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目的:研究p38和p42/p44 Ca~(2+).钙调蛋白依赖性蛋白激酶(CCDPK)信号通路对过氧化氢(H_2O_2)诱导牛主动脉内皮细胞(BAEC)凋亡的调节作用.方法:H_2O_2处理BAEC 24 h后,荧光显微镜下观察形态学变化及凋亡细胞计数.MTT法测定细胞活性,琼脂糖凝胶电泳分析DNA降解,蛋白质印迹法检测磷酸化p38和p42/p44 CCDPK表达.结果:H_2O_2诱导BAEC产生典型的凋亡细胞形态学变化(核浓染,核碎裂)和DNA断片.H_2O_2(100-500μmol·L~(-1))浓度依赖性刺激磷酸化p42/p44和p38 CCDPK的表达.p42/p44 CCDPK抑制剂U0126显著增强H_2O_2致凋亡作用;然而p38 CCDPK抑制剂SB203580可增强H_2O_2诱导的磷酸化p42/p44 CCDPK的表达,但不影响BAEC的存活.结论:p42/p44 CCDPK对H_2O_2诱导的BAEC凋亡起保护作用,而p38 CCDPK不是介导H_2O_2所致细胞凋亡的主要信号通路.
AIM: To investigate the regulatory effect of p38 and p42 / p44 Ca 2+ on the apoptosis of bovine aortic endothelial cells (BAEC) induced by hydrogen peroxide (H 2 O 2) mediated by CCDPK signal pathway.Methods : H 2 O 2 treatment of BAEC for 24 h, the morphological changes and the number of apoptotic cells were observed under a fluorescence microscope. The cell viability was determined by MTT assay, the DNA degradation was analyzed by agarose gel electrophoresis, and the expression of phosphorylated p38 and p42 / p44 CCDPK was detected by Western blotting. Results: H 2 O 2 induced typical morphological changes of apoptotic cells (nuclear staining, nuclear fragmentation) and DNA fragmentation.H 2 O 2 (100-500 μmol·L -1) stimulated the phosphorylation of p42 / p44 and p38 CCDPK.p42 / p44 CCDPK inhibitor U0126 significantly enhanced the apoptosis induced by H_2O_2; however p38 CCDPK inhibitor SB203580 enhanced the phosphorylation of p42 / p44 CCDPK induced by H_2O_2, but did not affect the survival of BAEC.Conclusion: p42 / p44 CCDPK protects BA apoptosis induced by H 2 O 2, while p38 CCDPK is not the main signal pathway that mediates apoptosis induced by H 2 O 2.