C_1抑制物对过氧化氢诱导培养乳鼠心肌细胞凋亡的影响

Effects of C_1-INH on Apoptosis of Neonate Rats’ Cardiomyocytes Induced by Hydrogen Peroxide

目的:观察C1-INH对H2O2诱导的乳鼠心肌细胞凋亡的影响。方法:采用培养的原代心肌细胞,随机分成3组:正常对照组(N组),无特殊处理;H2O2组(H组),加入终浓度0.3 mmol/L的H2O2;C1-INH+H2O2组(C组),加入终浓度0.15 g/L的C1-INH和终浓度0.3 mmol/L的H2O2。三组干预时间分别设为6,12,24 h。C3Elisa试剂盒检测N组和H组的细胞表面C3的沉积。各组心肌细胞用Hoechst 33258荧光剂染色,在荧光显微镜下观察心肌细胞凋亡形态学,按AnnexinⅤ-FITC试剂盒说明采用流式细胞仪检测细胞存活率和凋亡率。结果:在Hoechst 33258荧光剂染色下,N组心肌细胞凋亡极少,H组有较多心肌细胞发生凋亡,C组也有较多心肌细胞发生凋亡。干预6,12和24 h后,H组心肌细胞C3沉积较N组增多,均有显著性差异(P<0.05)。H组均较N组成活率降低,凋亡率增高,均有显著性差异(P<0.05)。除干预时间6 h外,干预12 h和24 h后,C组细胞存活率较H组高,凋亡率较H组低,均有显著性差异(P<0.05)。结论:H2O2损伤的心肌细胞表面C3的沉积较正常对照组明显增多,C1-INH减轻H2O2诱导的乳鼠心肌细胞凋亡。

Objective： To investigate the influence of C1-INH on apoptosis of neonate rats cardiomyocytes induced by hydrogen peroxide （H2O2 ）. Methods： Cultured cardiomyocytes were divided into three groups as the normal group （without any treatment, N group）, H2O2 group （treated with 0.3 mmol/L H2O2, H group）, and CI-INH＋ H2O2 group （treated with 0.3 mmol/L H2O2 and 0.15 g/L C1-INH, C group）. After 6, 12 and 24 h, cell-surface C3 deposition of N group and H group was detected by Elisa, morphological changes of apoptotic cardiomyocytes were observed, the apoptotic rate was determined by flow cytometry. Results： Apoptotic cardiomyocytes with fluorescein stain by Hoechst 33258 showed visible morphological changes. The apoptotic rate Of H group and C group was obviously higher than that of N group. After 6, 12 and 24 h, Cell-surface C3 deposition of H group was significantly higher than that of N group （P〈0. 05）, survival rate of H group was evidently lower than that of N group （P〈0.05） ,and apoptotic rate of H group was evidently higher than that of N group （P〈0.05）. After 12 and 24 h, survival rate of C group was evidently higher than that of H group（P〈0.05 ） ,and the apoptotic rate of C group was significantly lower than that of H group （P〈0.05）. Conclusion： Cell-surface C3 deposition of H group was evidently higher than that of N group, and C1-INH attenuated apoptosis of neonate rats＇ cardiomyocytes induced by H2O2.