X-染色体连锁的凋亡抑制蛋白对新生小鼠脑缺氧缺血后细胞色素C释放的影响

The influence of X-linked inhibitor of apoptosis protein on cytochrome C release after cerebral hypoxia- ischemia in neonatal mice

目的探讨X-染色体连锁的凋亡抑制蛋白(XIAP)对未成熟脑缺氧缺血(HI)后细胞色素C从线粒体释放的影响。方法新生9日龄转基因XIAP过度表达C57BL/6小鼠(XIAP组)及同期野生型9日龄C57BL/6小鼠(野生组)在HI后不同时间点处死取脑,部分脑组织进行手工匀浆后差速离心分离胞浆和线粒体成分进行Western蛋白印迹,部分脑组织进行细胞色素C免疫组化染色。结果Western蛋白印迹显示,HI后24h缺血侧脑组织线粒体中细胞色素C的含量较对侧减少,而胞浆中细胞色素C的含量较对侧增加(细胞色素C从线粒体释放);XIAP组脑组织线粒体中细胞色素C释放量明显少于野生组。免疫组化结果显示HI后3h、24h,XIAP组大脑皮层和海马CA1区细胞色素C阳性细胞数明显低于野生组。结论XIAP过度表达可抑制HI后细胞色素C从线粒体向胞浆的释放。

Objectives To explore the effect of X-linked inhibitor of apoptosis protein （XIAP） on cytochrome C release from mitochondria to cytosol after hypoxia-ischemia （HI） in neonatal mice. Methods Nine-day old transgenic XIAP overexpressing mice and wide-type C57BL/6 mice were subjected to left carotid artery ligation followed by 10% oxygen for 55 minutes. At different timepoints following HI, the mice were sacrificed. Brain tissues were homogenized. The mitochondria and cytosol fraction were isolated using high speed centrifugation. The cytochrome C was detected by Western blotting. Results The Western blotting assay showed that amount of cytochrome C in mitochondria was less and amount of cytochrome C in cytosol was more in ipsilateral than those in contralateral hemisphere respectively after 24 h HI （cytochrome C released from mitochondria）. Compared with wide type mice, XIAP overexpressing mice released less cytochrome C from mitochondria to cytosol after HI. The immunohistochemical results showed that the number of cytochrome C positive cells in the cortex and CA1 region of hippocampus were fewer in XIAP overexpressing mice than in wide type mice at 3 h and 24 h after HI. Conclusions XIAP overexpression could inhibit the release of cytochrome C from mitochondria to cytosol in neonatal mice brain after hypoxia-ischemia.