特应性皮炎免疫浸润相关miR-155靶基因的筛选及验证  

Screening and validation of miR-155 target genes associated with immune infiltration in atopic dermatitis

作　　者：王晓晨[1] 陈露 刘畅 陈晓青[2] 李芃[3] 邱文洪 WANG Xiaochen;CHEN Lu;LIU Chang;CHEN Xiaoqing;LI Peng;QIU Wenhong(Medical School,Jianghan University,Wuhan 430101,China;不详)

机构地区：[1]江汉大学医学部免疫学教研室,武汉430101 [2]江汉大学医学部实验中心 [3]武汉市中心医院皮肤科

出　　处：《山东医药》2023年第21期15-19,共5页Shandong Medical Journal

基　　金：国家自然科学基金资助项目(31671092)。

摘　　要：目的筛选与特应性皮炎(AD)免疫浸润相关的miR-155靶基因,并进行细胞实验验证。方法从GEO数据库获取AD基因表达谱数据集GSE121212和GSE157194,筛选出AD皮损区差异表达基因,并进行GO功能富集分析、KEGG通路富集分析,构建蛋白互作网络(PPI)。基于CIBERSORT算法,分析AD患者免疫细胞浸润情况。利用miRNet网站预测miR-155靶基因,并筛选与AD免疫浸润相关的miR-155靶基因。将人永生化角质形成细胞株HaCaT细胞分为control组和TI组,TI组细胞加入TNF-α和IFN-γ构建炎症模型,control组常规培养,采用qRT-PCR法检测TI组、control组细胞中miR-155。取TI组HaCaT细胞分成TI+miR-155 mimics组、TI+mimics NC组,TI+miR-155 mimics组转染miR-155过表达质粒miR-155 mimics,TI+mimics NC组转染过表达对照质粒mimics NC,采用qRT-PCR法检测TI+miR-155 mimics组、TI+mimics NC组细胞中CXCL1、CXCL8 mRNA。结果筛选得到519个AD皮损区差异表达基因。GO功能富集分析结果显示,差异表达基因主要参与免疫系统过程、免疫反应、对外部刺激的反应、防御反应、免疫系统的调节等生物过程,KEGG通路富集分析结果显示,通路主要富集在细胞因子—细胞因子受体相互作用、病毒蛋白与细胞因子及细胞因子受体相互作用、趋化因子信号通路、IL-17信号通路等炎症相关信号通路。成功构建出由67个节点、146条边组成的PPI网络图。树突状细胞、M2巨噬细胞和肥大细胞在表皮免疫微环境中占比最高,免疫浸润最明显。与AD免疫浸润相关性比较高的miR-155靶基因为CXCL1、CXCL8。TI组细胞中miR-155相对表达量高于control组(P<0.05),TI+miR-155 mimics组细胞中CXCL1、CXCL8 mRNA相对表达量高于TI+mimics NC组(P均<0.05)。结论筛选获得的与AD免疫浸润相关的miR-155靶基因是CXCL1、CXCL8基因。高表达miR-155可提高炎症时HaCaT细胞中CXCL1、CXCL8基因的表达水平。Objective To screen out the target genes of miR-155 associated with immune infiltration in atopic dermatitis(AD),and to verify them by cell experiments.Methods The gene expression profiles of AD(GSE121212 and GSE157194)were obtained from the GEO database,and the differentially expressed genes in AD lesions were screened out.GO functional enrichment analysis and KEGG pathway enrichment analysis were performed,and protein-protein interaction(PPI)network was constructed.Based on CIBERSORT algorithm,the infiltration of immune cells in AD patients was analyzed.The miRNet website was used to predict the target genes of miR-155,and the target genes related to AD immune infiltration were screened.Human immortalized epidermal cells(HaCaT cells)were divided into the control group and TI group.The cells in the TI group were treated with TNF-αand IFN-γto construct the inflammation models,and the cells in the control group were cultured routinely.The qRT-PCR was used to detect miR-155 in the TI group and control group.The HaCaT cells in the TI group were divided into the TI+miR-155 mimics group and TI+mimics NC group.The HacaT cells in the TI+miR-155 mimics group were transfected with miR-155 overexpression plasmid,and cells in the TI+mimics NC group were transfected with the control plasmid mimics NC.The mRNA levels of CXCL1 and CXCL8 in the TI+miR-155 mimics group and TI+mimics NC group were detected by qRT-PCR.Results A total of 519 differentially expressed genes were identified in AD lesions.GO functional enrichment analysis showed that the differentially expressed genes were mainly involved in biological processes such as immune system process,immune response,response to external stimuli,defense response,and regulation of the immune system.KEGG pathway enrichment analysis showed that,pathways were mainly enriched in cytokine-cytokine receptor interaction,viral protein-cytokine and cytokine receptor interaction,chemokine signaling pathway,IL-17 signaling pathway and other inflammation-related signaling pathways.A PPI ne

关 键 词：特应性皮炎 MIR-155 免疫浸润 CXCL1基因 CXCL8基因 生物信息学分析 细胞实验 

分 类 号：R392.8[医药卫生—免疫学]