滋肾活血方对糖氧剥夺/再灌注诱导的海马神经元线粒体自噬的影响

Effects of Zishen Huoxue Prescription on OGD/R-induced mitophagy in hippocampal neurons

目的探究滋肾活血方对糖氧剥夺/再灌注(oxygen-glucose deprivation/reoxygenation,OGD/R)诱导的原代海马神经元损伤的保护机制。方法分离的原代海马神经元经免疫荧光鉴定后,OGD/R诱导神经元损伤,激光共聚焦扫描显微镜观测不同再灌注时间自噬流的变化。OGD/R诱导的原代海马神经元再经滋肾活血方含药血清干预后,CCK-8检测细胞活力,流式细胞术检测细胞凋亡,透射电子显微镜观察自噬体,Western blot检测自噬相关蛋白表达。结果10%的滋肾活血方含药血清可以明显提升细胞活力及降低细胞凋亡比例,增加神经元中自噬体的数量,并上调自噬相关蛋白PINK1、Parkin、p-ATG16L1的表达。结论滋肾活血方能有效抵抗OGD/R诱导的原代海马神经元凋亡,其作用可能与调控PINK1-Parkin通路促进线粒体自噬有关。

Aim To explore the protective effect of Zishen Huoxue Prescription on OGD/R-induced primary hippocampal neuron damage in rats and the possible mechanism.Methods After the isolated primary hippocampal neurons were identified by immunofluorescence,OGD/R induced neuronal damage,and the changes of autophagic flux at different re-oxygenation time were observed by confocal laser scanning microscopy.After OGD/R-induced primary hippocampal neurons were intervened with serum containing Zishen Huoxue Prescription,cell viability was detected by CCK-8,cell apoptosis was detected by flow cytometry,autophagosomes were detected by transmission electron microscopy,and autophagy-related protein expressions were detected by Western blot.Results 10%Zishen Huoxue Prescription-containing serum could significantly improve cell viability and reduce the proportion of cell apoptosis,increase the number of autophagosomes in neurons,and up-regulate the expression of autophagy-related protein PINK1,Parkin,and p-ATG16L1.Conclusions Zishen Huoxue Prescription can effectively resist OGD/R-induced apoptosis of primary hippocampal neurons in rats,and its effect may be related to the regulation of PINK1-Parkin pathway to promote mitophagy.