亚低温对颅脑创伤后RIPK1诱导神经细胞坏死性凋亡的影响及其分子机制

Brain mild hypothermia treatment (HMT) has shown promise as therapy for neuronal protection in animal models and in patients with traumatic brain injury (TBI), however, the possible mechanisms of HMT for TBI is not conclusive. It has been suggested that HMT may modify a wide range of neuronal cell necrosis mechanism. Evidence now reveals that receptor-interacting protein kinase-1 (RIPK1), which is an essential regulator, induced the initiation of programmed necrosis, "necroptosis", after experimental traumatic brain injury. Previous studies from our laboratories have demonstrated that the expression of RIPK1 increased dramatically following by TBI, but degraded obviously after mild hypothermia treatment. HMT may act as neuroprotection by inhibiting the expression of RIPK1 and delayed the pathological process of necroptosis, which may be a new mechanism of treating TBI with MHT. To confirm this hypothesis, the fluid percussion injury (FPI) cell and animal models were established. The expressive changes and functional characteristics of RIPK1 in both uninjured, injured, and mild hypothermia tissues of each group were measured using molecular cloning, western blot, and flow cytometry techniques respectively, to investigate the effect of mild hypothermia treatment on the RIPK1-induced necroptosis and molecular mechanism of its signal transduction pathway. Necroptosis participates in the pathogenesis of traumatic brain injury, thereby representing an attractive target of the avoidance of unwarranted cell death.

亚低温治疗颅脑创伤（TBI）可增强神经保护作用的确切机制尚未阐明，多认为与减少神经细胞坏死有关。前期实验提示受体相互作用蛋白激酶-1（RIPK1）作为坏死性凋亡途径中的关键因子，在TBI后表达增加并参与细胞坏死的发生，而亚低温干预后其表达降至正常水平，据此推测亚低温可延缓神经细胞坏死性凋亡，可能是治疗TBI的一种新机制。为证实这一假说，本项目开展亚低温减少以RIPK1诱导的神经细胞坏死性凋亡的分子机制研究，拟建立液压冲击损伤的细胞及动物模型，从整体-组织-细胞三个层次，验证亚低温处理后RIPK1的表达变化及功能特性，探讨亚低温干预前后RIPK1表达对TBI引发的神经细胞坏死的影响及其信号转导通路的分子机制，为从新的视角阐明TBI发生的分子机制提供思路，为临床TBI的亚低温联合药物治疗奠定基础。

亚低温治疗(MHT)颅脑创伤(TBI)可增强神经保护作用的确切机制尚未阐明，多认为与减少神经细胞坏死有关。本项目针对亚低温增强TBI后神经保护作用的确切机制问题，开展亚低温减少以RIPK1 诱导的神经细胞坏死性凋亡的分子机制研究。预实验提示RIPK1 蛋白作为坏死性凋亡途径中的关键因子，在TBI 后表达增加并参与细胞坏死性凋亡的发生，而亚低温处理后其表达降低至正常水平，据此推测亚低温可延缓神经细胞坏死性凋亡，可能是亚低温治疗TBI 的一种新机制。方法：通过液压冲击损伤建立动物模型，通过细胞损伤控制仪建立细胞模型，采用分子克隆、免疫印迹、流式细胞仪等技术手段，从整体-组织-细胞三个层次，验证亚低温处理后RIPK1 的表达变化及功能特性，探讨亚低温处理前后RIPK1 表达对TBI 引起的神经细胞坏死的影响及其信号传导通路的分子机制，并拓展性的探索了低温对TBI后Aβ表达的影响。结果：成功建立了细胞和动物损伤模型，且损伤后RIPK1及相关的蛋白表达发生明显变化，细胞出现坏死性凋亡；低温治疗不仅可降低TBI后RIPK1及坏死性凋亡相关过程，还可降低TBI后海马Aβ的表达。. 本研究探索了TBI后的坏死性凋亡以及低温对坏死性凋亡的治疗效果，探索了低温对TBI后Aβ表达的影响，从多个角度阐明了低温对TBI后的脑保护作用。介于坏死性凋亡与炎症的关系以及Aβ对阿尔兹海默症的致病作用，本研究为TBI后炎症和阿尔兹海默症机制以及低温的治疗机制提供了理论基础。