曲美他嗪干预气虚血瘀证心衰大鼠心肌线粒体蛋白质组学研究

摘要

目的:研究曲美他嗪对气虚血瘀证心衰大鼠心肌线粒体蛋白质组学的影响.方法:SD大鼠随机分为造模组与假手术组,造模组采用结扎左冠前降支致心梗后气虚血瘀证心衰大鼠模型,术后随机分为模型组、曲美他嗪组,各组连续灌胃给药8 周后采集大鼠心肌,采用双向凝胶电泳和基质辅助激光解析电离飞行时间质谱技术,比较各组大鼠心肌线粒体蛋白质组学差异,并运用蛋白免疫印迹技术对差异蛋白进行回复性验证.结果:模型组与假手术组相比有差异而曲美他嗪可使差异趋势逆转的蛋白点为18 个,经质谱分析成功鉴定出10个蛋白,主要涉及能量代谢、氧化损伤、应激反应、细胞骨架、细胞分化增殖等功能.Western blot 结果显示模型组Stress-70、Nucleophosmin 表达上调,ATP-α表达下调,曲美他嗪组Stress-70、Nucleophosmin 表达下调,ATP-α表达上调,与蛋白质组学结果一致.结论:曲美他嗪可一定程度调节模型大鼠心肌线粒体蛋白质组学变化,其干预机制可能与改善能量代谢、抗氧化损伤、减轻应激反应及调节细胞分化增殖等功能相关.采用双向凝胶电泳和质谱技术研究曲美他嗪对气虚血瘀证心衰大鼠心肌线粒体蛋白质组学的影响,结果真实可靠.%Objective: To investigate the effects of Trimetazidine on mitochondrial proteomic alterations in heart failure rats with qi- deficiency and blood- stasis syndrome after myocardial infarction. Methods: Heart failure models were established by ligating the left anterior descending coronary artery of SD rats. Rats were randomly divided into sham group, model group, Trimetazidine group. 8 weeks after drug intervention, the mitochondrial proteomic alterations of myocardial tissue were detected by two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption/ ionization time- of- flight mass spectrometry (MALDI- TOF- MS). Furthermore, expression levels of part of the differentially expressed proteins were verified by western blot. Results: Compared with model group, 18 differentially expressed protein spots were detected in Trimetazidine group, 10 of which were successfully identified by MALDI-TOFMS. Bioinformatics analysis showed that these differential proteins were mainly associated with energy metabolism, stress reaction, oxidative damage, cyto-skeleton, cell differentiation and proliferation. Western blot results showed that the expression of Stress-70 protein and Nucleophosmin increased and the expression of ATP-αdecreased in model group. The expression of Stress- 70 protein and Nucleophosmin decreased and the expression of ATP- αincreased in Trimetazidine group, which showed the same results in proteomics. Conclusion: Trimetazidine can partly adjust proteomic alterations of myocardial mitochondrial in HF rats with qi-deficiency and blood-stasis syndrome, and its intervention mechanism may involve improving energy metabolism, relieving stress reaction and oxidative damage, as well as regulating cell differentiation and proliferation. The results of comparative proteomic analysis performed by using 2-DE and MALDI-TOF-MS are accurate, stable and reliable.