敲低长链非编码RNA H19(lncRNA H19)促进人结肠细胞自噬抑制脂多糖诱导的凋亡

Knock-down of long noncoding RNA H19(lncRNA H19)promotes human colorectal cell autophagy and inhibits cell apoptosis induced by lipopolysaccharide

目的 探讨lncRNA H19在溃疡性结肠炎(UC)患者中的表达水平及其在UC中的作用。方法 收集西藏军区总医院25名UC患者和25名健康体检者的结肠黏膜标本及血清标本,采用荧光定量PCR检测lncRNA H19表达水平,并对血清标本进行受试者工作特征(ROC)曲线分析。采用脂多糖(LPS)处理建立体外HT29结肠细胞炎性模型,并采用荧光定量PCR检测lncRNA H19在结肠细胞的表达。通过慢病毒技术构建lncRNA H19低表达的HT29细胞株,噻唑蓝(MTT)法检测HT29细胞存活率,流式细胞术检测细胞凋亡率,Western blot法检测凋亡和自噬蛋白表达情况。采用自噬激动剂雷帕霉素处理后,MTT法观察对HT29细胞存活的影响。结果 lncRNA H19在UC患者结肠黏膜组织和血清中高表达,ROC曲线下面积为0.786。LPS刺激后,HT29细胞lncRNA H19表达显著升高,并与刺激时间正相关。敲低lncRNA H19可促进HT29细胞存活并抑制细胞凋亡,促进细胞自噬水平增加;雷帕霉素处理后,细胞存活率明显升高。结论 敲低lncRNA H19促进结肠细胞自噬抑制LPS诱导的结肠细胞凋亡并促进其存活。

Objective To investigate the expression levels of lncRNA H19 in ulcerative colitis(UC)patients and its role in UC.Methods Colonic mucosa and serum samples were collected from 25 UC patients and 25 healthy individuals at the General Hospital of Xizang Military Region.The expression levels of lncRNA H19 were detected,and the receiver operating characteristic(ROC)curve analysis was performed using serum samples.An in vitro inflammatory model was established in HT29 colorectal cells under lipopolysaccharide(LPS)stimulation,and the expression levels of lncRNA H19 were observed in HT29 cells through fluorescence quantitative PCR.HT29 cells with downregulated lncRNA H19 was constructed using lentivirus-mediated shRNA.The effect of lncRNA H19 on cell survival was analyzed through MTT assay.Cell apoptosis was detected by flow cytometry,and the protein expression levels of apoptosis and autophagy markers were analyzed through Western blot.After treatment with rapamycin,the survival of HT29 cells was observed by MTT assay.Results lncRNA H19 was highly expressed in the colonic mucosa and serum samples of UC patients with the ROC area being 0.786.Following LPS stimulation,the expression levels of lncRNA H19 was significantly increased in a time-dependent manner.Downregulation of lncRNA H19 can promote cell survival,inhibit cell apoptosis and increase autophagy level in HT29 cells.Treatment with rapamycin significantly increased the cell survival rate.Conclusion Knock-down of lncRNA H19 increases autophagy levels,inhibits LPS-induced apoptosis and promotes the survival of colon cells.