牵正散合大补阴丸对蛋白酶抑制因子Ⅰ诱导的帕金森病体外细胞模型大鼠肾上腺嗜铬瘤细胞的增殖和凋亡的影响

The effect of Qianzheng San and Dabuyin Wan on the proliferation and apoptosis of pheochromocytoma cells in a vitro model of Parkinson’s disease induced by proteasome inhibitor Ⅰ

目的观察牵正散合大补阴丸水煎剂对蛋白酶抑制因子I(proteasome inhibitor I,PSI)诱导的帕金森病体外细胞模型大鼠肾上腺嗜铬瘤细胞(pheochromocy-toma cells,PC12)增殖、凋亡的影响,并探讨其作用机制。方法共设立9组,分别是模型组、模型加药组7组(PSI+Q0组,PSI+Q25组,PSI+D0组,PSI+D25组,P+D25+Q25组,P+D25+Q0组,P+Q25+D0组)以及空白对照组。流式细胞术检测牵正散合大补阴丸水煎剂对PC12细胞模型中细胞凋亡的影响;NBT核黄素比色法检测不同浓度药物处理PC12细胞模型后细胞上清液中总超氧化物歧化酶(superoxide dismutase,SOD)含量;蛋白免疫印迹法检测PC12细胞模型中泛素/蛋白酶体途径中相关因子TBP1-19、PA28α的表达水平。结果(1)与空白对照组相比,模型组与模型加药组细胞坏死、凋亡呈升高趋势;与模型组相比,P+Q25+D0组及P+Q0组、P+Q25组细胞坏死、凋亡趋势下降。(2)与空白对照组相比,模型组的SOD值降低(P<0.05);与模型组相比,各模型加药组SOD值升高(P<0.05)。(3)与空白对照组相比,模型组TBP1-19、PA28α蛋白的表达降低;与模型组相比,各模型加药组TBP1-19、PA28α蛋白的表达水平升高。结论牵正散、大补阴丸水煎剂及其合方均可改善PSI诱导的PC12细胞的细胞坏死及凋亡情况,可能是通过影响泛素/蛋白酶体途径中相关蛋白TBP1-19和PA28α的表达水平来改善PSI诱导的PC12细胞模型的凋亡和SOD活性,且不同浓度的合方对PSI诱导的PC12细胞模型保护程度不同。

Objective To observe the effects of Qianzheng San(QZS)and Dabuyin Wan(DBYW)on proliferation and apoptosis of PC12 cells induced by PSI in vitro model of Parkinson’s disease,and to explore its mechanism.Method 9 groups were set totally included PSI model group,7 model drug groups(PSI+Q0 group,PSI+Q25 group,PSI+D0 group,PSI+D25 group,P+D25+Q25 group,P+D25+Q0 group,P+Q25+D0 group)and control group.Flow cytometry was used to detect the apoptosis in PSI-induced PC12 cell model treated after QZS and DBYW.Cell liquid supernatant SOD activity was detected by NBT riboflavin colorimetry and the expressions of TBP1-19 and PA28α,ubiquitin-proteasome pathway related proteins were detected by Western bolt in PSI-induced PC12 cell model.Result Apoptosis detection results showed that compared with control group,the cell necrosis and apoptosis increased in model group and model drug groups.Compared with model group,necrosis and apoptosis decreased in P+Q25+D0 group,P+Q0 group and P+Q25 group.The results of SOD analysis indicated that the SOD activity of model group was lower than that of control group(P<0.05).Compared with model group,SOD value in model drug groups increased(P<0.05).Western blot results showed that the expressions of TBP1-19 and PA28αof model groups were both lower than control group.Compared with model group,the expression levels of TBP1-19 and PA28αprotein in model drug group increased.Conclusion Qianzheng San and Dabuyin Wan could improve necrosis and apoptosis in PSI-induced PC12 cells and ameliorate SOD activity may be by increasing the expressions of TBP1-19 and PA28αin ubiquitin-proteasome pathway.What’s more,different concentrations of Qianzheng Sa n and Dabuyin Wan played protection role for PSI-induced PC12 cell model in different degree.