基于转录组测序探究TGF-β对成纤维细胞作用的机制研究

Study on the mechanism of TGF-βaction on fibroblasts based on transcriptome squencing

目的基于转录组RNA测序筛选转化生长因子β(transforming growth factor-beta,TGF-β)刺激的成纤维细胞与正常成纤维细胞的差异mRNA。方法对正常人成纤维MRC-5细胞和经20 ng/mL TGF-β刺激24 h后的MRC-5细胞进行转录组RNA测序,筛选出两组之间的差异表达基因(DEGs),对DEGs进行生物信息学分析,包括基因功能(GO)、京都基因百科全书(KEGG)富集分析、蛋白质相互作用网络分析(PPI)以及相关基因对肿瘤生存预后的影响。结果筛选出在33692个基因中差异表达的mRNAs共161个,其中上调39个,下调122个;GO分析表明,DEGs主要富集的生物学过程包括核苷酸生物合成、代谢和神经元凋亡、坏死等过程;KEGG通路富集发现这些信号主要涉及核苷酸生物合成过程,核苷磷酸代谢过程,神经元凋亡等过程,其中核苷酸生物合成在6条信号通路中均被富集到;利用TCGA数据库对膀胱癌(BLCA)进行生存分析发现TGFB1I1、HYOU1和CAT基因同时高表达时,与患者较差预后显著相关。结论mRNA在TGF-β刺激后的成纤维细胞和正常成纤维细胞中存在表达差异,筛选出在BLCA病人中表达的TGFB1I1、HYOU1和CAT,有望作为患者的预后标志物。

Objective To screen the differential mRNA between transforming growth factor-beta(TGF-β)stimulated fibroblasts and normal fibroblasts based on transcriptome RNA sequencing.Methods Transcriptome RNA sequencing was performed on normal human fibroblasts MRC-5 cells and MRC-5 cells stimulated with 20 ng/mL TGF-βfor 24 hours,and differentially expressed genes(DEGs)were screened between the two groups.Bioinformatics analysis was performed on DEGs,including gene ontology(GO),kyoto encyclopedia of genes and genomes(KEGG)enrichment analysis,protein-protein interaction network analysis(PPI)and the effects of related genes on tumor survival and prognosis.Results A total of 161 differentially expressed mRNAs were screened out from 33692 genes,including 39 up-regulated and 122 down-regulated genes;GO analysis showed that biological processes mainly enriched by DEGs included nucleotide biosynthesis,metabolism,neuronal apoptosis and necrosis;KEGG pathway enrichment found that these signals were mainly involved in nucleotide biosynthesis,nucleotide phosphate metabolism and neuronal apoptosis,among which nucleotide biosynthesis was enriched in six signal pathways;survival analysis of bladder cancer(BLCA)by TCGA database found that simultaneous high expression of TGFB1I1,HYOU1,and CAT genes were significantly associated with poor prognosis in patients with BLCA.Conclusion mRNAs have differential expression between TGF-βstimulated fibroblasts and normal fibroblasts,and the screened TGFB1I1,HYOU1 and CAT genes are expected to be used as prognostic markers in patients with BLCA.