Preliminary Study On The Mechanism Of MiR-155 In Immune Infiltration Of Atopic Dermatitis

Background and objective: Atopic dermatitis(AD)is one of the major causes of the global burden of skin diseases.The core of the pathogenesis of AD is inflammation,but the specific regulatory mechanism is still unclear.Therefore,it is necessary to investigate the molecular mechanisms of the occurrence and development of AD to improve diagnostic and therapeutic strategies.As an important factor in the regulation of protein function,micro RNA can also mediate the inflammatory response of diseases.Studies have found that mi R-155 is significantly up-regulated in skin lesions of patients with AD.However,the mechanism of mi R-155 in the occurrence and development of AD is not very clear.In this study,the differentially expressed genes in the epidermal immune microenvironment were analyzed by bioinformatics methods,and a cell model was constructed to explore the mechanism of mi R-155 in the immune infiltration of AD.Methods: The transcriptome sequencing results of GSE121212 and GSE157194 were obtained from GEO data,and the differentially co-expressed genes in the epidermal immune microenvironment of patients with AD were screened by R software package and VENNY website.The R software package was used to perform GO and KEGG functional enrichment analysis of differentially expressed genes.Protein interaction network(PPI)was constructed based on STRING database and Cytoscape software.Based on CIBERSORT algorithm,the infiltration of immune cells in patients with AD was analyzed.mi RNet was used to predict the target genes of mi R-155.CCK8 was used to detect the effect of mi R-155 on the praoliferation of Ha Ca T cells.The m RNA levels of mi R-155,PI3,FOSL1,CXCL1 and CXCL8 were detected by q RTPCR.ELISA was used to detect the protein levels of IL-1β,IL-6,IL-10,IL-15,CXCL1 and CXCL8.The inflammatory cell model of Ha Ca T cells was induced by TNF-α and IFN-γ.Experimental grouping: control group,Ha Ca T cells;TNF-α and IFN-γ induction group,Ha Ca T cells+TNF-α and IFN-γ;TNF-α and IFN-γ induction+mi R-155 transfection group,Ha Ca T cells+TNF-α and IFN-γ+mi R-155 transfection group.ELISA was used to detect the protein levels of IL-1β,IL-6,IL-10,IL-15,CXCL1 and CXCL8.The m RNA levels of mi R-155,PI3,FOSL1,CXCL1 and CXCL8 were detected by q RT-PCR.Western Blot was used to detect the protein levels of PI3 and FOSL1.Results: A total of 519 differentially expressed genes were screened,of which 138 genes were down-regulated and 381 genes were up-regulated.A total of 1023 functions were enriched by GO analysis,including biological processes such as immune system process,immune response,response to external stimuli,and regulation of the immune system.A total of 44 pathways were enriched by KEGG analysis,and the differentially expressed genes were most related to cytokine-cytokine receptor interaction.According to the results of immune infiltration analysis,Dendritic cells(DC),Macrophages M2 and Mast cells activated immune infiltration were the most obvious in the epidermal immune microenvironment.According to the results of bioinformatics analysis,PI3,FOSL1,CXCL1 and CXCL8 were screened as potential target genes of mi R-155 in the epidermal immune microenvironment of AD patients.Ha Ca T cells were transfected with mi R-155.CCK8 assay showed that mi R-155 could inhibit the proliferation of Ha Ca T cells.q RT-PCR results showed that mi R-155 decreased the m RNA levels of PI3 and CXCL8,increased the m RNA level of FOSL1,and there was no change in the m RNA level of CXCL1.ELISA results showed that mi R-155 increased the secretion levels of IL-1β,IL-6,IL-15 and CXCL1,and decreased the secretion levels of IL-10 and CXCL8.In the inflammatory cell model of Ha Ca T cells induced by TNF-α and IFN-γ,the m RNA level of mi R-155 was significantly increased after TNF-α and IFN-γstimulation.CCK8 assay showed that mi R-155 significantly inhibited the proliferation of Ha Ca T cells during inflammation.ELISA results showed that mi R-155 significantly increased the secretion of IL-1β,IL-6,IL-10 and IL-15 by Ha Ca T cells in inflammation,and significantly increased the secretion of CXCL1 and CXCL8 by Ha Ca T cells.q RTPCR results showed that mi R-155 significantly increased the m RNA levels of PI3,FOSL1,CXCL1 and CXCL8 in inflammation cells.Western Blot results showed that mi R-155 significantly increased the expression levels of PI3 and FOSL1 proteins in inflammatory cells.Conclusion: mi R-155 promotes the production of inflammatory cytokines and proteins by Ha Ca T cells by inhibiting the proliferation of Ha Ca T cells,reducing the expression of anti-inflammatory gene PI3 and the secretion of anti-inflammatory cytokine IL-10,and increasing the expression of pro-inflammatory gene FOSL1 and the secretion of pro-inflammatory cytokines IL-1β,IL-6,IL-15 and CXCL1.It participates in the damage process of keratinocytes and promotes the infiltration of inflammatory cells in the epidermal microenvironment.When inflammation occurs,keratinocytes induce the expression of mi R-155,which further promotes the production of inflammatory cytokines and proteins by keratinocytes,induces the infiltration of immune cells in the epidermal microenvironment,and continues to secrete many proinflammatory cytokines,leading to the imbalance of inflammatory regulation mechanisms,aggravating the inflammatory response,and causing further damage to keratinocytes.Eventually,it leads to the destruction of skin protection function and causes atopic dermatitis.