干扰RNA METTL14对大鼠肺成纤维细胞向肌成纤维细胞转化的影响

Effect of RNA METTL14 on transformation of lung fibroblasts into myofibroblasts in rats

目的探究干扰RNA甲基转移酶样蛋白14(methyltransferase-like 14,METTL14)在转化因子β1(transforming growth factorβ1,TGF-β1)诱导的大鼠肺成纤维细胞向肌成纤维细胞转移中的作用。方法大鼠肺成纤维细胞中构建METTL14小干扰RNA(small interfering RNA METTL14,si-METTL14)序列(si-METTL14-122、si-METTL14-300、si-METTL14-450)及对照siRNA(Negative control siRNA,si-NC)。建立TGF-β1(20 ng/mL)诱导肺成纤维细胞向肌成纤维细胞转化模型,转染si-NC及si-METTL14序列至肺成纤维细胞,检测METTL14、肌成纤维细胞标志物Ⅰ型胶原蛋白α1(collagen typeⅠalpha 1,COL1A1)、α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)及肌成纤维细胞转化负向调控因子Smad家族7号蛋白(Smad family member 7,Smad7)的mRNA和蛋白水平。结果si-METTL14有效沉默肺成纤维细胞METTL14表达水平。肌成纤维细胞转化模型组中METTL14表达显著升高(P<0.05)。与control组相比,TGF-β1诱导model组COLIA1、α-SMA表达显著升高,Smad7表达显著降低,转染si-METTL14抑制COLIA1、α-SMA表达,促进Smad7表达(P<0.05)。结论METTL14抑制TGF-β1诱导大鼠肺成纤维细胞向肌成纤维细胞转化,且可能与METTL14上调Smad7表达有关。

Objective To investigate the effect of RNA methyltransferase-like 14(METTL14)interference on transforming growth factorβ1(TGF-β1)-induced transformation of lung fibroblasts into myofibroblasts in rats.Methods Small interfering RNA(siRNA)sequences of METTL14(si-METTL14-122,si-METTL14-300,si-METTL14-450)and negative control siRNA(si-NC)were constructed in lung fibroblasts in rats.Then TGF-β1(20 ng/mL)-induced lung fibroblast to myofibroblast transformation model was established.Si-NC and si-METTL14 sequences were transfected into lung fibroblasts.Finally,mRNA and protein expression levels of METTL14,myofibroblast markers including typeⅠalpha 1(COL1A1),α-smooth muscle actin(α-SMA)and a negative regulator of myofibroblast transformation of Smad family member 7(Smad7)were detected.Results Si-METTL14 silenced the expression level of METTL14 effectively in lung fibroblasts.The expression level of METTL14 increased significantly in the model of myofibroblast transformation(P<0.05).Compared with the control group,the expression of COL1A1 andα-SMA significantly increased in the TGF-β1-induced model group,while the expression of Smad7 decreased significantly(P<0.05).Transfection of si-METTL14 could significantly inhibit the expression of COL1A1,α-SMA,and promote Smad7 expression(P<0.05).Conclusion Interference of METTL14 can inhibit TGF-β1-induced transformation of rat lung fibroblasts into myofibroblast,which may be related to up-regulating Smad7 expression.