睾酮在自由基损伤的海马神经元保护作用中的机制

Mechanism underlying protective role of testosterone on primary cultured neuron damaged by free radical

用荧光染色和免疫印迹方法观察睾酮发挥神经保护作用中是否有抗神经元凋亡机制的参与。方法：大鼠原代培养10d海马神经元，按实验分为对照组、H202处理组、预先加入睾酮后再暴露于HzO2组。Hoechst 33258 显色，荧光显微镜下观察拍照。原代培养海马神经元Bcl-2和Bax蛋白进行免疫分析。结果：原代培养海马神经元，Hoechst 33258 显色后神经元呈均匀蓝色光，胞核明显。在H。02组（100脚）可见凋亡的细胞，胞核呈深染半月形或碎块状荧光，突起不明显。预先加入睾酮组再暴露于H2O2，海马神经元可见细胞核染色明显，胞核清楚，无明显核碎片。免疫印迹法显示对照组海马神经元有极少量Bcl-2与Bax表达。加入H：（]2后细胞Bax表达增加，而Bcl-2表达下降，与对照组相比，差异有统计学意义。提前加入睾酮后暴露于100um H2O2，可见Bcl-2表达增加，Bax表达下降，与HzO2组比较，差异有统计学意义。结论：自由基对原代培养海马神经元的损伤机制中，有细胞凋亡机制参与，预先给予睾酮后抗凋亡蛋白增加，而凋亡相关蛋白表达减少。

To identify whether anti-apoptosis effect participate in the neuroprotection of testosterone. Methods： The hippocampal neurons of rats were primary cultured for 10 days, and then divided into a control group, HzO2 group and testosterone preconditioned followed by expose to Hz 02. The morphology of neurons were observed using fluorescence mi- croscopy, and Bcl-2 and Bax expression in cultured hippocampal neurons were detected by Western blotting. Results： The apoptotic cells were found in H202 （100 /~m） group exhibited as semilunar or broken nucleus with deep dyeing, and the processes were not clear. And, cultured neurons preconditioned with testosterone were shown with clear nucleus and nuclear fragments were hardly observed. Compared with the control neurons that shown the weak expression of Bcl-2 and Bax, H2 02 led to a significant up-regulation in the expression of Bax and down-regulation in Bcl- 2 expression. The testosterone precondition statistically suppressed the overexpressed Bax and significantly recovered the down-regulation in Bcl-2 expres- sion caused by H2 02. Conclusion： These findings suggest that neuroproteetion of testosterone partially attribute to its capa- bility to protect the free radical iniured neurons against anontosis