连翘苷通过APPL1对缺氧/复氧诱导心肌细胞损伤的影响  

Effect of Phillyrin on Cardiomyocyte Injury Induced by Hypoxia/Reoxygenation through APPL1

作　　者：刘晓东 闫业军[1] LIU Xiao-dong;YAN Ye-jun(Department of Cardiology,Baoji Center Hospital,Baoji 721000,China)

机构地区：[1]陕西省宝鸡市中心医院心内科,宝鸡721000

出　　处：《微循环学杂志》2020年第2期12-19,共8页Chinese Journal of Microcirculation

摘　　要：目的:研究连翘苷通过转接蛋白APPL1对缺氧/复氧(H/R)诱导的心肌细胞氧化应激和凋亡的影响。方法:将大鼠心肌细胞H9C2分为Con组、H/R组、H/R+连翘苷低剂量组、H/R+连翘苷中剂量组、H/R+连翘苷高剂量组、H/R+pcDNA组、H/R+pcDNA-APPL1组、H/R+连翘苷+si-NC组和H/R+连翘苷+si-APPL1组。ELISA检测丙二醛(MDA)含量、超氧化物歧化酶(SOD)活性和谷胱甘肽过氧化物酶(GSH-Px)活性;流式细胞术检测心肌细胞凋亡率;蛋白质印迹法(Western blot)检测B细胞淋巴瘤/白血病-2(Bcl-2)、Bcl-2相关X蛋白(Bax)及APPL1蛋白;实时荧光定量PCR(qPCR)检测APPL1 mRNA表达。采用单因素方差分析和多重两两比较t检验进行统计学分析。结果:与Con组比较,H/R组心肌细胞MDA含量、凋亡率和Bax蛋白表达明显增加,SOD、GSH-Px活性、Bcl-2蛋白水平、APPL1 mRNA和蛋白表达显著降低(均P<0.01)。与H/R组比较,H/R+连翘苷各剂量组心肌细胞MDA含量、凋亡率、Bax蛋白表达明显减少,SOD、GSH-Px活性、Bcl-2蛋白水平、APPL1 mRNA和蛋白表达显著升高(均P<0.01)。连翘苷各剂量组APPL1 mRNA和蛋白表达高于H/R组(P<0.01)。与H/R+pcDNA组比较,H/R+pcDNA-APPL1组心肌细胞SOD和GSH-Px活性、Bcl-2蛋白水平显著升高,MDA含量、凋亡率、Bax蛋白表达显著降低(均P<0.01)。H/R+连翘苷+si-APPL1组与H/R+连翘苷+si-NC组比较,心肌细胞中MDA含量、凋亡率、Bax蛋白显著升高,SOD和GSH-Px活性、Bcl-2蛋白水平明显降低(P<0.05)。结论:连翘苷可能通过提高H/R心肌细胞的APPL1表达实现对H/R心肌细胞损伤的治疗作用。Objective:To investigate whether Phillyrin affects myocardial cell injury induced by hypoxia/reoxygenation(H/R)through APPL1(adaptor protein containing PH domain,PTB domain and leucine zipper motif 1).Method:Rat cardiomyocytes H9C2 were divided into Con group,H/R group,H/R+Phillyrin-low group,H/R+Phillyrin-middle group,H/R+Phillyrin-high group,H/R+pcDNA group,H/R+pcDNA-APPL1 group,H/R+Phillyrin+si-NC group,H/R+Phillyrin+si-APPL1 group.MDA content,SOD activity,GSH-Px activity were detected by ELISA Apoptosis of cardiomyocytes was analyzed by flow cytometry,Bcl-2,Bax and APPL1 protein expression were detected by Western blot,APPL1 mRNA expression was detected by qPCR.Results:Compared with Con group,MDA content,apoptosis rate and Bax protein expression in H/R treated cardiomyocytes were significantly increased,SOD,GSH-Px activity,Bcl-2 protein level,APPL1 mRNA and protein expression were obviously decreased(P<0.05).Compared with H/R group,MDA content,apoptosis rate and Bax protein expression in H/R+Phillyrin-low group,H/R+Phillyrin-middle group,H/R+Phillyrin-high group of cardiomyocytes greatly decreased,while SOD,GSH-Px activity,Bcl-2 protein level,APPL1 mRNA and protein expression were remarkably increased(P<0.05).Overex-pression of APPL1 clearly increased APPL1 protein,SOD activity,GSH-Px activity and Bcl-2 protein level in H/R injured cardiomyocytes,and evidently decreased MDA content,apoptosis rate and Bax protein expression(P<0.05).Inhibition of APPL1 expression reversed the effect of Phillyrin on APPL1 protein,MDA content,SOD activity,GSH-Px activity,apoptosis rate,Bcl-2 protein and Bax protein levels in H/R-induced cardiomyocytes(P<0.05).Conclusion:Phillyrin protects cardiomyocyte from injury induced by hypoxia/reoxygenation by regulating APPL1.

关 键 词：连翘苷 APPL1 缺氧/复氧 心肌细胞 氧化应激 凋亡 

分 类 号：R319[医药卫生—基础医学]