Network Construction For Specific Expression Of MiRNA-mRNA In The Process Of Keloid Formation

【Objective】Keloid is a benign skin tumor formed by excessive proliferation of fibrous tissues of the body secondary to repairing trauma after skin injury,which has a high incidence and recurrence rate during clinical treatment.Many studies have reported that a large number of RNA molecules that do not encode proteins,such as micro RNA(miRNA)and t RNA,are involved in the gene expression process of many tumors.Moreover,keloid has tumor-like characteristics,and the development of keloid cannot be separated from the influence of multiple micro RNAs.There are many studies on the role of multiple mRNAs and miRNAs in the growth process of keloid,but a complete miRNA-mRNA regulatory network has not been constructed in this area.In order to explore the genes,signaling pathways and potential regulatory relationship networks that are evident in high and low expression during keloid formation,we have compiled the data using bioinformatics analysis.【Methods】1.Collect genetic data on keloid studies in the GEO(Gene Expression Omnibus)database.The screening criteria were: data before and after the formation of keloid in normal subjects;data before and after the formation of keloid in patients with genetic predisposition to keloid.The mRNA and miRNA datasets GSE113621 and GSE113619 were obtained according to the criteria and grouped separately to design experimental and control groups.The control group was normal skin tissue(N0)from 6 normal individuals without genetic predisposition to keloid scarring,and post-keloid scar tissue(N42)formed at the same site 6 weeks after injury;the experimental group was normal skin tissue(K0)from 8 keloid patients with genetic predisposition to keloid scarring,and post-keloid scar tissue(k42)formed at the same site 6 weeks after injury.The selected data were first subjected to principal component analysis to determine that the selected data analysis and experimental design had its practical significance.The data of mRNA and miRNA datasets were initially organized using R software tools,such as normalized transformation,and then differential expression analysis was performed to derive the results,which were presented in volcano plots and clustered heat maps respectively.2.The results were obtained by functional annotation,pathway enrichment and gene set analysis using GO functional enrichment(Gene Ontology),Kyoto Encyclopedia of Genes and Genomes(KEGG)and GSEA gene set enrichment analysis,respectively,for the significantly increased and decreased mRNA expression of K0-42 from the screening.3.The miRDB database online analysis website was used to perform target gene prediction for the differentially expressed miRNAs obtained from our screening.Then the differentially expressed mRNAs obtained from the analysis and the predicted target gene results were taken as the overlap part,and the overlap part was the target gene we were looking for.4.The target genes as well as the differentially expressed miRNAs derived from the screening were initially constructed as miRNA-mRNA regulatory networks and visualized by the bioinfarmatic online analysis website mapping tool,so as to explore the molecular regulatory mechanism in the formation of keloid.【Results】1.Screening by dataset GSE113621 yielded 318 and 228 differentially expressed miRNAs up-and down-regulated by K0-K42 respectively;246 and 207 differentially expressed miRNAs up-and down-regulated by N0-N42 respectively;76 and 89 differentially expressed miRNAs up-and down-regulated by K42-N42 respectively.Screening by dataset GSE113619 yielded 783 and 711 differentially expressed mRNAs up-and down-regulated by K0-K42 respectively;753 and 547 differentially expressed mRNAs up-and down-regulated by N0-N42 respectively;74 and 63 differentially expressed mRNAs up-and down-regulated by K42-N42 respectively.2.The differentially expressed mRNAs up-and down-regulated by K0-K42 were analyzed by GO,KEGG and GSEA,respectively.the results of GO enrichment analysis mentioned that the main biological process they are involved in is extracellular matrix;the main cellular localization is collagen trimer;the main function they have is involved in collagen binding.The enrichment analysis of KEGG signaling pathway suggests that it is mainly involved in extracellular matrix receptor interaction,protein digestion and uptake,reticuloadhesive plaques,etc.The results of GSEA analysis show that the gene set is mainly enriched in extracellular matrix The results of GSEA analysis showed that the gene set was mainly enriched in extracellular matrix receptor interaction,and this pathway showed an overall trend of down-regulation.3.K0-42 up-regulates the unique vs K42 vs N42 up-regulates 5 differentially expressed miRNAs: hsa-miR-4521,hsa-miR-452-5p,hsa-miR-7846-3p,hsa-miR-331-3p,hsa-miR-4458;K0-42 down-regulates the unique vs K42 vs N42 down-regulation shared 2 differentially expressed miRNAs: hsa-miR-3646,hsa-miR-6800-3p.target gene prediction was performed for these 7 micro RNAs respectively,and the results of differentially expressed mRNAs and predicted target genes were taken as the overlap part,and the overlap part was the target gene we were looking for.4.We obtained seven sets of miRNA-mRNA relationship pairs: hsa-miR-4521 and COX15,hsa-miR-452-5p and SLC7A14,hsa-miR-4458 and GRID2 IP,hsa-miR-4458 and ITGB8,hsa-miR-4458 and SLC7A14,hsa-miR-3646 and SRGAP3,hsa-miR-3646 and PRPS1,to finally construct miRNA-mRNA regulatory relationships specifically expressed during keloid formation.【Conclusion】1.The miRNA-mRNA regulatory network specifically expressed during keloid formation was successfully constructed,providing new potential targets for the treatment of keloid.2.Screening obtained hsa-miR-4521,hsa-miR-452-5p,hsa-miR-4458,hsa-miR-3646,PRPS1,SLC7A14,GRID2 IP,ITGB8,SLC7A14,SRGAP3,COX15 and other micro RNAs and mRNAs that are significantly highly expressed and lowly expressed in keloid development.RNA and mRNA.