草苁蓉多糖下调miR-302a-3p表达对缺氧/复氧诱导的心肌细胞损伤的影响  

Effect of boschniakia rossica polysaccharides on hypoxia/reoxygenation-induced cardiomyocyte damage by down-regulating miR-302a-3p expression

作　　者：陈虹[1] 黄明伟[1] 程振东[1] 陈金成 卢辉耀[1] 许朝祥[1] 吴晶莹[1] CHEN Hong;HUANG Ming-wei;CHENG Zhen-dong;CHEN Jin-cheng;LU Hui-yao;XU Chao-xiang;WU Jing-ying(Dept of Cardiology,The Second Affiliated Hospital of Fujian Medical University,Quanzhou,Fujian 362000,China;Dept of Pharmacy,The Second Affiliated Hospital of Fujian Medical University,Quanzhou,Fujian 362000,China)

机构地区：[1]福建医科大学附属二院心血管内科,福建泉州362000 [2]福建医科大学附属二院药学部,福建泉州362000

出　　处：《中国药理学通报》2023年第7期1241-1247,共7页Chinese Pharmacological Bulletin

基　　金：福建省自然科学基金资助项目(No.2020J01224)。

摘　　要：目的探讨草苁蓉多糖(boschniakia rossica polysaccharides,BRPS)对缺氧/复氧(hypoxia/reoxygenation,H/R)诱导的心肌细胞损伤的影响及其作用机制。方法H/R诱导大鼠心肌细胞H9c2建立细胞损伤模型,不同剂量的BRPS处理H9c2细胞;酶联免疫吸附剂测定(enzyme-linked immunosorbent assay,ELISA)法检测丙二醛(malondialdehyde,MDA)的水平和超氧化物歧化酶(superoxide dismutase,SOD)、谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)的活性;流式细胞术检测凋亡率;qRT-PCR检测miR-302a-3p的表达量;anti-miR-NC、anti-miR-302a-3p分别转染至H9c2细胞后进行H/R处理,miR-NC、miR-302a-3p mimics分别转染至H9c2细胞后加入100 mg·L^(-1)BRPS处理24 h,进行H/R处理;Western blot检测Bcl-2、Bax蛋白表达;线软件预测和双荧光素酶报告实验检测miR-302a-3p和LRP8的靶向关系;Western blot检测低密度脂蛋白受体相关蛋白8(LRP8)的表达量。结果BRPS降低H/R诱导的H9c2细胞中MDA的水平、miR-302a-3p的表达量、细胞凋亡率、Bax表达(P<0.05),提高SOD、GSH-Px、Bcl-2(P<0.05);转染anti-miR-302a-3p后,MDA的水平、细胞凋亡率、Bax表达下调(P<0.05),SOD、GSH-Px、Bcl-2上调(P<0.05);转染miR-302a-3p mimics后,MDA的水平、细胞凋亡率和Bax蛋白水平升高(P<0.05),SOD、GSH-Px和Bcl-2降低(P<0.05)。miR-302a-3p靶向调节LRP8,且BRPS通过下调miR-302a-3p上调LRP8基因的表达。结论BRPS通过调节miR-302a-3p/LRP8轴减轻H/R诱导的心肌细胞损伤。Aim To explore the effect of boschniakia rossica polysaccharides(BRPS)on cardiomyocyte damage induced by hypoxia/reoxygenation(H/R)and its possible mechanism.Methods H/R was used to induce rat cardiomyocyte H9c2 to establish a cell injury model,and different doses of BRPS were used to treat H9c2 cells.ELISA method was used to detect the level of MDA and the activity of SOD and GSH-Px.Flow cytometry was used to detect the rate of apoptosis.qRT-PCR was used to detect the expression of miR-302a-3p.anti-miR-NC and anti-miR-302a-3p were respectively transfected into H9c2 cells and then subjected to H/R treatment.miR-NC and miR-302a-3p mimics were respectively transfected into H9c2 cells and treated with 100 mg·L^(-1) BRPS for 24 h,and then underwent H/R treatment.Western blot was used to detect the expression of Bcl-2 and Bax protein.The downstream target genes of miR-302a-3p were predicted by online software,and the targeting relationship between miR-302a-3p and LRP8 was detected by dual luciferase reporting assay.Western blot was used to measure the mRNA and protein level of low-density lipoprotein(LDL)receptor-related protein 8(LRP8)respectively.Results BRPS could reduce the level of MDA,the expression of miR-302a-3p,the rate of apoptosis and the protein level of Bax(P<0.05),while enhanced the activities of SOD,GSH-Px and the protein level of Bcl-2(P<0.05)in H9c2 cells induced by H/R.After transfection with anti-miR-302a-3p,the level of MDA,the rate of apoptosis and the protein level of Bax all decreased(P<0.05),while the activities of SOD,GSH-Px and the protein level of Bcl-2 increased(P<0.05).After transfection of miR-302a-3p mimics,the level of MDA,the rate of apoptosis and the protein level of Bax increased(P<0.05),while the activities of SOD,GSH-Px and the protein level of Bcl-2 decreased(P<0.05).LRP8 was a target of miR-302a-3p and BRPS upregulated LRP8 expression via downregulating miR-302a-3p level.Conclusions BRPS could inhibit the oxidative stress and apoptosis of cardiomyocytes by regulating the miR-302

关 键 词：草苁蓉多糖 缺氧/复氧 大鼠心肌细胞H9c2 miR-302a-3p 氧化应激 细胞凋亡 

分 类 号：R-332[医药卫生] R284.1R329.25R329.411R349.1