黄芪注射液在逆转心肌细胞肥大过程中对心肌细胞线粒体结构和功能的影响

目的:通过大鼠原代心肌细胞培养,探讨心肌肥大过程中有关心肌细胞线粒体改变的病理和治疗问题。方法:分离和培养大鼠原代心肌细胞,并与血管紧张素Ⅱ(AngⅡ)共培养72,96 h。通过BCA法检测细胞总蛋白含量;倒置显微镜拍摄并测量细胞直径,反映心肌细胞增殖情况;通过荧光显微镜测量线粒体内膜膜电位(ΔΨm);酶标仪检测线粒体单胺氧化酶(MAO)活性;分光光度计检测线粒体细胞色素C氧化酶(COX)活性和线粒体外膜损伤百分率;高效液相色谱法检测心肌细胞内ATP,ADP,AMP含量,反映心肌细胞线粒结构和功能在与AngⅡ共培养中的损伤和能量代谢情况。在此基础上给予黄芪注射液和缬沙坦,观察它们对心肌细胞重构中线粒体结构、功能的药理作用。结果:在72,96 h 2个时间点,模型组较空白组心肌细胞总蛋白含量和心肌细胞直径均显著性增加。在该增殖过程中,心肌细胞线粒体MAO活性和线粒体外膜损伤百分率均显著升高,线粒体COX活性和线粒体ΔΨm均显著减低,ATP,ADP含量减少,AMP含量增加。黄芪注射液和缬沙坦能抑制AngⅡ引起的心肌细胞蛋白质合成增加和细胞直径增大、改善线粒体外膜损伤和内膜膜电位及COX,MAO活性,增加ATP,ADP含量、降低AMP含量。结论:在心肌肥大过程中,存在线粒体的结构和功能的损害,心肌细胞能量代谢损伤变化是继心肌细胞线粒体结构损伤后发生的。黄芪注射液和缬沙坦在逆转血管紧张素Ⅱ所引起的心肌细胞肥大的过程中具有保护心肌细胞线粒体结构和功能的作用,逆转心肌细胞肥大、纠正心肌重构有利于心肌细胞能量的改善。

Effect of Astragali Radix injection on myocardial cell mitochondrial structure and function in process of reversing myocardial cell hypertrophy

Objective: To study pathological and therapeutical problems concerning myocardial cell mitochondria changes during myocardial cell hypertrophy by culturing rat primary myocardial cells.Method: Primary myocardial cells were seperated and cultured together with angiotensin II(Ang II) for 72 or 96 hours.The total protein content with the BCA method and the photography and measurement of cell diameter with inverted microscope reflected myocardial cell proliferation.The mitochondrial membrane potential(Δ Ψ m) with fluorescence microscope,the mitochondrial single amine oxidase(MAO) activity with spectrophotometer,the mitochondrial cytochrome oxidase(COX) activity and the injury percentage of mitochondrial outer membrane with microplate reader and the contents of ATP,ADP,AMP with high performance liquid chromatography reflected the injury and energy metabolism of myocardial cell mitochondrial structure and function when being cultured together with Ang II.On that basis,cells were treated with Astragali Radix injection and valsartan for observing pharmacological effects on mitochondrial structure and function in restructured myocardial cells.Result: In 72 h and 96 h,compare with the control group,the model group showed significantly increased total protein content and enlarged myocardial cell diameter.During the course of proliferation,the myocardial cell MAO activity and the injury percentage of mitochondrial outer membrane were significantly increased,with significant decrease in mitochondrial COX activity,mitochondrial Δ Ψ m and the content of ATP,ADP and rise in the content of AMP.Astragali Radix injection and valsartan reduced myocardial cell total protein content and cell diameter caused by Ang II,decreased myocardial cell MAO activity,significantly increased mitochondrial COX activity and the content of ATP and ADP,and decreased the content of AMP.Conclusion: During the process of myocardial hypertrophy,the injury of mitochondrial structure and function and the changes in myocardial cell energy metabolism injury occurred after the injury of mitochondria.Astragali Radix injection and valsartan can reverse myocardial cell mitochondrial structure and function during myocardial cell hypertrophy caused by Ang II.Reversion of myocardial cell hypertrophy and restructuring of myocardial cells helps improve energy metabolism of the myocardial cells.