lncRNA XIST靶向miR-150对LPS诱导的小鼠肺上皮MLE-12细胞凋亡和炎症因子分泌的影响  

The effect and mechanism of lncRNA XIST targeting miR-150 on LPS-induced apoptosis and secretion of inflammatory factors in mouse lung epithelial MLE-12 cells

作　　者：王慧敏[1] 蔡施霞[1] 周震[1] 隋娜[1] WANG Huimin;CAI Shixia;ZHOU Zhen;SUI Na(Departments of Critical Care Medicine, The Affiliated Hospital of Qingdao University, Qingdao 266003, China)

机构地区：[1]青岛大学附属医院重症医学科,山东青岛266003

出　　处：《西安交通大学学报（医学版）》2021年第4期522-528,共7页Journal of Xi’an Jiaotong University（Medical Sciences）

基　　金：国家自然科学基金青年科学基金资助项目(No.81701879)。

摘　　要：目的研究lncRNA XIST靶向miR-150对脂多糖(LPS)诱导的MLE-12细胞凋亡和炎症因子分泌的影响及机制。方法小鼠肺上皮MLE-12细胞分成Control、LPS(LPS处理)、sh-NC+LPS(转染shRNA control,给予LPS处理)、sh-XIST+LPS(转染XIST shRNA,给予LPS处理)组,采用qRT-PCR方法检测XIST表达水平,采用Annexin V-FITC/PI双染法检测细胞凋亡,采用Western blotting方法检测细胞中Bax、Bcl-2蛋白表达,利用TBA法检测MDA含量,利用黄嘌呤氧化法检测SOD活性,利用DCFH-DA法检测ROS水平,利用ELISA法分析培养液上清中IL-1β、TNF-α水平。利用生物信息学软件Starbase分析XIST的可能靶基因(miR-150),然后利用荧光素酶报告系统鉴定二者的靶向关系。在MLE-12细胞中将XIST shRNA、miR-150 inhibitor共转染,然后给予LPS处理,同样测定细胞凋亡、氧化损伤指标、炎症因子分泌变化。结果与Control组比较,LPS组MLE-12细胞中XIST水平升高,细胞凋亡率和Bax蛋白表达升高,Bcl-2蛋白表达降低,ROS和MDA水平升高,SOD水平降低,细胞分泌的IL-1β、TNF-α增多;与sh-NC+LPS组比较,sh-XIST+LPSMLE-12组细胞中XIST水平降低,细胞凋亡率和Bax蛋白表达降低,Bcl-2蛋白表达升高,ROS和MDA水平降低,SOD水平升高,细胞分泌的IL-1β、TNF-α减少。下调XIST靶向促进miR-150表达。miR-150 inhibitor能够逆转XIST shRNA对LPS诱导的MLE-12细胞凋亡、氧化损伤指标、炎症因子分泌的作用。结论下调lncRNA XIST靶向miR-150可抑制LPS诱导的小鼠肺上皮MLE-12细胞凋亡和炎症因子分泌作用。Objective To study the effect and mechanism of lncRNA XIST targeting miR-150 on LPS-induced apoptosis and secretion of inflammatory factors in MLE-12 cells.Methods Mouse lung epithelial MLE-12 cells were divided into control,LPS(LPS treatment),sh-NC+LPS(shRNA control transfection,LPS treatment),and sh-XIST+LPS(XIST shRNA transfection,LPS treatment)groups.We used qRT-PCR method to analyze and detect XIST expression level,Annexin V-FITC/PI double staining method to detect cell apoptosis,Western blotting method to analyze and detect the protein expressions of Bax and Bcl-2 in cells,TBA method to detect MDA content,Xanthine oxidation method to detect SOD activity,DCFH-DA method to detect ROS level,and ELISA method to analyze the levels of IL-1βand TNF-αin the culture supernatant.We used the bioinformatics software Starbase to analyze the possible target genes of XIST(miR-150),and then the luciferase reporter system to identify the target relationship between the two.In MLE-12 cells,XIST shRNA and miR-150 inhibitor were co-transfected,and thentreated with LPS.We also measured apoptosis,oxidative damage indicators,and inflammatory factor secretion changes.Results Compared with control group,XIST level in MLE-12 cells of LPS group increased,apoptosis rate and Bax protein expression level increased,Bcl-2 protein expression level decreased,ROS and MDA level increased,SOD level decreased,and the secretion of IL-1βand TNF-αincreased.Compared with the sh-NC+LPS group,the sh-XIST+LPSMLE-12 group had decreased XIST level,decreased apoptosis rate and Bax protein expression level,and increased Bcl-2 protein expression level,decreased ROS and MDA levels,increased SOD levels,and decreased IL-1βand TNF-αsecreted by cells.Down-regulating XIST targeting promoted miR-150 expression.miR-150 inhibitor could reverse the effects of XIST shRNA on LPS-induced MLE-12 cell apoptosis,oxidative damage indicators,and inflammatory factor secretion.Conclusion Down-regulation of lncRNA XIST targeting miR-150 inhibits LPS-induced apoptosis and

关 键 词：lncRNA XIST 凋亡 炎症因子 LPS 肺上皮MLE-12细胞 

分 类 号：R735.2[医药卫生—肿瘤]