中文摘要：

目的：研究蛋白激酶B（Akt）在促红细胞生成素（EPO）增强慢性缺氧环境中心肌细胞线粒体生物合成中的作用机制。方法：采用H9c2心肌细胞,将其于缺氧环境下培养7d（94%N2,1%O2,5%CO2）,建立心肌细胞慢性缺氧模型。将心肌细胞根据不同处理分为缺氧对照组（HC组）,20U/ml重组人EPO（rhEPO）处理缺氧组（HE组）和20U/ml rhEPO＋100nmol/ml wortmannin特异性阻断Akt缺氧组（HW组）。以电镜观察线粒体超微结构变化,计算线粒体的体密度（Vv）、数密度（Nv）;荧光探针观察检测线粒体数量变化;RT-PCR检测线粒体DNA相对拷贝数变化;Western blot检测Akt总蛋白表达及磷酸化水平变化。结果：在rhEPO作用后,Akt磷酸化显著增强（P〈0.05）,电镜观察显示线粒体Vv及Nv均显著升高（均P〈0.05）,线粒体数量及其DNA相对拷贝数也显著增加（P〈0.05）;采用wortmannin特异性阻断Akt使其磷酸化水平显著降低（P〈0.05）,同时使线粒体Vv、Nv、线粒体数量及其DNA相对拷贝数减少（均P〈0.05）。结论：EPO通过促进Akt磷酸化进而增强慢性缺氧心肌细胞线粒体生物合成。

英文摘要：

Objective：To study the mechanism that erythropoietin（EPO）enhances mitochondiral biogenesis in cardiomyocytes exposed to chronic hypoxia through phosphorylation of protein kinase B（Akt）.Method：H9c2cardiomyocytes were cultured in the environment of hypoxia for 1week（94%N2,1%O2,5%CO2）,establishing the chronic hypoxic cardiomyocyte model.All the cells were divided into 3groups：HC（chronic hypoxic control）,HE[（treated with chronic hypoxia and 20U/ml recombinant human EPO（rhEPO）]and HW（cells treated with both20U/ml rhEPO and 100nmol/ml wortmannin in chronic hypoxia）.Ultrastructure of mitochondria was observed with electron microscope.Mitochondrial volume density and numerical density were calculated simultaneously.Fluorescent probe was used to detect the changes of mitochondial number.Mitochondial DNA（mtDNA）relative copy number was assayed by RT-PCR.The expression and phosphorylation of Akt protein were analyzed with western blot.Result：rhEPO significantly increased the phosphorylation of Akt and elavated the mitochondial volume density,numerical density,mitochondial number and mtDNA relative copy number（P〈0.05）.Special blockade of Akt phosphorylation with wormannin significantly negativley affected all the above changes induced by rhEPO（P〈0.05）.Conclusion：EPO enhances mitochondrial biogenesis in cardiomyocytes exposed to chronic hypoxia at least partly through phosphorylating Akt.