6-OHDA对PC12细胞的凋亡及内质网应激的影响

Influence of 6-OHDA in apoptosis and endoplasmic reticuLum stress of PC12 cells

目的使用6-OHDA(6羟基多巴胺)作用于PC12细胞建立帕金森病细胞模型,探讨模型细胞的凋亡与HtrA2及Bip/Grp78表达的关系;方法将不同浓度(10-200μM)6-OHDA加入传代培养的PC12细胞中,作用24 h后采用CCK8法检测细胞活力,筛选细胞存活率为50%左右的6-OHDA浓度为造模浓度,并观察24 h内细胞活力的变化,使用Annexin V-PE/7AAD试剂盒检测细胞凋亡,并使用蛋白免疫印迹法检测HtrA2、Bip/Grp78的表达;结果在6-OHDA诱导下,PC12细胞活力呈剂量依赖性和时间剂量依赖性下降,其浓度为60μM时,细胞存活率为43.89%±0.90%,模型细胞的凋亡率、HtrA2和Bip/Grp78的表达较对照组明显增加(P<0.05);结论选择60μM 6-OHDA为PC12细胞帕金森病细胞模型的造模浓度,6-OHDA刺激PC12细胞的内质网应激,诱导PC12细胞的凋亡,HtrA2参与内质网应激且促进细胞凋亡。

Objective To induce the model cells for Parkinson＇s disease（PD） using 6-OHDA（6 hydroxydopamine） in PC12 cells,and investigate the relationship between apoptosis with expressing of HtrA2 and Bip/Grp78. Methods PC12 cells with different concentrations（10-200 μM） of 6-OHDA, using CCK8 to detect the cell vitality after 24 hours, screened the appropriate concentration of 6-OHDA that made the about 50% cells survive, and observed the change of the cell vitality within 24 hours, detected the cell apoptosis using the Annexin V-PE/7 AAD apoptosis kit, and detected the expression of HtrA2 and Bip/Grp78 proteins with the method of western blot; Results Under the inducing of 6-OHDA, cell vitality of PC12 presented dose and time dependent, when the concentration was 60 u M, cell vitality was 43.89 ±0.90%, the cell＇s apoptosis rate, expression of HtrA2 and Bip/Grp78 were significantly higher than that of the control group（P〈0.05）. Conclusion The research selected 60 uM 6-OHDA as the cell model＇s concentration for PD in PC12 cells, 6-OHDA stimulates the endoplasmic reticulum stress in PC12 cells, it induces apoptosis of PC12 cells, HtrA2 participates the endoplasmic reticulum stress and promoted cell＇s apoptosis.