下调miR-92a通过调控PIAS3抑制ox-LDL诱导RAW264.7细胞炎症介质分泌和脂质蓄积  

Down-regulation of miR-92a inhibits ox-LDL induced secretion of inflammatory mediators and lipid accumulation in RAW264.7 cells by regulating PIAS3

作　　者：胡威威 陈丽曼 刘智芬 HU Weiwei;CHEN Liman;LIU Zhifen(Department of Geriatrics,Harrison International Peace Hospital Affiliated to Hebei Medical University,Hengshui,Hebei 053000,China)

机构地区：[1]河北医科大学附属哈励逊国际和平医院老年病一科

出　　处：《中国动脉硬化杂志》2018年第11期1119-1126,共8页Chinese Journal of Arteriosclerosis

基　　金：衡水市科技计划项目(15018)

摘　　要：目的 探讨下调miR-92a对氧化型低密度脂蛋白(ox-LDL)诱导RAW264.7细胞炎症介质分泌和脂质蓄积的影响及其机制。方法 以不同浓度(0、25、50、100 mg/L)的ox-LDL处理RAW264.7细胞24 h或以100 mg/L ox-LDL分别处理RAW264.7细胞0、12、24 h后,采用定量实时聚合酶链反应(qRT-PCR)检测ox-LDL对miR-92a表达的影响。建立miR-92a低表达的RAW264.7细胞株,并给予100 mg/L ox-LDL处理后,用qRT-PCR、油红O染色、一氧化氮(NO)荧光探针(DAF-FMDA)和Western blot检测下调miR-92a表达对ox-LDL诱导的RAW264.7细胞中炎症因子诱导型一氧化氮合酶(iNOS)、白细胞介素6(IL-6)和IL-1β表达、脂质蓄积以及信号转导子和转录激活子3(STAT3)信号通路相关蛋白p-STAT3、STAT3蛋白表达的影响。建立活化STAT3蛋白抑制分子(PIAS3)和miR-92a低表达的RAW264.7细胞株,并给予100 mg/L ox-LDL处理后,观察沉默PIAS3表达对ox-LDL作用下miR-92a低表达的RAW264.7细胞中炎症因子、脂质蓄积以及STAT3信号通路的影响。TargetScan软件预测和双荧光素酶报告实验验证miR-92a和PIAS3的靶向关系。结果 随着ox-LDL作用时间和浓度的增加,RAW264.7细胞中miR-92a表达升高;双荧光素酶报告实验验证PIAS3是miR-92a的靶基因。采用ox-LDL处理后RAW264.7细胞中miR-92a、iNOS、IL-6、IL-1β、p-STAT3表达升高,NO释放增多和细胞内脂滴升高,而PIAS3的表达下降。这种调控作用可被anti-miR-92a所抑制;而沉默PIAS3后,anti-miR-92a的这一抑制作用得以恢复。结论 下调miR-92a可通过调控PIAS3抑制ox-LDL诱导RAW264.7细胞炎症介质分泌和脂质蓄积。Aim To investigate the effect of down-regulation of miR-92 a on the secretion of inflammatory mediators and lipid accumulation induced by oxidized low density lipoprotein( ox-LDL) in RAW264.7 cells and its mechanism.Methods RAW264.7 cells were treated with ox-LDL of different concentrations( 0,25,50,100 mg/L) for 24 hours or with 100 mg/L ox-LDL for 0,12 and 24 hours respectively. Quantitative real-time polymerase chain reaction( qRT-PCR)was used to detect the effect of ox-LDL on the expression of miR-92 a. A RAW264.7 cell line with low expression of miR-92 a was established and treated with 100 mg/L ox-LDL. Effects of down-regulation of miR-92 a expression on expressions of inflammatory factors inducible nitric oxide synthase( i NOS),interleukin-6( IL-6) and IL-1β,lipid accumulation and expressions of signal transducer and activator of transcription-3( STAT3) signal pathway-related proteins p-STAT3 and STAT3 in RAW264.7 cells induced by ox-LDL,were detected by qRT-PCR,oil red O staining,nitric oxide( NO) fluorescence probe( DAF-FMDA) and Western blot. A RAW264.7 cell line with low expressions of protein inhibitor of activated STAT3( PIAS3) and miR-92 a was established and treated with 100 mg/L ox-LDL. The effects of silencing PIAS3 expression on inflammatory factors,lipid accumulation and STAT3 signaling pathway were observed in RAW264.7 cells with low expression of miR-92 a induced by ox-LDL. Target Scan software prediction and dual luciferase reporting experiment were used to verify the target relationship between miR-92 a and PIAS3. Results With the increase of the action time and concentration of ox-LDL,the expression of miR-92 a was increased in RAW264.7 cells. The dual luciferase reporting experiment confirmed that PIAS3 was the target gene of miR-92 a. After ox-LDL treatment,the expressions of miR-92 a,i NOS,IL-6,IL-1β and p-STAT3 were increased,NO release and intracellular lipid droplets were increased in RAW264.7 cells,while the expression of PIAS3 was decreased. This regulatory effect could be inhibi

关 键 词：RAW264.7细胞 氧化型低密度脂蛋白 miR-92a 炎症介质 脂质蓄积 

分 类 号：R363[医药卫生—病理学]