miR-511-3p调节小儿咳嗽变异性哮喘患者巨噬细胞极化和过敏性炎症的机制

Mechanisms of miR⁃511⁃3p regulating macrophage polarization and allergic inflammation in children with cough variant asthma

目的探究miR-511-3p调节小儿咳嗽变异性哮喘(CVA)患者巨噬细胞极化和过敏性和M2细胞内TNF-α和TGF-β的分泌量;细胞转染法构建miR-511-3p表达细胞,RT-PCR和Western blot检测M1巨噬细胞相关标志物蛋白表达;用氢氧化铝和卵蛋白致敏并用的卵蛋白激活法构建CVA小鼠模型,用流式分选法分离小鼠巨噬细胞,用ELSA法检测灌洗液中巨噬细胞极化相关标志因子TNF-α和TGF-β及IRF3表达分泌量;用RT-PCR法和Western Blot法检测CVA患者血浆样本中miR-511-3p的转录水平。结果通过miR-511-3p转染发现,miR-511-3p过表达时,M1型巨噬细胞中TNF-α表达减少,CCL17表达增多(P <0.05);CVA小鼠模型中TNF-α和IRF3磷酸化水平减少,TGF-β增多(P <0.05);同时与非哮喘患者相比,CVA患者miR-511-3p表达减少(P <0.05)。结论 miR-511-3p可调节小儿CVA患者机体IRF3磷酸化水平,促使巨噬细胞向M2方向极化,抑制体内过敏性炎症。

Objective To explore the mechanism of miR⁃511⁃3p regulating macrophage polarization and allergic inflammation in children with cough variant asthma.Methods After THP⁃1 cells were induced to form M1 and M2 macrophages by LPS and IL⁃4 respectively,the secretion of TNF⁃αand TGF⁃βin M1 and M2 cells was detected by ELSA;mir⁃511⁃3p expression cells were constructed by cell transfection method,RT⁃PCR and Western blot were used.The expression of M1 macrophage related marker protein was detected by blot;the CVA mouse model was established by ovalbumin activation method sensitized by aluminum hydroxide and ovalbumin;the macrophages were isolated by flow cytometry;the expression and secretion of macrophage polarization related markers TNF⁃α,TGF⁃βand IRF3 in lavage fluid were detected by ELSA;RT⁃PCR and Western blot were used The transcription level of mir⁃511⁃3p in plasma samples of patients with CVA was detected by blot.Results By transfection of mir⁃511⁃3p,we found the relative expression of TNF⁃αdecreased and CCL17 increased in M1 macrophageswhen mir⁃511⁃3p was overexpressed(P<0.05);the phosphorylation levels of TNF⁃αand IRF3 decreased and TGF⁃βincreased in CVA mice model(P<0.05);at the same time,compared with non⁃asthma patients,the expression of mir⁃511⁃3p decreased in CVA patients(P<0.05).Conclusion MiR⁃511⁃3p could regulate the phosphorylation level of IRF3 in children with cough variant asthma,promote the polarization of macro⁃phages in the direction of M2 and inhibit allergic inflammation in the body.