miR-142-3p在感染后肠易激综合征中的表达及意义

Expression and significance of miR-142-3p in post-infectious irritable bowel syndrome

目的探讨微RNA(miR)-142-3p在感染后肠易激综合征(PI-IBS)中的表达及意义,进一步阐明miR-142-3p调控PI-IBS发生发展的分子机制。方法使用脂多糖(LPS)刺激后,在大鼠肠上皮细胞上考察miR-142-3p对高迁移率族蛋白B1(HMGB1)-Toll样受体4(TLR4)靶基因的作用;通过TargetScan预测软件分析发现HMGB1是miR-142-3p的靶基因,之后采用旋毛虫感染建立PI-IBS模型,原代肠上皮细胞进行小干扰RNA(siRNA)转染实验,提取肠上皮细胞总RNA,采用实时荧光定量PCR检测,检测细胞中miR-142-3p、HMGB1和TLR4的mRNA表达水平,考察miR-142-3p对HMGB1-TLR4靶基因的作用,并进一步验证HMGB1在miR-142-3p抗炎中的介导作用。结果炎症基因NOD样受体热蛋白结构域相关蛋白3(NLRP3)、IL-1β、IL-6、IL-18和TNF-α均为HMGB1-TLR4的靶基因。使用LPS刺激后,施加miR-142-3p大大抑制了HMGB1-TLR4靶基因(NLRP3、IL-1β、IL-6、IL-18和TNF-α)的表达(P均<0.01)。使用siRNA对肠上皮细胞中的HMGB1靶向敲低后,细胞中的HMGB1表达降低超过80%,HMGB1-TLR4靶基因(NLRP3、IL-1β、IL-6、IL-18和TNF-α)的表达均无明显变化(P均>0.05)。结论miR-142-3p在肠上皮细胞中具有抗炎作用,其抗炎作用依赖于HMGB1。

Objective To explore the expression and significance of miR-142-3 p in post-infectious irritable bowel syndrome(PI-IBS),and to further clarify the molecular mechanism of miR-142-3 p in regulating the incidence and development of PI-IBSMethods After stimulation using the lipopolysaccharide(LPS),the effect of miR-142-3 p on high mobility group box 1(HMGB1)-toll-like receptor 4(TLR4)target genes was investigated in the rat intestinal epithelial cells.TargetScan prediction software analysis showed that HMGB1 was a target gene of miR-142-3 p.Subsequently,the PI-IBS model was established using trichinella infection.Primary intestinal epithelial cells were subjected to the siRNA transfection assay.Total RNA was extracted from intestinal epithelial cells.The expression levels of miR-142-3 p,HMGB1 and TLR4 mRNA in cells were quantitatively determined by real-time PCR.The effect of miR-142-3 p on the HMGB1-TLR4 target genes was evaluated.The mediating role of HMGB1 in the anti-inflammation of miR-142-3 p was further validated.Results Inflammatory genes NLRP3,IL-1β,IL-6,IL-18 and TNF-αwere all the target genes of HMCB1-TLR4.After the LPS stimulation,administration of miR-142-3 p significantly inhibited the expression levels pf HMGB1-TLR4 target genes(NLRP3,IL-1β,IL-6,IL-18 and TNF-α)(all P<0.01).After the targeted knockdown of HMCB1 in intestinal epithelial cells using siRNA,the HMGB1 expression level in the cells was down-regulated by more than 80%,whereas no significant changes were observed in the expression levels of HMGB1-TLR4 target genes(NLRP3,IL-1β,IL-6,IL-18 and TNF-α)(all P>0.05).Conclusion miR-142-3 p plays an anti-inflammatory role in intestinal epithelial cells,which is dependent on HMGB1.