参麦注射液保护氧化损伤心肌细胞线粒体的机制研究

目的: 探究参麦注射液对氧化损伤心肌细胞的保护作用及其对线粒体能量代谢过程的影响。方法: 采用叔丁基过氧化氢（t-BHP）诱导构建H9c2心肌细胞氧化损伤模型。分别采用MTT法和化学发光法检测参麦注射液对细胞存活率和ATP水平的影响；采用Clark氧电极检测参麦注射液对细胞有氧呼吸速率的调节作用；采用ELISA法检测丙酮酸及丙酮酸脱氢酶（PDH）的含量及活性；采用蛋白质印迹法检测丙酮酸脱氢酶亚基（PDHA1）含量；采用实时定量RT-PCR检测丙酮酸脱氢酶激酶1（PDK1）mRNA的表达。结果: 与氧化损伤模型比较，参麦注射液可有效提高心肌细胞存活率和ATP水平（均P < 0.01），提高细胞氧呼吸速率，减少丙酮酸含量，并提高PDH活性（P < 0.05或P < 0.01）。在分子水平上，参麦注射液可提高PDHA1蛋白表达（P < 0.05），并具有降低PDK1 mRNA表达的趋势（P>0.05）。结论: 参麦注射液可有效保护氧化损伤心肌细胞，其机制可能与提高PDH活性、减少丙酮酸堆积，从而维持线粒体功能稳定和细胞能量代谢稳定有关。

Shenmai injection protects mitochondria from oxidative injury in myocardial cells and its mechanism

Objective: To investigate the effect of Shenmai injection on myocardial cells with oxidative injury and the underlying mechanisms. Methods: Tert-butyl hydroperoxide (t-BHP) was used to induce the oxidative stress in H9c2 myocardial cells. The cell viability and ATP level were evaluated using MTT-colorimetric method and CellTiter-Glo luminescent cell viability assay. The oxygen respiration rate was examined by Clark oxygen electrode. Pyruvate and pyruvate dehydrogenase (PDH) levels were evaluated by ELISA kit. Western blot and quantitative real-time RT-PCR were employed to evaluate the expression of pyruvate dehydrogenase alpha 1(PDHA1) and pyruvate dehydrogenase kinase 1(PDK1). Results: Shenmai injection significantly improved viability and respiration of H9c2 myocardial cells after t-BHP injury (P < 0.05 or P < 0.01). It increased ATP contents by consuming pyruvate and increasing PDH level (P < 0.05 or P < 0.01). Furthermore, Shenmai injection had the tendency to increase protein expression of PDHA1(P < 0.05) and decrease mRNA expression of PDK1 (P>0.05). Conclusion: Shenmai injection protects mitochondria from oxidative stress by increasing PDH level, which indicates that it may improve energy metabolism of myocardial cells.