Background Extracellular cold-inducible RNA-binding protein (eCIRP) plays a vital role in the inflammatory response during cerebral ischaemia. However, the potential role and regulatory mechanism of eCIRP in traumatic brain injury (TBI) remain unclear. Here, we explored the effect of eCIRP on the development of TBI using a neural-specific CIRP knockout (KO) mouse model to determine the contribution of eCIRP to TBI-induced neuronal injury and to discover novel therapeutic targets for TBI. Methods TBI animal models were generated in mice using the fluid percussion injury method. Microglia or neuron lines were subjected to different drug interventions. Histological and functional changes were observed by immunofluorescence and neurobehavioural testing. Apoptosis was examined by a TdT-mediated dUTP nick end labelling assay in vivo or by an annexin-V assay in vitro. Ultrastructural alterations in the cells were examined via electron microscopy. Tissue acetylation alterations were identified by non-labelled quantitative acetylation via proteomics. Protein or mRNA expression in cells and tissues was determined by western blot analysis or real-time quantitative polymerase chain reaction. The levels of inflammatory cytokines and mediators in the serum and supernatants were measured via enzyme-linked immunoassay. Results There were closely positive correlations between eCIRP and inflammatory mediators, and between eCIRP and TBI markers in human and mouse serum. Neural-specific eCIRP KO decreased hemispheric volume loss and neuronal apoptosis and alleviated glial cell activation and neurological function damage after TBI. In contrast, eCIRP treatment resulted in endoplasmic reticulum disruption and ER stress (ERS)-related death of neurons and enhanced inflammatory mediators by glial cells. Mechanistically, we noted that eCIRP-induced neural apoptosis was associated with the activation of the protein kinase RNA-like ER kinase-activating transcription factor 4 (ATF4)-C/EBP homologous protein signalling pathway, and that eCIRP-induced microglial inflammation was associated with histone H3 acetylation and the α7 nicotinic acetylcholine receptor. Conclusions These results suggest that TBI obviously enhances the secretion of eCIRP, thereby resulting in neural damage and inflammation in TBI. eCIRP may be a biomarker of TBI that can mediate the apoptosis of neuronal cells through the ERS apoptotic pathway and regulate the inflammatory response of microglia via histone modification.

中文翻译：

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创伤性脑损伤后细胞外冷诱导RNA结合蛋白介导的神经炎症和神经元凋亡


背景细胞外冷诱导RNA结合蛋白（eCIRP）在脑缺血期间的炎症反应中发挥着至关重要的作用。然而，eCIRP 在创伤性脑损伤（TBI）中的潜在作用和调节机制仍不清楚。在这里，我们使用神经特异性 CIRP 敲除 (KO) 小鼠模型探讨了 eCIRP 对 TBI 发展的影响，以确定 eCIRP 对 TBI 诱导的神经元损伤的贡献，并发现 TBI 的新治疗靶点。方法采用液体冲击损伤法建立小鼠TBI动物模型。小胶质细胞或神经元系接受不同的药物干预。通过免疫荧光和神经行为测试观察组织学和功能变化。通过体内 TdT 介导的 dUTP 缺口末端标记测定或体外膜联蛋白-V 测定来检查细胞凋亡。通过电子显微镜检查细胞超微结构的改变。通过蛋白质组学通过非标记定量乙酰化来鉴定组织乙酰化改变。通过蛋白质印迹分析或实时定量聚合酶链反应测定细胞和组织中的蛋白质或mRNA表达。通过酶联免疫分析测定血清和上清液中炎症细胞因子和介质的水平。结果人和小鼠血清中eCIRP与炎症介质、eCIRP与TBI标志物之间均呈密切正相关。神经特异性 eCIRP KO 减少了 TBI 后的半球体积损失和神经元凋亡，并减轻了胶质细胞活化和神经功能损伤。 相比之下，eCIRP 治疗导致内质网破坏和内质网应激 (ERS) 相关的神经元死亡，并增强神经胶质细胞的炎症介质。从机制上讲，我们注意到 eCIRP 诱导的神经细胞凋亡与蛋白激酶 RNA 样 ER 激酶激活转录因子 4 (ATF4)-C/EBP 同源蛋白信号通路的激活相关，并且 eCIRP 诱导的小胶质细胞炎症与具有组蛋白 H3 乙酰化和 α7 烟碱乙酰胆碱受体。结论 这些结果提示TBI明显增强eCIRP的分泌，从而导致TBI的神经损伤和炎症。 eCIRP可能是TBI的生物标志物，可以通过ERS凋亡途径介导神经元细胞凋亡，并通过组蛋白修饰调节小胶质细胞的炎症反应。