丹红注射液抗H9c2细胞凋亡的作用机制研究

Mechanisms research of anti-apoptosis by Danhong Injection in H9c2 myocardial cells

关键词:  丹红注射液  心肌细胞凋亡  线粒体膜电位  ROS  钙离子

Key words:  Danhong Injection  myocardial cell apoptosis  mitochondrial membrane potential  ROS  Ca²⁺

摘要:

[目的]从线粒体损伤角度研究丹红注射对异丙肾上腺素（ISO）诱导的心肌细胞H9c2凋亡的影响。[方法]培养H9c2细胞，分为对照组、10 μmol/L ISO模型组，10 μmol/L ISO和20、200 μL/L丹红注射液组。按相应给药时间后采用流式细胞术检测细胞凋亡，明确药物对细胞凋亡的作用。活细胞荧光成像仪检测细胞线粒体膜电位变化、激光共聚焦检测细胞内活性氧簇（ROS）的产生、Fluo-3 AM ester荧光染料染色检测细胞内钙离子浓度研究丹红注射液抗心肌细胞凋亡的相关作用机制。[结果]与对照组（1.33%±0.42%）相比，ISO显著上升细胞凋亡率（5.13%±0.15%）（P<0.01）；与ISO组比较，20、200 μL/L丹红注射液后，细胞凋亡率分别下降为（4.23%±0.15%）和（2.37%±0.38%）。与对照组相比，给予ISO后细胞内线粒体膜电位去极化程度显著增加，为对照的（151.76%±16.01%）（P<0.01），20、200 μL/L丹红注射液显著抑制ISO诱导的细胞内线粒体膜去极化，分别为对照的（121.07%±11.53%）和（117.15%±12.91%）。此外，丹红注射液显著抑制ISO诱导的细胞内ROS浓度及钙离子浓度的升高，差异具有统计学意义。[结论]丹红注射液可通过抑制ISO引起的细胞内线粒体膜电位去极化、降低细胞内ROS和钙离子浓度而发挥其抗H9c2细胞凋亡作用。

Abstract:

[Objective] The aim of this article is to study the effects of Danhong injection on apoptosis induced by isoproterenol(ISO) in H9c2 myocardial cells from the perspective of mitochondrial damage.[Methods] H9c2 cells were cultured and there were control group,10 μmol/L ISO model group,10 μmol/L ISO group,20 μL/L Danhong injection group,200 μL/L Danhong injection group. Cell apoptosis was detected by flow cytometry. Mitochondrial membrane potential was detected by living cell fluorescence imaging. Production of reactive oxygen spices (ROS) was detected by laser focal microscope. Fluo-3,AM ester fluorescent dye were used to detect intracellular calcium concentration.[Results] Compared with control group (1.33±0.42)%,the apoptotic rate in ISO group increased (5.13±0.15)% (P<0.01);Compared with ISO group,the apoptotic rate in 20 and 200 μL/L Danhong injection groups decreased to(4.23±0.15)% and (2.37±0.38)% respectively. Compared with the control group,the degree of mitochondrial membrane potential depolarization increased significantly after ISO treatment,which was (151.76±16.01)% (P<0.01). 20 and 200 mL/L Danhong injection significantly inhibited the mitochondrial membrane depolarization induced by ISO,which were (121.07±11.53)% and (117.15±12.91)%. In addition,Danhong injection significantly inhibited the increase of ROS and Ca²⁺ concentration induced by ISO,and the differences were statistically significant.[Conclusion] Danhong injection can inhibit the depolarization of mitochondrial membrane potential induced by ISO and decrease the concentration of ROS and Ca²⁺ in cells,thus exerting its anti-apoptotic effect on H9c2 cells.