肥大细胞microRNA-126表达差异对白介素-4分泌影响观察

摘要

Objective To observe the expression features of peripheral blood microRNA (miR)-126 in children with asthma and to investigate the influence of miR-126 downregulation on release of interleukin(IL)-4 in mast cells and its possible mechanism.Methods The peripheral blood of healthy subjects (control group) and astham children (asthma group) were enrolled.The asthma group included 6 children who were treated in Nanjing Children's Hospital from September to December of 2015.The levels of serum miR-126 were measured in both groups by quantitative real-time polymerase chain reaction (RT-PCR).After stimulation with lipopolysaccharide (LPS) on P815,miR-126 expression was detected by RT-PCR.P815 were transfected with miR-126 negative control (negative control group) and miR-126 inhibitor (miR-126 inhibitor group) respectively.The levels of IL-4 in supematants were detect by ELISA,and the levels of phosphorylated P38 were detected by Western Blot in two groups.Results The levels of peripheral blood miR-126 in asthma group were significantly higher than those in control group (P ＜ 0.05).In LPS stimulated P815,miR-126 expression increased (P ＜ 0.05).The levels of IL-4 and phosphorylated P38 in the miR-126 inhibitor group were lower than those in the negative control group,with statistical significance (P ＜ 0.05).Conclusion The expression of peripheral blood miR-126 increases in asthma children.Downregulation of miR-126 may suppress IL-4 release from mast cells by reducing the expression of phosphorylated P38.%目的 观察microRNA (miR)-126在哮喘患儿外周血中的表达特点,检测其下调对肥大细胞白介素(IL)-4分泌的影响及可能机制.方法 以2015年9～12月在南京医科大学附属南京儿童医院哮喘专科门诊就诊的哮喘患儿6例作为观察组,收集同期健康体检儿童作为对照组.应用实时荧光定量聚合酶链反应(RT-PCR)方法检测2组外周血miR-126的表达差异.采用RT-PCR检测小鼠肥大细胞株(P815)中miR-126的变化.将P815细胞分别转染miR-126阴性对照序列(阴性对照组)及miR-126抑制物(miR-126下调组),应用酶联免疫吸附实验(ELISA)检测2组细胞上清IL-4水平,应用免疫印迹法(Western Blot)验证2组细胞磷酸化P38水平.结果 哮喘组外周血miR-126水平显著高于对照组(P＜0.05).在细胞水平的研究发现,脂多糖(LPS)刺激后P815中miR-126表达量增加(P＜0.05).而miR-126下调组的IL-4及磷酸化P38水平均低于阴性对照组,差异均有统计学意义(P＜0.05).结论 miR-126在哮喘患儿外周血表达升高.下调miR-126可能通过减少磷酸化P38蛋白表达抑制肥大细胞IL-4分泌.