虎杖甙抑制自噬及PI3K-Akt通路对缺氧诱导的心肌细胞损伤的保护作用

Polydatin Protects Against Hypoxia-induced Cardiomyocyte Injury by Inhibiting Cell Autophagy and PI3 K/AKT Pathway

目的 探究虎杖甙(polydatin,PDT)对缺氧心肌细胞损伤的影响及机制.方法 H9C2心肌细胞在1%O2的条件下培养12 h后复氧,建立缺氧心肌细胞模型.将缺氧心肌细胞随机分成4组:缺氧组(Hypoxia)、 虎杖甙低剂量组(PDT 5μmol/L)、 虎杖甙中剂量组(PDT 10μmol/L)、 虎杖甙高剂量组(PDT 20μmol/L);另外设正常对照组(H9C2).CCK8检测0、1、2、3、4 d细胞增殖倍数.相关试剂盒检测细胞中超氧化物歧化酶(superoxide dismutase,SOD)和丙二醛(malondialdehyde,MDA)的含量.赫斯特(Hoechst)染色检测细胞凋亡情况.免疫印迹(Western印迹)检测自噬标记蛋白Beclin 1、P62、LC3Ⅱ/LC3Ⅰ、 磷脂酰肌醇3-羟激酶(PI3K)和蛋白激酶B(AKT)的表达.结果 与对照组相比,Hy-poxia组细胞增殖速率显著降低;氧化应激水平上升;凋亡细胞的比率显著上升(P<0.01);细胞自噬水平显著降低,PI3K通路活性升高.缺氧心肌细胞经虎杖甙处理后,细胞增殖速率显著提高(P<0.01);氧化应激水平受抑;凋亡细胞的比率显著降低(P<0.01);细胞自噬和PI3K通路活性均受到抑制.结论 虎杖甙抑制缺氧心肌细胞自噬和PI3K-AKT通路,对缺氧诱导的心肌细胞损伤具有保护作用.

Objective To explore the effect of polydatin (PDT) on myocardial cell injury induced by hypoxia. Methods The hypoxic myocardial cell model was established by culturing H9 C2 cardiomyocytes in 1 % O2 for 12h and then reoxygenation. The anoxic cardiomyocytes were randomly divided into 4 groups: hypoxia group, low-dose PDT group (5μmol/L), moderate-dose PDT group (10μmol/L), high-dose PDT group (20μmol/L) and normal control group (H9 C2).The proliferation of 0, 1, 2, 3 , 4 d cells was detected by CCK8. The contents of superoxide dismutase (SOD) and malondialdehyde ( MDA) in the cells were detected by using the related kits. Hoechst staining was used to detect cell apoptosis. The expression levels of autophagic marker proteins Beclin 1, P62, LC3 II/LC3 I,< phosphatidyl inositol 3-hydroxykinase (PI3 K) and protein kinase B ( AKT) were detected by Western blotting. Results The proliferation rate of cells was significantly decreased, the level of oxidative stress significantly increased, the ratio of apoptot-ic cells significantly increased, the level of autophagy significantly decreased , and the activity of PI3 K pathway markedly increased in the hypoxia group when compared with those in the control group (P<0.01). After hypoxic cardiomyocytes were treated with polydatin, the cell proliferation rate was significantly increased (P<0.01), the oxidative stress level was profoundly inhibited, the apoptotic cell rate was significantly reduced (P<0.01) and the cell autophagy and PI3 K pathway activity were inhibited. Conclusion Polydatin inhibits autophagy and PI3 K-AKT pathway in hypoxic cardiomyocytes, and it has protective effects on hypoxia-induced cardiomyocyte injury.