detected by qRT-PCR as well as ELISA. Receptors of prostanoid E2 (PGE2), known as EP1-4, on CD4 T cells were tested with RT-PCR and FACS. Proportion of CD4CXCR5ICOS T cells was determined after EP2/EP4 antagonists and anti-IL-6R antibody was added into co-cultured system, respectively. ER-stressed MSCs further down-regulated peripheral CD4CXCR5ICOS T cells compared with Tg-stimulated MSCs and CD4 T co-cultured group. PGE2 and IL-6 increased obviously in the supernatants. EP2/EP4 could be detected on CD4 T cells and frequencies of CD4CXCR5ICOS T cells were upregulated when EP2 and/or EP4 antagonists rather than anti-IL-6R antibody were added. ER-stressed MSCs exhibited better inhibition effect on RA CD4CXCR5ICOS T cells by releasing PGE2, indicating the immunosuppressive effect of MSCs could be enhanced by induction of ER stress.
Correlation between CXCR4, CXCR5 and CCR7 expression and survival outcomes in patients with clinical T1N0M0 non-small cell lung cancer.
Yue Zhao,Ningning Ding,Lin Yang,Jianming Ying,Hongtu Zhang,Ligong Yuan,Feng Li,Shuaibo Wang,Yousheng Mao
BACKGROUND:Lung cancer is the leading cause of cancer-related death. Even if early detection and treatment have proven to be effective, the survival outcomes are still poor. METHODS:Tissue samples and clinicopathological data of 244 patients with clinical T1N0M0 NSCLC were collected. We investigated CXCR4, CXCR5 and CCR7 expression levels using the immunohistochemical method and analyzed their correlations with clinicopathological characteristics and survival outcomes. RESULTS:Elevated expression levels of CXCR4, CXCR5 and CCR7 were found in tumor tissues (P < 0.001). The expression levels were remarkably different in histological type (CXCR4, P = 0.032; CXCR5, P < 0.001; CCR7, P < 0.001) and LVI (CXCR4, P = 0.017; CXCR5, P = 0.030; CCR7, P < 0.001). In addition, CXCR4 and CXCR5 expression were significantly different in tumor differentiation (CXCR4, P < 0.001; CXCR5, P < 0.001). Survival analysis showed that patients with positive CXCR4 expression had a significantly lower five-year DFS (P = 0.007) and a lower five-year OS (P = 0.010). Patients in the CXCR5 positive group had a significantly lower five-year DFS (P = 0.038) and a lower five-year OS (P = 0.220), which were statistically insignificant. However, five-year DFS and five-year OS of patients with positive CCR7 expression were significantly higher (DFS: P < 0.001; OS: P < 0.001). CXCR5 and CCR7 expression were found to be independent prognostic factors through multivariate analysis. CONCLUSIONS:Expression levels of CXCR4, CXCR5 and CCR7 were significantly higher in tumor tissues, and expression of CXCR5 and CCR7 were independent prognostic factors for survival. Moreover, all three chemokines were correlated to the survival outcomes of patients with clinical T1N0M0 NSCLC, providing potential prognosticators and therapy targets for lung cancer treatment.
The CXCL13/CXCR5-chemokine axis in neuroinflammation: evidence of CXCR5+CD4 T cell recruitment to CSF.
Harrer Christine,Otto Ferdinand,Pilz Georg,Haschke-Becher Elisabeth,Trinka Eugen,Hitzl Wolfgang,Wipfler Peter,Harrer Andrea
Fluids and barriers of the CNS
BACKGROUND:C-X-C chemokine ligand 13 (CXCL13) is frequently elevated in cerebrospinal fluid (CSF) in a variety of inflammatory central nervous system (CNS) diseases, has been detected in meningeal B cell aggregates in brain tissues of multiple sclerosis patients, and proposedly recruits B cells into the inflamed CNS. Besides B cells also follicular helper T (Tfh) cells express the cognate receptor C-X-C chemokine receptor type 5 (CXCR5) and follow CXCL13 gradients in lymphoid tissues. These highly specialized B cell helper T cells are indispensable for B cell responses to infection and vaccination and involved in autoimmune diseases. Phenotypically and functionally related circulating CXCR5+CD4 T cells occur in blood. Their co-recruitment to the inflamed CSF is feasible but unresolved. METHODS:We approached this question with a retrospective study including data of all patients between 2017 and 2019 of whom immune phenotyping data of CXCR5 expression and CSF CXCL13 concentrations were available. Discharge diagnoses and CSF laboratory parameters were retrieved from records. Patients were categorized as pyogenic/aseptic meningoencephalitis (ME, n = 29), neuroimmunological diseases (NIMM, n = 22), and non-inflammatory neurological diseases (NIND, n = 6). ANOVA models and Spearman's Rank-Order correlation were used for group comparisons and associations of CXCL13 levels with immune phenotyping data. RESULTS:In fact, intrathecal CXCL13 elevations strongly correlated with CXCR5+CD4 T cell frequencies in the total cohort (p < 0.0001, r = 0.59), and ME (p = 0.003, r = 0.54) and NIMM (p = 0.043, r = 0.44) patients. Moreover, the ratio of CSF-to-peripheral blood (CSF/PB) frequencies of CXCR5+CD4 T cells strongly correlated with CXCL13 levels both in the total cohort (p = 0.001, r = 0.45) and ME subgroup (p = 0.005, r = 0.50), indicating selective accumulation. ME, NIMM and NIND groups differed with regard to CSF cell counts, albumin quotient, intrathecal IgG, CXCL13 elevations and CXCR5+CD4 T cells, which were higher in inflammatory subgroups. CONCLUSION:The observed link between intrathecal CXCL13 elevations and CXCR5+CD4 T cell frequencies does not prove but suggests recruitment of possible professional B cell helpers to the inflamed CSF. This highlights CSF CXCR5+CD4 T cells a key target and potential missing link to the poorly understood phenomenon of intrathecal B cell and antibody responses with relevance for infection control, chronic inflammation and CNS autoimmunity.
CXCL13/CXCR5 are potential biomarkers for diagnosis and prognosis for breast cancer.
Jiang Li,Wang Dong,Sheng Mengfei,Tong Deyong,Liu Hanchen,Dong Liangliang,Ma Jianjun
Journal of B.U.ON. : official journal of the Balkan Union of Oncology
PURPOSE:Breast cancer is known as the second frequent cancer in the world, even as the most common cancer among women. This study aimed to explore the correlation of CXCL13/CXCR5 expression with clinical characteristics in breast cancer and evaluate their potential to be used as biomarkers in diagnosis and prognosis of this disease. METHODS:A total of 133 female patients diagnosed with breast cancer were collected. The expression of CXCL13 and CXCR5 mRNA was analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemical staining. RESULTS:The expression of CXCL13 and CXCR5 was significantly higher in breast cancer tissue than in normal breast tissues, with a high correlation coefficient of 0.9973. Positive cell numbers and positive expression rates of CXCL13 and CXCR5 in cancer breast tissue were much higher than those in normal breast tissue, and raised with increase of cancer stage. The high expression of CXCL13 and CXCR5 in breast cancer tissue was notably associated with lymph node metastasis, distant metastasis, disease stage, but not with age, Her-2 status, histological type or tumor size. Immunohistochemistry analysis showed that the positive expression of CXCL13 and CXCR5 was related with cancer stage, Also, positive expression of CXCL13 was correlated with positive expression of CXCR5. Patient age, Her-2 status, tumor size, histological type, and lymph node metastasis were independent factors for the 5-year survival rate of breast cancer patients, whereas the 5-year survival was correlated with distant metastasis and expression of CXCL13 and CXCR5. CONCLUSIONS:These results suggested that CXCL13 and CXCR5 expressions could act as potential biomarkers for breast cancer diagnosis and prognosis.
Association of the CXCL9-CXCR3 and CXCL13-CXCR5 axes with B-cell trafficking in giant cell arteritis and polymyalgia rheumatica.
Graver Jacoba C,Abdulahad Wayel,van der Geest Kornelis S M,Heeringa Peter,Boots Annemieke M H,Brouwer Elisabeth,Sandovici Maria
Journal of autoimmunity
OBJECTIVE:B-cells are present in the inflamed arteries of giant cell arteritis (GCA) patients and a disturbed B-cell homeostasis is reported in peripheral blood of both GCA and the overlapping disease polymyalgia rheumatica (PMR). In this study, we aimed to investigate chemokine-chemokine receptor axes governing the migration of B-cells in GCA and PMR. METHODS:We performed Luminex screening assay for serum levels of B-cell related chemokines in treatment-naïve GCA (n = 41), PMR (n = 31) and age- and sex matched healthy controls (HC, n = 34). Expression of chemokine receptors on circulating B-cell subsets were investigated by flow cytometry. Immunohistochemistry was performed on GCA temporal artery (n = 14) and aorta (n = 10) and on atherosclerosis aorta (n = 10) tissue. RESULTS:The chemokines CXCL9 and CXCL13 were significantly increased in the circulation of treatment-naïve GCA and PMR patients. CXCL13 increased even further after three months of glucocorticoid treatment. At baseline CXCL13 correlated with disease activity markers. Peripheral CXCR3+ and CXCR5+ switched memory B-cells were significantly reduced in both patient groups and correlated inversely with their complementary chemokines CXCL9 and CXCL13. At the arterial lesions in GCA, CXCR3+ and CXCR5+ B-cells were observed in areas with high CXCL9 and CXCL13 expression. CONCLUSION:Changes in systemic and local chemokine and chemokine receptor pathways related to B-cell migration were observed in GCA and PMR mainly in the CXCL9-CXCR3 and CXCL13-CXCR5 axes. These changes can contribute to homing and organization of B-cells in the vessel wall and provide further evidence for an active involvement of B-cells in GCA and PMR.
The age of the bone marrow microenvironment influences B-cell acute lymphoblastic leukemia progression via CXCR5-CXCL13.
Zanetti Costanza,Kumar Rahul,Ender Joscha,Godavarthy Parimala S,Hartmann Mark,Hey Joschka,Breuer Kersten,Weissenberger Eva S,Minciacchi Valentina R,Karantanou Christina,Gu Zhaohui,Roberts Kathryn G,Metzler Markus,Stock Wendy,Mullighan Charles G,Bloomfield Clara D,Filmann Natalie,Bankov Katrin,Hartmann Sylvia,Hasserjian Robert P,Cousins Antony F,Halsey Christina,Plass Christoph,Lipka Daniel B,Krause Daniela S
B-cell acute lymphoblastic leukemia (B-ALL) occurs most commonly in children, while chronic myeloid leukemia (CML) is more frequent in adults. The myeloid bias of hematopoiesis in elderly individuals has been considered causative, but the age of the bone marrow (BM) microenvironment (BMM) may be contributory. Using various murine models of B-ALL in young versus old mice, we recapitulated B-ALL preponderance in children versus adults. We showed differential effects of young versus old BM macrophages on B-ALL cell function. Molecular profiling using RNA- and ATAC-seq revealed pronounced differences in young versus old BMM-derived macrophages and enrichment for gene sets associated with inflammation. In concordance with the role of C-X-C motif chemokine (CXCL) 13 for disease-associated B cell chemoattraction, we found CXCL13 to be highly expressed in young macrophages on a translational compared to a transcriptional level. Inhibition of CXCL13 in BM macrophages impaired leukemia cell migration and decreased the proliferation of cocultured B-ALL cells, while recombinant CXCL13 increased pAKT and B-ALL cell expansion. Pretreatment of B-ALL-initiating cells with CXCL13 accelerated B-ALL progression. Deficiency of Cxcr5, the receptor for CXCL13, on B-ALL-initiating cells prolonged murine survival, while high expression of CXCR5 in pediatric B-ALL may predict central nervous system relapse. CXCL13 staining was increased in bone sections from pediatric compared to adult B-ALL patients. Taken together, our study shows that the age of the BMM and, in particular, BM macrophages influence the leukemia phenotype. The CXCR5-CXCL13-axis may act as prognostic marker and an attractive novel target for the treatment of B-ALL.
S-nitrosylation-mediated coupling of G-protein alpha-2 with CXCR5 induces Hippo/YAP-dependent diabetes-accelerated atherosclerosis.
Chao Meng-Lin,Luo Shanshan,Zhang Chao,Zhou Xuechun,Zhou Miao,Wang Junyan,Kong Chuiyu,Chen Jiyu,Lin Zhe,Tang Xin,Sun Shixiu,Tang Xinlong,Chen Hongshan,Wang Hong,Wang Dongjin,Sun Jin-Peng,Han Yi,Xie Liping,Ji Yong
Atherosclerosis-associated cardiovascular disease is one of the main causes of death and disability among patients with diabetes mellitus. However, little is known about the impact of S-nitrosylation in diabetes-accelerated atherosclerosis. Here, we show increased levels of S-nitrosylation of guanine nucleotide-binding protein G(i) subunit alpha-2 (SNO-GNAI2) at Cysteine 66 in coronary artery samples from diabetic patients with atherosclerosis, consistently with results from mice. Mechanistically, SNO-GNAI2 acted by coupling with CXCR5 to dephosphorylate the Hippo pathway kinase LATS1, thereby leading to nuclear translocation of YAP and promoting an inflammatory response in endothelial cells. Furthermore, Cys-mutant GNAI2 refractory to S-nitrosylation abrogated GNAI2-CXCR5 coupling, alleviated atherosclerosis in diabetic mice, restored Hippo activity, and reduced endothelial inflammation. In addition, we showed that melatonin treatment restored endothelial function and protected against diabetes-accelerated atherosclerosis by preventing GNAI2 S-nitrosylation. In conclusion, SNO-GNAI2 drives diabetes-accelerated atherosclerosis by coupling with CXCR5 and activating YAP-dependent endothelial inflammation, and reducing SNO-GNAI2 is an efficient strategy for alleviating diabetes-accelerated atherosclerosis.
CXC chemokine ligand-13 promotes metastasis via CXCR5-dependent signaling pathway in non-small cell lung cancer.
Chao Chia-Chia,Lee Wei-Fang,Wang Shih-Wei,Chen Po-Chun,Yamamoto Ayaho,Chang Tsung-Ming,Weng Shun-Long,Liu Ju-Fang
Journal of cellular and molecular medicine
The CXC chemokine ligand-13 (CXCL13) is a chemoattractant of B cells and has been implicated in the progression of many cancers. So far, CXCL13 and its related receptor CXCR5 have been proved to regulate cancer cell migration as well as tumour metastasis. However, the role of CXCL13-CXCR5 axis in metastasis of lung cancer is still poorly understood. In this study, we found that CXCL13 and CXCR5 were commonly up-regulated in lung cancer specimens compared with normal tissues among different cohorts. Our evidence showed that CXCL13 obviously promoted migration of lung cancer cells, and this effect was mediated by vascular cell adhesion molecule-1 (VCAM-1) expression. We also confirmed that CXCR5, the major receptor responsible for CXCL13 function, was required for CXCL13-promoted cell migration. We also test the candidate components which are activated after CXCL13 treatment and found that phospholipase C-β (PLCβ), protein kinase C-α (PKCα) and c-Src signalling pathways were involved in CXCL13-promoted cell migration and VCAM-1 expression in lung cancer cells. Finally, CXCL13 stimulated NF-κB transcription factor in lung cancer cells, contributing to VCAM-1 expression in translational level. These evidences propose a novel insight into lung cancer metastasis which is regulated by CXCL13.
Unexpected enhancement of FVIII immunogenicity by endothelial expression in lentivirus-transduced and transgenic mice.
Shi Qizhen,Carman Christopher V,Chen Yingyu,Sage Peter T,Xue Feng,Liang Xin M,Gilbert Gary E
Factor VIII (FVIII) replacement therapy for hemophilia A is complicated by development of inhibitory antibodies (inhibitors) in ∼30% of patients. Because endothelial cells (ECs) are the primary physiologic expression site, we probed the therapeutic potential of genetically restoring FVIII expression selectively in ECs in hemophilia A mice (FVIIInull). Expression of FVIII was driven by the Tie2 promoter in the context of lentivirus (LV)-mediated in situ transduction (T2F8LV) or embryonic stem cell-mediated transgenesis (T2F8Tg). Both endothelial expression approaches were associated with a strikingly robust immune response. Following in situ T2F8LV-mediated EC transduction, all FVIIInull mice developed inhibitors but had no detectable plasma FVIII. In the transgenic approach, the T2F8Tg mice had normalized plasma FVIII levels, but showed strong sensitivity to developing an FVIII immune response upon FVIII immunization. A single injection of FVIII with incomplete Freund adjuvant led to high titers of inhibitors and reduction of plasma FVIII to undetectable levels. Because ECs are putative major histocompatibility complex class II (MHCII)-expressing nonhematopoietic, "semiprofessional" antigen-presenting cells (APCs), we asked whether they might directly influence the FVIII immune responses. Imaging and flow cytometric studies confirmed that both murine and human ECs express MHCII and efficiently bind and take up FVIII protein in vitro. Moreover, microvascular ECs preconditioned ex vivo with inflammatory cytokines could functionally present exogenously taken-up FVIII to previously primed CD4+/CXCR5+ T follicular helper (Tfh) cells to drive FVIII-specific proliferation. Our results show an unanticipated immunogenicity of EC-expressed FVIII and suggest a context-dependent role for ECs in the regulation of inhibitors as auxiliary APCs for Tfh cells.
Enhanced IgG -mediated antibody response towards thymus-dependent immunization in CXCR1-deficient mice.
Jaufmann Jennifer,Carevic Melanie,Tümen Leyla,Eliacik Derya,Schmitt Fee,Hartl Dominik,Beer-Hammer Sandra
Immunity, inflammation and disease
BACKGROUND:Chemokine receptors and their corresponding ligands are key players of immunity by regulation of immune cell differentiation and migration. CXCR1 is a high-affinity receptor for CXCL8. Differential expression of CXCR1 is associated with a variety of human pathologies including cancer and inflammatory diseases. While various studies have highlighted the importance of CXCR1-mediated CXCL8-sensing for neutrophil trafficking and function, its role in B-cell responses remains unsolved. Therefore, our aim was to investigate innate and adaptive antibody responses in CXCR1-deficient mice. METHODS:Cell populations of the spleen and the peritoneal cavity were identified and quantified via flow cytometry. To investigate thymus-independent (TI) and thymus-dependent (TD) antibody responses, mice were immunized intraperitoneally with TNP-Ficoll, Pneumovax23, and TNP-Chicken Gamma Globulin. Mice were bled before as well as 7 and 14 days after vaccination to collect serum. Serum antibody levels overtime were analyzed according to their specificity by enzyme-linked immunosorbent assay. B-1 cell functionality was examined by IL-5/IL-5Rα-dependent stimulation of peritoneal and splenic cells in vitro. To analyze CXCR1/2-expression, CD19 splenocytes were enriched by magnetic-activated cell sorting before isolation of total RNA contents, followed by reverse transcription and real-time polymerase chain reaction. RESULTS:The distribution of natural B-1 cell populations was disturbed in the absence of CXCR1, while their responsiveness towards TI antigens and in vitro stimulation remained functional. Besides, CXCR1-deficiency was accompanied by increased frequencies of follicular B-2 cells in the spleen. Interestingly, these mice produced elevated levels of antigen-specific IgG upon TD immunization and harbored a significantly enlarged proportion of CXCR5-expressing T helper (H) cells. CXCR1-expression was detectable in CD19 splenocytes derived from wild-type, but not CXCR1-deficient mice. CONCLUSION:Our data demonstrate a previously unknown relevance of CXCR1 for the production of specific IgG in response to vaccination. These findings identify CXCR1 as a promising candidate for future studies on the regulation of adaptive antibody responses.
Whole-blood phenotyping to assess alloimmunization status in transfused sickle cell disease patients.
Tamagne Marie,Pakdaman Sadaf,Bartolucci Pablo,Habibi Anoosha,Galactéros Frédéric,Pirenne France,Vingert Benoît
It is essential to limit hemolytic transfusion reactions in polytransfused individuals, and the prevention of alloimmunization is a key solution. CD4+ T lymphocyte (TL) markers, particularly follicular T helper (Tfh) cells, may differentiate between responder and nonresponder alloimmunization statuses. We tested this hypothesis by studying the phenotype of CXCR5+PD1+ TLs in whole blood. Our results suggest that high levels of CXCR5+PD1+CD4+ TLs in whole blood may be a characteristic of nonalloimmunized patients. However, these cells did not display the phenotypic characteristics of active Tfh cells. Instead, a decrease in blood quiescent Tfh-cell levels was observed in nonalloimmunized polytransfused patients. High levels of CXCR5+PD1+CD4+ TLs may be associated with inhibitory signaling functions of T cells, as reflected by the low levels of PD1+ICOS+ cells in the nonalloimmunized polytransfused group. The description of these particular phenotypes, and their comparison among groups of patients, responders, and nonresponders, suggests that new immunological components should be considered when trying to understand posttransfusion alloimmunization.
Antitumor CD8 T cell responses in glioma patients are effectively suppressed by T follicular regulatory cells.
Lu Lenian,Sun Jie,Su Hang,Luo Shi,Chen Jianmin,Qiu Shengcong,Chi Yajie,Lin Jiye,Xu Xiaobing,Zheng Dahai
Experimental cell research
Regulatory T (Treg) cells are thought to contribute to tumor pathogenesis by suppressing tumor immunosurveillance and antitumor immunity. T follicular regulatory (Tfr) cells are a recently characterized Treg subset that expresses both the Treg transcription factor (TF) Foxp3 and the T follicular helper (Tfh) TF Bcl-6. The role of Tfr cells in glioma patients remains unclear. In this study, we found that the level of Tfr cells, identified as Foxp3Bcl-6 CD4 T cells, was significantly elevated in tumor-infiltrating CD4 T cells from resected glioma tumors. Both Tfr cells and Treg cells significantly suppressed the proliferation and the cytotoxic capacity of CD8 T cells toward glioma tumor cells, and the suppression was positively associated with the proportion of Tfr cells and Treg cells, respectively. Tfr and Treg cells from glioma tumor samples demonstrated higher suppression potency than those from healthy blood samples and glioma blood samples. Interestingly, canonical CXCR5 Treg cells could suppress both CXCR5 and CXCR5 CD8 T cells, albeit with stronger potency toward CXCR5 CD8 T cells. However, Tfr cells presented much higher suppression potency toward CXCR5 CD8 T cells, whereas CXCR5 CD8 T cells are a potent CD8 T cell subset previously described to have antiviral and antitumor roles. Overall, these data indicate that Tfr cells are enriched in glioma tumors and have suppressive capacity toward CD8 T cell-mediated effector functions.
CXCL13/CXCR5 axis facilitates endothelial progenitor cell homing and angiogenesis during rheumatoid arthritis progression.
Tsai Chun-Hao,Chen Chao-Ju,Gong Chi-Li,Liu Shan-Chi,Chen Po-Chun,Huang Chien-Chung,Hu Sung-Lin,Wang Shih-Wei,Tang Chih-Hsin
Cell death & disease
Angiogenesis is a critical process in the formation of new capillaries and a key participant in rheumatoid arthritis (RA) pathogenesis. The chemokine (C-X-C motif) ligand 13 (CXCL13) plays important roles in several cellular functions such as infiltration, migration, and motility. We report significantly higher levels of CXCL13 expression in collagen-induced arthritis (CIA) mice compared with controls and also in synovial fluid from RA patients compared with human osteoarthritis (OA) samples. RA synovial fluid increased endothelial progenitor cell (EPC) homing and angiogenesis, which was blocked by the CXCL13 antibody. By interacting with the CXCR5 receptor, CXCL13 facilitated vascular endothelial growth factor (VEGF) expression and angiogenesis in EPC through the PLC, MEK, and AP-1 signaling pathways. Importantly, infection with CXCL13 short hairpin RNA (shRNA) mitigated EPC homing and angiogenesis, articular swelling, and cartilage erosion in ankle joints of mice with CIA. CXCL13 is therefore a novel therapeutic target for RA.