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Cold-inducible RNA-binding protein (CIRP) induces translation of the cell-cycle inhibitor p27Kip1. Roilo Martina,Kullmann Michael K,Hengst Ludger Nucleic acids research The CDK inhibitor p27Kip1 plays a central role in controlling cell proliferation and cell-cycle exit. p27Kip1 protein levels oscillate during cell-cycle progression and are regulated by mitogen or anti-proliferative signaling. The abundance of the protein is frequently determined by post-transcriptional mechanisms including ubiquitin-mediated proteolysis and translational control. Here, we report that the cold-inducible RNA-binding protein (CIRP) selectively binds to the 5' untranslated region of the p27Kip1 mRNA. CIRP is induced, modified and relocalized in response to various stress stimuli and can regulate cell survival and cell proliferation particularly during stress. Binding of CIRP to the 5'UTR of the p27Kip1 mRNA significantly enhanced reporter translation. In cells exposed to mild hypothermia, the induction of CIRP correlated with increased translation of a p27Kip1 5'UTR reporter and with the accumulation of p27Kip1 protein. shRNA-mediated CIRP knockdown could prevent the induction of translation. We found that p27Kip1 is central for the decreased proliferation at lower temperature, since p27Kip1 KO mouse embryonic fibroblasts (MEFs) hardly increased their doubling time in hypothermic conditions, whereas wild-type MEFs significantly delayed proliferation in response to cold stress. This suggests that the CIRP-dependent p27Kip1 upregulation during mild hypothermia contributes to the cold shock-induced inhibition of cell proliferation. 10.1093/nar/gkx1317
Posttranscriptional Regulation of 14q32 MicroRNAs by the CIRBP and HADHB during Vascular Regeneration after Ischemia. Downie Ruiz Velasco Angela,Welten Sabine M J,Goossens Eveline A C,Quax Paul H A,Rappsilber Juri,Michlewski Gracjan,Nossent A Yaël Molecular therapy. Nucleic acids After induction of ischemia in mice, 14q32 microRNAs are regulated in three distinct temporal patterns. These expression patterns, as well as basal expression levels, are independent of the microRNA genes' order in the 14q32 locus. This implies that posttranscriptional processing is a major determinant of 14q32 microRNA expression. Therefore, we hypothesized that RNA binding proteins (RBPs) regulate posttranscriptional processing of 14q32, and we aimed to identify these RBPs. To identify proteins responsible for this posttranscriptional regulation, we used RNA pull-down SILAC mass spectrometry (RP-SMS) on selected precursor microRNAs. We observed differential binding of cold-inducible RBP (CIRBP) and hydroxyacyl-CoA dehydrogenase trifunctional multienzyme complex subunit beta (HADHB) to the precursors of late-upregulated miR-329-3p and unaffected miR-495-3p. Immunohistochemical staining confirmed expression of both CIRBP and HADHB in the adductor muscle of mice. Expression of both CIRBP and HADHB was upregulated after hindlimb ischemia in mice. Using RBP immunoprecipitation experiments, we showed specific binding of CIRBP to pre-miR-329 but not to pri-miR-329. Finally, using CRISPR/Cas9, we generated HADHB 3T3 cells, which display reduced expression of miR-329 and miR-495 but not their precursors. These data suggest a novel role for CIRBP and HADHB in posttranscriptional regulation of 14q32 microRNAs. 10.1016/j.omtn.2018.11.017
Cold-inducible RNA binding protein in cancer and inflammation. Lujan Daniel A,Ochoa Joey L,Hartley Rebecca S Wiley interdisciplinary reviews. RNA RNA binding proteins (RBPs) play key roles in RNA dynamics, including subcellular localization, translational efficiency and metabolism. Cold-inducible RNA binding protein (CIRP) is a stress-induced protein that was initially described as a DNA damage-induced transcript (A18 hnRNP), as well as a cold-shock domain containing cold-stress response protein (CIRBP) that alters the translational efficiency of its target messenger RNAs (mRNAs). This review summarizes recent work on the roles of CIRP in the context of inflammation and cancer. The function of CIRP in cancer appeared to be solely driven though its functions as an RBP that targeted cancer-associated mRNAs, but it is increasingly clear that CIRP also modulates inflammation. Several recent studies highlight roles for CIRP in immune responses, ranging from sepsis to wound healing and tumor-promoting inflammation. While modulating inflammation is an established role for RBPs that target cytokine mRNAs, CIRP appears to modulate inflammation by several different mechanisms. CIRP has been found in serum, where it binds the TLR4-MD2 complex, acting as a Damage-associated molecular pattern (DAMP). CIRP activates the NF-κB pathway, increasing phosphorylation of Iκκ and IκBα, and stabilizes mRNAs encoding pro-inflammatory cytokines. While CIRP promotes higher levels of pro-inflammatory cytokines in certain cancers, it also decreases inflammation to accelerate wound healing. This dichotomy suggests that the influence of CIRP on inflammation is context dependent and highlights the importance of detailing the mechanisms by which CIRP modulates inflammation. WIREs RNA 2018, 9:e1462. doi: 10.1002/wrna.1462 This article is categorized under: RNA in Disease and Development > RNA in Disease RNA Interactions with Proteins and Other Molecules > Protein-RNA Interactions: Functional Implications. 10.1002/wrna.1462
Extracellular cold inducible RNA-binding protein mediates binge alcohol-induced brain hypoactivity and impaired cognition in mice. Molecular medicine (Cambridge, Mass.) BACKGROUND:Alcohol abuse affects the brain regions responsible for memory, coordination and emotional processing. Binge alcohol drinking has shown reductions in brain activity, but the molecular targets have not been completely elucidated. We hypothesized that brain cells respond to excessive alcohol by releasing a novel inflammatory mediator, called cold inducible RNA-binding protein (CIRP), which is critical for the decreased brain metabolic activity and impaired cognition. METHODS:Male wild type (WT) mice and mice deficient in CIRP (CIRP) were studied before and after exposure to binge alcohol level by assessment of relative brain glucose metabolism with fluorodeoxyglucose (FDG) and positron emission tomography (PET). Mice were also examined for object-place memory (OPM) and open field (OF) tasks. RESULTS:Statistical Parametric Analysis (SPM) of FDG-PET uptake revealed marked decreases in relative glucose metabolism in distinct brain regions of WT mice after binge alcohol. Regional analysis (post hoc) revealed that while activity in the temporal (secondary visual) and limbic (entorhinal/perirhinal) cortices was decreased in WT mice, relative glucose metabolic activity was less suppressed in the CIRP mice. Group and condition interaction analysis revealed differing responses in relative glucose metabolism (decrease in WT mice but increase in CIRP mice) after alcohol in brain regions including the hippocampus and the cortical amygdala where the percent changes in metabolic activity correlated with changes in object discrimination performance. Behaviorally, alcohol-treated WT mice were impaired in exploring a repositioned object in the OPM task, and were more anxious in the OF task, whereas CIRP mice were not impaired in these tasks. CONCLUSION:CIRP released from brain cells could be responsible for regional brain metabolic hypoactivity leading to cognitive impairment under binge alcohol conditions. 10.1186/s10020-019-0092-3
Cold-inducible RNA-binding protein-derived peptide C23 attenuates inflammation and tissue injury in a murine model of intestinal ischemia-reperfusion. Surgery BACKGROUND:Cold-inducible RNA-binding protein is a novel damage-associated molecular pattern that causes inflammation. C23, a short peptide derived from cold-inducible RNA-binding protein, has been found to have efficacy in blocking cold-inducible RNA-binding protein's activity. We hypothesized that C23 reduces inflammation and tissue injury induced by intestinal ischemia-reperfusion. METHODS:Male C57BL/6 mice were subjected to 60 minutes of intestinal ischemia by clamping the superior mesenteric artery. Immediately after reperfusion, either normal saline (vehicle) or C23 peptide (8 mg/kg body weight) was injected intraperitoneally. Four hours after reperfusion, blood, intestinal, and lung tissues were collected for analysis of inflammatory and tissue injury parameters. RESULTS:Cold-inducible RNA-binding protein levels in the intestinal tissues were significantly increased following intestinal ischemia-reperfusion. Histologic examination of the intestine revealed a significant reduction in injury score in the C23 group by 48% as compared with the vehicles after intestinal ischemia-reperfusion. The serum levels of lactate dehydrogenase and aspartate aminotransferase were increased in animals that underwent vehicle-treated intestinal ischemia-reperfusion, whereas C23-treated animals exhibited significant reductions by 48% and 53%, respectively. The serum and intestinal tissue levels of tumor necrosis factor α were elevated in vehicle-treated intestinal ischemia-reperfusion mice but decreased by 72% and 69%, respectively, in C23-treated mice. Interleukin-6 mRNA levels in the lungs were reduced by 86% in the C23-treated group in comparison to the vehicle-treated group after intestinal ischemia-reperfusion. Expression of macrophage inflammatory protein 2 and level of myeloperoxidase activity in the lungs were dramatically increased after intestinal ischemia-reperfusion and significantly reduced by 91% and 25%, respectively, in the C23-treated group. CONCLUSION:C23 has potential to be developed into a possible therapy for reperfusion injury after mesenteric ischemia and reperfusion. 10.1016/j.surg.2018.06.048
Association between CIRP expression and hypoxic-ischemic brain injury in neonatal rats. Chen Lifang,Tian Qiaohuan,Wang Weihua Experimental and therapeutic medicine The role of cold inducible RNA-binding protein (CIRP) in mediating ischemic brain injury in neonatal rats under chronic hypobaric hypoxia was investigated. The neonatal rat model of chronic hypobaric hypoxia and the cell culture model of SH-SY5Y cells exposed to hypoxia (1% O) were constructed. The expression of CIRP and hypoxia-inducible factor-1α (HIF-1α) was detected after hypoxic exposure, and the apoptosis-related proteins were analyzed via terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) and western blot analysis to detect neuronal apoptosis. Moreover, the effects of CIRP overexpression on HIF-1α and neuronal apoptosis were identified. Chronic hypobaric hypoxia can lead to HIF-1α expression and neuronal apoptosis in the body. CIRP was induced at early exposure (3 d/7 d). However, the CIRP level in the hypoxic group was obviously lower than that in the control group with the prolongation of exposure time (21 d). In addition, the knockdown of HIF-1α significantly reduced the neuronal apoptosis under hypoxic conditions, indicating that HIF-1α may promote apoptosis during exposure. The overexpression of CIRP significantly inhibited the upregulation of HIF-1α during hypoxia and the HIF-1α-mediated neuronal apoptosis. Results of the current study showed that, CIRP is involved in the ischemic brain injury induced by chronic hypoxia through downregulation of HIF-1α expression. 10.3892/etm.2019.7767
CIRP downregulation renders cardiac cells prone to apoptosis in heart failure. Chen Minxiao,Fu Hui,Zhang Jingjing,Huang He,Zhong Peng Biochemical and biophysical research communications OBJECTIVES:Cold-inducible RNA binding protein (CIRP) is a stress protein which is involved in regulating multiple cellular processes. However, its role in pathological heart diseases is still unknown. Our current study was aimed at addressing the response and functional role of CIRP in heart failure. METHODS:CIRP protein level was evaluated in heart samples from patients with heart failure and mice with post-myocardial infarction (post-MI). Cardiac-derived H9C2 cells were utilized to test the effects of CIRP deficiency on cell survival and apoptosis in response to HO treatment. RESULTS:Reduced expression of cardiac CIRP was observed in patients with heart failure, mice with post-MI. In addition, knockdown of CIRP exacerbated cell apoptosis and cell death in response to HO treatment, suggesting a protective role of CIRP in cell apoptosis induced by oxidative stress in the heart. CONCLUSIONS:Our findings suggest that altered expression of CIRP may be involved in the pathogenesis of heart failure and downregulation of CIRP may render cardiac cells prone to apoptosis in heart failure. 10.1016/j.bbrc.2019.05.012
Extracellular CIRP (eCIRP) and inflammation. Aziz Monowar,Brenner Max,Wang Ping Journal of leukocyte biology Cold-inducible RNA-binding protein (CIRP) was discovered 2 decades ago while studying the mechanism of cold stress adaptation in mammals. Since then, the role of intracellular CIRP (iCIRP) as a stress-response protein has been extensively studied. Recently, extracellular CIRP (eCIRP) was discovered to also have an important role, acting as a damage-associated molecular pattern, raising critical implications for the pathobiology of inflammatory diseases. During hemorrhagic shock and sepsis, inflammation triggers the translocation of CIRP from the nucleus to the cytosol and its release to the extracellular space. eCIRP then induces inflammatory responses in macrophages, neutrophils, lymphocytes, and dendritic cells. eCIRP also induces endoplasmic reticulum stress and pyroptosis in endothelial cells by activating the NF-κB and inflammasome pathways, and necroptosis in macrophages via mitochondrial DNA damage. eCIRP works through the TLR4-MD2 receptors. Studies with CIRP mice reveal protection against inflammation, implicating eCIRP to be a novel drug target. Anti-CIRP Ab or CIRP-derived small peptide may have effective therapeutic potentials in sepsis, acute lung injury, and organ ischemia/reperfusion injuries. The current review focuses on the pathobiology of eCIRP by emphasizing on signal transduction machineries, leading to discovering novel therapeutic interventions targeting eCIRP in various inflammatory diseases. 10.1002/JLB.3MIR1118-443R
CIRBP protects H9C2 cells against myocardial ischemia through inhibition of NF-κB pathway. Long T Y,Jing R,Kuang F,Huang L,Qian Z X,Yang T L Brazilian journal of medical and biological research = Revista brasileira de pesquisas medicas e biologicas Myocardial ischemia is a major cause of death and remains a disease with extremely deficient clinical therapies and a major problem worldwide. Cold inducible RNA-binding protein (CIRBP) is reported to be involved in multiple pathological processes, including myocardial ischemia. However, the molecular mechanisms of myocardial ischemia remain elusive. Here, we first overexpressed CIRBP by transfection of pc-CIRBP (pcDNA3.1 containing coding sequenced for CIRBP) and silenced CIRBP by transfection of small interfering RNA targeting CIRBP (siCIRBP). pcDNA3.1 and the negative control of siCIRBP (siNC) were transfected into H9C2 cells to act as controls. We then constructed a cell model of myocardial ischemia through culturing cells in serum-free medium with hypoxia in H9C2 cells. Subsequently, AlamarBlue assay, flow cytometry and western blot analysis were used, respectively, to assess cell viability, reactive oxygen species (ROS) level and apoptosis, and expression levels of IκBα, p65 and Bcl-3. We demonstrated that CIRBP overexpression promoted cell proliferation (P<0.001), inhibited cell apoptosis (P<0.05), reduced ROS level (P<0.001), down-regulated phosphorylated levels of IκBα and p65 (P<0.01 or P<0.001), and up-regulated expression of Bcl-3 (P<0.001) in H9C2 cells with myocardial ischemia. The influence of CIRBP knockdown yielded opposite results. Our study revealed that CIRBP could protect H9C2 cells against myocardial ischemia through inhibition of NF-κB pathway. 10.1590/1414-431X20175861
The Mechanism of CIRP in Regulation of STAT3 Phosphorylation and Bag-1/S Expression Upon UVB Radiation. Photochemistry and photobiology Cold-inducible RNA binding protein (CIRP) is a stress-inducible protein, which could be activated by various cellular stresses, such as hypothermia, hypoxia and UV irradiation. Our previous study indicated that UVB radiation (3 mJ cm ) induces CIRP expression, which promotes keratinocytes growth, survival and eventually transformation via activation of STAT3-Bag-1/S signaling cascade. However, the mechanism(s) of CIRP in regulating p-STAT3 activation and Bag-1/S expression have not been fully elucidated. In this study, we demonstrate that repeated exposure of UVB radiation (3 mJ cm ) or overexpression of CIRP could lead to an elevation of the phosphorylation of Janus kinase (JAK) family proteins (JAK2 and JAK3) in HaCaT cells. The increased phosphorylation of the JAKs correlates to an increased phosphorylation of STAT3 (p-STAT3) in the cells; inhibiting JAKs using JAK inhibitor I lead to a reduction of STAT3 phosphorylation and Bag-1/S expression in wild type HaCaT and CIRP stably transfected HaCaT cells with or without UVB exposure. Furthermore, our data indicated that inhibiting the downstream factor of CIRP, NF-κB, using BAY 11-7085 could also decrease the p-STAT3. These results lead us to propose that CIRP mediates the activation of STAT3-Bag-1/S signaling cascade via activating the JAKs and NF-κB signaling pathways. 10.1111/php.12981
Exogenous hydrogen sulfide ameliorates high glucose-induced myocardial injury & inflammation via the CIRP-MAPK signaling pathway in H9c2 cardiac cells. Zhao Hong-Lei,Wu Bao-Quan,Luo Ying,Zhang Wen-Ying,Hao Yun-Ling,Liang Jin-Jie,Fang Fang,Liu Wei,Chen Xie-Hui Life sciences AIMS:Hydrogen sulfide (HS) is a novel signaling molecule with potent cytoprotective actions. In this study, we hypothesize that exogenous HS may protect cardiac cells against high glucose (HG)-induced myocardial injury and inflammation with the involvement of the CIRP-MAPK signaling pathway. MAIN METHODS:H9c2 cardiac cells cultured under HG conditions were transfected with siRNA and different inhibitor for detecting the effects of sodium hydrogen sulfide (NaHS) (a HS donor) on cell biological processes. The cardiac cell viability and LDH activity were determined by CCK-8 and LDH kit. ELISA was employed to measure the levels of inflammatory factors, while 2',7'-dichlorofluorescein diacetate (DCFH-DA) to evaluate reactive oxygen species (ROS). Mitochondrial membrane potential (MMP) was identified by rhodamine 123 staining. TUNEL staining and Hoechst 33258 staining were employed to observe cardiac cell apoptosis. Besides, we determined the expression of CIRP-MAPK signaling pathway- and apoptosis-related factors by protein immunoblot analysis. KEY FINDINGS:HG culturing induced toxicity, LDH, higher level of inflammatory factors, ROS, MMP, and apoptosis in cardiac cells, attenuated the viability of cardiac cells, and activated the CIRP-MAPK signaling pathway. Notably, CIRP silencing aggravated the above condition. HS or blockade of the MAPK signaling pathway reversed the above conditions induced by HG. SIGNIFICANCE:The present study provides evidence for the protective effect of exogenous HS on HG-induced myocardial injury and inflammation in H9c2 cardiac cells and suggests that the activation of CIRP-MAPK signaling pathway might be one of the mechanisms underlying the protective effect of HS. 10.1016/j.lfs.2018.05.051
Neuroprotective effects of cold-inducible RNA-binding protein during mild hypothermia on traumatic brain injury. Wang Guan,Zhang Jian-Ning,Guo Jia-Kui,Cai Ying,Sun Hong-Sheng,Dong Kun,Wu Cheng-Gang Neural regeneration research Cold-inducible RNA-binding protein (CIRP), a key regulatory protein, could be facilitated by mild hypothermia in the brain, heart and liver. This study observed the effects of mild hypothermia at 31 ± 0.5°C on traumatic brain injury in rats. Results demonstrated that mild hypothermia suppressed apoptosis in the cortex, hippocampus and hypothalamus, facilitated CIRP mRNA and protein expression in these regions, especially in the hypothalamus. The anti-apoptotic effect of mild hypothermia disappeared after CIRP silencing. There was no correlation between mitogen-activated extracellular signal-regulated kinase activation and CIRP silencing. CIRP silencing inhibited extracellular signal-regulated kinase-1/2 activation. These indicate that CIRP inhibits apoptosis by affecting extracellular signal-regulated kinase-1/2 activation, and exerts a neuroprotective effect during mild hypothermia for traumatic brain injury. 10.4103/1673-5374.182704
Deficiency of cold-inducible ribonucleic acid-binding protein reduces renal injury after ischemia-reperfusion. Cen Cindy,Yang Weng-Lang,Yen Hao-Ting,Nicastro Jeffrey M,Coppa Gene F,Wang Ping Surgery BACKGROUND:Renal ischemia-reperfusion injury, commonly caused by major operation and shock, leads to acute kidney injury and is associated with high morbidity and mortality. Cold-inducible ribonucleic acid-binding protein, a cold shock protein, has recently been identified as a damage-associated molecular pattern. We hypothesized that cold-inducible ribonucleic acid-binding protein exacerbates severity of injury in renal ischemia-reperfusion. METHODS:Renal ischemia was induced in an 8-week-old male C57BL/6 wild-type mice and Cirp(-/-) mice via bilateral clamping of renal pedicles for 30 minutes, followed by reperfusion for 5 or 24 hours and harvest of blood and renal tissue for analysis. Anti-cold-inducible ribonucleic acid-binding protein antibody or non-immunized immunoglobulin G (IgG) was injected intravenously (10 mg/kg body weight) at time of reperfusion. RESULTS:After renal ischemia-reperfusion, Cirp(-/-) mice demonstrated a reduction of blood urea nitrogen and creatinine of 53% and 60%, respectively, compared with wild-type mice. Serum IL-6 levels were reduced significantly: 70% in Cirp(-/-) mice compared with wild-type mice after renal ischemia-reperfusion. Levels of nitrotyrosine, an oxidatively modified protein marker, and cyclooxygenase-2, an inflammatory mediator, also were significantly decreased in the kidneys of the Cirp(-/-) mice compared with wild-type mice after renal ischemia-reperfusion. Renal caspase-3 activity was decreased in Cirp(-/-) mice compared with wild-type mice after renal ischemia-reperfusion, which corresponded to the reduction of apoptotic cells determined by terminal deoxynucleotidyl transferase dUTP nick-end labeling assay. Injection of neutralizing anti-cold-inducible ribonucleic acid-binding protein antibody into wild-type mice led to an 82% reduction in blood urea nitrogen compared with the vehicle after renal ischemia-reperfusion. CONCLUSION:Deficiency of cold-inducible ribonucleic acid-binding protein results in less renal injury after renal ischemia-reperfusion by attenuating inflammation and oxidative stress. Furthermore, blockade of cold-inducible ribonucleic acid-binding protein shows a protective effect, indicating cold-inducible ribonucleic acid-binding protein as a target in the treatment of renal ischemia-reperfusion. 10.1016/j.surg.2016.04.014
RBM3 and CIRP expressions in targeted temperature management treated cardiac arrest patients-A prospective single center study. Rosenthal Lisa-Maria,Leithner Christoph,Tong Giang,Streitberger Kaspar Josche,Krech Jana,Storm Christian,Schmitt Katharina Rose Luise PloS one BACKGROUND:Management of cardiac arrest patients includes active body temperature control and strict prevention of fever to avoid further neurological damage. Cold-shock proteins RNA-binding motif 3 (RBM3) and cold inducible RNA-binding protein (CIRP) expressions are induced in vitro in response to hypothermia and play a key role in hypothermia-induced neuroprotection. OBJECTIVE:To measure gene expressions of RBM3, CIRP, and inflammatory biomarkers in whole blood samples from targeted temperature management (TTM)-treated post-cardiac arrest patients for the potential application as clinical biomarkers for the efficacy of TTM treatment. METHODS:A prospective single center trial with the inclusion of 22 cardiac arrest patients who were treated with TTM (33°C for 24 hours) after ROSC was performed. RBM3, CIRP, interleukin 6 (IL-6), monocyte chemotactic protein 1 (MCP-1), and inducible nitric oxide synthase (iNOS) mRNA expressions were quantified by RT-qPCR. Serum RBM3 protein concentration was quantified using an enzyme-linked immunosorbent assay (ELISA). RESULTS:RBM3 mRNA expression was significantly induced in post-cardiac arrest patients in response to TTM. RBM3 mRNA was increased 2.2-fold compared to before TTM. A similar expression kinetic of 1.4-fold increase was observed for CIRP mRNA, but did not reached significancy. Serum RBM3 protein was not increased in response to TTM. IL-6 and MCP-1 expression peaked after ROSC and then significantly decreased. iNOS expression was significantly increased 24h after return of spontaneous circulation (ROSC) and TTM. CONCLUSIONS:RBM3 is temperature regulated in patients treated with TTM after CA and ROSC. RBM3 is a possible biomarker candidate to ensure the efficacy of TTM treatment in post-cardiac arrest patients and its pharmacological induction could be a potential future intervention strategy that warrants further research. 10.1371/journal.pone.0226005
Cold-inducible RNA-binding protein as a novel target to alleviate blood-brain barrier damage induced by cardiopulmonary bypass. Liu Mingyue,Li Yongnan,Liu Yiwei,Yan Shujie,Liu Gang,Zhang Qiaoni,Ji Bingyang The Journal of thoracic and cardiovascular surgery BACKGROUND:Hypothermia provides neuroprotection and alleviates cerebral injury after cardiopulmonary bypass (CPB). The mechanism of cold-inducible RNA-binding protein (CIRP), which has been reported to be facilitated by hypothermia and act as a critical regulatory protein in the brain, remains unclear in CPB. Hence, the role of CIRP on hypothermia CPB-induced brain injury was investigated in a rat model. METHODS:Cirp rats were generated using the transcription activator-like effector nucleases-based genome editing technique. The animals were randomly allocated to 3 groups (n = 5, each group): sham group, CPB group, and CPB in Cirp group (Cirp group). Three biological replicates received RNA sequencing in the CPB and Cirp groups. The relative protein expression of the hippocampus was detected. The integrity of the blood-brain barrier (BBB) was measured using transmission electron microscopy and immunoglobulin G immunostaining. Glial fibrillary acidic protein in serum was detected. The brain was fixed for histopathological assessment. RESULTS:More differentially expressed genes of BBB leakage were clustered functionally by gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses. Transforming growth factor-β1, matrix metallopeptidase-9, tumor necrosis factor-α, and malondialdehyde in the hippocampus were higher in the Cirp group, whereas the interleukin-4 level was opposite. Furthermore, more serious BBB disruption in the Cirp group was shown using transmission electron microscopy and immunoglobulin G extravasation. Moreover, Cirp showed enhanced tight junction protein degradation and histopathologic injury in the hippocampus (pathological score, surviving hippocampal neurons, and terminal deoxynucleotidyl transferase-mediated 2'-deoxyuridine, 5'-triphosphate nick-end labeling-positive neurons). Therefore, CIRP significantly alleviated neurologic injury. CONCLUSIONS:CIRP exerted important neuroprotective effects by alleviating BBB breakdown, which might be associated with transforming growth factor-β1-matrix metallopeptidase-9 signals in hypothermia CPB. 10.1016/j.jtcvs.2018.08.100
Cold-Inducible Protein RBM3 Protects UV Irradiation-Induced Apoptosis in Neuroblastoma Cells by Affecting p38 and JNK Pathways and Bcl2 Family Proteins. Zhuang Rui-Juan,Ma Jian,Shi Xiang,Ju Fei,Ma Shuang-Ping,Wang Lei,Cheng Bin-Feng,Ma Yan-Wen,Wang Mian,Li Tong,Feng Zhi-Wei,Yang Hai-Jie Journal of molecular neuroscience : MN Induced by hypothermia, cold-inducible protein RBM3 (RNA-binding protein motif 3), has been implicated in neuroprotection against various toxic insults such as hypoxia and ischemia. However, whether mild hypothermia and RBM3 prevent neural cells from UV irradiation-elicited apoptosis is unclear. In the present study, human neuroblastoma cell line SH-SY5Y was used as a cell model for neural cell death, and it was demonstrated that mild hypothermia protects SH-SY5Y cells from UV irradiation-induced apoptosis. However, the protective effect of mild hypothermia was abrogated when RBM3 was silenced. Conversely, the overexpression of RBM3 rescued SH-SY5Y cells from UV-induced apoptosis, as indicated by the decreased levels of cleaved caspase-3 and PARP, and increased cell survival. The analysis on the mechanism underlying RBM3-mediated neuroprotection against UV insult showed that RBM3 could substantially block the activation of p38 and JNK signaling pathways. In addition, the overexpression of RBM3 reduced the expression of pro-apoptotic proteins Bax and Bad, leaving the pro-survival protein Bcl-2 unaffected. In conclusion, RBM3 is the key mediator of mild hypothermia-related protection against UV in neuroblastoma cells, and the neuroprotective effect might be exerted through interfering with pro-apoptotic signaling pathways p38 and JNK and regulating pro-apoptotic proteins Bax and Bad. 10.1007/s12031-017-0964-3
A novel target to reduce microglial inflammation and neuronal damage after deep hypothermic circulatory arrest. Liu Mingyue,Li Yongnan,Gao Sizhe,Yan Shujie,Zhang Qiaoni,Liu Gang,Ji Bingyang The Journal of thoracic and cardiovascular surgery BACKGROUND:Neuroinflammation acts as a contributor to neurologic deficits after deep hypothermic circulatory arrest. However, the molecular mechanism remains unclear. This study postulates that cold-inducible RNA-binding protein can promote deep hypothermic circulatory arrest-induced neuroinflammation. METHODS:Rats were randomly assigned into 3 groups (n = 5, each group): sham group, deep hypothermic circulatory arrest group, and deep hypothermic circulatory arrest + Cirp group (Cirp group). Murine microglial BV2 cells were administered by adeno-associated viral vectors containing cold-inducible RNA-binding protein small interference RNA or negative control small interference RNA at 2 days before 4-hour oxygen-glucose deprivation at 18°C. Microglial activation, cell death, neuroinflammation, and related protein expression were assessed in tissue samples and cell cultures. RESULTS:Cold-inducible RNA-binding protein was elevated along with evident neuroinflammation and neuronal damage in rats exposed to deep hypothermic circulatory arrest. In Cirp rats, histologic injury (3.00 [interquartile range, 2.00-3.00] vs 1.00 [interquartile range, 1.00-1.50] neuropathological score, P < .001) and microglial activation (40 ± 4 vs 13 ± 7 CA1 area, P < .001) were alleviated after deep hypothermic circulatory arrest. With RNA-sequencing analysis, this associated with reduction of key proinflammatory cytokines induced by inhibiting Brd2-NF-κB signals. In BV2 cells treated with small interference RNA-cold-inducible RNA-binding protein, similar protective effects were observed, including decreased proinflammatory cytokines and cytotoxicity. Brd2-NF-κB signals were confirmed by the addition of Brd2 inhibitor JQ1. Notably, the conditioned medium from BV2 cells transfected with small interference RNA cold-inducible RNA-binding protein significantly reduced apoptosis in neural SH-SY5Y cells after oxygen-glucose deprivation, which was similar to that after JQ1 administration. CONCLUSIONS:Enhanced cold-inducible RNA-binding protein in microglia aggravates neuronal injury by promoting the release of proinflammatory cytokines, which might be mediated through Brd2-NF-κB signals during deep hypothermic circulatory arrest. 10.1016/j.jtcvs.2019.06.115
Cold-inducible protein RBM3 mediates hypothermic neuroprotection against neurotoxin rotenone via inhibition on MAPK signalling. Journal of cellular and molecular medicine Mild hypothermia and its key product, cold-inducible protein RBM3, possess robust neuroprotective effects against various neurotoxins. However, we previously showed that mild hypothermia fails to attenuate the neurotoxicity from MPP , one of typical neurotoxins related to the increasing risk of Parkinson disease (PD). To better understand the role of mild hypothermia and RBM3 in PD progression, another known PD-related neurotoxin, rotenone (ROT) was utilized in this study. Using immunoblotting, cell viability assays and TUNEL staining, we revealed that mild hypothermia (32°C) significantly reduced the apoptosis induced by ROT in human neuroblastoma SH-SY5Y cells, when compared to normothermia (37°C). Meanwhile, the overexpression of RBM3 in SH-SY5Y cells mimicked the neuroprotective effects of mild hypothermia on ROT-induced cytotoxicity. Upon ROT stimulation, MAPK signalling like p38, JNK and ERK, and AMPK and GSK-3β signalling were activated. When RBM3 was overexpressed, only the activation of p38, JNK and ERK signalling was inhibited, leaving AMPK and GSK-3β signalling unaffected. Similarly, mild hypothermia also inhibited the activation of MAPKs induced by ROT. Lastly, it was demonstrated that the MAPK (especially p38 and ERK) inhibition by their individual inhibitors significantly decreased the neurotoxicity of ROT in SH-SY5Y cells. In conclusion, these data demonstrate that RBM3 mediates mild hypothermia-related neuroprotection against ROT by inhibiting the MAPK signalling of p38, JNK and ERK. 10.1111/jcmm.14588
Recent progress in the research of cold-inducible RNA-binding protein. Zhong Peng,Huang He Future science OA Cold-inducible RNA-binding protein (CIRP) is a cold-shock protein which can be induced after exposure to a moderate cold-shock in different species ranging from amphibians to humans. Expression of CIRP can also be regulated by hypoxia, UV radiation, glucose deprivation, heat stress and HO, suggesting that CIRP is a general stress-response protein. In response to stress, CIRP can migrate from the nucleus to the cytoplasm and regulate mRNA stability through its binding site on the 3'-UTR of its targeted mRNAs. Through the regulation of its targets, CIRP has been implicated in multiple cellular process such as cell proliferation, cell survival, circadian modulation, telomere maintenance and tumor formation and progression. In addition, CIRP can also exert its functions by directly interacting with intracellular signaling proteins. Moreover, CIRP can be secreted out of cells. Extracellular CIRP functions as a damage-associated molecular pattern to promote inflammatory responses and plays an important role in both acute and chronic inflammatory diseases. Here, we summarize novel findings of CIRP investigation and hope to provide insights into the role of CIRP in cell biology and diseases. 10.4155/fsoa-2017-0077
Involvement of Cold Inducible RNA-Binding Protein in Severe Hypoxia-Induced Growth Arrest of Neural Stem Cells In Vitro. Zhang Qian,Wang Ya-Zhou,Zhang Wenbin,Chen Xiaoming,Wang Jiye,Chen Jingyuan,Luo Wenjing Molecular neurobiology Neonatal hypoxia is the leading cause of brain damage with birth complications. Many studies have reported proliferation-promoting effect of mild hypoxia on neural stem cells (NSCs). However, how severe hypoxia influences the behavior of NSCs has been poorly explored. In the present study, we investigated the effects of 5, 3, and 1 % oxygen exposure on NSCs in vitro. MTT, neurosphere assay, and 5-ethynyl-2'-deoxyuridine (EdU) incorporation revealed a quick growth arrest of C17.2 cells and primary NSCs induced by 1 % oxygen exposure. Cell cycle analysis showed that this hypoxia exposure caused a significant increase of cells in G0/G1 phase and decrease of cells in S phase that is associated with decrease of Cyclin D1. Interestingly, the expression of cold inducible RNA-binding protein (CIRBP), a cold responsive gene reacting to multiple cellular stresses, was decreased in parallel with the 1 % oxygen-induced proliferation inhibition. Forced expression of CIRBP under hypoxia could restore the proliferation of NSCs, as showed by EdU incorporation and cell cycle analysis. Furthermore, the expression of Cyclin D1 under hypoxia was also restored by CIRBP overexpression. Taken together, these data suggested a growth-suppressing effect of severe hypoxia on NSCs and, for the first time, revealed a novel role of CIRBP in hypoxia-induced cell cycle arrest, suggesting that modulating CIRBP may be utilized for preventing hypoxia-induced neonatal brain injury. 10.1007/s12035-016-9761-1
Cold-inducible RNA-binding protein maintains intestinal barrier during deep hypothermic circulatory arrest. Li Yongnan,Liu Mingyue,Gao Sizhe,Cai Liming,Zhang Qiaoni,Yan Shujie,Liu Gang,Ji Bingyang Interactive cardiovascular and thoracic surgery OBJECTIVES:The intestinal injury during deep hypothermic circulatory arrest (DHCA) is harmful to clinical outcomes. Cold-inducible RNA-binding protein (CIRBP) plays a protective role in hypothermia. The aim of this study was to explore the effects of CIRBP on intestinal barrier during DHCA. METHODS:Sprague-Dawley (wild type, n = 13) and knockout of Cirbp (Cirbp-/-, n = 8) rats were used in the model of DHCA. The histomorphology of the epithelial barrier was evaluated by haematoxylin-eosin, Chiu's scores, Gram's stain and Ki67. The function of the intestinal barrier was evaluated by serum intestinal fatty acid-binding protein, diamine oxidase and d-lactate. The structure of the epithelial barrier, phosphocreatine-creatine kinase system and adenosine triphosphate were assessed in the intestine. RESULTS:The expression of CIRBP significantly increased in the intestine during DHCA. Cirbp-/- rats showed obvious destruction of intestinal barrier after DHCA. Chiu's scores, intestinal fatty acid-binding protein, diamine oxidase and d-lactate significantly increased in the Cirbp-/- group. Ki67 showed that cell proliferation decreased in the Cirbp-/- rats. In the Cirbp-/- group, zonula occludens-1, E-cadherin and occludin levels were significantly decreased, and these proteins either disappeared or redistributed in the monolayer. Besides, Cirbp-/- resulted in decreased levels of creatine kinase B, glycine amidinotransferase, adenosine triphosphate and creatine contents in the intestine, affecting energy metabolism and balance, which is associated with the maintenance of epithelial barrier during acute injury. CONCLUSIONS:CIRBP is related to the maintenance of the intestinal epithelial barrier during DHCA, which is expected to be a new target for the prevention of intestinal injury. 10.1093/icvts/ivz147