二甲基甲酰胺致心肌细胞凋亡线粒体通路的体外实验研究

In vitro study of mitochondrial pathway involved cardiomyocytes apoptosis induced by N,N-dimethylformamide

目的探讨二甲基甲酰胺(dimethylformamide,DMF)致H9c2心肌细胞凋亡线粒体通路相关蛋白的影响。方法采用蛋白免疫印迹法(western blot,WB)检测不同剂量DMF(0 mM、50 mM、100 mM、200 mM)染毒24h后胞浆内活化的Caspase-3(cleaved caspase-3)、Bax、Bcl-2以及细胞色素C(Cyt-c)水平的改变。结果不同剂量DMF(0 mM,50 mM,100 mM,200 mM)染毒24 h后胞浆Cleaved caspase-3、Bax和Cyt-c水平升高,Bcl-2水平降低,组间差异均有统计学意义(Cleaved caspase-3:F=23.33,P<0.001;Bax:F=60.31,P<0.001;Bcl-2:F=577.8,P<0.001;Cyt-c:F=104.0,P<0.001)。Cleaved caspase-3和Cyt-c水平在100 mM、200 m M染毒组较对照组升高,差异均有统计学意义(均有P<0.05)。Bax在50 mM、100 mM染毒组较对照组升高,差异均有统计学意义(均有P<0.05),200 mM染毒组与对照组相比差异无统计学意义(P=1.000)。Bcl-2水平在50 mM、100 mM、200 mM染毒组较对照组均降低,差异均有统计学意义(均有P<0.05)。Bax/Bcl-2比值增加,100 mM,200 mM组较对照组差异均有统计学意义(均有P<0.05)。结论 DMF染毒后可上调H9c2心肌细胞Cleaved caspase-3、Cyt-c以及Bax/Bcl-2水平,下调Bcl-2水平。线粒体通路参与了DMF诱导的H9c2心肌细胞凋亡的过程。

Objective To investigate Dimethylformamide（ DMF） on apoptosis-related proteins about mitochondrial pathway in cardiomyocytes. Methods Western Blotting was used to detect the the levels of Cleaved caspase-3,Bax,Bcl-2 and Cytochrome C（ Cyt-c） in cytoplasm after different doses of DMF（ 0 mmol / L,50 mmol / L,100 mmol / L,200 mmol / L） exposure for 24 h. Results After different doses of DMF（ 0 mM,50 mM,100 mM,200 mM） exposure for24 h,the levels of Cleaved caspase-3,Bax and Cyt-c increased,while the level of Bax decreased. The differences between groups were statistically different（ Cleaved caspase-3： F = 23. 33,P〈0.001; Bax： F = 60. 31,P〈0.001; Bcl-2： F =577. 8,P〈0.001; Cyt-c： F = 104. 0,P〈0.001）. Cleaved caspase-3 and Cyt-c increased after 100 m M and 200 m M exposure compared with control group,the differences were significant（ all P〈0.05）. The levels of Bax increased after 50 m M and 100 m M DMF exposure compared with control group（ all P〈0.05）,while the difference between 200 mM DMF exposure and control group were not statistically significant（ P = 1. 000）. The level of Bcl-2 exposed under 50 mM,100 mM and 200 mM DMF decreased compared with the control group,the differences were statistically significant（ all P〈0.05）. The ratio of Bax / Bcl-2 in 100 mM and 200 mM exposure group increased compared with control group,the differences were significant（ all P〈0.05）. Conclusions DMF up-regulated the level of Cleaved caspase-3,Cyt-c and Bax /Bcl-2,meanwhile down-regulated the level of Bcl-2. Mitochondrial pathway was involved in H9c2 cardiomyocytes apoptosis induced by DMF.