TMEM97蛋白在蛛网膜下腔出血早期脑损伤中抑制线粒体碎片化的作用机制研究

Subarachnoid hemorrhage (SAH) is a severe neurological disorder which still lacks effective treatment. Important pathological features of SAH include apoptosis, autophagy, inflammation and BBB destruction. Recently, the study of cellular sub-organelles after SAH has got more attention. Our previous study has demonstrated that excessive mitochondrial fragmentation could lead to apoptosis and oxidative stress after SAH. However, the pathway involved in mitochondrial fragmentation is still not clear. As indicated by one of our most recent findings, TMEM97 protein may play a role in regulating mitochondrial dynamic and fragmentation. Thus, we hypothesize that activation of TMEM97 could alleviate early brain injury after SAH via promoting mitophagy and inhibiting excessive mitochondrial fission. Based on our previous study and some preliminary data, we plan to establish endovascular puncture SAH rat models and use hemin-stimulated primary neuron cultures as SAH in vitro model, then use pharmacological agonists as well as adenovirus to amplify or knockdown the expression of target proteins. Furthermore, we plan to explore the correlation between TMEM97 level in blood or in cerebrospinal fluid and the prognosis of SAH patients in order to identify whether TMEM97 can be a biomarker for the treatment of SAH patients, with hope to find a new strategy for SAH treatment. Methods to be used include ultra-high resolution electron microscope, electron microscopy, Duolink®PLA®, etc.

蛛网膜下腔出血（SAH）是常见神经急危重症，缺乏有效内科治疗。其致病机制复杂，在传统凋亡、自噬、炎症、BBB破坏等机制的研究基础上，SAH中亚细胞器的变化逐渐成为研究热点。我们前期研究证实SAH后神经元细胞存在线粒体过度碎片化可诱发凋亡和氧化应激，但上游调控因素尚不清楚。本项目预实验发现TMEM97 蛋白可能参与线粒体碎片化的调控，并提出假说：SAH后激活TMEM97蛋白能通过抑制线粒体过度分裂和促进线粒体自噬两方面减轻线粒体碎片化及其引起的脑损伤。我们建立SAH颈动脉穿刺模型和Hemin刺激神经元SAH体外模型，以药物激动和基因干预手段，应用超高分辨电子显微镜、透射电镜、Duolink® PLA®技术等方法验证假说；同时结合临床资料探讨SAH患者脑脊液及血清中TMEM97表达水平与患者预后之间的关系，分析其作为SAH后脑损伤预后判断及治疗效果标志物的可行性，为今后SAH的治疗提供新策略。

蛛网膜下腔出血（SAH）是常见神经急危重症，缺乏有效内科治疗。其致病机制复杂，在传统凋亡、自噬、炎症、BBB破坏等机制的研究基础上，SAH中亚细胞器的变化逐渐成为研究热点。我们前期研究证实SAH后神经元细胞存在线粒体过度碎片化可诱发凋亡和氧化应激，但上游调控因素尚不清楚。本项目预实验发现TMEM97 蛋白可能参与线粒体碎片化的调控，并提出假说：SAH后激活TMEM97蛋白能通过抑制线粒体过度分裂和促进线粒体自噬两方面减轻线粒体碎片化及其引起的脑损伤。我们建立SAH颈动脉穿刺模型和Hemin刺激神经元SAH体外模型，以药物激动和基因干预手段，应用超高分辨电子显微镜、透射电镜、Duolink® PLA®技术等方法验证假说；本项目研究成果证明以下几点：1.细胞氧化应激反应和凋亡与SAH大鼠早期脑损伤密切相关；2.TMEM97/NPC1信号通路在SAH大鼠早期脑损伤过程中发挥重要作用，通过对此通路的调节可以减轻SAH大鼠神经元氧化应激反应和凋亡；3.Siramesine能够通过TMEM97/NPC1信号通路减轻SAH大鼠氧化应激反应和细胞凋亡。本研究证明了SAH患者脑脊液及血清中TMEM97表达水平与患者预后之间的关系，分析其作为SAH后脑损伤预后判断及治疗效果标志物的可行性，为今后SAH的为防治早期脑损伤及后续治疗提供了新的理论依据和治疗靶点。