长爪沙鼠脑缺血/再灌注神经损伤模型体系的建立及半胱氨酸蛋白酶抑制剂C的神经保护作用与分子机制

我们前期成功培育了长爪沙鼠脑缺血模型近交系，但有关其脑缺血/再灌注模型的观测时机、脑损伤程度、表型变化及之间的关联尚不清楚，成为推广应用的瓶颈。在培育近交系过程中我们发现半胱氨酸蛋白酶抑制剂C（CysC）与血管形成相关，文献和我们前期工作均提示CysC具有脑缺血神经保护作用；而脑缺血所导致的神经元持续性损伤与自噬和凋亡有关；CysC可通过p53等途径抑制自噬和凋亡；并且外泌体CysC作用明显。据此提出科学假说：CysC和外泌体CysC在脑缺血/再灌注中通过抑制凋亡和自噬缓解神经元持续性损伤。本项目拟利用长爪沙鼠脑缺血模型近交系和CysC敲除沙鼠建立标准化的脑缺血/再灌注模型体系，在此基础上通过体内外实验研究CysC和外泌体CysC通过抑制神经细胞自噬和凋亡缓解神经损伤，发挥对神经细胞保护作用和机制。本研究将为长爪沙鼠脑缺血模型近交系的推广应用提供依据，为寻求脑缺血神经保护新靶点奠定基础。

Previously, we successfully bred an inbred strain of high incidence of cerebral ischemic Mongolian gerbil (CIMG). However, the best observation time-point, severity of neurological injury, symptom score and the relations have not been explored clearly. This problem make it a big challenge for the wide application of this strain. During the breeding process, we found that cystatin C (CysC) is related with angiogenesis. The references and our primary studies implied that CysC exhibited a neurological protective effect in cerebral ischemia. It is also shown that persistent neurological injury resulted from cerebral ischemia was related to apoptosis and autophagy. On the other hand, CysC could inhibit apoptosis and autophagy via p53 pathway. Especially, the form of exosomal CysC indicated better effect than CysC protein in rescue of neurons and primary cortical smooth muscle cells from starvation-induced death. According to these researches, we propose a scientific hypothesis: CysC and exosomal CysC alleviate persistent neuronal damage by inhibiting apoptosis and autophagy during cerebral ischemia/reperfusion. In the present study, we will establish a standardized cerebral ischemia/reperfusion gerbil model system using CIMG and CysC knockout gerbils. Based on these work, we will study the effect and mechanism of CysC protein and exosomal CysC on amending persistent damage of the cerebral nerves by inhibiting the autophagy and apoptosis, consequently exert the protective effect in cerebral ischemia. This study will effectively promote the standardization and extensive use of cerebral ischemia/reperfusion model in gerbils and provide the consolidation of seeking new target for neurological protection in stroke treatment.