Role of NKp46 natural killer cells in house dust mite-driven asthma.
Haspeslagh Eline,van Helden Mary J,Deswarte Kim,De Prijck Sofie,van Moorleghem Justine,Boon Louis,Hammad Hamida,Vivier Eric,Lambrecht Bart N
EMBO molecular medicine
House dust mite (HDM)-allergic asthma is driven by T helper 2 (Th2) lymphocytes, but also innate immune cells control key aspects of the disease. The precise function of innate natural killer (NK) cells during the initiation and propagation of asthma has been very confusing, in part because different, not entirely specific, strategies were used to target these cells. We show that HDM inhalation rapidly led to the accumulation of NK cells in the lung-draining lymph nodes and of activated CD69 NK cells in the bronchoalveolar lumen. However, genetically engineered -DTA or -DTR mice that constitutively or temporarily lack NK cells, still developed all key features of acute or chronic HDM-driven asthma, such as bronchial hyperreactivity, Th2 cytokine production, eosinophilia, mucus overproduction, and Th2-dependent immunoglobulin serum titers. The same results were obtained by administration of conventional NK1.1 or asialo-GM1 NK cell-depleting antibodies, antibody-mediated blocking of the NKG2D receptor, or genetic NKG2D deficiency. Thus, although NK cells accumulate in allergen-challenged lungs, our findings comprehensively demonstrate that these cells are not required for HDM-driven asthma in the mouse.
Allergen-induced activation of natural killer cells represents an early-life immune response in the development of allergic asthma.
Altman Matthew C,Whalen Elizabeth,Togias Alkis,O'Connor George T,Bacharier Leonard B,Bloomberg Gordon R,Kattan Meyer,Wood Robert A,Presnell Scott,LeBeau Petra,Jaffee Katy,Visness Cynthia M,Busse William W,Gern James E
The Journal of allergy and clinical immunology
BACKGROUND:Childhood asthma in inner-city populations is a major public health burden, and understanding early-life immune mechanisms that promote asthma onset is key to disease prevention. Children with asthma demonstrate a high prevalence of aeroallergen sensitization and T2-type inflammation; however, the early-life immune events that lead to T2 skewing and disease development are unknown. OBJECTIVE:We sought to use RNA sequencing of PBMCs collected at age 2 years to determine networks of immune responses that occur in children with allergy and asthma. METHODS:In an inner-city birth cohort with high asthma risk, we compared gene expression using RNA sequencing in PBMCs collected at age 2 years between children with 2 or more aeroallergen sensitizations, including dust mite, cockroach, or both, by age 3 years and asthma by age 7 years (cases) and matched control subjects who did not have any aeroallergen sensitization or asthma by age 7 years. RESULTS:PBMCs from the cases showed higher levels of expression of natural killer (NK) cell-related genes. After cockroach or dust mite allergen but not tetanus antigen stimulation, PBMCs from the cases compared with the control subjects showed differential expression of 244 genes. This gene set included upregulation of a densely interconnected NK cell-like gene network reflecting a pattern of cell activation and induction of inflammatory signaling molecules, including the key T2-type cytokines IL9, IL13, and CCL17, as well as a dendritic cell-like gene network, including upregulation of CD1 lipid antigen presentation molecules. The NK cell-like response was reproducible in an independent group of children with later-onset allergic sensitization and asthma and was found to be specific to only those children with both aeroallergen sensitization and asthma. CONCLUSION:These findings provide important mechanistic insight into an early-life immune pathway involved in T2 polarization, leading to the development of allergic asthma.
Epithelium-derived cystatin SN enhances eosinophil activation and infiltration through IL-5 in patients with chronic rhinosinusitis with nasal polyps.
Yan Bing,Lou Hongfei,Wang Yang,Li Ying,Meng Yifan,Qi Sihan,Wang Ming,Xiao Lei,Wang Chengshuo,Zhang Luo
The Journal of allergy and clinical immunology
BACKGROUND:The interaction between epithelial cells and immune cells plays an important role in the pathogenesis of chronic rhinosinusitis with nasal polyps (CRSwNP); however, the mechanism or mechanisms underlying T-biased inflammation in this process are largely unknown. Profiling protein expression in patients with CRSwNP by using shotgun proteomics suggested that cystatin SN (CST1), a type 2 cysteine protease inhibitor, might play a role because this was expressed with the greatest difference in patients with eosinophilic chronic rhinosinusitis with nasal polyps (ECRSwNP) and those with noneosinophilic chronic rhinosinusitis with nasal polyps (nonECRSwNP). OBJECTIVES:We sought to investigate the expression and role of CST1 in modulating eosinophilic inflammation in patients with CRSwNP. METHODS:Sinonasal tissues were collected from 192 patients with ECRSwNP, 52 patients with nonECRSwNP, and 40 control subjects. CST1 mRNA expression, localization, and concentration in the tissues were measured by using real-time PCR, in situ hybridization, immunohistochemistry, and an ELISA. Recombinant CST1 was used to further explore the function of the molecule in dispersed nasal polyp cells and eosinophils extracted from polyp tissues and peripheral blood. RESULTS:CST1 was mainly expressed by epithelial cells and significantly increased in patients with ECRSwNP but decreased in patients with nonECRSwNP compared with that in control subjects. CST1 expression was further increased in patients with ECRSwNP and comorbid asthma and correlated with eosinophil percentages in tissue samples. CST1 was induced by IL-4 and IL-13 in tissue from both patients with ECRSwNP and those with nonECRSwNP and repressed by IL-17A in patients with nonECRSwNP in the presence of neutrophils. CST1 enhanced eosinophil activation and recruitment through induction of IL-5. CONCLUSION:Epithelium-derived CST1 modulates eosinophil activation and recruitment, expression of which could be regulated by T2 and T17 cytokines.
Cross-talk between T2 and T17 pathways in patients with chronic rhinosinusitis with nasal polyps.
Wang Min,Zhang Nan,Zheng Ming,Li Ying,Meng Lingling,Ruan Yu,Han Jinbo,Zhao Na,Wang Xiangdong,Zhang Luo,Bachert Claus
The Journal of allergy and clinical immunology
BACKGROUND:Chronic rhinosinusitis with nasal polyps (CRSwNP) is a heterogeneous disease with a spectrum of endotypes. T2- and T17-related cytokines are 2 central regulators involved in the inflammation associated with CRSwNP. OBJECTIVE:We sought to investigate the interregulation of T2 and T17 pathways in Chinese patients with CRSwNP. METHODS:Levels of key T2- and T17-related factors were measured in homogenates of polyp tissue obtained from patients with CRSwNP. The relationship of these factors and their expression in groups classified according to tissue IL-5 and IL-17 concentrations were analyzed. Cross-regulation of T2 and T17 cytokines and the effects of dexamethasone treatment were studied in dispersed nasal polyp cells. Associations between T2- and T17 related factors and comorbid atopic status and asthma, disease recurrence, and edema scores were also explored. RESULTS:Four CRSwNP groups were classified based on expression or nonexpression of mutually exclusive T2- and T17-related factors. The T2 cytokines IL-4 and IL-13 inhibited expression of T17-related factors, whereas the T17 cytokines IL-17 and TGF-β1 enhanced expression of T2-related factors. Dexamethasone treatment inhibited both the T2 and T17 pathways. A patient's atopic status was related to their T2 immune response. Edema scores were positively correlated with the T2 pathway and negatively correlated with the T17 pathway. CONCLUSION:The T2 and T17 pathways are mutually exclusive and regulate each other, favoring the development of a T2 immune response in Chinese patients with CRSwNP.
Th2 cytokines orchestrate the secretion of MUC5AC and MUC5B in IL-5-positive chronic rhinosinusitis with nasal polyps.
Zhang Y,Derycke L,Holtappels G,Wang X D,Zhang L,Bachert C,Zhang N
BACKGROUND:Mucin over-secretion is a significant characteristic of chronic rhinosinusitis with nasal polyps (CRSwNP). This study aimed to investigate the relationship between Th2 cytokines and MUC5AC or MUC5B, and the mechanism of mucin over-secretion in the type-2 inflammatory endotype of CRSwNP. METHODS:Main Th-cell cytokines, associated mediators, and mucins were determined in the homogenates of nasal polyp samples from 21 CRSwNP patients and inferior turbinate samples from 8 controls, by ELISA or UniCAP system. Secretion of MUC5AC and MUC5B was measured in the supernatants of IL-5, IL-4, or IL-13 primed nasal polyp fragments. Co-localization of MUC5AC, MUC5B, and IL-4 receptor α (IL-4Rα) in CRSwNP and controls was evaluated by immunohistochemistry. Gene expression of IL-4Rα in the samples was measured by real-time reverse transcription-polymerase chain reaction. RESULTS:Baseline protein levels of the Th2-cytokines IL-4, IL-5, and IL-13, and mucins MUC5AC and MUC5B were significantly higher in the IL-5(+) CRSwNP group, compared to control and IL-5(-) CRSwNP groups. MUC5AC and MUC5B secretions were significantly increased in IL-4- or IL-13-primed, but not IL-5-primed fragments of nasal polyps. Immuno-stained serial sections demonstrated that IL-4Rα was widely expressed in the epithelium and submucosal glands in control and nasal polyp tissues. Gene expression of IL-4Rα was elevated in nasal polyp tissues, specifically in the IL-5(+) CRSwNP group. CONCLUSIONS:In type-2 inflammatory nasal polyps, characterized by the tissue expression of IL-5, MUC5AC and MUC5B are overexpressed. Both IL-4 and IL-13 may upregulate mucin expression via IL-4Rα, which is also overexpressed in IL-5(+) CRSwNP.
Interleukin-8 gene expression in chronic sinusitis.
Takeuchi K,Yuta A,Sakakura Y
American journal of otolaryngology
PURPOSE:Interleukin-8 (IL-8), a monocyte-derived and macrophage-derived cytokine, displays potent chemotactic activating properties toward neutrophils and thus may contribute to the pathogenesis of chronic sinusitis. The object of this investigation was to show the expression of the IL-8 gene in chronic sinusitis by Northern blot analysis and a reverse transcription-polymerase chain reaction (RT-PCR). MATERIALS AND METHODS:For Northern blot analysis, RNAs were extracted from maxillary mucosa and nasal polyps from two patients with chronic sinusitis, respectively, and from the inferior turbinate of a nasal allergy patient. For RT-PCR, RNAs were extracted from 11 patients with chronic sinusitis, 8 patients with allergic rhinitis, and 4 patients with hypertrophic rhinitis. RESULTS:Whereas IL-8 mRNA was expressed in the maxillary mucosa, IL-8 transcript was not detected in the inferior turbinate by Northern blot analysis. IL-8 transcripts were detected in 45% of chronic sinusitis RNAs (5/11) and in 50% of allergic rhinitis RNAs (5/10) by RT-PCR. CONCLUSION:These data suggest IL-8 may contribute to neutrophil involvement in chronic sinusitis.
The Role of Macrolides in Chronic Rhinosinusitis (CRSsNP and CRSwNP).
Oakley Gretchen M,Harvey Richard J,Lund Valerie J
Current allergy and asthma reports
PURPOSE OF REVIEW:We assess the literature on the pharmacokinetics, indications, important considerations, and effectiveness of long-term, low-dose macrolide antibiotics in chronic rhinosinusitis (CRS). RECENT FINDINGS:The key to effective implementation of macrolide therapy in CRS is appropriate patient selection. Macrolides have demonstrated the most benefit in Th1-mediated non-eosinophilic CRS when used for durations of at least 3 months. Macrolide antibiotics have demonstrated great benefit when used for their anti-inflammatory or immunomodulatory properties, which include the blockage of pro-inflammatory cytokines, such as interleukin (IL)-8 and tumor necrosis factor-α (TNF-α). They have been used in CRS patients not responding to traditional corticosteroid-based treatment regimens, but appear to be most effective specifically in Th1-mediated non-eosinophilic CRS in long durations and low doses. Further research is needed to better identify characteristics known to correlate with macrolide response so early directed therapy can be implemented.
Airway epithelial cell apoptosis and inflammation in COPD, smokers and nonsmokers.
Comer David M,Kidney Joseph C,Ennis Madeleine,Elborn J Stuart
The European respiratory journal
We hypothesised that primary bronchial epithelial cells (PBECs) from subjects with chronic obstructive pulmonary disease (COPD) respond differently to Pseudomonas aeruginosa lipopolysaccharide (LPS) after cigarette smoke extract (CSE) exposure than PBECs obtained from smokers without airflow obstruction and nonsmokers. PBECs from 16 COPD subjects, 10 smokers without airflow obstruction and nine nonsmokers were cultured at air-liquid interface. Cultures were incubated with CSE prior to stimulation with P. aeruginosa LPS. Interleukin (IL)-6 and IL-8 were measured by ELISA and Toll-like receptor (TLR)-4 expression by fluorescence-activated cell sorter. Activation of nuclear factor (NF)-κB was determined by Western blotting and ELISA, and MAPK and caspase-3 activity by Western blotting. Apoptosis was evaluated using Annexin-V staining and the terminal transferase-mediated dUTP nick end-labelling methods. Constitutive release of IL-8 and IL-6 was greatest from the COPD cultures. However, CSE pretreatment followed by P. aeruginosa LPS stimulation reduced IL-8 release from COPD PBECs, but increased it from cells of smokers without airflow obstruction and nonsmokers. TLR-4 expression, MAPK and NF-κB activation in COPD cultures were reduced after CSE treatment, but not in the smokers without airflow obstruction or nonsmoker groups, which was associated with increased apoptosis. CSE attenuates inflammatory responses to LPS in cells from people with COPD but not those from nonsmoking individuals and those who smoke without airflow obstruction.
Inflammatory immune response in rabbits with Staphylococcus aureus biofilm-associated sinusitis.
Wu Xianmin,Zhang Yue,Chen Xiaoyun,Chen Jun,Jia Minghui
International forum of allergy & rhinology
BACKGROUND:Staphylococcus aureus is the most commonly isolated bacterium from patients with surgically recalcitrant chronic rhinosinusitis (CRS). Understanding the immune responses to S aureus biofilms will provide insights into how the host response may be manipulated by therapeutic agents to improve the chances of successfully preventing and treating these infections. In this study, we investigated the inflammatory immune response in a rabbit model of S aureus biofilm-related sinusitis by analyzing the levels of some major inflammatory cytokines. METHODS:Eighteen New Zealand white rabbits were randomly divided into 3 groups: a blank-control group; a negative-control group; and a model group. Four weeks after the biofilm-associated sinusitis models were established, the sinus mucosa was harvested and examined using hematoxylin-eosin (H&E) staining, scanning electron microscopy (SEM), reverse transcription polymerase chain reaction (RT-PCR), and western blotting. The expression levels of inflammatory cytokines were analyzed statistically. RESULTS:Interleukin (IL)-1β, IL-8, and tumor necrosis factor (TNF)-α expression levels were significantly higher in the model group than in the blank-control group (p < 0.05); mRNA levels were increased by 1600%, 230%, and 130%, respectively, and the protein levels were increased by 180%, 100%, and 100%, respectively. In contrast, IL-4 and IL-5 mRNA levels were reduced by 44% and 70%, respectively, compared with the blank-control group (p < 0.05). CONCLUSION:S aureus biofilms in the rabbit maxillary sinus mucosa were associated with increased IL-1β, IL-8, and TNF-α expression, and decreased IL-4 and IL-5 expression.
Oxygen matters: hypoxia as a pathogenic mechanism in rhinosinusitis.
Cho Hyung-Ju,Kim Chang-Hoon
The airway epithelium is the first place, where a defense mechanism is initiated against environmental stimuli. Mucociliary transport (MCT), which is the defense mechanism of the airway and the role of airway epithelium as mechanical barriers are essential in innate immunity. To maintain normal physiologic function, normal oxygenation is critical for the production of energy for optimal cellular functions. Several pathologic conditions are associated with a decrease in oxygen tension in airway epithelium and chronic sinusitis is one of the airway diseases, which is associated with the hypoxic condition, a potent inflammatory stimulant. We have observed the overexpression of the hypoxia-inducible factor 1 (HIF-1), an essential factor for oxygen homeostasis, in the epithelium of sinus mucosa in sinusitis patients. In a series of previous reports, we have found hypoxia-induced mucus hyperproduction, especially by MUC5AC hyperproduction, disruption of epithelial barrier function by the production of VEGF, and down-regulation of junctional proteins such as ZO-1 and E-cadherin. Furthermore, hypoxia-induced inflammation by HMGB1 translocation into the cytoplasm results in the release of IL-8 through a ROS-dependent mechanism in upper airway epithelium. In this mini-review, we briefly introduce and summarize current progress in the pathogenesis of sinusitis related to hypoxia. The investigation of hypoxia-related pathophysiology in airway epithelium will suggest new insights on airway inflammatory diseases, such as rhinosinusitis for clinical application and drug development. [BMB Reports 2018; 51(2): 59-64].
Immunological Characteristics in Refractory Chronic Rhinosinusitis with Nasal Polyps Undergoing Revision Surgeries.
Ryu Gwanghui,Kim Dong Kyu,Dhong Hun Jong,Eun Kyoung Mi,Lee Kyung Eun,Kong Il Gyu,Kim HyoYeol,Chung Seung Kyu,Kim Dong Young,Rhee Chae Seo,Cho Seong Ho,Hong Sang Duk,Kim Dae Woo
Allergy, asthma & immunology research
PURPOSE:Despite medical and surgical treatments, some cases of nasal polyps (NP) exhibit recidivism. However, the endotype of refractory chronic rhinosinusitis with NP (CRSwNP) remains unclear. Therefore, the objective of this study was to characterize the immunological profile of refractory CRSwNP. METHODS:The control (n =23), primary NP group (pNP, n =70) and refractory NP group (rNP, n =86) were enrolled in this study. Patients who underwent revision surgeries due to failed maximal medical treatment after primary surgery were defined as the rNP group. A total of 18 inflammatory markers were investigated in nasal tissues using multiplex cytokine assay or enzyme-linked immunosorbent assay. RESULTS:The clinical characteristics of rNP included more extensive disease and worse clinical course after surgery. Additionally, rNP subjects showed higher infection rate (mucopurulence and culture-positive rate), more frequent use of antibiotics and suffered from symptomatic bacterial infection, increased asthma morbidity compared to pNP. Cytokine profile analysis showed that levels of Th17-associated mediators (myeloperoxidase, interleukin (IL)-8, IL-17A and IL-23), B-cell activating factor (BAFF) and Th1 cytokine (interferon-γ) were up-regulated in rNP compared to controls and pNP. Human neutrophil elastase-positive cells were also enhanced in rNP compared with pNP. Upregulation of Th17/Th1mediators and BAFF were observed in rNP, regardless of tissue eosinophilia or asthmatic comorbidity. Interestingly, eosinophilic markers, such as eosinophil cationic protein and C-C motif chemokine ligand 24, were up-regulated in asthmatic rNP compared to pNP and controls. Levels of anti-dsDNA immunoglobulin (Ig) G and IgA were up-regulated in rNP and highest in asthmatic eosinophilic rNP among subtypes of rNP. CONCLUSIONS:Our results suggest that Th17/Th1-associated mediators and BAFF may play a role and be a potential therapeutic target in refractory CRSwNP. Additionally, eosinophilic markers and autoantibodies may contribute to refractoriness in asthmatic rNP.
Distinct mucosal immunopathologic profiles in atopic and nonatopic chronic rhinosinusitis without nasal polyps in Central China.
Wang Bao-Feng,Cao Ping-Ping,Long Xiao-Bo,Zhang Xin-Hao,Xu Kai,Cui Yong-Hua,Liu Zheng
International forum of allergy & rhinology
BACKGROUND:The role of atopy to aeroallergens in chronic rhinosinusitis without nasal polyps (CRSsNP) remains unclear. This study aimed to investigate the mucosal immunopathologic characteristics of CRSsNP with and without atopy to inhalant allergens. METHODS:Thirteen nonatopic CRSsNP patients, 9 atopic CRSsNP patients, and 11 nonatopic control subjects were enrolled in this study. The expression of type 1, 2, and 17 cytokines, growth factors, and chemokines for T cell subsets and granulocytes in sinonasal mucosa was detected using Bio-Plex suspension chip technology or enzyme-linked immunosorbent assay (ELISA). Subjective symptoms were scored on a visual analogue scale (VAS), while disease severity on computed tomography (CT) was graded by the Lund-Mackay CT scoring system. RESULTS:There was no significant difference in VAS and CT scores between atopic and nonatopic CRSsNP. Compared with control, both atopic and nonatopic CRSsNP demonstrated increased interferon γ (IFN-γ) levels in sinonasal mucosa. In contrast, although no difference in interleukin 5 (IL-5), IL-13 and eotaxin-1 expression, or mucosal eosinophil infiltration, was found between the control and whole CRSsNP group, atopic CRSsNP manifested an increased expression of IL-5, IL-13 and eotaxin-1, as well as an enhanced infiltration of mucosal eosinophils in comparison with control and nonatopic CRSsNP. Mucosal eosinophil infiltration correlated with IL-5 and eotaxin-1 expression. No difference in IL-12, IL-4, IL-6, IL-17A, IL-8, myeloperoxidase, "regulated upon activation normal T cell expressed and presumably secreted" (RANTES), or chemokine (C-X-C motif) ligand 10 (CXCL10) protein expression was found among control, atopic CRSsNP, and nonatopic CRSsNP. CONCLUSION:Atopic and nonatopic CRSsNP have distinct mucosal immunopathologic profiles. CRSsNP is a heterogeneous disorder consisting of multiple groups of biological subtypes, or "endotypes," which may argue for different therapeutic strategies.
Activation of Activin receptor-like kinases curbs mucosal inflammation and proliferation in chronic rhinosinusitis with nasal polyps.
Tengroth Lotta,Arebro Julia,Larsson Olivia,Bachert Claus,Georén Susanna Kumlien,Cardell Lars-Olaf
Chronic rhinosinusitis with nasal polyps (CRSwNP) is a widespread disease causing obstruction of the nasal cavity. Its cause remains unclear. The transforming growth-factor beta (TGF-β) superfamily and their receptors, termed Activin receptor-like kinases (ALKs), have recently been suggested to play a role in local airway inflammation, but have so far not been evaluated in human nasal epithelial cells (HNECs) from CRSwNP patients. We demonstrated that ALK1-7 were expressed in the nasal polyp epithelium, and the expression of ALK1-6 was markedly elevated in polyps compared to nasal mucosa from healthy controls. Stimulation with the ALK ligand TGF-β1 decreased Ki67 expression in HNECs from CRSwNP patients, not evident in controls. Likewise, TGF-β1, Activin A and Activin B, all ALK ligands, decreased IL-8 release and Activin A and Activin B reduced ICAM1 expression on HNECs from CRSwNP patients, not seen in controls. Pre-stimulation with TGF-β1, Activin A, BMP4 and Activin B attenuated a TNF-α-induced ICAM1 upregulation on HNECs of CRSwNP. No effect was evident in controls. In conclusion, an increased expression of ALK1-6 was found on polyp epithelial cells and ligand stimulation appeared to reduce proliferation and local inflammation in polyps.
The Role of NF-κB in Chronic Rhinosinusitis With Nasal Polyps.
Jung Hahn Jin,Zhang Yu Lian,Kim Dong Kyu,Rhee Chae Seo,Kim Dong Young
Allergy, asthma & immunology research
PURPOSE:Whereas the majority of nasal polyps observed in Western populations are eosinophilic, non-eosinophilic nasal polyps are significantly more frequent in Asian countries. Given the importance of nuclear factor-kappa B (NF-κB) in inflammation, this study focused on the role of NF-κB in the pathogenesis of chronic rhinosinusitis with nasal polyps (CRSwNPs) in Asian patients. METHODS:A total of 46 patients were enrolled in this study (22 diagnosed with CRSwNPs, 10 with chronic rhinosinusitis without nasal polyps [CRSsNP], and 14 control subjects). Nasal polyps and uncinate tissues (UTs) were collected and the tissues prepared for hematoxylin-eosin staining and immunohistochemistric (IHC) analysis. Total RNA was isolated for real-time polymerase chain reaction for p65, interleukin (IL)-6, IL-8, intracellular adhesion molecule (ICAM)-1, IL-1β, tumor necrosis factor (TNF)-α, and eotaxin. RESULTS:In the CRSwNPs group, 50% of nasal polyps were non-eosinophilic. IHC revealed a significantly higher fraction of NF-κB p65-positive cells in nasal polyps of the CRSwNPs group than in the UTs of control and CRSsNP groups. No difference in NF-κB p65-positive cell fraction was observed between eosinophilic and non-eosinophilic nasal polyps. The mRNA expression of p65, IL-6, IL-8, and eotaxin was significantly higher in nasal polyps of the CRSwNPs than in the UTs of control and CRSsNP group. However, no difference in expression was observed between eosinophilic and non-eosinophilic nasal polyps, with the exception of IL-1β expression. CONCLUSIONS:Elevated expression of NF-κB- and NF-κB-associated inflammatory cytokines suggests NF-κB as the key factor for CRSwNPs pathogenesis in Asian patients. Understanding NF-κB-associated mechanisms will provide a deeper insight into CRSwNPs pathogenesis and ultimately improve therapeutic strategies for CRSwNPs.
The theory of a "staphylococcus superantigen" in chronic rhinosinusitis with nasal polyps: myth or reality?
Dobretsov K,Negm H,Ralli M,Passali D
European review for medical and pharmacological sciences
OBJECTIVE:The aim of our study was to search for evidence of a "staphylococcus superantigen" in chronic rhinosinusitis with nasal polyps. PATIENTS AND METHODS:Sixty-nine patients with chronic rhinosinusitis with nasal polyps and 45 healthy controls were included in the study. All patients in the study and control groups underwent bacteriological and immunological examination on nasal smear samples. Total IgE and the following cytokines were tested in all patients: tumor necrosis factor (TNF), interleukin-1 (IL1), interleukin-6 (IL6), interleukin-8 (IL8). RESULTS:The concentration of bacteria in the nasal cavity was much higher in patients in the study group compared to those in the control group, mainly due to staphylococci. In species identification of staphylococci, bacteria most represented were S. aureus and S. epidermidis. The greater the concentration of S. aureus, the lower the level of IgE. Proinflammatory cytokines were uniformly increased in patients with nasal polyps. The level of IgE was maximal in patients with chronic rhinosinusitis with nasal polyps with a poor growth of culture and minimal in patients with abundant growth, suggesting that in the latter the effect of eosinophilic inflammation on the disease was reduced, and conversely, the activity of eosinophilic inflammation was maximal with a poor seeding of the nasal cavity. CONCLUSIONS:Although this study has some limits, our findings do not support the theory of a staphylococcus superantigen in which the IgE level and eosinophilic inflammation should increase with increasing activity of Staphylococcus aureus. Further research supported by a larger sample of patients is required to better delineate the role of a staphylococcus superantigen in the pathogenesis of patients with chronic rhinosinusitis with nasal polyps.
Anti-Inflammatory Effects of EM900 on Cultured Human Nasal Epithelial Cells.
Wakayama Nozomu,Matsune Shoji,Takahara Eriko,Sekine Kuwon,Yoshioka Yuma,Ishida Mariko,Yamaguchi Satoshi,Okubo Kimihiro,Sunazuka Toshiaki,Ōmura Satoshi
Journal of Nippon Medical School = Nippon Ika Daigaku zasshi
OBJECTIVES:Macrolide therapy is an important conservative therapy for chronic rhinosinusitis, especially in Japan. The mechanism underlying this therapy involves anti-inflammatory and not antimicrobial activity. However, the administration of long-term low-dose macrolides (LTLMs) causes an increase in the number of antibiotic-resistant bacteria. EM900 is a derivative of erythromycin (EM), with anti-inflammatory but not antibacterial effects. It does not induce macrolide-resistant bacteria as shown by LTLM. In the present study, we analyzed the inhibitory effects of EM900 in comparison with those of clarithromycin (CAM) on inflammatory cytokine production in human nasal epithelial cells (HNEpCs). METHODS:After HNEpCs were cultured for 4 days, CAM or EM900 was added into the culture, followed by stimulation with tumor necrosis factor (TNF)-α. Interleukin (IL)-8 and vascular endothelial growth factor (VEGF) levels were measured using real-time polymerase chain reaction (RT-PCR) and an enzyme-linked immunosorbent assay (ELISA). RESULTS:Both the ELISA and RT-PCR showed that EM900 and CAM significantly inhibited IL-8 production in HNEpCs. In contrast, EM900 and CAM did not suppress the increased VEGF production when HNEpCs were stimulated with TNF-α. CONCLUSION:EM900 showed an anti-inflammatory effect, such as that of CAM, due to the inhibitory effect on IL-8 production in HNEpCs.
Thromboxane A2 Regulates CXCL1 and CXCL8 Chemokine Expression in the Nasal Mucosa-Derived Fibroblasts of Chronic Rhinosinusitis Patients.
Tsai Yih-Jeng,Hao Sheng-Po,Chen Chih-Li,Wu Wen-Bin
BACKGROUND:Chronic rhinosinusitis without nasal polyps (CRSsNP) is a common chronic disease and the etiology remains unclear. Thromboxane A2 (TXA2) participates in platelet aggregation and tissue inflammation. In this study, the CXCL1/8 chemokine and TXA2-TP receptor expression in the CRSsNP mucosa was investigated. EXPERIMENTAL APPROACH:Immunohistochemistry, chemokine release assay by ELISA, RT-PCR, Real-time PCR, Western blotting, pharmacological and siRNA knockdown analysis were applied in the CRSsNP tissue specimen and cultured nasal mucosa-derived fibroblasts. RESULTS:The immunohistochemistry results indicated that CXCL1 and CXCL8 were highly expressed in the CRSsNP mucosa compared with the controls; however, the TP receptors were expressed in both mucosa. Therefore, U46619 and IBOP, a TXA2 analog and TP agonist, were used to explore the role of TP activation in CXCL1/8 expression; both of these induced CXCL1/8 mRNA and protein expression in CRSsNP mucosa-derived fibroblasts. U46619 phosphorylated PI-3K, cyclic AMP (cAMP)/PKA, PKC, and cAMP response element (CREB). Activation of cAMP/PKA, PKC, and CREB was the major pathway for cxcl1/8 gene transcription. Pharmacological and siRNA knockdown analyses revealed that activation of cAMP/PKA and PKCμ/PKD pathways were required for CREB phosphorylation and PKA/C crosstalked with the PI-3K pathway. CONCLUSION AND IMPLICATIONS:Our study provides the first evidence for abundant TP receptor and CXCL1/8 expression in human CRSsNP mucosa and for TXA2 stimulation inducing CXCL1/8 expression in nasal fibroblasts primarily through TP receptor, cAMP/PKA, PKCμ/PKD, and CREB-related pathways.
Diesel Exhaust Particles Upregulate Interleukins IL-6 and IL-8 in Nasal Fibroblasts.
Kim Jin Ah,Cho Jae Hoon,Park Il-Ho,Shin Jae-Min,Lee Seoung-Ae,Lee Heung-Man
BACKGROUND:Diesel exhaust particles (DEP) are a major source of air pollution. Nasal fibroblasts are known to produce various cytokines and chemokines. The aim of this study was to evaluate DEP-induced cytokines and chemokines in nasal fibroblasts and to identify the signaling pathway involved. METHODS:A cytokine and chemokine array performed after stimulation of nasal fibroblasts with DEP revealed that levels of IL-6 and IL-8 were increased most significantly among various cytokines and chemokines. RT-PCR and ELISA were used to determine the mRNA and protein expression levels of IL-6 and IL-8. Signaling pathways of p-38, Akt, and NF-κB were analyzed by western blotting, luciferase assay, and ELISA. Organ cultures of nasal interior turbinate were also developed to demonstrate the ex vivo effect of DEP on the expression of IL-6 and IL-8 and the associated signaling pathway. RESULTS:DEP increased the expressions of IL-6 and IL-8 in nasal fibroblasts at mRNA and protein levels. DEP induced phosphorylation of p38, Akt, and NF-κB, whereas inhibitors of p38, Akt, and NF-κB blocked these phophorylations and the expressions of IL-6 and IL-8. These findings were also observed in ex vivo organ culture of nasal inferior turbinate. CONCLUSIONS:DEP induces expression of IL-6 and IL-8 via p38, Akt, and NF-κB signaling pathways in nasal fibroblasts. This finding suggests that air pollution might induce or aggravate allergic rhinitis or chronic rhinosinusitis.
HMGB1-TLR4 signaling contributes to the secretion of interleukin 6 and interleukin 8 by nasal epithelial cells.
Shimizu Shino,Kouzaki Hideaki,Kato Tomohisa,Tojima Ichiro,Shimizu Takeshi
American journal of rhinology & allergy
BACKGROUND:Alarmins play important roles in the pathogenesis of inflammatory and autoimmune diseases. However, the role of the alarmin protein high-mobility group box 1 (HMGB1) in upper airway inflammation is unclear. OBJECTIVE:To determine if HMGB1 is present in the nasal mucosa and, if so, to elucidate its role in upper airway inflammation. METHODS:Nasal secretions were collected from a total of 32 patients with chronic rhinosinusitis with nasal polyp, allergic rhinitis, and control subjects. The concentration of HMGB1 in nasal secretions and its tissue and cellular localization were examined by enzyme immunoassays and immunofluorescent staining of nasal polyps and cultured nasal epithelial cells. We then examined whether nasal epithelial cells secrete HMGB1 after inflammatory stimulation by tumor necrosis factor (TNF) α. The effects of HMGB1 on the production and secretion of interleukin (IL) 6 and IL-8 were also examined in cultured nasal epithelial cells. RESULTS:Significantly higher concentrations of HMGB1 were found in nasal secretions from patients with chronic rhinosinusitis with nasal polyp or allergic rhinitis compared with the control subjects. HMGB1 expression was localized in the nuclei of epithelial cells and other constitutive cells in nasal polyps and in the nuclei of cultured nasal epithelial cells. TNF-α stimulated the production and secretion of HMGB1 by cultured nasal epithelial cells. HMGB1 stimulated the production and secretion of IL-6 and IL-8 by cultured nasal epithelial cells, and anti-toll-like receptor 4 blocking antibody significantly inhibited HMGB1-induced secretion of IL-6 and IL-8. CONCLUSIONS:Nasal secretions contain substantial amounts of HMGB1. TNF-α stimulates the production of HMGB1, which, in turn, upregulates the production and secretion of IL-6 and IL-8 by nasal epithelial cells via toll-like receptor 4, which indicated that HMGB1 plays an important role in the pathogenesis of upper airway inflammation.
ROS-dependent HMGB1 secretion upregulates IL-8 in upper airway epithelial cells under hypoxic condition.
Min H J,Kim J-H,Yoo J E,Oh J-H,Kim K S,Yoon J-H,Kim C-H
High-mobility group box 1 (HMGB1) mediates various functions according to the location. We tried to investigate the role of HMGB1 in upper airway under hypoxic conditions. We cultured primary normal human nasal epithelium (NHNE) cells under hypoxic conditions and evaluated the movement of HMGB1 by western blotting, immunofluorescence, and enzyme-linked immunosorbent assay (ELISA). Reactive oxygen species (ROS) level was evaluated to estimate the translocation mechanism of HMGB1. The role of secreted HMGB1 was evaluated by ELISA assay. Furthermore, we collected human nasal mucosa samples and nasal lavage fluids from patients conditioned under hypoxic and non-hypoxic environment, and compared the expression of HMGB1 in human nasal mucosa samples by immunohistochemistry and the levels of HMGB1 in lavage fluids using ELISA assay. Hypoxia induced translocation of HMGB1 into the extracellular area and it was dependent on ROS produced by dual oxidase 2. Secreted HMGB1 was involved in the upregulation of interleukin (IL)-8. In human samples, HMGB1 was translocated from nucleus to the cytoplasm in hypoxic-conditioned nasal mucosa. HMGB1 was increased in nasal lavage samples of chronic rhinosinusitis patients, whose sinus mucosa was supposed to be hypoxic as compared with controls. We suggest that HMGB1 is secreted in hypoxic condition via ROS-dependent mechanism and secreted HMGB1 participates in IL-8 upregulation mediating inflammatory response.
Predictive markers of long-term recurrence in chronic rhinosinusitis with nasal polyps.
Rosati Davide,Rosato Chiara,Pagliuca Giulio,Cerbelli Bruna,Della Rocca Carlo,Di Cristofano Claudio,Martellucci Salvatore,Gallo Andrea
American journal of otolaryngology
BACKGROUND:In last years, many attempts were made to recognize chronic rhinosinusitis with nasal polyps (CRSwNP) phenotypes focusing on identifying relevant key pathogenic molecules. Polyps recurrence rate ranges from 4% to 60%, so it's clear that not all clinical and immunologic factors associated with recurrence are known. OBJECTIVE:We investigate the inflammatory profile in patients with long term recurrent and non-recurrent CRSwNPs and if a specific profile is associated with recurrence, comparing eosinophilic, neutrophilic and lymphocytic infiltration, as well as IL-5 and IL-8 expression to long term recurrence rate. METHODS:This prospective study included 44 adult patients with CRSwNP treated with endoscopic sinus surgery between 2008 and 2010. Long term follow-up data (8-10 years) indicated that among 44 patients, 18 (40.1%) experienced long term recurrence of nasal polyposis needing maximal medical treatment or revision surgery. We realized two groups: one with patients who didn't present long term recurrence (26 patients) and another with patients who presented long term recurrence (18 patients) and in both groups eosinophilic, neutrophilic and lymphocytic infiltration and IL-5 and IL-8 expression were measured. RESULTS:The parameters that reached statistical significance (p < 0.05) comparing the two groups were eosinophilic infiltration and IL-5 expression, whereas neutrophilic and lymphocytic infiltration, as IL-8 expression didn't show any significant difference. Asthma and aspirin intolerance seemed significantly more frequent in patients with recurrence, while allergy presented not statistically significant difference between two groups. CONCLUSIONS:We can conclude that high eosinophilic infiltration and high IL-5 expression in CRSwNP correlate with higher rate of long term recurrence, while neutrophilic and lymphocytic infiltration, and IL-8 expression don't correlate with it. These findings provide the opportunity to improve our ability to predict the prognosis of surgical intervention, although it is still needed to explore the optimal predictor of outcome in CRSwNP.
Chronic rhinosinusitis is associated with higher prevalence and severity of bronchiectasis in patients with COPD.
Yang Xia,Xu Yali,Jin Jianmin,Li Ruimin,Liu Xiaofang,Sun Yongchang
International journal of chronic obstructive pulmonary disease
BACKGROUND AND PURPOSE:Bronchiectasis revealed by high-resolution computed tomography (HRCT) is common in chronic obstructive pulmonary disease (COPD), but the causes and risk factors remain to be determined. Chronic rhinosinusitis (CRS) is closely associated with bronchiectasis or COPD, but whether it is associated with comorbid bronchiectasis in COPD (COPD-Bx) is unknown. PATIENTS AND METHODS:Patients with stable COPD were enrolled consecutively and evaluated for the presence of CRS by questionnaire and paranasal sinus computed tomography. The presence and severity of bronchiectasis on lung HRCT were evaluated by the Smith and severity scores. COPD symptoms were evaluated by COPD Assessment Test (CAT) and Modified British Medical Research Council Questionnaire. The sputum cell differentials and concentrations of interleukin (IL)-6, IL-8, IL-5, matrix metalloproteinases-9 (MMP-9), and tissue inhibitor of matrix metalloproteinases-1 were measured. RESULTS:We enrolled 136 patients with stable COPD, of which 66 (48.5%) were diagnosed with CRS according to the European Position Paper on Rhinosinusitis and Nasal Polyps (EPOS) criteria. The prevalence of bronchiectasis was 57.6% in patients with CRS, but 37.1% in those without CRS (=0.017). COPD-Bx patients with CRS showed a significantly higher severity score of bronchiectasis than those without CRS (=0.034). COPD patients with CRS had a higher percentage of eosinophils, higher levels of IL-8, IL-6, and MMP-9 in sputum as compared to those without CRS. In COPD-Bx patients with CRS, the percentage of eosinophils and the levels of IL-6 and MMP-9 in sputum were increased as compared to those without CRS. In all the subjects, Sino-Nasal Outcome Test-20 correlated with CAT score (=0.315, <0.01) and in COPD patients with CRS, Lund-MacKay scores correlated with forced expiratory volume in 1 s (% pred) (=-0.251, <0.05). CONCLUSIONS:CRS was associated with COPD-Bx and this was probably due to increased airway inflammation.
HSP70 is associated with the severity of inflammation in chronic rhinosinusitis.
Min Hyun Jin,Yoon Joo-Heon,Kim Chang-Hoon
American journal of rhinology & allergy
BACKGROUND:Nasal secretions include cytokines and inflammatory mediators that are involved in the pathogenesis of upper airway inflammation. OBJECTIVE:We tried to find unknown biomolecules that are involved in the pathogenesis of chronic rhinosinusitis (CRS). METHODS:We collected nasal mucosal secretions from patients who were diagnosed as having CRS and who underwent endoscopic sinus surgery. A total of 63 patients who underwent nasal secretion collection were reviewed. Enzyme-linked immunosorbent assay was performed by using nasal lavage samples to evaluate which biomolecules were associated with the severity of inflammation based on the Lund-Mackay score. By using human nasal epithelial cells, we performed Western blot, real-time polymerase chain reaction, and enzyme-linked immunosorbent assay to evaluate the secretory mechanism of heat shock protein (HSP) 70. RESULTS:We found that the level of interleukin 8 and HSP70 were significantly associated with the Lund-Mackay score and interleukin 17C, C-X-C motif chemokine 10, and HSP27 were not significantly associated. HSP70 was also significantly associated with the surgical outcome of the enrolled patients. Furthermore, we found that exposure to hypoxia and treatment of lipoteichoic acid induced the secretion of HSP70 but that lipopolysaccharide did not induce the secretion of HSP70 in human nasal epithelial cells. CONCLUSION:Our findings indicated that HSP70 might play a role in the pathogenesis of CRS and the possibility of HSP70 as a biomolecule that represents the severity of CRS.
Superoxide dismutase reduces the inflammatory response to Aspergillus and Alternaria in human sinonasal epithelial cells derived from patients with chronic rhinosinusitis.
Lawrence Lauren A,Mulligan Jennifer K,Roach Catherine,Pasquini Whitney N,Soler Zachary M,Banglawala Sarfaraz M,Karnezis Tom T,Gudis David A,Schlosser Rodney J
American journal of rhinology & allergy
BACKGROUND:Aspergillus fumigatus and Alternaria alternata are ubiquitous environmental fungal allergens that can exacerbate airway inflammation and contribute to the disease process in patients with chronic rhinosinusitis (CRS). These antigens have been shown to induce human sinonasal epithelial cells (HSNECs) to promote a proinflammatory response, but what is unclear is a means by which to reduce these effects. Inhaled pathogens can induce HSNECs to produce reactive oxygen species (ROS) that trigger cytokine production. OBJECTIVE:This study aimed to determine whether the free radical scavenger superoxide dismutase (SOD) could reduce HSNEC-derived inflammation, as measured by interleukin (IL)-6 and IL-8 production, in response to Aspergillus or Alternaria exposure. METHODS:Sinus tissue explants were collected at the time of surgery from control patients (n = 7) and patients with CRS with nasal polyps (CRSwNP) (n = 9). HSNECs were cultured from the explants and treated with Aspergillus, Alternaria, and SOD for 24 hours. Cell supernatants and lysates were collected, and IL-6 and IL-8 concentrations were measured using enzyme-linked immunosorbent assay. RESULTS:In control and CRSwNP HSNECs, Aspergillus and Alternaria both increased cytokine production (p < 0.05), as measured by IL-6 and IL-8 concentration. SOD treatment reduced the inflammatory response to fungal antigen exposure from CRSwNP HSNECs but not control HSNECs. In CRSwNP patients, SOD significantly decreased IL-6 and IL-8 production after Alternaria exposure and IL-8 after Aspergillus exposure (p < 0.05). CONCLUSIONS:When HSNECs from CRSwNP patients are treated with SOD concurrently with Aspergillus or Alternaria, SOD treatment decreases the fungal antigen-induced inflammatory response. The ability to attenuate inflammation induced by common fungal allergens with SOD treatment could provide a novel therapeutic or preventative approach for patients with CRS or other allergic inflammatory airway diseases.
Cytokines in Chronic Rhinosinusitis. Role in Eosinophilia and Aspirin-exacerbated Respiratory Disease.
Stevens Whitney W,Ocampo Christopher J,Berdnikovs Sergejs,Sakashita Masafumi,Mahdavinia Mahboobeh,Suh Lydia,Takabayashi Tetsuji,Norton James E,Hulse Kathryn E,Conley David B,Chandra Rakesh K,Tan Bruce K,Peters Anju T,Grammer Leslie C,Kato Atsushi,Harris Kathleen E,Carter Roderick G,Fujieda Shigeharu,Kern Robert C,Schleimer Robert P
American journal of respiratory and critical care medicine
RATIONALE:The mechanisms that underlie the pathogenesis of chronic rhinosinusitis without nasal polyps (CRSsNP), chronic rhinosinusitis with nasal polyps (CRSwNP), and aspirin-exacerbated respiratory disease (AERD) are not clear. OBJECTIVES:To first evaluate the inflammatory profiles of CRSsNP and CRSwNP tissues and then to investigate whether clinical differences observed between CRSwNP and AERD are in part secondary to differences in inflammatory mediator expression within nasal polyp (NP) tissues. METHODS:Expression levels of numerous inflammatory mediators were determined by quantitative real-time polymerase chain reaction, ELISA, and multiplex immunoassay. MEASUREMENTS AND MAIN RESULTS:CRSwNP NP had increased levels of type 2 mediators, including IL-5 (P < 0.001), IL-13 (P < 0.001), eotaxin-2 (P < 0.001), and monocyte chemoattractant protein (MCP)-4 (P < 0.01), compared with sinonasal tissue from subjects with CRSsNP and control subjects. Expression of IFN-γ messenger RNA or protein was low and not different among the chronic rhinosinusitis subtypes examined. Compared with CRSwNP, AERD NP had elevated protein levels of eosinophil cationic protein (ECP) (P < 0.001), granulocyte-macrophage colony-stimulating factor (GM-CSF) (P < 0.01), and MCP-1 (P = 0.01), as well as decreased gene expression of tissue plasminogen activator (tPA) (P = 0.02). Despite the higher eosinophilia in AERD, there was no associated increase in type 2 mediator protein levels observed. CONCLUSIONS:CRSwNP was characterized by a predominant type 2 inflammatory environment, whereas CRSsNP did not reflect a classic type 1 milieu, as has been suggested previously. AERD can be distinguished from CRSwNP by elevated ECP levels, but this enhanced eosinophilia is not associated with elevations in traditional type 2 inflammatory mediators associated with eosinophil proliferation and recruitment. However, other factors, including GM-CSF, MCP-1, and tPA, may be important contributors to AERD pathogenesis.
Cytokine correlation between sinus tissue and nasal secretions among chronic rhinosinusitis and controls.
Oyer Samuel L,Mulligan Jennifer K,Psaltis Alkis J,Henriquez Oswaldo A,Schlosser Rodney J
OBJECTIVES/HYPOTHESIS:Compare cytokine levels in sinus tissue to sinus secretions from controls and chronic rhinosinusitis patients. STUDY DESIGN:In vitro. METHODS:Polyurethane foam sponges were placed into middle meati of patients with chronic rhinosinusitis without nasal polyps (CRSsNP), with polyps (CRSwNP), and controls. Sinus biopsies were then taken from the same location. Protein levels of tumor necrosis factor-alpha (TNF-α), interferon-γ (IFN-γ), and interleukins (IL) 2, 4, 6, 8, 10, and 17A were measured via cytometric bead assay for each sample. Protein values from sinus tissue and secretions were compared with Pearson's correlation between samples as well as one-way ANOVA with posthoc t test between groups. RESULTS:Samples from 43 patients in total were examined. Mucus was measured from 10 controls, 11 CRSsNP and 10 CRSwNP, and sinus tissue was measured from 10 controls, 15 CRSsNP and 10 CRSwNP. IL-8 and IFN-γ levels were outside of the detectable range of the assay. Levels of secreted IL-2, 4, 6, 10, and 17A correlated with tissue levels (P < 0.05 for all, r > 0.49) while TNF-α did not (P = 0.71). CRSsNP had elevated mucus levels of IL-2, 4, 6, 10, and 17A compared to controls. CRSwNP had elevated mucus levels of IL-4, 6, 10, 17A, and TNF-α compared to controls. CONCLUSIONS:Cytokine levels in sinus secretions correlate with levels in sinus tissue and are elevated in CRS versus control based on Th1/Th2 skewing.
Immunological profiling of key inflammatory drivers of nasal polyp formation and growth in chronic rhinosinusitis.
Biggs T C,Hayes S M,Harries P G,Allan R N,Walls A F,Pender S L F,Salib R J
BACKGROUND:Chronic rhinosinusitis (CRS) is a chronic inflammatory condition of the upper airways, often associated with the formation of nasal polyps (CRSwNP). It is well established that macroscopically normal (non-polypoidal) sinonasal mucosa in CRSwNP patients can undergo polypoidal change over time, turning into frank polyps. However, little is known about what drives this process. This study aimed to investigate potential drivers of nasal polyp formation or growth through comparison of the immunological profiles of nasal polyps with contiguous non-polypoidal sinonasal mucosa, from the same patients. METHODS:The immune profiles of three types of tissue were compared; nasal polyps and adjacent non-polypoidal sinonasal mucosa from 10 CRSwNP patients, and sinonasal mucosa from 10 control patients undergoing trans-sphenoidal pituitary surgery. Nasal polyp and control samples were also stimulated with Staphylococcus aureus enterotoxin B (SEB) using a nasal explant model, prior to cytokine analysis. Real time quantitative polymerase chain reaction (IL-5, T-bet, IL-17A, FoxP3, TLR-4, IL-8, IL-1beta and IL-6) and Luminex (IFNgamma, IL-5 and IL-17A) were used to quantify pro-inflammatory responses. RESULTS:Nasal polyps and contiguous non-polypoidal sinonasal mucosa from CRSwNP patients displayed a very similar pro-inflammatory profile. When stimulated with SEB, nasal polyps displayed a Th2/Th17 mediated response when compared to controls. CONCLUSIONS:In CRSwNP, nasal polyps and non-polypoidal sinonasal mucosa from the same patient displayed a similar pro-inflammatory profile skewed towards the Th2/Th17 pathway in nasal polyps following SEB stimulation, with evidence of disordered bacterial clearance. These factors may contribute to enhanced survival of bacteria and development of a chronic inflammatory milieu, potentially driving new polyp formation and recurrence following surgical removal.
Cytokine profiles in Japanese patients with chronic rhinosinusitis.
Sejima Takayuki,Holtappels Gabriele,Kikuchi Hisashi,Imayoshi Shoichiro,Ichimura Keiichi,Bachert Claus
Allergology international : official journal of the Japanese Society of Allergology
BACKGROUND:Chronic rhinosinusitis (CRS) is classified in CRS without nasal polyp (CRSsNP) and CRS with nasal polyp (CRSwNP) in western countries, whereas this classification was not common so far in Japan. Studying inflammatory mediators in clearly defined disease subgroups may lead to a better differentiation of chronic sinus diseases. METHODS:Homogenates of sinonasal mucosal tissue from 14 controls, 9 CRSsNP patients, and 19 CRSwNP patients were assayed for transforming growth factor (TGF)-β, interleukin (IL)-5, immunoglobulin E (IgE), Staphylococcus enterotoxin (SAE)-IgE, eosinophil-catioic protein (ECP), myeloperoxidase (MPO), IL-1β, IL-6, and IL-8 by enzyme-linked immunosorbent assay or UNICAP system. RESULTS:CRSwNP had significantly higher levels of IL-5, IgE, SAE-IgE, and ECP compared with CRSsNP and controls. CRSsNP was characterized by high levels of TGF-β, while CRSwNP showed a Th2 polarization and lower levels of TGF-β. Especially, in CRSwNP samples, 68.4% were eosinophilic (ECP/MPO ratio >1), and 52.6% were SAE-IgE positive. On the other hand, in 9 CRSsNP patients, eosinophilic or SAE-IgE positive sample was only one sample respectively. Additionally, 31.6% of CRSwNP were asthmatic patients, while none of CRSsNP patient was suffering from bronchial asthma. CONCLUSIONS:This study is thought to be the first one that showed the cytokine profiles in Japanese CRSs/wNP similar to those of European CRS. Based on mediator profiles, we suggest that CRSsNP and CRSwNP are distinct disease entities within the group of chronic sinus diseases.
Features of airway remodeling in different types of Chinese chronic rhinosinusitis are associated with inflammation patterns.
Shi L-L,Xiong P,Zhang L,Cao P-P,Liao B,Lu X,Cui Y-H,Liu Z
BACKGROUND:The remodeling patterns in different types of chronic rhinosinusitis (CRS) have rarely been compared, particularly the difference between eosinophilic and noneosinophilic CRS with nasal polyps (CRSwNP). Moreover, whether there is a link between remodeling and inflammation remains controversial. OBJECTIVE:To directly compare the remodeling features of different CRS and to explore their relationship with inflammation in Chinese patients. METHODS:Histologic characteristics of surgical samples were analyzed in 33 controls, 72 eosinophilic and 76 noneosinophilic CRSwNP, and 72 CRS without nasal polyps (CRSsNP) patients. Tissue samples from 38 controls, 26 eosinophilic and 26 noneosinophilic CRSwNP, and 32 CRSsNP patients were measured for mRNA and/or protein expression of profibrotic growth factors, metalloproteinases (MMPs), tissue inhibitor of metalloproteinases (TIMPs), hypoxia-inducible factor (HIF)-1α, interleukin (IL)-8, eosinophil cationic protein (ECP), and myeloperoxidase (MPO). RESULTS:The amount of collagen decreased, whereas the edema scores increased, from CRSsNP to noneosinophilic CRSwNP and to eosinophilic CRSwNP. Transforming growth factor (TGF)-β2 protein levels were enhanced in CRSsNP compared with CRSwNP. TIMP-4 protein levels decreased in eosinophilic CRSwNP compared with noneosinophilic CRSwNP and CRSsNP. The number of neutrophils decreased from CRSsNP to noneosinophilic CRSwNP and to eosinophilic CRSwNP. ECP levels were only up-regulated in eosinophilic CRSwNP. ECP levels and neutrophil number correlated positively with the severity of edema and fibrosis, respectively. Neutrophils were the major sources of TGF-β2 in CRSsNP and noneosinophilic CRSwNP. CONCLUSION:Distinct remodeling patterns are revealed for different types of CRS, particularly for eosinophilic and noneosinophilic CRSwNP. Tissue remodeling associates with inflammation in CRS.
Nasal fluid cytology and cytokine profiles of eosinophilic and non-eosinophilic chronic rhinosinusitis with nasal polyps.
Zhu Z,Wang W,Zhang X,Wang X,Zha Y,Chen Y,Zhou L,Lv W
BACKGROUND:Chronic rhinosinusitis with nasal polyps (CRSwNP) is a heterogeneous disease with different clinical characteristics and different treatment responsiveness. The aims of this study were to compare the nasal fluid cytology and cytokines between eosinophilic CRSwNP (eCRSwNP) and non-eosinophilic CRSwNP (neCRSwNP) and establish a new multivariate model to predict eCRSwNP before surgery to improve personalized treatment for CRSwNP patients. METHODS:Eighty-six consecutive patients with CRSwNP and sixteen healthy controls were recruited in this study. Nasal fluid (NF) was collected from all subjects and nasal polyp tissue was collected during the surgery. The differential cell counts and concentrations of IL-6, IL-8, TNF-77; and IL-10 in NF were measured. Univariate and multivariate logistic regression were used to identify predictors for eCRSwNP. RESULTS:There were more inflammatory cells in NF of CRSwNP than controls. The eosinophil percentage was significantly higher in eCRSwNP than neCRSwNP and controls. The level of IL-8 was significantly higher in neCRSwNP than in eCRSwNP and controls. Blood eosinophilia, nasal fluid eosinophilia, higher total ethmoid score / total maxillary score (E/M ratio) and higher visual analogue scale (VAS) score of CRS were associated with eCRSwNP, the area under receiver operating characteristic curve (AUC) was 0.800, 0.755, 0.703 and 0.648, respectively. Using the coefficients of multivariate regression, we set up a scoring system to predict eCRSwNP with three of the variates and the AUC was 0.883. CONCLUSION:ECRSwNP, neCRSwNP and healthy controls demonstrated different cytology and cytokine profiles in NF. A new preoperational multivariate prediction model for eCRSwNP with NF eosinophilia, blood eosinophilia and higher E/M ratio was established.
Diversity of T cytokine profiles in patients with chronic rhinosinusitis: A multicenter study in Europe, Asia, and Oceania.
Wang Xiangdong,Zhang Nan,Bo Mingyu,Holtappels Gabriele,Zheng Ming,Lou Hongfei,Wang Hong,Zhang Luo,Bachert Claus
The Journal of allergy and clinical immunology
BACKGROUND:To date, no study has evaluated the diversity of T cell cytokine patterns of patients with chronic rhinosinusitis (CRS) among centers in different continents using identical methods. OBJECTIVE:We sought to assess T cytokine profiles in patients with CRS from Europe, Asia, and Australia. METHODS:Patients with chronic rhinosinusitis with nasal polyps (CRSwNP) and without nasal polyps (CRSsNP; n = 435) and control subjects (n = 138) were recruited from centers in Adelaide, Benelux, Berlin, Beijing, Chengdu, and Tochigi. Nasal mucosal concentrations of T2, T17, and T1 cytokines; eosinophilic cationic protein (ECP); myeloperoxidase (MPO); IL-8; and tissue total and Staphylococcus aureus enterotoxin (SE)-specific IgE were measured by using identical tools. RESULTS:Combinations of T1/T2/T17 cytokine profiles in patients with CRSwNP varied considerably between regions. CRSwNP tissues from patients from Benelux, Berlin, Adelaide, and Tochigi were T2 biased, whereas those from Beijing mainly demonstrated T2/T1/T17 mixed patterns, and patients from Chengdu showed an even lower T2 expression. Concentrations of IL-8 and tissue total IgE in patients with CRSwNP were significantly higher than those in control subjects in all regions. More than 50% of patients with CRSwNP in Benelux, Berlin, Adelaide, and Tochigi showed a predominantly eosinophilic endotype compared with less than 30% of patients in Beijing and Chengdu. SE-specific IgE was found in significantly greater numbers in patients with CRSwNP from Benelux, Adelaide, and Tochigi and significantly lower numbers in patients from Beijing and Chengdu. Moreover, the T1/T2/T17 cytokine profiles in patients with CRSsNP showed diversity among the 6 regions. CONCLUSION:T cytokine levels, eosinophilic/neutrophilic patterns, and SE-specific IgE expressions show extreme diversity among patients with CRS from Europe, Asia, and Oceania.
Olfactory and middle meatal cytokine levels correlate with olfactory function in chronic rhinosinusitis.
Wu Jeffanie,Chandra Rakesh K,Li Ping,Hull Benjamin P,Turner Justin H
OBJECTIVES/HYPOTHESIS:The etiology of chronic rhinosinusitis (CRS)-associated olfactory loss is unclear, but may result from inflammatory changes in the olfactory epithelium that result in signaling dysfunction or loss of olfactory neurons. Several proinflammatory cytokines have been associated with CRS, but their expression within the olfactory cleft microenvironment and association with olfactory function is unknown. STUDY DESIGN:Prospective case-control study. METHODS:Mucus was collected from the olfactory cleft and middle meatus of 31 CRS without nasal polyps subjects, 36 CRS with nasal polyps (CRSwNP) subjects, and 12 healthy controls. Olfactory function was assessed using the validated Smell Identification Test (SIT). Site-specific levels of 14 cytokines/chemokines (interleukin [IL]-1β, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12, IL-13, IL-17A, tumor necrosis factor-α, Eotaxin, RANTES [regulated on activation, normal T cell expressed and secreted]) were assessed using a multiplex flow cytometric bead assay and correlated with SIT scores. RESULTS:Mucus cytokine levels in the olfactory cleft were strongly or moderately correlated with levels in the middle meatus for all but one measured inflammatory mediators. SIT scores were inversely correlated with levels of IL-2 (P = .006), IL-5 (P < .0001), IL-6 (P = .0009), IL-10 (P < .0001), and IL-13 (P < .0001), with significance largely driven by CRSwNP patients. CONCLUSIONS:The inflammatory microenvironment within the olfactory cleft mirrors that within the middle meatus. Elevated levels of IL-2, IL-5, IL-6, IL-10, and IL-13 in olfactory cleft mucus are associated with reduced olfactory identification scores in CRS patients. Altered levels of select olfactory mucus cytokines could potentially have deleterious effects on olfactory neuron function and turnover. LEVEL OF EVIDENCE:NA. Laryngoscope, 128:E304-E310, 2018.
Inflammatory endotypes of chronic rhinosinusitis based on cluster analysis of biomarkers.
Tomassen Peter,Vandeplas Griet,Van Zele Thibaut,Cardell Lars-Olaf,Arebro Julia,Olze Heidi,Förster-Ruhrmann Ulrike,Kowalski Marek L,Olszewska-Ziąber Agnieszka,Holtappels Gabriele,De Ruyck Natalie,Wang Xiangdong,Van Drunen Cornelis,Mullol Joaquim,Hellings Peter,Hox Valerie,Toskala Elina,Scadding Glenis,Lund Valerie,Zhang Luo,Fokkens Wytske,Bachert Claus
The Journal of allergy and clinical immunology
BACKGROUND:Current phenotyping of chronic rhinosinusitis (CRS) into chronic rhinosinusitis with nasal polyps (CRSwNP) and chronic rhinosinusitis without nasal polyps (CRSsNP) might not adequately reflect the pathophysiologic diversity within patients with CRS. OBJECTIVE:We sought to identify inflammatory endotypes of CRS. Therefore we aimed to cluster patients with CRS based solely on immune markers in a phenotype-free approach. Secondarily, we aimed to match clusters to phenotypes. METHODS:In this multicenter case-control study patients with CRS and control subjects underwent surgery, and tissue was analyzed for IL-5, IFN-γ, IL-17A, TNF-α, IL-22, IL-1β, IL-6, IL-8, eosinophilic cationic protein, myeloperoxidase, TGF-β1, IgE, Staphylococcus aureus enterotoxin-specific IgE, and albumin. We used partition-based clustering. RESULTS:Clustering of 173 cases resulted in 10 clusters, of which 4 clusters with low or undetectable IL-5, eosinophilic cationic protein, IgE, and albumin concentrations, and 6 clusters with high concentrations of those markers. The group of IL-5-negative clusters, 3 clusters clinically resembled a predominant chronic rhinosinusitis without nasal polyps (CRSsNP) phenotype without increased asthma prevalence, and 1 cluster had a TH17 profile and had mixed CRSsNP/CRSwNP. The IL-5-positive clusters were divided into a group with moderate IL-5 concentrations, a mixed CRSsNP/CRSwNP and increased asthma phenotype, and a group with high IL-5 levels, an almost exclusive nasal polyp phenotype with strongly increased asthma prevalence. In the latter group, 2 clusters demonstrated the highest concentrations of IgE and asthma prevalence, with all samples expressing Staphylococcus aureus enterotoxin-specific IgE. CONCLUSION:Distinct CRS clusters with diverse inflammatory mechanisms largely correlated with phenotypes and further differentiated them and provided a more accurate description of the inflammatory mechanisms involved than phenotype information only.
Chronic Rhinosinusitis without Nasal Polyps in Asian Patients Shows Mixed Inflammatory Patterns and Neutrophil-Related Disease Severity.
Kim Dae Woo,Eun Kyoung Mi,Roh Eun Youn,Shin Sue,Kim Dong-Kyu
Mediators of inflammation
Chronic rhinosinusitis (CRS) shows heterogeneous immunologic features. Western studies revealed that CRS without nasal polyps (CRSsNP) showed a predominantly type 1 immune response and CRS with nasal polyps (CRSwNP) was characterized by type 2 immune response; however, the detailed immunologic profile of CRSsNP in Asian patients has not been thoroughly investigated. Therefore, we investigated the inflammatory endotypes of CRSsNP in Asian patients. Patients with CRSsNP ( = 57), patients with CRSwNP ( = 13), and a control group ( = 10), who underwent endoscopic sinus surgery, were enrolled; uncinate process (UP) tissues were harvested from all patients. Homogenates were prepared from the UP of each group, and immunologic profiles were analyzed, including major cytokines (32 inflammatory mediators). When comparing the UPs between groups, CRSsNP patients showed higher levels of Th2 cytokines (IL-4 and IL-13), eosinophilic chemokines (CCL-11 and CCL-24), ECP, and total IgE expression than control subjects. In addition, several neutrophilic markers (IL-1, IL-6, IL-8, CXCL-1, CXCL-2, and MPO), IL-17A, IL-22, and TNF- were dominant in CRSsNP patients. Among these inflammatory mediators, IL-17A showed higher expression levels in CRSsNP patients than in the control group and CRSwNP patients. However, IFN- expression was not significantly elevated in CRSsNP patients. The levels of neutrophil-associated cytokines were well correlated with each other; of which, CXCL2, IL-8, and MMP-9/TIMP-1 levels were significantly correlated with disease extent ( = 0.338, = 0.317, and = 0.424, respectively). However, the levels of eosinophil-associated cytokines showed little correlation with each other and were not correlated with disease extent. Our study revealed that Asian CRSsNP patients showed a mixed (types 2 and 17) immune response, but neutrophil-related markers were dominant and associated with disease extent. Knowledge of this immunologic feature may help clinicians make better individual treatment decisions for Asian CRSsNP patients.
Contrasting the microbiomes from healthy volunteers and patients with chronic rhinosinusitis.
Aurora Rajeev,Chatterjee Dhrubamitra,Hentzleman Joshua,Prasad Gaurav,Sindwani Raj,Sanford Thomas
JAMA otolaryngology-- head & neck surgery
IMPORTANCE:Chronic rhinosinusitis (CRS) is the persistent inflammation of the sinus and nasal passages lasting over 3 months. The etiology of CRS is not well understood. OBJECTIVE:To obtain insights into the disease process, we contrasted the microbiome and immune response from patients with CRS and healthy controls. DESIGN, SETTING, AND PARTICIPANTS:A case vs control design was used. Samples were collected in the operating room in an institutional hospital or clinic. Thirty patients with CRS and 12 healthy controls undergoing surgery were recruited. MAIN OUTCOMES AND MEASURES:The microbiome was analyzed by deep sequencing of the bacterial 16S and fungal 18S ribosomal RNA genes. Immune response was measured by quantification of 30 different cytokines by multiplexed enzyme-linked immunosorbent assay, and immune cells in the lavage were identified by flow cytometry. The immune response of peripheral blood leukocytes to the lavage microbiota was assessed by interleukin (IL)-5 enzyme-linked immunospot assay. RESULTS:While quantitative increase in most bacterial and fungal species was observed in patients with CRS relative to controls, the microbiomes of patients with CRS were qualitatively similar to the controls. Because these results indicated that bacteria and fungi are not triggering CRS, we undertook a more detailed characterization of the immune response. Patients with CRS had increased levels of the following cytokines: IL-4, IL-5, IL-8, and IL-13, along with increased levels of eosinophils and basophils in the lavage. Importantly, peripheral blood leukocytes isolated from patients with CRS responded to control lavage samples (ie, to commensals) to produce IL-5. In contrast, the same lavage sample evoked no IL-5 production in leukocytes from healthy controls. CONCLUSIONS AND RELEVANCE:These findings support the theory that in some cases CRS results from an immune hyperresponsiveness to commensal organisms.
The expression of epithelial intercellular junctional proteins in the sinonasal tissue of subjects with chronic rhinosinusitis: a histopathologic study.
Li Ying,Wang Xiangdong,Wang Ruihan,Bo Mingyu,Fan Erzhong,Duan Su,Zhang Luo
ORL; journal for oto-rhino-laryngology and its related specialties
OBJECTIVE:To evaluate the expression of five epithelial intercellular junctional proteins in the sinonasal tissue of subjects with chronic rhinosinusitis (CRS). METHODS:Forty-one samples of nasal polyp tissue of CRS patients with nasal polyps (wNP), 20 ethmoid sinus mucosa of CRS patients without nasal polyps (sNP) and 19 nasal mucosa of controls were collected and assessed for the expression of zonulae occludens (ZO-1), claudin-1, E-cadherin and desmoglein-1 and -2 (DSG1, DSG2) using immunohistochemical staining. Interleukin (IL)-5, IL-6 and IL-8 concentrations in the tissues were also measured using ELISA. RESULTS:The expression of ZO-1, claudin-1, DSG1 and DSG2 in the CRSwNP patient group and the expression of claudin-1, DSG1 and DSG2 of the CRSsNP patient group was significantly lower compared to that of the control group. Furthermore, the expression of DSG1 in the CRSwNP patient group was also significantly lower than in the CRSsNP patient group. In contrast, the expression of E-cadherin in the CRSwNP and the CRSsNP patient groups was significantly greater compared to the controls. The assessment of associations between the expression of the intercellular junctional proteins and cytokines demonstrated negative correlations between IL-5 and claudin-1, IL-6 and claudin-1, IL-6 and DSG2, IL-8 and DSG1, and IL-8 and DSG2. In contrast, a positive correlation was found between IL-8 and E-cadherin. CONCLUSIONS:Differences in the expression of epithelial intercellular junctional proteins may play an important role in the pathogenesis of CRS.
Differences in Expression of Inflammatory Mediator in Mucosal and Polyp Tissue between Chronic Rhinosinusitis and Recurrent Chronic Rhinosinusitis.
Huriyati Effy,Darwin Eryati,Yanwirasti Yanwirasti,Wahid Irza
Open access Macedonian journal of medical sciences
BACKGROUND:Chronic rhinosinusitis with nasal polyps (CRSwNP) remains a challenging clinical entity with its propensity for recurrence. This disease decreases the patients' quality of life and creates a high economic burden. An effort to investigate the aetiology of recurrent polyps has to be more alert. AIM:This study aims to prove the differences in expression of IL-5, IL-8, IL-17A and TGF-β1 in mucosal and polyp tissue between CRSwNP and recurrent CRSwNP and also to determine which expression of cytokines that have the main role in mucosal and polyp tissue in recurrent CRSwNP. MATERIAL AND METHODS:An observational study was conducted with a comparative cross-sectional design of CRS patients with 15 recurrent CRSwNP and CRSwNP who had never undergone surgery for as many as 15 polyps. Mucosal specimens of nasal polyps are taken by brushing, and polyp tissue specimens are taken during surgical removal of nasal polyps. Specimens from the polyp mucosa were examined by ELISA while the polyp tissue specimens were carried out immunohistochemistry (IHC). RESULTS:The result showed that there is a significant difference in IL-5 expression in the polyp mucosal between CRSwNP with recurrent CRSwNP, where expression is higher in recurrent CRSwNP. The expression of IL-8, IL-17 and TGF-β1 were lower in recurrent CRSwNP, but the difference was not significant. In nasal polyp tissue, there is a significant difference in TGF-β1 and IL-8 expression between CRSwNP and recurrent CRSwNP, where the expression of both cytokines is lower in recurrent CRSwNP. Interleukin-5 expression was higher in recurrent CRSwNP than CRSwNP, but the difference was not significant. In the polyps mucosal, IL-5 has the main role in recurrent CRSwNP polyp, whereas TGF-β has the main role in polyp tissue. CONCLUSION:This study concluded that the expression of IL-5 in the mucosa could be examined with simple techniques like brushing before polypectomy or FESS was performed to determine the possibility of polyps recurrencies.
Hypomethylation of the IL8 promoter in nasal epithelial cells of patients with chronic rhinosinusitis with nasal polyps.
Li Jingyun,Jiao Jian,Wang Ming,Gao Yunbo,Li Ying,Wang Yang,Zhang Yuan,Wang Xiangdong,Zhang Luo
The Journal of allergy and clinical immunology
BACKGROUND:IL-8 is an important chemokine implicated in the pathogenesis of chronic rhinosinusitis (CRS), but little is known about epigenetic regulation of IL8 in the pathogenesis of CRS. OBJECTIVE:We sought to investigate the relationship between the DNA methylation level in the IL8 proximal promoter and CRS in Han Chinese subjects. METHODS:Patients with chronic rhinosinusitis with nasal polyps (CRSwNP; n = 187), patients with chronic rhinosinusitis without nasal polyps (CRSsNP; n = 89), and control subjects (n = 57) were enrolled in 2 independent cohorts. Purified human nasal epithelial cells from each participant were assessed for percentage DNA methylation of CpG sites in the IL8 proximal promoter by using bisulfite pyrosequencing and for functional aspects of methylation status by using in vitro assays. RESULTS:DNA methylation of CpG sites 1, 2, and 3, respectively, in the IL8 proximal promoter was significantly decreased in human nasal epithelial cells of patients with CRSwNP compared with that in patients with CRSsNP (P < .001) and control subjects (P < .001). Percentage of DNA methylation of the CpG3 site was correlated negatively with both tissue eosinophilic cationic protein (P < .01) and myeloperoxidase (P < .05) levels. IL-1β (P < .001) and TNF-α (P < .01) significantly increased IL8 expression accompanied by a reduction in methylation at the CpG3 site (P < .001). Electrophoretic mobility shift assays demonstrated that methylation status of CpG3 changed the binding of octamer-binding transcription factor 1 and nuclear factor κB. CONCLUSION:Decreased DNA methylation of particularly CpG sites in the IL8 proximal promoter might play a role in the pathogenesis of CRSwNP.
Expression of E-prostanoid receptors in nasal polyp tissues of smoking and nonsmoking patients with chronic rhinosinusitis.
Xie Li,Liu Ai-Guo,Peng Li-Yan,Wang Su-Jie,Zhang Yin-Ping,Wang Xian-Song
BACKGROUND:Different inflammatory reactions have been observed in the polyp tissues of nonsmokers and smokers with chronic rhinosinusitis (CRS). E-prostanoid (EP) receptors play a role in the inflammatory processes. Cigarette smoke (CS) exposure regulates EP-receptor expression levels promoting inflammatory mediator release from various inflammatory cells. In this study, we characterize the EP-receptor expression profiles in the polyps of nonsmoking and smoking CRS patients to explore the possible role of CS in the pathogenesis of chronic rhinosinusitis with nasal polyps (CRSwNP). METHODS:Polyp biopsies were obtained from 28 non-smoking and 21 smoking CRSwNP patients. Histopathological characteristics were observed under a light microscope. The prostaglandin E2 (PGE2), TNF-α, and IL-8 contents in polyp tissues were detected using enzyme-linked immunosorbent assay. Immunostaining was used to locate EP receptors in polyps. Messenger RNA and protein expression of EP receptors were examined using quantitative real-time polymerase chain reaction and Western blot, respectively. RESULTS:More severe inflammatory reactions occurred in polyp tissues of smoking CRSwNP patients. The PGE2, TNF-α, and IL-8 in tissue homogenate levels were significantly higher in smoking CRSwNP patients than those in nonsmoking CRSwNP patients. Moreover, the distribution of each EP receptor subtype was similar in both groups. Compared with the EP-receptor expression in nonsmokers, messenger RNA and protein of EP2 and EP4 receptor were significantly down-expressed in smoking patients, but EP1 and EP3 receptors did not show significant differences. CONCLUSION:CS exposure downregulates the expression levels of EP2 and EP4 receptors and stimulates the production of PGE2 and the proinflammatory cytokine IL-8 and TNF-α in polyp tissues of CRS patients. The down-expressed EP2 and EP4 receptors might be associated with severe inflammatory reactions in smoking CRSwNP patients.
Up-regulation of microRNA-335-5p reduces inflammation via negative regulation of the TPX2-mediated AKT/GSK3β signaling pathway in a chronic rhinosinusitis mouse model.
Gu Xiao,Yao Xiaocui,Liu Dengtao
Chronic rhinosinusitis (CRS) is featured with chronic symptoms of inflammation or infection in the nasal and sinus tissues. MicroRNAs (miRNAs/miRs), such as dysregulated expression of miR-125b and miR-26a, has been previously demonstrated to be related to CRS. The present study is intended to define the role of miR-335-5p in inflammation and the related mechanism in a mouse model of CRS. The differentially expressed genes associated with CRS were screened by microarray analysis. The targeting relationship between miR-335-5p and TPX2 was analyzed by target prediction program and dual luciferase reporter gene assay. The mouse model of CRS was established, and mice were introduced with miR-335-5p mimics, miR-335-5p inhibitors, or siRNA against TPX2 to explore the regulatory functions of miR-335-5p. The regulatory effect of miR-335-5p on inflammation with the involvement of the AKT signaling pathway was also analyzed with the expression of inflammatory cytokines and AKT signaling pathway-related factors measured. It was indicated that miR-335-5p regulated the TPX2 gene-mediated AKT signaling pathway. TPX2 was identified as a target gene of miR-335-5p, and miR-335-5p elevation inhibited the activation of the AKT signaling pathway. In mice with CRS, up-regulation of miR-335-5p or silence of TPX2 inhibited the inflammation, as evidenced by decreased levels of TNF-α, IL-6 and IL-8, and higher levels of GSK3β and IL-10. Collectively, miR-335-5p inhibits the activation of AKT signaling pathway by negatively mediating TPX2, which may confer anti-inflammatory protection in CRS.
Multidimensional endotypes of chronic rhinosinusitis and their association with treatment outcomes.
Liao B,Liu J-X,Li Z-Y,Zhen Z,Cao P-P,Yao Y,Long X-B,Wang H,Wang Y,Schleimer R,Liu Z
BACKGROUND:The expression of chronic rhinosinusitis (CRS) is multidimensional. Disease heterogeneity in patients with CRS remains poorly understood. This study aimed to identify endotypes of CRS using cluster analysis by integrating multidimensional characteristics and to explore their association with treatment outcomes. METHODS:A total of 28 clinical variables and 39 mucosal cellular and molecular variables were analyzed using principal component analysis. Cluster analysis was performed on 246 prospectively recruited Chinese CRS patients with at least 1-year postoperative follow-up. Difficult-to-treat CRS was characterized in each generated cluster. RESULTS:Seven subject clusters were identified. Cluster 1 (13.01%) was comparable to the classic well-defined eosinophilic CRS with polyps, having severe disease and the highest proportion of difficult-to-treat CRS. Patients in cluster 2 (16.26%) and cluster 4 (13.82%) had relatively lower proportions of presence of polyps and presented mild inflammation with moderate proportions of difficult-to-treat cases. Subjects in cluster 2 were highly atopic. Cluster 3 (7.31%) and cluster 6 (21.14%) were characterized by severe or moderate neutrophilic inflammation, respectively, and with elevated levels of IL-8 and high proportions of difficult-to-treat CRS. Cluster 5 (4.07%) was a unique group characterized by the highest levels of IL-10 and lacked difficult-to-treat cases. Cluster 7 (24.39%) demonstrated the lowest symptom severity, a low proportion of difficult-to-treat CRS, and low inflammation load. Finally, we found that difficult-to-treat CRS was associated with distinct clinical features and biomarkers in the different clusters. CONCLUSIONS:Distinct clinicopathobiologic clusters of CRS display differences in clinical response to treatments and characteristics of difficult-to-treat CRS.
Anticytokine autoantibodies in chronic rhinosinusitis.
Tsybikov Namjil N,Egorova Elena V,Kuznik Boris I,Fefelova Elena V,Magen Eli
Allergy and asthma proceedings
BACKGROUND:Anticytokine autoantibodies (AAbs) involve a great panel of cytokines both in healthy subjects and in patients with various diseases, but their incidence and pathophysiologic role are widely debated. The issue of AAbs in chronic rhinosinusitis (CRS) has never been explored. OBJECTIVE:The aim of the study was to check AAbs in patients with CRS and with nasal polyps (CRSwNP) and patients with CRS and without nasal polyps (CRSsNP). METHODS:One-hundred subjects with CRS and 40 healthy controls were included. CRS severity was determined by the 22-item Sino-Nasal Outcome Test and Lund-Mackay scores. Levels of immunoglobulin A (IgA), secretory IgA, IgG, IgE, interleukin (IL) 1β, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, and IL-17A, and AAbs levels in subjects' serum and nasal secretions (NS) were measured by the enzyme-linked immunosorbent assay. RESULTS:Forty-six patients with CRSsNP, 25 women (54.3%) and 21 men (45.7%), mean (standard deviation [SD]) ages 34.1 ± 12.3 years; and 54 patients with CRSwNP, 24 women (44.4%) and 30 men (55.6%), mean (SD) ages 37.9 ± 17.5 years. A group of 40 healthy subjects served as controls. In both CRSsNP and CRSwNP groups, serum and NS IL-1β, IL-4, IL-5, IL-6, IL-8, IL-10, and IL-17A levels were higher compared with healthy controls, but there was no difference in the serum levels of cytokines between the CRSsNP and CRSwNP groups. Binding IgA antibodies against IL-1β, IL-2, IL-5, and IL-8 were found at low levels in NS of both patients with CRSsNP and patients with CRSwNP. The highest levels of AAbs were detected against IL-5 (0.43 ± 0.38 optical density values) and IL-17A (0.51 ± 0.32 optical density values) in NS of patients with CRSwNP. In the CRSwNP group, a positive correlation was found between NS IL-5 and anti-IL-5 AAbs (r = 545; p < 0.001). Positive correlations between anti-IL-5 AAbs with NS total IgE (r = 0.424; p = 0.001) and with NS secretory IgA (r = 0.545; p < 0.001) were noted in the CRSwNP group. CONCLUSIONS:In patients with CRS, IgA class AAbs were detected in NS, whereas the highest levels of anti-IL-5 and anti-IL-17A AAbs were detected in patients with CRSwNP. Maybe these AAbs indicate disruption of immune tolerance and mucosal autoimmunity.
MicroRNA-761 suppresses remodeling of nasal mucosa and epithelial-mesenchymal transition in mice with chronic rhinosinusitis through LCN2.
Cheng Jinzhang,Chen Junjun,Zhao Yin,Yang Jingpu,Xue Kai,Wang Zonggui
Stem cell research & therapy
BACKGROUND:Chronic rhinosinusitis (CRS) is characterized by persistent symptomatic inflammation of the nasal passage and sinus mucosa. Various microRNAs (miRs) have been implicated in CRS. Hence, the current study was conducted to explore the effect of microRNA-761 (miR-761) on remodeling of nasal mucosa and epithelial-mesenchymal transition (EMT). METHODS:Bioinformatics analysis was initially performed to predict the differentially expressed genes (DEGs) associated with CRS. Gene targeting relationship between miR-761 and lipocalin 2 (LCN2) was analyzed by bioinformatics analysis and verified using dual-luciferase reporter gene assay. Histopathological analyses of the nasal mucosa tissues were conducted via hematoxylin-eosin (HE) and alcian blue (AB)-periodic acid Schiff (PAS) staining. ELISA was employed to determine the IL-8 and MMP-9 levels. To define downstream pathway of miR-761, levels of proteins related to LCN2/Twist1 signaling pathway were assessed. Additionally, the effects of miR-761 on EMT, proliferation, and apoptosis were determined. RESULTS:LCN2 was highly expressed in CRS. LCN2 was a target of miR-761. miR-761 overexpression or LCN2 silencing decreased IL-8 and MMP-9 levels and morphological changes in nasal epithelial tissue from CRS mice. Overexpressed miR-761 or silenced LCN2 decreased the expression of LCN2 and Twist1, indicating LCN2/Twist1 signaling pathway was inactivated. Moreover, miR-761 overexpression or LCN2 silencing reduced the expression of N-cadherin and vimentin, while increased that of E-cadherin, suggesting inhibition of EMT. Furthermore, miR-761 overexpression or LCN2 silencing promoted cell proliferation and inhibited cell apoptosis in CRS. CONCLUSION:Taken together, miR-761 suppressed the remodeling of nasal mucosa through inhibition of LCN2 and the LCN2/Twist1 signaling pathway.
Desmoglein 3 Silencing Inhibits Inflammation and Goblet Cell Mucin Secretion in a Mouse Model of Chronic Rhinosinusitis via Disruption of the Wnt/β-Catenin Signaling Pathway.
Cheng Jinzhang,Yang Jingpu,Xue Kai,Zhao Yin,Zhao Chang,Li Song,Wang Zonggui
Chronic rhinosinusitis (CRS) is a common disease characterized by inflammation of the nose and paranasal sinuses lasting over 12 weeks. This study aims to evaluate the effect of desmoglein 3 (DSG3) on inflammatory response and goblet cell mucin secretion in a mouse model of CRS. The CRS-related differentially expressed genes and disease genes were screened using microarray-based gene expression analysis. Subsequently, CRS mouse models were established. The levels of pro-inflammatory factors TNF-α, IL-6, and IL-8 were measured by ELISA. In addition, loss-of-function experiment was conducted using siRNAs targeting DSG3 and β-catenin. The secretion of mucins MUC5B and MUC5AC in goblet cells was detected, and the apoptosis of goblet cells was assessed. The regulatory effect of DSG3 on the Wnt/β-catenin signaling pathway was analyzed by determining the mRNA and protein levels of DSG3, Wnt, β-catenin, and GSK3β. DSG3 was identified to be an upregulated gene in CRS, which was further documented in CRS mice models. Elevated inflammation and mucin production were noted in CRS mice models. Also, it was found that DSG3 or β-catenin silencing could decrease the levels of TNF-α, IL-6, and IL-8, and the positive rates of MUC5B and MUC5AC while enhancing goblet cell apoptosis. The Wnt/β-catenin signaling pathway was blocked by DSG3, evidenced by downregulated Wnt and β-catenin as well as upregulated GSK3β mRNA and protein levels. Overall, this study provides evidence that silencing DSG3 could inhibit the activation of the Wnt/β-catenin signaling pathway, thus alleviating CRS.
Involvement of B2 receptor in bradykinin-induced proliferation and proinflammatory effects in human nasal mucosa-derived fibroblasts isolated from chronic rhinosinusitis patients.
Tsai Yih-Jeng,Hao Sheng-Po,Chen Chih-Li,Lin Brian J,Wu Wen-Bin
Chronic rhinosinusitis (CRS) is a chronic inflammatory disease of the sinonasal mucosa either accompanied by polyp formation (CRSwNP) or without polyps (CRSsNP). CRSsNP accounts for the majority of CRS cases and is characterized by fibrosis and neutrophilic inflammation. However, the pathogenesis of CRS, especially CRSsNP, remains unclear. Immunohistochemistry of CRSsNP specimens in the present study showed that the submucosa, perivascular areas, and the mucous glands were abundant in fibroblasts. Therefore, we investigated the effects bradykinin (BK), an autacoid known to participate in inflammation, on human CRSsNP nasal mucosa-derived fibroblasts (NMDFs). BK increased CXCL1 and -8 secretion and mRNA expression with EC50 ranging from 0.15~0.35 μM. Moreover, BK enhanced cell proliferation and upregulated the expressions of proinflammatory molecules, including cell adhesion molecules (CAMs) and cyclooxygenase (COX)-1 and -2. These functionally caused an increase in monocyte adhesion to fibroblast monolayer. Using pharmacological intervention and BKR siRNA knockdown, we demonstrated that the BK-induced CXCL chemokine release, cell proliferation and COX and CAM expressions were mainly through the B2 receptor (B2R). Accordingly, the B2R was preferentially expressed in the NMDFs than B1R. The B2R was highly expressed in the CRSsNP than the control specimens, while the B1R and kininogen (KNG)/BK expression slightly increased in the CRSsNP mucosa. Collectively, we report here for the first time that fibroblasts, KNG/BK, and BKRs are overexpressed in CRSsNP mucosa and BK upregulates chemokine expression, proliferation, and proinflammatory molecule expression in NMDFs via B2R activation, which lead to a functional increase in monocyte-fibroblast interaction. Our findings reveal a critical role of fibroblast, KNG/BK, and BKRs in the development of CRSsNP.
Cigarette smoke combined with Toll-like receptor 3 signaling triggers exaggerated epithelial regulated upon activation, normal T-cell expressed and secreted/CCL5 expression in chronic rhinosinusitis.
Yamin Moshe,Holbrook Eric H,Gray Stacey T,Harold Rachel,Busaba Nicolas,Sridhar Avinash,Powell Katia J,Hamilos Daniel L
The Journal of allergy and clinical immunology
BACKGROUND:Chronic rhinosinusitis (CRS) is characterized by persistent mucosal inflammation and frequent exacerbations. OBJECTIVE:To determine whether innate epithelial responses to cigarette smoke or bacterial or viral pathogens may be abnormal in CRS leading to an inappropriate inflammatory response. METHODS:Primary nasal epithelial cells (PNECs) were grown from middle turbinate biopsies of 9 healthy controls and 11 patients with CRS. After reaching 80% to 90% confluence, PNECs were exposed to medium or cigarette smoke extract (CSE) 5% (vol/vol) for 1 hour, washed, then stimulated with staphylococcal lipoteichoic acid, LPS, or double-stranded RNA (dsRNA). After 24 hours, gene expression was quantified by QRT-PCR. RESULTS:At baseline, PNECs revealed elevated TNF-alpha and growth-related oncogene-alpha (a C-X-C chemokine)/CXCL1 (GRO-alpha) (4-fold increase, P = .02; and 16-fold increase, P = .004, respectively) in subjects with CRS compared with controls with normal levels of IL-1beta, IL-6, IL-8/CXCL8, human beta-defensin-2, monocyte chemoattractant protein 2/CCL8, monocyte chemoattractant protein 3/CCL7, and regulated upon activation, normal T-cell expressed and secreted (RANTES)/CCL5. Immunostaining of nasal biopsies, however, revealed comparable epithelial staining for TNF-alpha, GRO-alpha, and RANTES. There were no differences in mRNA induction by CSE, TNF-alpha, lipoteichoic acid, LPS, or dsRNA alone. The combination of CSE+dsRNA induced exaggerated RANTES (12,115-fold vs 1500-fold; P = .03) and human beta-defensin-2 (1120-fold vs 12.5-fold; P = .05) in subjects with CRS. No other genes were differentially induced. Furthermore, CSE+dsRNA induced normal levels of IFN-beta, IFN-lambda1, and IFN-lambda2/3 mRNA in subjects with CRS. CONCLUSION:Cigarette smoke extract plus dsRNA induces exaggerated epithelial RANTES expression in patients with CRS. We propose that an analogous response to cigarette smoke plus viral infection may contribute to acute exacerbations and eosinophilic mucosal inflammation in CRS.
Sinonasal epithelial cell response to Staphylococcus aureus burden in chronic rhinosinusitis.
Kohanski Michael A,Lane Andrew P
JAMA otolaryngology-- head & neck surgery
IMPORTANCE:Chronic rhinosinusitis (CRS) is an inflammatory disorder of the nose and paranasal sinuses. Staphylococcus aureus is increasingly linked with CRS exacerbations. Little is known about how bacteria activate inflammatory pathways that contribute to CRS. OBJECTIVE:To develop an in vitro coculture system to explore how infection with S aureus stimulates innate immune responses of sinonasal epithelial cells (SNECs). DESIGN, SETTING, AND PARTICIPANTS:Sinonasal epithelial cells were collected from 13 patients during endoscopic sinus surgery and grown in culture at the air-liquid interface from July 2014 through December 2014. INTERVENTIONS:Differentiated SNECs from control individuals, patients with CRS with nasal polyps (CRSwNPs), and patients with CRS without nasal polyps (CRSsNPs) were infected with S aureus at 3 different concentrations for 24 hours. MAIN OUTCOMES AND MEASURES:Growth of S aureus and viability of SNECs were measured. Expression of inflammatory markers and innate immune genes was measured by reverse transcription-polymerase chain reaction. Basal secretion of interleukin 8 was determined by enzyme-linked immunosorbent assay. RESULTS:Cultured SNECs from patients with CRSsNPs demonstrated a significant increase (P < .05) in expression of interleukin 8 (23-fold to 82-fold) and tumor necrosis factor (11-fold to 61-fold) at all the tested concentrations of S aureus. Control or CRSwNP SNECs demonstrated a significant increase (P < .05) in expression of interleukin 8 (47-fold and 50-fold, respectively) and tumor necrosis factor (106-fold and 58-fold, respectively) at the higher inoculum of S aureus. Basal secretion of inflammatory markers correlated with expression changes. No significant changes in expression were observed for the helper T cell, subtype 2, inflammatory mediators tested. CONCLUSIONS AND RELEVANCE:In this study, we developed a model to study early innate immune-mediated changes in SNECs cocultured at an air-liquid interface with bacteria. We also demonstrated that bacterial burden can be detected by SNECs in the absence of adaptive immune-mediated responses. The CRSsNP SNECs are more sensitive to S aureus burden than control or CRSwNP SNECs. Future studies will further develop this infection model and explore the SNEC innate immune response to bacteria.
Primary role of growth-related oncogene-alpha and granulocyte chemotactic protein-2 as neutrophil chemoattractants in chronic rhinosinusitis.
Rudack C,Sachse F,Alberty J
Clinical and experimental allergy : journal of the British Society for Allergy and Clinical Immunology
BACKGROUND:The aetiology of chronic rhinosinusitis (CRS) remains unclear. The purpose of this study was to investigate neutrophil-attracting chemokine patterns in CRS without nasal polyposis. METHODS:The biological activity of the chemokines was identified using a two-step high-performance liquid chromatography (HPLC) technique combined with a bioassay in extracts from 55 CRS patients, and in the turbinate mucosa (TM) of patients (N=51) undergoing septumplasty. The biologic activity of each chemokine was assessed using blocking antibodies to chemokines. Immunolocalization of detected neutrophil chemokines was performed by quantitative evaluation of immunohistochemistry. Besides, PCR analysis was performed to quantify neutrophil chemokine mRNA. RESULTS:In CRS, the chemokines primarily detected by two-step HPLC were growth-related oncogene-alpha (GRO-alpha) and the granulocyte chemotactic protein-2 (GCP-2). Blocking of GCP-2 and GRO-alphad each resulted in chemotaxis inhibition rates of 43.3% and 35.9%, respectively, whereas anti-IL-8 and anti-ENA-78 had no effect. Both GCP-2 and GRO-alphad were generally synthesized by the surface epithelium and mucosal glands while GRO-alpha in particular was synthesized by endothelial cells, as shown by immunohistochemistry. The concentrations of the chemokines IL-8 and epithelial cell-derived neutrophil attractant-78 (ENA-78) were low in CRS and TM. CONCLUSION:It appears that both GRO-alpha and GCP-2 contribute to neutrophil chemotaxis in CRS, whereas IL-8 and ENA-78 appear to be of secondary importance for the chemotaxis of neutrophils in this condition. The expression of chemokines in mucosal gland cells is the main phenomenon involved in constitutive neutrophil chemotaxis in the TM.
Role of interleukins and transforming growth factor-beta in chronic rhinosinusitis and nasal polyposis.
Bradley Dewayne T,Kountakis Stilianos E
OBJECTIVES:To determine the role of interleukin (IL)-4, IL-4 receptor (R), IL-6, IL-8, IL-11, and transforming growth factor (TGF)-beta in chronic rhinosinusitis (CRS) and chronic rhinosinusitis with nasal polyposis (CRS/NP). METHODS:Sinus tissue from patients undergoing endoscopic sinus surgery for CRS and CRS/NP was collected. Sinus tissue was then analyzed using reverse-transcription polymerase chain reaction (RT-PCR) to detect transcription of IL-4R, IL-6, IL-8, and IL-11. Sinus tissue samples were also cultured in vitro, treated with IL-4 for 24 hours, and real-time PCR was used to quantify the transcription of TGF-beta. RESULTS:Twenty patients were evaluated, 9 with CRS/NP and 11 with CRS alone. The mean age was 43 (20-74) years, with 13 females and 7 males. IL-4R, IL-6, IL-8, and IL-11 were identified by RT-PCR in all 20 patients. The transcription of TGF-beta was found to be 3.2 times greater in patients with CRS/NP than in patients with CRS alone (P = .047). CONCLUSION:IL-6, IL-8, and IL-11 are nonspecific markers of sinus inflammation being transcribed in patients with CRS and patients with CRS/NP. However, patients with CRS/NP demonstrate increased transcription of TGF-beta in response to IL-4 treatment, suggesting an IL-4 mediated mechanism for stromal proliferation in the formation of nasal polyposis.
One airway: Biomarkers of protection from upper and lower airway injury after World Trade Center exposure.
Cho Soo Jung,Echevarria Ghislaine C,Kwon Sophia,Naveed Bushra,Schenck Edward J,Tsukiji Jun,Rom William N,Prezant David J,Nolan Anna,Weiden Michael D
BACKGROUND:Firefighters exposed to World Trade Center (WTC) dust have developed chronic rhinosinusitis (CRS) and abnormal forced expiratory volume in 1 s (FEV1). Overlapping but distinct immune responses may be responsible for the clinical manifestations of upper and lower airway injury. We investigated whether a panel of inflammatory cytokines, either associated or not associated with WTC-LI, can predict future chronic rhinosinusitis disease and its severity. METHODS:Serum obtained within six months of 9/11/2001 from 179 WTC exposed firefighters presenting for subspecialty evaluation prior to 3/2008 was assayed for 39 cytokines. The main outcomes were medically managed CRS (N = 62) and more severe CRS cases requiring sinus surgery (N = 14). We tested biomarker-CRS severity association using ordinal logistic regression analysis. RESULTS:Increasing serum IL-6, IL-8, GRO and neutrophil concentration reduced the risk of CRS progression. Conversely, increasing TNF-α increased the risk of progression. In a multivariable model adjusted for exposure intensity, increasing IL-6, TNF-α and neutrophil concentration remained significant predictors of progression. Elevated IL-6 levels and neutrophil counts also reduced the risk of abnormal FEV1 but in contrast to CRS, increased TNF-α did not increase the risk of abnormal FEV1. CONCLUSIONS:Our study demonstrates both independent and overlapping biomarker associations with upper and lower respiratory injury, and suggests that the innate immune response may play a protective role against CRS and abnormal lung function in those with WTC exposure.
P-glycoprotein regulates Staphylococcus aureus enterotoxin B-stimulated interleukin-5 and thymic stromal lymphopoietin secretion in organotypic mucosal explants.
Bleier Benjamin S,Singleton Amy,Nocera Angela L,Kocharyan Armine,Petkova Victoria,Han Xue
International forum of allergy & rhinology
BACKGROUND:T-helper 2 (Th2) inflammation is a hallmark of chronic rhinosinusitis with nasal polyps (CRSwNP) although the pathogenesis is poorly understood. P-glycoprotein (permeability glycoprotein, P-gp) is an efflux pump that is capable of regulating cytokine transport and is expressed within sinonasal mucosa. The purpose of this study was to examine if the oversecretion of interleukin 5 (IL-5) and thymic stromal lymphopoietin (TSLP) in CRSwNP could be explained through P-gp-mediated secretory pathways. METHODS:Fifteen ethmoid mucosal explants were harvested from patients with CRS (n = 10) and CRSwNP (n = 10) and stimulated with Staphylococcus aureus enterotoxin B (SEB). P-gp was inhibited using zosuquidar trihydrochloride (herein Zosuquidar). P-gp expression was measured using real-time polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). IL-5, IL-8, and TSLP secretion were quantified using ELISA. RESULTS:P-gp protein was overexpressed in CRSwNP (28.32 ± 25.94 ng/mL per mg explant) as compared to CRS (10.74 ± 8.61; p = 0.01, 2-tailed Mann-Whitney U test). There was no difference in messenger RNA (mRNA) expression. SEB induced a significant increase in IL-5 and TSLP but not IL-8 secretion relative to control in the CRSwNP explants only. Subsequent P-gp inhibition significantly reduced IL-5 and TSLP secretion (p = 0.04 for both, 2-tailed Student t test) to control levels. The concentration of IL-5 and TSLP secretion were strongly and significantly correlated to the concentration of P-gp within the same explant (IL-5: r = 0.791, p = 0.001; TSLP: r = 0.687, p = 0.003; 2-tailed Spearman's rank-order correlation). CONCLUSION:P-gp protein is expressed at higher concentrations in CRSwNP as compared to CRS. This overexpression directly contributes to the relative hypersecretion of IL-5 and TSLP. These findings suggest a novel mechanism for Th2 skewing in CRSwNP.
Role of TLRs in the production of chemical mediators in nasal polyp fibroblasts by fungi.
Shin Seung-Heon,Ye Mi-Kyung,Kim Yee-Hyuk,Kim Jeong-Kyu
Auris, nasus, larynx
OBJECTIVE:Fibroblasts are major supporting cells in nasal mucosa and can induce inflammatory process with recruitment of inflammatory cells. Airborne fungi have been suggested as an etiologic factor of chronic rhinosinusitis (CRS). The aim of this study was to investigate the interaction between airborne fungi and pattern recognition receptors (PRRs) in nasal fibroblasts. METHODS:Primary nasal polyp fibroblasts were cultured with Alternaria and Aspergillus for 48h. To determine the production of chemical mediators interleukine-6 (IL-6), IL-8, granulocyte-macrophage colony stimulating factor (GM-CSF), eotaxin, and regulated on activation normal T expressed and secreted (RANTES) were measured with enzyme immunoassay methods. PRRs for toll-like receptors (TLRs) and protease-activated receptors (PARs) mRNA were determined with reverse transcription polymerase chain reaction (RT-PCR). To determine the role of PRRs, fibroblasts were treated with small interfering RNA (siRNA). RESULTS:IL-6 and IL-8 productions were significantly increased by 50 and 100μg/ml of Alternaria. However, GM-CSF, eotaxin, and RANTES productions did not change. Aspergillus did not influence the production of chemical mediators from nasal polyp fibroblasts. TLR2 and TLR5 mRNA expressions were significantly increased by fungi and these two TLRs were associated with the production of IL-6 and IL-8. CONCLUSION:Alternaria interacts as a pathogen-associated molecular pattern with the PRRs, such as TLR2 and TLR5, which induce the production of inflammatory chemical mediators from nasal polyp fibroblasts. Airborne fungi enhance the innate immune defense mechanism and may be associated with the pathogenesis of nasal inflammatory diseases.
Proinflammatory impact of Staphylococcus epidermidis on the nasal epithelium quantified by IL-8 and GRO-alpha responses in primary human nasal epithelial cells.
Sachse F,von Eiff C,Becker K,Steinhoff M,Rudack C
International archives of allergy and immunology
BACKGROUND:Bacterial etiology of chronic rhinosinusitis (CRS) still remains controversial. Whereas Staphylococcus aureus enterotoxins have been detected in CRS, the impact of Staphylococcus epidermidis, a major commensal inhabitant of the nose, has not been studied. Among others, serine and cysteine proteases have been identified as factors of virulence in S. epidermidis. METHODS:S. epidermidis was examined in tissue biopsies of 30 CRS patients (16 with nasal polyposis) using standard procedures. Primary human nasal epithelial cells from inferior nasal turbinates (HNECs), from nasal polyps (NPECs) and A549 airway epithelial cells were stimulated with S. epidermidis supernatants DSM20044 or ATCC35984 and the IL-8 and GRO-alpha response was quantified by ELISA. Protease-triggered chemokine responses and involvement of NF-kappaB were investigated by addition of protease or NF-kappaB inhibitors. Activation of NF-kappaB was demonstrated by quantitative DNA binding assay. RESULTS:S. epidermidis was the most frequently isolated bacteria in the majority of CRS patients. HNECs and NPECs revealed no different IL-6 and IL-8 synthesis following stimulation with DSM20044 or ATCC35984. Stimulation of HNECs and A549 cells with S. epidermidis supernatants resulted in increased IL-8 and GRO-alpha expression which could be suppressed by the serine protease inhibitor AEBSF and the NF-kappaB inhibitor BAY 11 but not by the cysteine protease inhibitor E64. Results obtained for A549 cells were similar to HNECs. CONCLUSION:S. epidermidis was present in the majority of CRS specimens. Proinflammatory impact of S. epidermidis supernatants on nasal epithelial cells was demonstrated by serine protease-triggered and NF-kappaB-dependent chemokine responses.
Induction of interleukin-8 from nasal epithelial cells during bacterial infection: the role of IL-8 for neutrophil recruitment in chronic rhinosinusitis.
Yoon Bit-Na,Choi Nan-Geum,Lee Hyun-Sun,Cho Kyu-Sup,Roh Hwan-Jung
Mediators of inflammation
OBJECTIVES:The aim of this study was to elucidate the role of IL-8 for neutrophil recruitment in nonallergic CRS patients. METHODS:After coculture of Streptococcus pneumoniae (SP) with the mucosal epithelial cells (MECs) from non-CRS patients, at three different SP/MEC (1/1, 10/1, 100/1) ratios, the expression of IL-8 mRNA and the concentration of IL-8 were measured by RT-PCR and ELISA. The expression of CD11b/CD18 on neutrophils and E-selectin/ICAM-1 on endothelial cells and the adherence between neutrophils and human umbilical vascular endothelial cells (HUVECs) were determined by flow cytometric analysis, ELISA, and RIA, respectively. RESULTS:IL-8 concentration and IL-8 mRNA expression continued to increase from 3 hours after incubation in SP number-dependent manner. The expression of CD11b/CD18 on neutrophils and E-selectin/ICAM-1 on HUVECs, and the adherence between neutrophils and HUVECs were significantly increased in 10 SP/MEC-CM, and the increments were significantly blocked by anti-IL-8 antibody. CONCLUSION:MEC and IL-8 are major factors for neutrophil recruitment in nonallergic CRS.