TRPV1对H9c2心肌细胞缺氧复氧后凋亡的影响及机制
|
摘要
目的探讨瞬时受体电位阳离子通道V亚族成员1(TRPV1)对H9c2心肌细胞缺氧复氧后凋亡的保护作用及可能机制。方法将H9c2心肌细胞随机分为对照组、模型组(缺氧/复氧组)、辣椒素(TRPV1激活剂)组(缺氧/复氧+辣椒素组)、辣椒素+LY组[缺氧/复氧+辣椒素+LY294002(PI3k抑制剂)组]。采用CCK-8法测定心肌细胞存活率,采用流式细胞仪测定各组心肌细胞凋亡率,采用Western blot法和逆转录-聚合酶链反应(RTPCR)法测定各组细胞caspase-3、B淋巴细胞瘤-2基因(Bcl-2)、Bcl-2相关X蛋白(Bax)、磷酸化-蛋白激酶B(pAkt)、蛋白激酶B(Akt)水平。结果与对照组比较,模型组、辣椒素组、辣椒素+LY组心肌细胞存活率和Bcl-2水平降低(均P<0.05),细胞凋亡率和caspase-3、Bax、p-Akt水平升高(均P<0.05);与模型组比较,辣椒素组心肌细胞存活率和Bcl-2、p-Akt水平升高(均P<0.05),细胞凋亡率和caspase-3、Bax水平降低(均P<0.05);模型组与辣椒素+LY组各指标比较差异均无统计学意义(均P>0.05)。结论 TRPV1可抑制缺氧复氧后H9c2心肌细胞凋亡,其机制可能与TRPV1激活磷脂酰肌醇3-激酶(PI3k)/Akt信号通路有关。
Objective To investigate the protective effect and possible mechanism of transient receptor potential cation channel subfamily V member 1(TRPV1) on apoptosis of H9c2 cardiomyocyte after hypoxia-reoxygenation injury.Methods H9c2 cardiomyocyte were randomly divided into control group,model group(hypoxia/reoxygenation),capsaicin group(hypoxia/reoxygenation + capsaicin),capsaicin + LY group(hypoxia/reoxygenation + capsaicin +LY294002).The cell survival rate of cardiomyocyte was determined by CCK-8 method.The apoptosis rate of cardiomyocyte was determined by flow cytometry.Western blot and reverse transcription-polymerase chain reaction(RT-PCR) were used to determine caspase-3,B-cell lymphoma 2 gene(Bcl-2),Bcl-2 related X protein(BAX),phosphoric acid-protein kinase B(p-Akt) and protein kinase B(Akt) levels.Results Compared with the control group,the cell survival rate of cardiomyocyte and the Bcl-2 level in the model group,capsaicin group and capsaicin + LY group decreased(all P < 0.05),the apoptosis rate and the levels of caspase-3,Bax,p-Akt increased(all P < 0.05);Compared with the model group,the cell survival rate of cardiomyocyte and the levels of Bcl-2,p-Akt in the capsaicin group were increased(all P < 0.05),and the apoptosis rate and the levels of caspase-3 and Bax were decreased(all P < 0.05).There were no significant difference in each indexes between the model group and the capsaicin + LY group(all P > 0.05).Conclusion TRPV1 can inhibit H9c2 cardiomyocyte apoptosis after hypoxia-reoxygenation inury,and its mechanism may be related to TRPV1 activation of phosphatidylinositol 3-kinase(PI3k)/Akt signaling pathway.
Objective To investigate the protective effect and possible mechanism of transient receptor potential cation channel subfamily V member 1(TRPV1) on apoptosis of H9c2 cardiomyocyte after hypoxia-reoxygenation injury.Methods H9c2 cardiomyocyte were randomly divided into control group,model group(hypoxia/reoxygenation),capsaicin group(hypoxia/reoxygenation + capsaicin),capsaicin + LY group(hypoxia/reoxygenation + capsaicin +LY294002).The cell survival rate of cardiomyocyte was determined by CCK-8 method.The apoptosis rate of cardiomyocyte was determined by flow cytometry.Western blot and reverse transcription-polymerase chain reaction(RT-PCR) were used to determine caspase-3,B-cell lymphoma 2 gene(Bcl-2),Bcl-2 related X protein(BAX),phosphoric acid-protein kinase B(p-Akt) and protein kinase B(Akt) levels.Results Compared with the control group,the cell survival rate of cardiomyocyte and the Bcl-2 level in the model group,capsaicin group and capsaicin + LY group decreased(all P < 0.05),the apoptosis rate and the levels of caspase-3,Bax,p-Akt increased(all P < 0.05);Compared with the model group,the cell survival rate of cardiomyocyte and the levels of Bcl-2,p-Akt in the capsaicin group were increased(all P < 0.05),and the apoptosis rate and the levels of caspase-3 and Bax were decreased(all P < 0.05).There were no significant difference in each indexes between the model group and the capsaicin + LY group(all P > 0.05).Conclusion TRPV1 can inhibit H9c2 cardiomyocyte apoptosis after hypoxia-reoxygenation inury,and its mechanism may be related to TRPV1 activation of phosphatidylinositol 3-kinase(PI3k)/Akt signaling pathway.
引文
[1]YU L M,DI W C,DONG X,et al.Diallyl trisulfide exerts cardioprotection against myocardial ischemia-reperfusion injury in diabetic state,role of AMPK-mediated AKT/GSK-3β/HIF-1αactivation[J].Oncotarget,2017,8(43):74791-74805.
[2]RONG R,XIJUN X.Erythropoietin pretreatment suppresses inflammation by activating the PI3K/Akt signaling pathway in myocardial ischemia-reperfusion injury[J].Exp Ther Med,2015,10(2):413-418.
[3]GAO Y,SONG J,CHEN H,et al.TRPV1 activation is involved in the cardioprotection of remote limb ischemic postconditioning in ischemiareperfusion injury rats[J].Biochem Biophys Res Commun,2015,463(4):1034-1039.
[4]姜小雪,刘关羽,雷寒,等.PI3K/Akt通路介导TRPV1抑制离体小鼠心脏缺血再灌注后的细胞凋亡[J].基础医学与临床,2016,36(9):1187-1192.
[5]RANDHAWA P K,JAGGI A S.Investigating the involvement of TRPV1 ion channels in remote hind limb preconditioning-induced cardioprotection in rats[J].Naunyn Schmiedebergs Arch Pharmacol,2017,390(2):117-126.
[6]SINGH A,RANDHAWA P K,BALI A,et al.Exploring the role of TRPV and CGRP in adenosine preconditioning and remote hind limb preconditioning-induced cardioprotection in rats[J].Cardiovasc Drugs Ther,2017,31(2):133-143.
[7]王挺,杨怡,田玥,等.瞬时受体电位香草醛亚家族1对高脂处理肝细胞凋亡的保护作用[J].华西药学杂志,2017,32(1):25-27.
[8]VEMULA P,GAUTAM B,ABELA G S,et al.Myocardial ischemia/reperfusion injury:potential of TRPV1 agonists as cardioprotective agents[J].Cardiovasc Hematol Disord Drug Targets,2014,14(1):71-78.
[9]YIN X,WANG X,FAN Z,et al.Hyperbaric oxygen preconditioning attenuates myocardium ischemia-reperfusion injury through upregulation of heme oxygenase 1 expression:PI3K/Akt/Nrf2 pathway involved[J].J Cardiovasc Pharmacol Ther,2015,20(4):428-438.
[10]WANG J,JI S Y,LIU S Z,et al.Cardioprotective effect of breviscapine:inhibition of apoptosis in H9c2 cardiomyocytes via the PI3K/Akt/e NOS pathway following simulated ischemia/reperfusion injury[J].Pharmazie,2015,70(9):593-597.
[11]LI C M,SHEN S W,WANG T,et al.Myocardial ischemic post-conditioning attenuates ischemia reperfusion injury via PTEN/Akt signal pathway[J].Int J Clin Exp Med,2015,8(9):15801-15807.
[12]WANG X,CHEN J,WANG H,et al.Memantine can reduce ethanolinduced caspase-3 activity and apoptosis in H4 cells by decreasing intracellular calcium[J].J Mol Neurosci,2017,62(3-4):402-411.
[13]SERGEEVA T F,SHIRMANOVA M V,ZLOBOVSKAYA O A,et al.Relationship between intracellular pH,metabolic co-factors and caspase-3 activation in cancer cells during apoptosis[J].Biochim Biophys Acta,2017,1864(3):604-611.
[14]仇志富,吴晓光,孟杰,等.补阳还五汤提取物灌胃对脑出血大鼠脑组织中PI3K、AKT、Caspase-3表达的影响[J].山东医药,2016,56(9):29-31.
[15]BAO RK,ZHENG S F,WANG X Y.Selenium protects against cadmium-induced kidney apoptosis in chickens by activating the PI3K/AKT/Bcl-2 signaling pathway[J].Environ Sci Pollut Res Int,2017,24(25):20342-20353.
[16]WANG C,WEI Z,JIANG G,et al.Neuroprotective mechanisms of miR-124 activating PI3K/Akt signaling pathway in ischemic stroke[J].Exp Ther Med,2017,13(6):3315-3318.
[17]XI H,FAN X,ZHANG Z,et al.Bax and Bcl-2 are involved in the apoptosis induced by local testicular heating in the boar testis[J].Reprod Domest Anim,2017,52(3):359-365.
[18]YANG Z,GENG Y,YAO Z,et al.Spatiotemporal expression of Bcl-2/Bax and neural cell apoptosis in the developing lumbosacral spinal cord of rat fetuses with anorectal malformations[J].Neurochem Res,2017,42(11):3160-3169.
[19]LIN C H,WU M R,LI C H,et al.Editor's highlight:Periodic exposure to smartphone-mimic low-luminance blue light induces retina damage through Bcl-2/BAX-dependent apoptosis[J].Toxicol Sci,2017,157(1):196-210.
[20]黎萍,郑百红,陈鹏,等.硒对病毒性心肌炎小鼠心肌细胞凋亡的抑制作用及其作用机制[J].吉林大学学报(医学版),2016,42(1):54-58.
[2]RONG R,XIJUN X.Erythropoietin pretreatment suppresses inflammation by activating the PI3K/Akt signaling pathway in myocardial ischemia-reperfusion injury[J].Exp Ther Med,2015,10(2):413-418.
[3]GAO Y,SONG J,CHEN H,et al.TRPV1 activation is involved in the cardioprotection of remote limb ischemic postconditioning in ischemiareperfusion injury rats[J].Biochem Biophys Res Commun,2015,463(4):1034-1039.
[4]姜小雪,刘关羽,雷寒,等.PI3K/Akt通路介导TRPV1抑制离体小鼠心脏缺血再灌注后的细胞凋亡[J].基础医学与临床,2016,36(9):1187-1192.
[5]RANDHAWA P K,JAGGI A S.Investigating the involvement of TRPV1 ion channels in remote hind limb preconditioning-induced cardioprotection in rats[J].Naunyn Schmiedebergs Arch Pharmacol,2017,390(2):117-126.
[6]SINGH A,RANDHAWA P K,BALI A,et al.Exploring the role of TRPV and CGRP in adenosine preconditioning and remote hind limb preconditioning-induced cardioprotection in rats[J].Cardiovasc Drugs Ther,2017,31(2):133-143.
[7]王挺,杨怡,田玥,等.瞬时受体电位香草醛亚家族1对高脂处理肝细胞凋亡的保护作用[J].华西药学杂志,2017,32(1):25-27.
[8]VEMULA P,GAUTAM B,ABELA G S,et al.Myocardial ischemia/reperfusion injury:potential of TRPV1 agonists as cardioprotective agents[J].Cardiovasc Hematol Disord Drug Targets,2014,14(1):71-78.
[9]YIN X,WANG X,FAN Z,et al.Hyperbaric oxygen preconditioning attenuates myocardium ischemia-reperfusion injury through upregulation of heme oxygenase 1 expression:PI3K/Akt/Nrf2 pathway involved[J].J Cardiovasc Pharmacol Ther,2015,20(4):428-438.
[10]WANG J,JI S Y,LIU S Z,et al.Cardioprotective effect of breviscapine:inhibition of apoptosis in H9c2 cardiomyocytes via the PI3K/Akt/e NOS pathway following simulated ischemia/reperfusion injury[J].Pharmazie,2015,70(9):593-597.
[11]LI C M,SHEN S W,WANG T,et al.Myocardial ischemic post-conditioning attenuates ischemia reperfusion injury via PTEN/Akt signal pathway[J].Int J Clin Exp Med,2015,8(9):15801-15807.
[12]WANG X,CHEN J,WANG H,et al.Memantine can reduce ethanolinduced caspase-3 activity and apoptosis in H4 cells by decreasing intracellular calcium[J].J Mol Neurosci,2017,62(3-4):402-411.
[13]SERGEEVA T F,SHIRMANOVA M V,ZLOBOVSKAYA O A,et al.Relationship between intracellular pH,metabolic co-factors and caspase-3 activation in cancer cells during apoptosis[J].Biochim Biophys Acta,2017,1864(3):604-611.
[14]仇志富,吴晓光,孟杰,等.补阳还五汤提取物灌胃对脑出血大鼠脑组织中PI3K、AKT、Caspase-3表达的影响[J].山东医药,2016,56(9):29-31.
[15]BAO RK,ZHENG S F,WANG X Y.Selenium protects against cadmium-induced kidney apoptosis in chickens by activating the PI3K/AKT/Bcl-2 signaling pathway[J].Environ Sci Pollut Res Int,2017,24(25):20342-20353.
[16]WANG C,WEI Z,JIANG G,et al.Neuroprotective mechanisms of miR-124 activating PI3K/Akt signaling pathway in ischemic stroke[J].Exp Ther Med,2017,13(6):3315-3318.
[17]XI H,FAN X,ZHANG Z,et al.Bax and Bcl-2 are involved in the apoptosis induced by local testicular heating in the boar testis[J].Reprod Domest Anim,2017,52(3):359-365.
[18]YANG Z,GENG Y,YAO Z,et al.Spatiotemporal expression of Bcl-2/Bax and neural cell apoptosis in the developing lumbosacral spinal cord of rat fetuses with anorectal malformations[J].Neurochem Res,2017,42(11):3160-3169.
[19]LIN C H,WU M R,LI C H,et al.Editor's highlight:Periodic exposure to smartphone-mimic low-luminance blue light induces retina damage through Bcl-2/BAX-dependent apoptosis[J].Toxicol Sci,2017,157(1):196-210.
[20]黎萍,郑百红,陈鹏,等.硒对病毒性心肌炎小鼠心肌细胞凋亡的抑制作用及其作用机制[J].吉林大学学报(医学版),2016,42(1):54-58.