大鼠海马神经元缺氧/复氧后caspase-3活性的变化

Changes of caspase-3 activities in rat hippocampal neurons during hypoxia/reoxygenation

目的:研究caspase-3在体外培养的海马神经元缺氧/复氧损伤中的作用及其抑制剂的干预效果,为缺血性脑损伤的治疗提供实验资料。方法:诱导体外培养的大鼠海马神经元缺氧3 h,再复氧12～48 h,并在缺氧前1 h用不同剂量的caspase-3抑制剂Ac-DEVD-CMK进行干预处理,应用MTT比色法测定细胞存活率,底物裂解法检测caspase-3活性。结果:缺氧/复氧后大鼠海马神经元存活率逐渐降低,caspase-3活性明显增强,复氧24 h达高峰;用Ac-DEVD-CMK预处理的海马神经元caspase-3活性降低,细胞存活率增高,并呈剂量依赖性。结论:体外培养的大鼠海马神经元缺氧及缺氧/复氧后出现迟发性死亡,caspase-3介导的细胞凋亡机制在这种损伤过程中起一定的作用,通过抑制其活性可产生神经保护作用。

Objective: To study caspase-3 activities and the neuroprotective effect of caspase-3 inhibitor Ac-DEVD-CMK in primary cultures of rat hippocampal neurons during hypoxia/reoxygenation (H/R). Methods: After pretreated with Ac-DEVD-CMK or the vector DMSO fo 1 h, primary cultures of rat hippocampal neurons were induced to hypoxia for 3 h, followed by 12 to 48 h of reoxygenation. The neuronal viability was estimated by MTT assay, and caspase-3 cellular activities were measured by a colorimetric method using Ac-DEVD-pNA as a substrate. Results: During H/R, the cultured rat hippocampal neuronal viability gradually decreased, and caspase-3 activities in the primary cultures of rat hippocampal neurons were significantly increased, and peaked at 24 h of reoxygenation. Caspase-3 activities were decreased in the neurons pretreated with Ac-DEVD-CMK in a dose dependent manner. Conclusion: Delayed neuronal death is detected in cultured hippocampal neurons during H/R, and apoptosis mediated by caspase-3 may play an important role in it, and caspase-3 inhibitor has a neuroprotective effect on them.