芪苈强心提取物对缺氧诱导的大鼠心肌细胞葡萄糖代谢异常的改善作用  

Qiliqiangxin ameliorates hypoxia-induced glucose metabolic disorders in rat cardiomyocytes

作　　者：汪菁峰 李福海 沈冬丽 宋昱 韩雪婷 周京敏 葛均波 WANG Jingfeng;LI Fuhai;SHEN Dongli;SONG Yu;HAN Xueting;ZHOU Jingmin;GE Junbo(Department of Cardiology,Zhongshan Hospital,Fudan University,Shanghai 200032,China)

机构地区：[1]复旦大学附属中山医院心内科,上海200032

出　　处：《上海医学》2022年第6期387-393,共7页Shanghai Medical Journal

基　　金：国家自然科学基金(81870177);上海市卫生和计划生育委员会中医药科研课题(2018JP002)。

摘　　要：目的观察芪苈强心提取物对缺氧状态下大鼠心肌细胞葡萄糖代谢异常的改善作用。方法采用酶消化法+差速贴壁法分离培养Sprague Dawley(SD)大鼠乳鼠原代心肌细胞,随机分为对照组、缺氧组、芪苈强心+缺氧组、曲美他嗪+缺氧组。干预12 h后,行心肌细胞凋亡检测[TUNEL染色及天冬氨酸蛋白水解酶3(caspase3)活力测定],采用荧光法测定细胞葡萄糖摄取,酶法检测心肌细胞ATP水平及细胞上清液乳酸脱氢酶(LDH)活性,免疫印迹法检测心肌细胞内葡萄糖转运子-4(Glut-4)、乳酸脱氢酶(LDH)A、丙酮酸脱氢酶(PDH)、柠檬酸合酶(CS)等蛋白质表达水平。结果与对照组比较,缺氧组TUNEL阳性细胞比例显著增高[(15.11±2.30)%比(2.85±0.95)%,P<0.01],caspase3活性显著增强[(81.53±13.78)U/mg比(27.61±5.82)U/mg,P<0.01],细胞葡萄糖摄取率显著降低[(66.82±10.88)%比(100.19±1.40)%P<0.01],ATP生成显著减少[(4.87±1.02)nmol/mg比(8.21±1.40)nmol/mg,P<0.01],细胞上清液LDH活性显著增强[(246.92±24.66)U/L比(149.70±20.17)U/L,P<0.01];同时,细胞LDH A蛋白质相对表达量显著升高(P<0.01),Glut-4、PDH及CS蛋白质相对表达量显著降低(P<0.01)。与缺氧组比较,芪苈强心干预可显著降低TUNEL阳性细胞比例[(8.23±1.11)%比(15.11±2.30)%,P<0.01]和caspase3活性[(56.91±6.29)U/mg比(81.53±13.78)U/mg,P<0.01],提高葡萄糖摄取率[(82.75±15.09)%比(66.82±10.88)%,P<0.01],增加ATP生成[(6.61±1.39)nmol/mg比(4.87±1.02)nmol/mg,P<0.01],降低细胞上清液LDH活性[(216.76±15.41)U/L比(246.92±24.66)U/L,P<0.01],减少细胞LDH A蛋白质相对表达量(P<0.01),增加细胞Glut-4、PDH及CS蛋白质相对表达量(P<0.01),其效应与曲美他嗪相当。结论芪苈强心提取物通过调控葡萄糖代谢相关酶表达,可改善缺氧心肌细胞葡萄糖代谢,从而减少心肌细胞凋亡。Objective To observe the efficacy of Qiliqiangxin(QL)extract on glucose metabolic disorders in rat cardiomyocytes exposed to hypoxia.Methods Primary cardiomyocytes were isolated from neonatal Sprague Dawley(SD)rats using enzyme digestion and differential adhesion methods,and were randomized into control group,hypoxia group,QL+hypoxia group,and trimetazidine+hypoxia group.After 12-hour intervention,cell apoptosis was detected by TUNEL and caspase 3 activity assay.Glucose uptake was estimated by fluorometric assay.Enzyme labeling was applied for ATP concentration and lactic dehydrogenase(LDH)activity.Western Blot were used to measure protein expression of intracellular glucose transporter 4(Glut-4),LDH A,pyruvate dehydrogenase(PDH)and citrate synthase(CS).Results Compared with control group,cardiomyocytes in hypoxia group showed significantly increased apoptosis[(15.11±2.30)%vs.(2.85±0.95)%,P<0.01]and caspase3 activity[(81.53±13.78)U/mg vs.(27.61±5.82)U/mg,P<0.01],decreased glucose uptake[(66.82±10.88)%vs.(100.19±1.40)%,P<0.01]and ATP production[(4.87±1.02)nmol/mg vs.(8.21±1.40)nmol/mg,P<0.01],increased supernatant LDH activity[(246.92±24.66)U/L vs.(149.70±20.17)U/L,P<0.01]and intracellular LDH A expression(P<0.01),and decreased intracellular expression of Glut-4,PDH and CS(P<0.01).Cell apoptosis[(8.23±1.11)%vs.(15.11±2.30)%,P<0.01]and caspase3 activity[(56.91±6.29)U/mg vs.(81.53±13.78)U/mg,P<0.01]were significantly attenuated by QL treatment.Meanwhile,QL increased glucose uptake[(82.75±15.09)%vs.(66.82±10.88)%,P<0.01]and ATP production[(6.61±1.39)nmol/mg vs.(4.87±1.02)nmol/mg,P<0.01],reduced supernatant LDH activity[(216.76±15.41)U/L vs.(246.92±24.66)U/L,P<0.01]and intracellular LDH A level(P<0.01),and up-regulated the protein expression levels of Glut-4,PDH and CS in cardiomyocytes exposed to hypoxia(P<0.01).The effects were similar to trimetazidine.Conclusion QL extract can ameliorate glucose metabolism and attenuate apoptosis in cardiomyocytes exposed to hypoxia by regulating related enzyme

关 键 词：芪苈强心 心肌细胞 缺氧 葡萄糖代谢 凋亡 

分 类 号：R285[医药卫生—中药学]