摘要
目的探讨过表达miR-203a-3p对脂多糖(LPS)致大鼠急性肺损伤后肺纤维化的影响及其机制。方法采用多次小剂量注射LPS制备大鼠急性肺损伤后肺纤维化模型,并将造模成功的大鼠随机分成模型组(Model组)、agomir阴性对照组(agomir-NC组)和miR-203a-3p agomir组(agomir组),另设对照组(Control组),每组15只。造模前1 d及造模开始后1周,给予尾静脉注射miR-203a-3p agomir干预。造模2周后,测定肺组织湿/干比重(W/D)及肺组织羟脯胺酸(Hyp)含量;HE染色和Masson染色观察肺组织病理变化及肺纤维化程度;ELISA法检测各组大鼠肺泡灌洗液(BALF)中白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)和肿瘤坏死因子-α(TNF-α)水平;qRT-PCR检测大鼠肺组织中miR-203a-3p和卵泡抑素样蛋白1(Fstl1)mRNA表达水平;Western blot检测大鼠肺组织中Fstl1蛋白表达水平;双荧光素酶报告系统检测miR-203a-3p和Fstl1的靶向关系。结果与Control组比较,Model组大鼠肺损伤及肺纤维化程度较为严重,肺组织W/D值及Hyp含量增加(P<0.05),BALF中IL-1β、IL-6和TNF-α水平升高(P<0.05),肺组织中miR-203a-3p表达水平降低(P<0.05),而Fstl1 mRNA和蛋白表达水平升高(P<0.05)。与Model组比较,agomir组大鼠肺损伤及纤维化得到改善,肺组织W/D值及Hyp含量降低(P<0.05),BALF中IL-1β、IL-6和TNF-α水平降低(P<0.05),肺组织中miR-203a-3p表达水平升高(P<0.05),而Fstl1 mRNA和蛋白表达水平降低(P<0.05)。荧光素酶报告基因实验证实,Fstl1是miR-203a-3p靶基因。结论miR-203a-3p过表达可改善LPS诱导的急性肺损伤后肺纤维化,其机制可能与靶向下调Fstl1表达有关。
Objective To investigate the effects of miR-203a-3p overexpression on pulmonary fibrosis after acute lung injury induced by lipopolysaccharide(LPS)in rats and its possible mechanism.Methods The rat model of pulmonary fibrosis after acute lung injury was established by multiple low-dose injections of LPS.The successfully modeled rats were randomly divided into model group,agomir-negative control group(agomir-NC group)and miR-203a-3p agomir group(agomir group),and another control group was set,with 15 rats in each group.One day before modeling and one week after modeling,miR-203a-3p agomir was injected by caudal vein for intervention.Two weeks after modeling,the wet/dry specific gravity(W/D)and hydroxyprolinic acid(Hyp)content of lung tissue were determined.The pathological changes and the degree of pulmonary fibrosis of lung tissue were observed by HE staining and Masson staining.The levels of interleukin-1β(IL-1β),interleukin-6(IL-6)and tumor necrosis factor-α(TNF-α)in broncholveolr lavage fluid(BALF)were determined by ELISA.The expression levels of miR-203a-3p and follicle-statin like 1(Fstl1)mRNA in the lung tissues of rats were detected by qRT-PCR.The protein expression level of Fstl1 in lung tissue of rats was detected by Western blot.The targeting relationship between miR-203a-3p and Fstl1 was detected by dual luciferase reporting system.Results Compared with control group,the degree of lung injury and pulmonary fibrosis in model group were relatively serious,the W/D value and Hyp content of lung tissue increased(P<0.05),the levels of IL-1β,IL-6 and TNF-αin BALF increased(P<0.05),the expression of miR-203a-3p in lung tissue decreased(P<0.05),while the mRNA and protein expression levels of Fstl1 increased(P<0.05).Compared with model group,the degree of lung injury and pulmonary fibrosis were improved in agomir group,the W/D value and Hyp content of lung tissue decreased(P<0.05),the levels of IL-1β,IL-6 and TNF-αin BALF decreased(P<0.05),the expression of miR-203a-3p in lung tissue increased(P<0.05),while the mRNA and protein expression levels of Fstl1 decreased(P<0.05).The luciferase reporter gene assay confirmed that Fstl1 was the target gene of miR-203A-3p.Conclusion Overexpression of miR-203a-3p improves pulmonary fibrosis after LPS-induced acute lung injury,and the mechanism may be related to the targeted down-regulation of Fstl1 expression.
作者
任亦频
周厚荣
李亚骐
黄佳
Ren Yipin;Zhou Hourong;Li Yaqi;Huang Jia(Dept of Emergency Medicine,Guizhou Provincial People′s Hospital,Guiyang 550002;Dept of General Practice,Guizhou Provincial People′s Hospital,Guiyang 550002)
出处
《安徽医科大学学报》
CAS
北大核心
2022年第4期563-568,共6页
Acta Universitatis Medicinalis Anhui
基金
贵州省科技计划项目(编号:黔科合LH字[2016]7153号)。
