基于线粒体凋亡途径介导的骨骼肌细胞凋亡探究参芪复方对糖尿病骨骼肌病变的干预机制  

Exploring Intervention Mechanism of Skeletal Muscle Cell Apoptosis in Diabetic Skeletal Myopathy Based on Mitochondrial Apoptosis Pathway-Mediated Skeletal Muscle Cell Apoptosis with Shenqi Compound(参芪复方)

作　　者：朱海燕 万李 丁若兰[3] 张宏[3] ZHU Haiyan;WAN Li;DING Ruolan;ZHANG Hong(Clinical Medical College and The First Affiliated Hospital of Chengdu Medical College,Chengdu 610500,Sichuan,China;Sports Medicine and Health College of Chengdu Sports University,Chengdu 610041,Sichuan,China;Hospital of Chengdu University of Traditional Chinese Medicine,Chengdu 610072,Sichuan,China)

机构地区：[1]成都医学院临床医学院第一附属医院,四川成都610500 [2]成都体育学院运动医学与健康学院,四川成都610041 [3]成都中医药大学附属医院,四川成都610072

出　　处：《中华中医药学刊》2023年第8期66-72,I0024,共8页Chinese Archives of Traditional Chinese Medicine

基　　金：国家自然科学基金项目(81660788);四川省干部保健科研基金项目(川干研2021-513)。

摘　　要：目的 探讨参芪复方通过调控线粒体凋亡途径介导的骨骼肌细胞凋亡对糖尿病骨骼肌病变的作用。方法 以含5.56 mmol/L葡萄糖(Glucose, Glu)培养基及含25 mmol/L Glu的培养基分别培养L6细胞,模拟正常及糖尿病的生理环境。实验设置6组,为空白组、模型组、参芪复方低剂量组、参芪复方中剂量组、参芪复方高剂量组、罗格列酮组,分别给予含5.56 mmol/L Glu培养基稀释的10%空白血清、含25 mmol/L Glu培养基稀释的10%空白血清、5%参芪复方含药血清、10%参芪复方含药血清、15%参芪复方含药血清、300 ng/mL罗格列酮混悬液进行干预。药物干预24 h后收集细胞,以流式细胞术检测各组细胞凋亡率,以RT-qPCR和Western blot检测各组细胞内B淋巴细胞瘤-2基因(Bcl-2)、B淋巴细胞瘤-xl基因(Bcl-xl)、Bcl-2同源拮抗剂/杀伤剂(Bak)、Bcl-xl/Bcl-2相关死亡启动因子(Bad)、Bcl-2相关X蛋白(Bax)、细胞色素C(Cyt c)、第二个线粒体来源的胱氨酸酶激活剂/低等电点IAP直接结合蛋白(Smac/Diablo)、细胞凋亡诱导因子(AIF)、半胱氨酸蛋白酶-3(Caspase-3)、半胱氨酸蛋白酶-9(Caspase-9)的基因与蛋白表达水平。结果 与空白组相比,模型组细胞凋亡率显著升高(P<0.05),细胞内Bak、Bax、Cyt c、Smac/Diablo、AIF、Caspase-3、Caspase-9的基因与蛋白表达显著增加(P<0.05),Bad蛋白表达显著增加(P<0.05),Bcl-2的基因与蛋白表达显著降低(P<0.05),Bcl-xl基因表达显著降低(P<0.05);与模型组相比,参芪复方组Bak、Bax、Caspase-9、Caspase-3的基因与蛋白表达显著降低(P<0.05),Bad、Cyt c、Smac/Diablo的蛋白表达显著降低,Bcl-2、Bcl-xl的基因与蛋白表达显著升高(P<0.05)。结论 参芪复方糖尿病骨骼肌病变的干预作用可能是通过调控线粒体凋亡途径,降低高糖环境下骨骼肌细胞凋亡率来实现的。Objective To investigate the effect of skeletal muscle cell apoptosis mediated by the mitochondrial apoptosis pathway in diabetic skeletal myopathy through the regulation of Shenqi Compound(参芪复方).Methods L6 cells were cultured with medium containing 5.56 mmol/L glucose(Glu)and medium containing 25 mmol/L Glu to simulate the normal and diabetic physiological environments,respectively.Six groups were set up,namely blank group,model group,Shenqi Compound low dose group,Shenqi Compound medium dose group,Shenqi Compound high dose group and rosiglitazone group,which were given 10%blank serum diluted with 5.56 mmol/L Glu medium,10%blank serum diluted with 25 mmol/L Glu medium,5%Shenqi Compound medicated serum,10%Shenqi Compound medicated serum,15%Shenqi Compound medicated serum and 300 ng/mL rosiglitazone suspension for intervention.The cells were collected 24 h after drug intervention,and the apoptosis rate of each group was detected by flow cytometry,and the gene and protein expressions of B-cell lymphoma-2(Bcl-2),B-cell lymphoma-extra large(Bcl-xl),Bcl-2 homologous antagonist/kller(Bak),Bcl-xl/Bcl-2 associated death promoter(Bad),Bcl-2-associ-ated-X-protein(Bax),Cytochrome C(Cyt c),second mitochondria-derived activator of caspase/dDirect IAP-binding protein with low PI(Smac/Diablo),apoptosis-inducing factor(AIF),Caspase-3 and Caspase-9 were detected by RT-qPCR and Western blot in each group.Results Compared with that of the blank group,the apoptosis rate was significantly higher in the model group(P<0.05),the intracellular gene and protein expressions of Bak,Bax,Cyt c,Smac/Diablo,AIF,Caspase-3 and Caspase-9 were significantly increased(P<0.05)in the model group,Bad protein expression was significantly increased(P<0.05)in the model group,the gene and protein expression of Bcl-2 were significantly decreased(P<0.05)in the model group,and Bcl-xl gene expression was significantly decreased(P<0.05)in the model group.Compared with those of the model group,the gene and protein expressions of Bak,Bax,Caspase-9 and Cas

关 键 词：糖尿病骨骼肌病变 细胞凋亡 线粒体凋亡途径 参芪复方 

分 类 号：R256.1[医药卫生—中医内科学]