KLF5与NF-κB p50相互作用促进高糖诱导的血管平滑肌细胞炎症  

Interaction between KLF5 and NF-κB p50 promotes high glucose-induced VSMCs inflammation

作　　者：张曼莉[1] 张曼娜 佟飞[1] 温进坤 ZHANG Man-lii;ZHANG Man na;TONG Fei;WEN Jin-kun(Department of Emergency,the Second Hospital of Hebei Medical University,Shi.jiazhuang 050000 China;Department of Clinical Laboratory,the Second Hospital of Hebei Medical University,Shijiazhuang 050000,China;Department of Biochemistry and Molecular Biology,School of Basic Medical Sciences,Hebei Medical University,Shijiazhuang 050017,China)

机构地区：[1]河北医科大学第二医院急诊科,河北石家庄050000 [2]河北医科大学第二医院检验科,河北石家庄050000 [3]河北医科大学基础医学院生物化学与分子生物学教研室,河北石家庄050017

出　　处：《河北医科大学学报》2018年第12期1371-1375,1379,共6页Journal of Hebei Medical University

基　　金：河北省医学科学研究重点课题(20180314)

摘　　要：目的探讨锌指转录因子Krüppel样因子5(Krüppel-like factor 5,KLF5)在高糖诱导的血管平滑肌细胞(vascular smooth muscle cells,VSMCs)炎症中的作用及机制。方法体外培养VSMCs,实时定量反转录聚合酶链反应(quantitative real-time polymerase chain reaction,qRT-PCR)、蛋白质印迹法(Western blot)检测高糖对VSMCs炎症基因肿瘤坏死因子α(tumor necrosis factor-α,TNF-α)、KLF5和核因子κB p50(nuclear factorκB p50,NF-κB p50)表达的影响。用小干扰RNA(small interfering RNA,siRNA)内源性敲低KLF5,Western blot检测VSMCs中TNF-α的表达。免疫共沉淀(co-immunoprecipitation,CoIP)检测VSMCs中高糖对KLF5和NF-κB p50相互作用的影响。结果qRT-PCR和Western blot结果显示,在mRNA和蛋白质水平,高糖显著上调炎症基因TNF-α、KLF5和NF-κB p50的表达,呈剂量及时间依赖性(P<0.05)。用siRNA内源性敲低KLF5后,高糖不能上调炎症基因TNF-α的表达。CoIP结果显示,高糖显著增强KLF5和NF-κB p50的相互作用。结论 KLF5通过与NF-κB p50相互作用促进高糖诱导的VSMCs炎症。Objective To explore whether Krüppel-like factor 5（KLF5）was involved and the mechanism in high glucose-induced inflammation in vascular smooth muscle cells（VSMCs）.Methods VSMCs were cultured with different concentrations of glucose or treated with high glucose for various times,quantitative real-time polymerase chain reaction（qRT-PCR）and Western blot were performed to examine the expression of tumor necrosis factor-α（TNF-α）,KLF5 and NF-κB p50.Endogenous KLF5 was knocked down by transfecting VSMCs with KLF5-specific siRNA or nonspecific siRNA,and then treated the cells with or without high glucose,Western blot was performed to examine the expression of TNF-α.VSMCs were cultured in medium containing 5.5 mmol/L or 25 mmol/L glucose.The cell lysates were immunoprecipitated With anti-KLF5 or anti-NF-κB p50 antibody,and the resulting precipitates were analyzed by Western blot using anti-NF-κB p50 or anti-KLF5 antibody.Results In mRNA and protein levels,high glucose increased the level of TNF-α,KLF5 and NF-κB p50 in a dose-and time-dependent manner in VSMCs（P〈0.05）.When knocked down endogenous KLF5,the results of Western blot suggested that high glucose-induced TNF-αexpression was mediated by KLF5.The results of co-immunoprecipitation experiments showed that exposure of high glucose increased the interactions of KLF5 and NF-κB p50.Conclusion KLF5 and NF-κB p50 interactions induced by high glucose cooperatively induce pro-inflammatory gene expression.

关 键 词：血管平滑肌细胞 Krüppel样因子5 核因子ΚB P50 

分 类 号：R513[医药卫生—内科学]