Bioinformatics Analysis Of Differentially Expressed Gene In Keloid Fibroblasts And Keratinocytes

Background:Keloid is a kind of benign tumor of connective tissue,which is mainly manifested by the excessive proliferation of connective tissue after the damaged wound has healed,which grows with the passage of time,and extends even beyond the original site of the injury.Keloids can cause pain and itching;moreover,it can bring discomfort and psychological pressure to patients,and hence affect the quality of life of patients.Moreover,keloid rarely resolves spontaneously and has a high recurrence rate after treatment.To date,with the latest advances in molecular biology and genetics,people have gained new understanding of the complex process of scar formation.However,the pathogenesis of keloid has not been fully elucidated,and the mechanism of its occurrence and development still needs to be further studied.Studies have shown that several transcription factors in fibroblasts and keratinocytes are involved in the regulation of keloids.For example,activated transcription factor 3 regulates growth,apoptosis,invasion,and collagen synthesis of keloid fibroblasts through the transforming growth factor(TGF-?)/Smad signaling pathway.Smad7 gene regulation can affect the EMT process of normal skin keratinocytes and keloid keratinocytes,and then regulate cell proliferation,migration and apoptosis.Runx2 siRNA may reverse keloid pathological phenotypes by inhibiting the PI3K/AKT signaling pathway.The downregulation of Epac gene can reduce the fibrosis and induce apoptosis of keloid fibroblasts through the AKT signaling pathway.It has also been found that miR-194-5p plays an inhibitory role in human keloid fibroblasts by targeting NR2F2.Although some genes and pathways have been reported to be related to keloid fibroblasts and keratinocytes,the key molecular mechanisms of keloid pathology and genes for targeted therapy are still not fully understood.In this study,we used bioinformatics to discover the possible regulatory pathways involved in keloids formation,explore the key genes differentially expressed in keloid fibroblasts and keratinocytes,and potential drugs for keloid treatment.Objective:1)To explore the potential key genes and signaling pathways involved in regulating keloid formation in keloid derived fibroblasts;2)To study the potential key genes and signaling pathways involved in the regulation of keloid formation in keloid derived keratinocytes;3)To explore the potential interactions between key genes and drugs in keloid derived fibroblasts or keratinocytes.Methods:The data was obtained from the Gene Expression Comprehensive Database(GEO),using the dataset of GSE44270.The analysis groups were set as:keloid fibroblasts(n=9)vs non-damaging fibroblasts(n=4)and keloid keratinocytes(n=9)vs non-damaging keratinocytes(n=4).The classical Bayesian method provided by the limma package(Version 3.10.3)was used to analyze the differential expressed genes(DEGs)of keloid fibroblasts and keratinocytes.Next,GO enrichment analysis,KEGG pathway and Uniport function analysis was conducted on the DEGs using the String(count>=2,and P.Value<0.05 was considered as a significant enrichment result),and the result was visualized using R language.DEGs were represented by volcano plots and cluster plots.Combining with String database,the protein-protein interaction(PPI)network was constructed.Meanwhile,the PPI results were imported into Cytoscape,and the interaction between the most significant clustered module proteins in PPI network was analyzed using Cytoscape plug-in MCODE method.Finally,existing resources were mined through DGIdb to predict all gene-related drug-gene pairs in key subnetwork modules,Cytoscape was to construct the drug-gene network.Results:(1)Compared with the non-injured group,343 DEGs were analyzed in keloid fibroblasts,of which 273 were up-regulated and 70 were down-regulated.The enrichment results of KEGG pathway mainly included ubiquitin mediated proteolysis and endocytosis pathways.Five hub genes(UBB,POLR2C,COPB1,ANAPC10 and RAB1B)and one significant module(score=9)were obtained from all selected PPI networks.ANAPC10 is found to have close interactions in this important module.(2)Compared with the non-injured group,752 DEGs were analyzed in keloid keratinocytes,of which 602 were up-regulated genes and 150 were down-regulated genes.The enrichment results of KEGG pathway mainly include cell cycle,oocyte meiosis,progesterone-mediated oocyte maturation,RNA transport pathways.Five hub genes(CDK1,CCNB1,CDC20,PLK-1 and AURKB)and one significant module(score=44.514)were obtained from all selected PPI networks.PPP2R5B is found to have close interactions in this important module and is the only down-regulated gene in the significant module.(3)No significant results were obtained when the drug-gene interaction relationship was analyzed for the 9 key genes in the key sub-network of fibroblasts.A total of 11 of the 75 genes in the keratinocyte key network were predicted.Among them,TOP2A gene was regarded as a key gene targeted by drugs,which corresponded to 28 drug-gene interaction relationships.AURKA,CHEK1 and TTK genes were followed by 28,3,3,and 2 drug-gene interaction pairs,respectively.Analysis conclusions:ANAPC10 and PPP2R5B were the key genes involved in the formation of keloid fibroblasts and keratinocytes,respectively,which may be involved in ubiquitin mediated proteolysis and cell cycle signaling pathways to affect the occurrence and development of keloid.Additionally,the TOP2A gene was found to be a key potential gene locus for keloid drug targeting.