H_(2)S减少H_(2)O_(2)诱发H9c2心肌细胞内质网应激的机制研究  

A study of mechanism for H_(2)S reducing endoplasmic reticulum stress induced by H_(2)O_(2) in H9c2 cardiomyocytes

作　　者：武丹[1] 祝德秋[1] WU Dan;ZHU Deqiu(Department of Pharmacy,Affiliated Tongji Hospital,Tongji University,Shanghai 200065,CHINA)

机构地区：[1]同济大学附属同济医院药剂科,上海200065

出　　处：《江苏医药》2021年第12期1198-1201,共4页Jiangsu Medical Journal

基　　金：上海市临床药学重点专科建设项目(沪卫计药政[2018]8号)。

摘　　要：目的探讨硫化氢(H_(2)S)减少过氧化氢(H_(2)O_(2))诱发H9c2心肌细胞内质网应激的作用机制。方法H_(2)O_(2)150μmol/L诱发制备H9c2心肌细胞内质网应激模型后分为三组,A组为模型对照组,B组和C组分别采用硫氢化钠25 μmol/L和S-炔丙基半胱氨酸25μmol/L处理;另设空白对照(D)组。CCK-8法检测细胞生存率,Hoechst染色法检测凋亡细胞,荧光分析法检测Caspase-12活性,Western blot检测转录因子C/EBP同源蛋白(CHOP)、免疫球蛋白结合蛋白(BiP)、钙/钙调蛋白依赖性蛋白激酶Ⅱ(CaMKⅡ)、受磷蛋白(PLN)和心肌肌浆网钙ATP酶2a(SERCA2a)表达,免疫共沉淀法检测CaMKⅡ与PLN以及SERCA2a与PLN的结合。结果A组细胞存活率低于D组,而B组和C组高于A组(P<0.05)。A组凋亡细胞数多于D组,而B组和C组均少于A组(P<0.05)。A组Caspase-12活性、PLN磷酸化水平、CHOP、BiP、CaMKⅡ以及CaMKⅡ与PLN结合表达均高于D组,B组和C组则均低于A组(P<0.05)。A组SERCA2a与PLN结合的表达低于D组,B组和C组则均高于A组(P<0.05)。结论H_(2)S可以通过CaMKⅡ/PLN/SERCA2a途径减少内质网应激,对H_(2)O_(2)诱导的心肌细胞损伤发挥保护作用。Objective To investigate the mechanism for hydrogen sulfide(H_(2)S)reducing endoplasmic reticulum stress induced by hydrogen peroxide(H_(2)O_(2))in H9c2 cardiomyocytes.Methods The endoplasmic reticulum stress models of H9c2 cardiomyocytes were prepared by H_(2)O_(2)150 μmol/L,which were divided into three groups of A(model control),B(treated with sodium hydride 25 μmol/L),and C(treated with S-propargyl cysteine 25 μmol/L).Blank control group(group D)was set up.The survival rate of the cells was detected by CCK-8,apoptotic cells were detected by Hoechst staining,and Caspase-12 activity was detected by fluorescence analysis.The expressions of transcription factor C/EBP homologous protein(CHOP),binding immunoglobulin protein(BiP),calcium/calmodulin-dependent protein kinase Ⅱ(CaMK Ⅱ),phosphoprotein(PLN)and cardiac sarcoplasmic reticulum calcium-ATPase 2a(SERCA2a)were detected by Western blot.The binding of CaMK Ⅱ to PLN and binding of SERCA2a to PLN were detected by immunoprecipitation.Results The cell survival rate in group A was lower than that in group D,which in groups of B and C was higher than that in group A(P<0.05).The number of apoptotic cells,in group A was more than that in group D,which in groups of B and C was less than that in group A(P<0.05).The activity of Caspase-12,PLN phosphorylation level,and the expressions of CHOP,BiP,CaMK Ⅱ,CaMK Ⅱ bound with PLN in group A were higher than those in group D,which in groups of B and C were lower than those in group A(P<0.05).The expression of SERCA2a bound with PLN in group A was lower than that in group D,which in groups of B and C was higher than that in group A(P<0.05).Conclusion H_(2)S can reduce the endoplasmic reticulum stress through the CaMK Ⅱ/PLN/SERCA2a pathway,by which H_(2)S plays a protective role in H_(2)O_(2)-induced myocardial cell injury.

关 键 词：硫化氢 内质网应激 钙/钙调蛋白依赖性蛋白激酶Ⅱ 受磷蛋白 心肌肌浆网钙三磷酸腺苷酶2a 

分 类 号：R542[医药卫生—心血管疾病]