激活M_3受体减轻CoCl_2诱导的大鼠心肌细胞系H9c2低氧损伤  

Activation of M_3-mAChR attenuates hypoxia injury induced by CoCl_2 in rat myocardial cell line H9c2

作　　者：彭源[1] 赵延礼[1,2] 苏晓灵 沈国平[1] 龙启福[1,2] 秦海兰[1] 王嵘 PENG Yuan;ZHAO Yanli;SU Xiaoling;SHEN Guoping;LONG Qifu;QIN Hailan;WANG Rong(Medical College of Qinghai University,Xining 810000;Basic Science Research Center,Medical College of Qinghai University,Xining 810000;Qinghai Provincial People s Hospital,Xining 810000,China)

机构地区：[1]青海大学医学院,青海西宁810000 [2]青海大学医学院基础医学研究中心,青海西宁810000 [3]青海省人民医院,青海西宁810000

出　　处：《基础医学与临床》2018年第4期458-463,共6页Basic and Clinical Medicine

基　　金：国家自然科学基金(81460051);青海省自然科学基金(2015-ZJ-744)

摘　　要：目的探讨卡巴胆碱(CCH,毒蕈碱受体亚型3特异性激动剂)激活毒蕈碱受体亚型3(M_3-mAChR)在氯化钴(CoCl_2)诱导的大鼠心肌细胞系H9c2低氧损伤中的作用及机制。方法正常大鼠心肌细胞系H9c2作为对照组,利用CoCl_2建立低氧损伤模型,选用M_3受体特异性激动剂CCH及其特异性阻断剂4-二苯乙酰氧基-N-甲基-哌啶甲碘化物(4-DAMP)对低氧损伤组进行干预。噻唑蓝比色法(MTT)检验细胞增殖活力;流式细胞计量术检测细胞凋亡;Western blot检测M_3受体、HIF-1α、HO-1和caspase-3蛋白表达水平。结果低氧模型组细胞凋亡率显著增高(P<0.01),细胞增殖显著降低(P<0.01);HIF-1α、caspase-3和HO-1蛋白表达水平显著上调(P<0.01)。用CCH干预后细胞凋亡率明显下降(P<0.01),细胞增殖活力明显增加(P<0.01);M_3受体、HIF-1α和HO-1蛋白表达水平显著上升(P<0.01);caspase-3蛋白表达水平明显降低(P<0.01)。应用4-DAMP干预后,由CCH介导的上述相关作用被抑制。结论 CCH激活M_3受体抑制CoCl_2诱导的大鼠心肌细胞系H9c2的低氧损伤,机制可能与HIF-1α和HO-1的蛋白表达水平上调有关。Objective To investigate the cytoprotection and mechanism of carbachol(CCH)to stimulate M 3 muscarinic acetylcholine receptor(M3-mAChR)against hypoxia injury induced by cobaltous chloride hexahydrate(CoCl2)in rat cardiomyocyte line H9c2.Methods Select the normal rat cardiomyocyte line H9c2 as the control group,rat cardiomyocyte line H9c2 was managed with CoCl2 to develop hypoxia injury model,M3-mAChR specific agonist CCH and antagonist 4-diphenyl-acetoxy-N-methyl-piperidine methiodide(4-DAMP)were used for intervention.The cell viability was tested by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide(MTT);The apoptosis in cardiomyocyte was detected by flow cytomery(FCM);The expression of M 3-mAChR,caspase-3,HIF-1αand HO-1 proteins was measured by Western blot assay.Results In hypoxia group,the apoptosis rate was significantly increased while cell proliferation decreased,the expression of HIF-1α,caspase-3 and HO-1 proteins were up-regulated obviously;After treatment with CCH,the apoptosis and cell proliferation of cardiomyocytes were significantly decreased,while the proliferation of cells increased,and the expression of M3-mAChR,HIF-1αand HO-1 proteins increased,the expression of caspase-3 protein was significantly decreased.Moreover,when applying 4-DAMP as intervention,these effects mediated by CCH was abolished.Conclusions CCH stimulates M3-mAChR against hypoxia injury induced by CoCl2 in rat cardiomyocyte strain H9c2,and the mechanism may be related to down-regulation of caspase-3 expression and up-regulation of HIF-1αand HO-1 protein expression.

关 键 词：毒蕈碱受体亚型3 低氧损伤 低氧诱导因子-1Α 血红素加氧酶-1 

分 类 号：R541[医药卫生—心血管疾病]