树脑缺血后海马线粒体应激与神经元损伤机制研究

目的研究光化学诱导树脑缺血后不同时间海马神经元细胞色素C(CytC)表达及caspase mRNA含量的改变;探讨脑缺血时神经元线粒体应激导致海马继发性损伤的分子机制。方法免疫组化法检测树缺血后不同时间缺血侧海马神经元CytC蛋白表达;低温差速离心分离海马脑组织线粒体和细胞质部分,western blot法检测其CytC的含量变化;实时荧光PCR检测海马组织caspase-3及caspase-9 mRNA。结果光化学诱导树脑缺血后,海马神经元CytC于24h时由线粒体释放入胞质,而caspase-3、caspase-9 mRNA显著升高,caspase-3与caspase-9之间具有相关性。结论光化学诱导树脑缺血后,海马神经元线粒体应激,促凋亡蛋白CytC从线粒体释放入胞质,改变了空间分布,启动caspase级联反应,是脑缺血后海马神经元继发性损伤的病理生理机制之一。

The mechanisms of hippocampal mitochondrial stress and neuronal injury after thrombotic cerebral ischemia in Tree Shrews

Objective To explore the changes in expression of cytochrome C protein of hippocampal neurons and the contents of caspase mRNA after thrombotic cerebral ischemia,we will provide a molecular mechanism of mitochondrial stress resulting in the hippocampal neurons secondary injury.Methods The expression of cytochrome C was observed in ischemic lateral hippocampi at different times by immunochemistry.Also,the hippocampus was removed,then mitochondria and cytoplasmic fragment were divided by low temperature centrifugation and the distribution of cytochrome C was assessed through western blot.We used real time fluorescence polymerase chain reaction to evaluate the contents of caspase-3 and caspase-9 mRNA.Results Cytochrome C was released from mitochondria into the cytosol at 24h.The contents of caspase-3 and caspase-9 mRNA increased.There was a positive correlation between caspase-3 and caspase-9.Conclusion After photochemical inducing cerebral ischemia in tree shrews,hipppocampal neuronal mitochondrias begin to stress and proapoptotic protein cytochrome C released from mitochondria into cytoplasm,which trigger caspase cascade reaction.This is one of the pathophysiological mechanism about hippocampal neuronal secondary injury after cerebral ischemia.