吡格列酮对过氧化氢诱导的乳鼠心肌细胞凋亡的抑制作用

目的观察吡格列酮对乳鼠心肌细胞凋亡的抑制作用及其可能作用机制。方法采用培养的新生大鼠乳鼠心肌细胞制备H2O2200μmol·L-1氧化损伤模型。实验分为正常对照组、模型组、吡格列酮组(0.1,1及10μmol·L-1)、吡格列酮(10μmol.L-1)+GW9662(10μmol·L-1)组、维拉帕米1μmol.L-1组。MTT法测心肌细胞的活力;采用硫代巴比妥酸比色法、黄嘌呤氧化酶法分别测定心肌细胞中脂质过氧化产物丙二醛(MDA)含量及超氧化物歧化酶(SOD)活性;流式细胞仪检测心肌细胞的凋亡率;采用Till阳离子测定系统,以Fura-2/AM为荧光探针,观察心肌细胞Ca2+]i瞬间变化。结果H2O2200μmo.lL-1作用于心肌细胞12h能引起心肌细胞凋亡。吡格列酮浓度依赖性地增加受损心肌细胞的活力及SOD活性,降低MDA含量。吡格列酮10μmol·L-1抑制作用最明显,其与维拉帕米1μmol·L-1有相似的抑制作用,二者都可以减少心肌细胞的凋亡率,降低由H2O2诱导的升高的Ca2+]i静息水平及频率。过氧化物酶体增殖物激活受体PPARγ受体特异性拮抗剂GW966210μmol.L-1能拮抗吡格列酮的部分抑制作用,但不影响吡格列酮对Ca2+]i的瞬变作用。结论吡格列酮可以抑制H2O2诱导的乳鼠心肌细胞凋亡,其作用机制可能与其抗脂质氧化和减少细胞内钙超载有关,其抑制作用部分是通过PPARγ受体介导的。

Inhibitory effect of pioglitazone against hydrogen peroxide induced cardiomyocytes apoptosis in neonatal rats

AIM To observe the protective effect of pioglitazone against apoptosis induced by hydrogen peroxide(H₂O₂) in neonatal rat cardiomyocytes and investigate the possible mechanism. METHODS H₂O₂ 200 μmol·L^-1 were used to set up an oxidative stress-induced injury model in neonatal rat cardiomyocytes. There were normal control, model, pioglitazone(0.1, 1 and 10 μmol·L^-1), pioglitazone (10 μmol·L^-1)+GW9662 (10 μmol·L^-1), and verapamil 1 μmol·L^-1 groups in the experiment. The cellular survival was determined with MTT. The content of malondialdehyde (MDA) was determined by measuring thiobarbituric acid-reactive substances and the activity of superoxide dismutase (SOD) was examined by xanthine oxidase method. Apoptotic rates were determined by flow cytometry. ［Ca²⁺］_i transient was measured by Till image system by cell loading Fura-2/AM. RESULTS Application of H2O2 200 μmol·L^-1 to myocytes for 12 h can cause cardiomyocytes apoptosis. Pioglitazone concentration-depndently increased the cell viabilty and SOD activity, decreased the content of MDA. Pioglitazone 10 μmol·L^-1 was the most obvious concentration, whose protective effect was similar to verapamil 1 μmol·L^-1. Both could decrease apoptotic rates of cardiomyocytes and ［Ca²⁺］_i resting level and frequency induced by H₂O₂. Peroxisome proliferator-activated receptor gamma (PPARγ) receptor antagonist GW9662 10 μmol·L^-1 inhibited partial promoting effects of pioglitazone, but not inhibit its effect on ［Ca²⁺］_i transient. CONCLUSION Pioglitazone can inhibit apoptosis induced by H₂O₂ in neonatal rat cardiomyocytes. The protective mechanism could be related to its ability to reduce lipid peroxidation and the calcium overload of the cardiomyocytes, which is partially mediated by PPARγ receptor.