抑制PARP-1表达在棕榈酸诱导大鼠心肌细胞凋亡中的保护作用

Protective effect of inhibiting PARP-1 expression in rat cardiomyocyte apoptosis induced by palmitic acid

目的探讨抑制PARP-1表达缓解棕榈酸诱导的H9C2心肌细胞凋亡效果及其作用机制。方法取H9C2细胞随机分成正常组、PA组和抑制组。PA组接受0.4μmol/L PA干预,抑制组接受20 nmol/L PARP-1抑制剂+0.4μmol/L PA干预,正常组不接受任何干预。干预24 h后测定各组细胞凋亡情况、线粒体电位、ROS水平和凋亡相关蛋白。结果抑制组细胞存活抑制率明显高于PA组(P<0.05)。ROS测定结果显示,抑制组ROS水平低于PA组。线粒体膜电位结果显示,抑制组细胞线粒体跨膜电位高于PA组,而弱于对照组。流式细胞仪检测结果显示,PA组活细胞率明显低于对照组(P<0.01),而细胞凋亡率明显高于对照组(P<0.01),抑制组活细胞率明显高于PA组(P<0.01),而细胞凋亡率明显低于PA组(P<0.01)。抑制组H9C2细胞的PARP-1蛋白表达(25.84±6.47)明显低于对照组(96.94±8.74)%和PA组(89.72±7.55)%(P<0.05),抑制组H9C2细胞的Caspase-3蛋白(44.66±7.72)%和Bax蛋白(48.62±7.58)%表达明显低于PA组(95.44±12.58)%和(88.28±10.25)%(P<0.05),但高于对照组H9C2细胞(23.45±6.25)%和(20.57±6.22)%(P<0.05)。结论抑制PARP-1表达能缓解PA脂毒性引起H9C2心肌细胞凋亡,其作用机制可能为提升心肌细胞线粒体活性,抑制细胞ROS水平和细胞凋亡蛋白表达。

Objective To investigate the effect of PARP-1 on palmitic acid-induced apoptosis of H9 C2 cardiomyocytes and its mechanism.Methods H9 C2 cells were randomly divided into normal group,PA group and inhibition group.PA group received the intervention of 0.4 micromol/L PA,inhibition group received the intervention of 20 nmol/L PARP-1 inhibitor and 0.4 micromol/L PA,normal group did not accept any intervention.Apoptotic status,mitochondrial potential,ROS level and apoptotic related proteins were measured 24 hours after intervention.Results The inhibition rate of cell survival in inhibition group was significantly higher than that in PA group(P<0.05).The result of ROS showed the number of H9 C2 cells with green fluorescence in the inhibition group was more than that in the control group,but less than that in the PA group,suggesting that the ROS level in the inhibition group was lower than that in the PA group.The result of mitochondrial membrane potential showed that H9 C2 cells in the control group were mainly J-aggregates cells,and their mitochondrial membrane potential was higher;H9 C2 cells in the PA group were mainly monomer cells with low mitochondrial membrane potential;the red fluorescence of H9 C2 cells in the inhibition group was stronger than that in the PA group,but weaker than that in the control group,indicating that the mitochondrial transmembrane potential in the inhibition group was higher than that in the PA group and weaker than that in the control group.The result of flow cytometry showed that the living cell rate of PA group was significantly lower than that of control group(P<0.01),while the apoptotic rate of PA group was significantly higher than that of control group(P<0.01),the living cell rate of inhibition group was significantly higher than that of PA group(P<0.01),and the apoptotic rate of PA group was significantly lower than that of PA group(P<0.01).The expression of PARP-1 protein in H9 C2 cells in inhibited group was significantly lower than that in control group(25.84±6.47)%and PA group(89.72±7.55)%(P<0.05).The expressions of Caspase-3 protein and Bax protein in H9 C2 cells in inhibited group(44.66±7.72)%and(48.62±7.58)%were significantly lower than those in PA group(95.44±12.58)%and(88.28±10.25)%(P<0.05),but higher those that in control group(23.45±6.25)%(P<0.05).And(20.57±6.22)%(P<0.05).Conclusion Inhibition of PARP-1 expression can alleviate apoptosis of H9 C2 cardiomyocytes induced by PA lipotoxicity,and its mechanism may be to enhance mitochondrial activity,inhibit ROS level and apoptotic protein expression in cardiomyocytes.