miR-181a通过调控Toll样受体参与儿童过敏性紫癜肾炎PS/iPS的激活

The miR-181a participates in PS/iPS activation by regulating Toll-like receptors in children with allergic purpura nephritis

目的探究miR-181a在儿童过敏性紫癜肾炎中的免疫调节机制。方法收集我院儿童过敏性紫癜(HSP)患儿和正常健康体检儿童的外周血,分离外周血单核细胞(PBMC),采用实时定量qRT-PCR检测样本PBMC中miR-181a表达水平。分析非肾型过敏性紫癜患儿(HSP-nonN)、儿童过敏性紫癜肾炎(HSPN)患儿和健康对照儿童(HC)间不同miRNAs的表达差异,利用miRNA数据库对其靶基因进行预测。采用qRT-PCR定量检测HSPN,HSP-nonN和HC儿童PBMC中Toll样受体(TLR)、低分子量蛋白2(LMP2)/β1亚基表达水平;蛋白质免疫印迹(Western blot)检测TLR的表达水平。利用TLR2抑制剂(INH14)处理各组PBMC,qRT-PCR检测LMP2/β1亚基比值和P65磷酸化水平。结果HSP组血清miR-181a表达高于健康对照组(P<0.005),HSPN组miR-181a表达高于HSP-nonN组(P<0.005)。miR-181a的靶基因高度预测为Toll样受体。qRT-PCR和Western blot结果显示,HSP-nonN组和HSPN组TLR2 mRNA水平均高于HC组(P<0.05),HSPN组LMP2/β1亚基比值高于HSP-nonN组(P<0.05);INH14处理后HSPN组LMP2/β1亚基比值明显低于HSP-nonN组(P<0.05)。HSPN组PBMC的P65磷酸化多于HSP-nonN组和HC组,INH14处理后,HSPN组的P65磷酸化明显降低。结论miR-181a通过TLR2激活NF-κB信号通路参与儿童过敏性紫癜肾炎的PS/iPS转换。

Objective To explore the immune regulatory mechanism of miR-181a in children with allergic purpura nephritis.Methods The peripheral blood of children with allergic purpura and children with normal physical examination were collected.The peripheral blood mononuclear cells(PBMC)were isolated to detect the expression level of miR-181a by real-time quantitative PCR.The expression of different miRNAs was compared among Henoch-Schonlein purpura without nephritis(HSP-nonN),Henoch-Schonlein purpura nephritis(HSPN)and healthy controls(HC),and their target genes were predicted using the database.The qRT-PCR was used to quantitatively detect the levels of Toll-like receptor(TLR)and LMP2/β1 expression in PBMC from HSPN,HSP-nonN and HC,and Western blot was used to detect the expression level of TLR.After PBMCs from patients and HC were treated with TLR2 inhibitors,LMP2/β1 conversion levels and the P65 phosphorylation were detected by qRT-PCR.Results The expression of miR-181a in HSP group was higher than that in HC group(P<0.005),and the expression of miR-181a in HSPN group was higher than that in HSP-nonN group(P<0.005).The target gene of miR-181a was highly predicted to be Toll-like receptor.The mRNA level of TLR2 in HSP-nonN group and HSPN group was higher than that in HC group(P<0.05),and the ratio of LMP2/β1 subunit in HSPN group was higher than that in HSP-nonN group(P<0.05).After INH14 treatment,the ratio of LMP2/β1 subunit in HSPN group was significantly lower than that in HSP-nonN group(P<0.05).The P65 phosphorylation of PBMC in HSPN group was higher than that in HSP-nonN group and HC group,and the P65 phosphorylation in HSPN group was significantly reduced after INH14 treatment.Conclusion The miR-181a may be involved in PS/iPS conversion in children with allergic purpura nephritis by activating the NF-κB signaling pathway through TLR2.