微小核糖核酸-146a对寻常型银屑病患儿CD4＋T细胞的调控作用及其临床意义

Role of miRNA-146a in modulation of CD4＋ T cells and its clinical significance in children with psoriasis vulgaris

目的：探讨微小核糖核酸（miRNA）-146a在寻常型银屑病患儿外周血CD4＋T淋巴细胞的调控机制。方法采用实时荧光定量PCR检测2013年10月至2015年12月就诊于本院皮肤性病科门诊的寻常型银屑病患儿30例和30名正常对照儿童外周血CD4＋T淋巴细胞miRNA-146a表达水平，免疫磁珠法分选外周血CD4＋T淋巴细胞，将分离的细胞分为对照组、miRNA-146a模拟体组和miRNA-146a抑制物组，并进行细胞培养；流式细胞术检测辅助性T细胞Th1数量，免疫印迹法和荧光定量PCR分别检测IFN-γ受体α（IFN-γRα）蛋白和mRNA的表达，酶联免疫吸附试验检测血浆和体外细胞培养上清液干扰素（IFN）-γ表达水平。结果寻常型银屑病患儿外周血CD4＋T淋巴细胞miRNA-146a与血浆IFN-γ表达较正常对照组升高[（2.43±0.94）vs（1.05±0.23），（27.69±7.64）ng/L vs（9.75±2.81）ng/L]，均P〈0.01，且miRNA-146a的表达与血清IFN-γ呈正相关（r=0.837，P〈0.01）。体外CD4＋T淋巴细胞培养结果显示，与对照组比较，miRNA-146a模拟体组Th1数量、培养液上清IFN-γ水平显著增加，IFN-γRα蛋白表达降低，均P〈0.01。miRNA-146a抑制物可有效地阻断miRNA-146a对Th1细胞的调控作用。结论 miRNA-146a通过影响Th1细胞的分化和功能参与对寻常型银屑病患儿外周血CD4＋T淋巴细胞的调控，在寻常型银屑病发病过程中可能发挥着一定的作用。

Objective To investigate the mechanism by which the miRNA-146a modulates peripheral blood CD4＋T lymphocytes in children with psoriasis vulgaris. Methods Real-time quantitative reverse transcription polymerase chain reaction（RT-PCR）was used to detect the expression level of miRNA-146a in peripheral CD4＋ T lymphocytes in 30 children with psoriasis vulgaris who visited the Clinic of Dermatology in our hospital and another 30 normal control children between October 2013 and December 2015. The peripheral blood CD4＋T lymphocytes were sorted by immunomagnetic beads method. The isolated cells were divided into control group,miRNA-146a mimic group and miRNA-146a inhibitor group,and subject to cell culture. Flow cytometry analysis（FACS）was used to measure the number of T helper（Th1） cells. The expression of IFN-γRα protein and mRNA was detected by Western blotting and real-time quantitative PCR. The levels of IFN-γin plasma and supernatant of in vitro culture medium were detected by enzyme-linked immunosorbent assay（ELISA）. Results The expression levels of miRNA-146a and IFN-γin peripheral CD4＋T lymphocytes were significantly higher in children with psoriasis vulgaris than those in control group[（2.43±0.94）vs（1.05±0.23）,and（27.69±7.64）ng/L vs（9.75±2.81）ng/L,both P〈0.01）;the expression of miRNA-146a was positively correlated with serum IFN-γ（r=0.837,P〈0.01）. As shown by in vitro culture of CD4＋T lymphocytes：compared with the control group,the Th1 counts and the level of IFN- γin the culture medium supernatant in the miRNA-146a mimic group were significantly increased,and the expression level of IFN-γRαprotein was significanlty decreased（all P〈0.01）. miRNA-146a inhibitor could effectively block the modulatory effect of miRNA-146a on Th1 cells. Conclusion miRNA-146a may be playing an important role in the pathogenesis of psoriasis vulgaris by modulating the differentiation and functioning of peripheral CD4＋T lymphocytes in children with psoriasis vulgaris.