代森锰锌对PC-12细胞凋亡的影响

Effects of mancozeb on apoptosis of PC-12 cells

目的探讨代森锰锌对PC-12细胞凋亡的影响及其机制。方法 采用体外细胞培养方法,PC-12细胞加入代森锰锌0,1,10,30,60和120μmol.L-1,培养24h后,应用WST-8法检测PC-12细胞增殖;PC-12细胞中加入代森锰锌0,1,30和120μmol.L-1,培养24h后,流式细胞术FITC-AnnexinⅤ/PI双染检测细胞凋亡率;Hoechst33258染色及倒置荧光显微镜观察细胞形态学改变;Western印迹法检测Bcl-2和Bax的表达以及ERK蛋白磷酸化水平。结果与正常对照组相比,随着代森锰锌浓度增加,代森锰锌组晚期凋亡率升高,呈浓度依赖关系,IC50为49.95μmol.L-1。代森锰锌120μmol.L-1组细胞晚期凋亡率为(90±4)%(P<0.05);Hoechst33258染色可见细胞核膨大、染色质边集浓染等凋亡特征;与正常对照组相比,Bcl-2逐渐降低,Bax和p-ERK1/2表达增高(P<0.05),代森锰锌120μmol.L-1组p-ERK1/2积分吸光度分别为128.0±2.5和178.4±4.0。结论代森锰锌能够诱导PC-12细胞凋亡,ERK信号通路可能在此过程中发挥作用。

OBJECTIVE To investigate the effect of mancozeb on the apoptosis of PC-12 cells.METHODS The PC-12 cells were cultured for 24 h after mancozeb 0,1,10,30,60 and 120 μmol·L-1 were added.Cell Counting Kit-8 was used to assess the proliferation and toxicity induced by mancozeb.Twenty-four hours after mancozeb 0,50 μmol·L-1 was added to PC-12 cells,the morphological changes of PC-12 cells were observed by a microscope,and cell apoptosis rates were detected by FITC-AnnexinⅤ/PI flow cytometry.The expression of Bcl-2,Bax and p-ERK1/2 was determined by Western blot.RESULTS Compared with normal control group,PC-12 cells in mancozeb groups showed higher ap＊optosis rates and evident morphological changes that became more evident with the dose of mancozeb.The IC50 was 49.95 μmol·L-1,the apoptosis rate of PC-12 cells in mancozeb 120 μmol·L-1 group was（90±4）%.The Bax protein levels increased and the Bcl-2 protein levels in mancozeb groups were decreased.Compared with normal control group,the p-ERK1/2 expression was significantly up-regulated.The grey value of p-ERK1/2 in mancozeb 120 μmol·L-1 group was 128.0±2.5 and 178.4±4.0.CONCLUSION Mancozeb can induce apoptosis of PC-12 cells,in which the expression of p-ERK1/ 2 proteins may play a role in PC-12 apoptosis.