肥大细胞microRNA-126表达差异对白介素-4分泌影响观察

Expression differences of micro RNA-126 in mast cell and its influence on release of IL-4

目的观察micro RNA(mi R)-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mi R-126的表达差异。采用RT-PCR检测小鼠肥大细胞株(P815)中mi R-126的变化。将P815细胞分别转染mi R-126阴性对照序列(阴性对照组)及mi R-126抑制物(mi R-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mi R-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mi R-126表达量增加(P<0.05)。而mi R-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 mi R-126在哮喘患儿外周血表达升高。下调mi R-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。

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 ceils 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 supernatants 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. Down- regulation of miR-126 may suppress IL-4 release from mast cells by reducing the expression of phosphorylated P38.