参附益心颗粒通过线粒体KATP通道改善缺氧H2C9心肌细胞能量代谢的作用  

Study on Effects of Shenfu Yixin Granules Improving Energy Metabolism of Hypoxic H2C9 Cardiomyocytes through Mitochondrial KATP Channel

作　　者：郑荣菲 曹亚选 司春婴[2] 陈玉善[2] 李彬[2,3] 崔琳[4] 关怀敏[2] 朱明军[2] 王贺[2] Zheng Rongfei;Cao Yaxuan;Si Chunying;Chen Yushan;Li Bin;Cui Lin;Guan Huaimin;Zhu Mingjun;Wang He(The First School of Clinical Medicine,Henan University of Chinese Medicine,Zhengzhou 450099,China;Center of Cardiology,The First Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450099,China;Graduate Division,The First Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450099,China;Central Laboratory,The First Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450099,China)

机构地区：[1]河南中医药大学第一临床医学院,郑州450099 [2]河南中医药大学第一附属医院心脏中心,郑州450099 [3]河南中医药大学第一附属医院研究生科,郑州450099 [4]河南中医药大学第一附属医院中心实验室,郑州450099

出　　处：《世界科学技术-中医药现代化》2020年第11期3906-3912,共7页Modernization of Traditional Chinese Medicine and Materia Medica-World Science and Technology

基　　金：国家自然科学基金委员会青年项目(81603432):参附益心颗粒通过调节心肌ATP敏感钾通道改善心肌梗死后心衰大鼠的能量代谢研究,负责人:王贺;国家自然科学基金委员会青年项目(81603431):桂枝甘草汤通过AC-cAMP-PKA通路改善窦房结增龄性重构的作用机制研究,负责人:曹英杰;河南省中医管理局河南省中医药科学研究项目(2019ZY2139):参附益心颗粒通过调控心肌细胞线粒体自噬改善心肌梗死大鼠心室重构的机制研究,负责人:王贺。

摘　　要：目的探索参附益心颗粒改善缺氧心肌细胞能量代谢的机制。方法采用CCK-8法筛选出参附益心颗粒含药血清实验研究的药物浓度。H2C9心肌细胞随机分为正常组,缺氧组,缺氧+二氮嗪组,缺氧+参附益心颗粒组,缺氧+参附益心颗粒+5-羟基癸酸组.除正常组外,根据实验分组,缺氧处理前预先对细胞进行二氮嗪、5-羟基癸酸和参附益心颗粒含药血清处理,后将细胞置于缺氧的37℃的细胞培养箱中连续培养6 h。采用荧光素酶发光法检测细胞ATP的浓度,CCK-8检测细胞的增殖能力,流式细胞术检测细胞的凋亡水平,Western blot测定线粒体KATP(mitoKATP)的蛋白Kir6.2和SUR2A的表达。结果根据CCK-8实验结果选择250μM的参附益心颗粒含药血清作为实验药物浓度。与正常组相比,缺氧H2C9心肌细胞ATP水平显著降低,细胞的增殖能力降低,凋亡率增加,mitoKATP通道蛋白的Kir6.2和SUR2A的表达水平均显著降低(均P<0.01)。与缺氧组相比,二氮嗪和参附益心颗粒组的ATP浓度,细胞增殖能力和Kir6.2,SUR2A蛋白表达水平均显著升高,细胞的凋亡率显著下降(均P<0.01)。而参附益心颗粒经mitoKATP阻断剂5-羟基癸酸作用后,细胞的ATP浓度,增殖能力和Kir6.2,SUR2A的表达均显著下降(均P<0.01),凋亡率明显升高(P<0.01)。结论参附益心颗粒可能通过mitoKATP通道改善缺氧心肌细胞的能量代谢。Objective To explore the mechanism of Shenfu Yixin granules in improving energy metabolism of hypoxic cardiomyocytes.Methods MTT assay was used to screen the drug concentration of Shenfu Yixin granules in serum.H2 C9 myocardial cells were randomly divided into normal group,hypoxia group,hypoxia+diazoxide group,hypoxia+Shenfu Yixin granules group,hypoxia+5-hydroxydecanoate+Shenfu Yixin granules group.Except for that in the normal group,the cells in the other groups were prepared respectively with diazoxide,5-hydroxydecanoate,and Shenfu Yixin granules before hypoxia treatment,then the cells were cultured in an oxygen-deficient incubator at 37℃ for 6 hours.The concentration of ATP was detected by luciferase luminescence method,the proliferation ability was detected by CCK-8,the apoptosis level was detected by flow cytometry,and the expression of mitokatp protein KATP(mitoKATP)proteins Kir6.2 and SUR2 A were determined by Western blot.Results According to MTT results,250 μM serum containing Shenfu Yixin granules was selected as the experimental drug concentration.Compared with the normal group,ATP level of H2 C9 myocardial cells decreased significantly,cell proliferation ability decreased,apoptosis rate increased,and the expression levels of Kir6.2 and SUR2 A of mitoKATP channel protein decreased significantly(all P <0.01).Compared with the hypoxia group,ATP concentration,cell proliferation ability,Kir6.2 and SUR2 A protein expression levels of the diazoxide and Shenfu Yixin granules groups increased,and cell apoptosis rate decreased(all P <0.01).However,after Shenfu Yixin granules were treated with mitoKATP channel blocker 5-hydroxydecanoate,ATP concentration,proliferation ability,Kir6.2 and SUR2 A expression decreased significantly(all P <0.01),and apoptosis rate increased(P <0.01).Conclusion Shenfu Yixin granules may improve the energy metabolism of hypoxic cardiomyocytes through mitoKATP channel.

关 键 词：参附益心颗粒 能量代谢 线粒体KATP通道 缺氧心肌细胞 

分 类 号：R285.5[医药卫生—中药学]