基于转录组学探究T细胞衰老的分子机制

Molecular mechanisms of T cell senescence through transcriptomics

背景T细胞衰老是免疫衰老的关键环节之一,与多种老年相关疾病和肿瘤的发生发展密切相关,但缺乏基于转录组学探究T细胞衰老过程中分子机制的研究。目的建立T细胞衰老模型并进行转录组测序,利用生物信息学方法分析T细胞衰老过程中的潜在分子机制并筛选关键基因。方法从人类外周血单个核细胞中分选出T细胞,在体外经过短暂培养扩增后随机分为两组(衰老T细胞组使用含有20 mg/mL D-半乳糖的培养基,正常对照组使用不含有D-半乳糖的培养基),在培养48 h后,检测T细胞衰老前后端粒酶活性、端粒长度、衰老相关分泌表型、β-半乳糖苷酶活性等改变情况。通过对细胞模型的mRNA转录组测序并利用生物信息学方法分析显著差异表达基因(differentially expressed genes,DEGs),通过对DEGs进行富集分析和蛋白质-蛋白质互作(protein-protein interaction,PPI)网络构建,分析T细胞衰老过程中DEGs的功能状态并筛选关键基因。结果D-半乳糖处理48 h后,T细胞发生端粒功能障碍,p21基因及蛋白质表达升高,分泌细胞因子增多,β半乳糖苷酶活性增加和细胞内脂质合成增加(P<0.05)。富集分析结果显示T细胞衰老后细胞因子-细胞因子受体相互作用、p53、PI3K/Akt/mTOR、NF-kappaB等信号通路活动增强;PPI网络分析发现ISGs和RPs基因家族可能在T细胞衰老过程中发挥关键作用。结论衰老T细胞端粒功能障碍、脂质代谢紊乱、促炎能力和细胞衰老相关信号通路活动增强,并且ISGs和RPs基因家族可能参与到T细胞衰老的关键过程之中。

Background T cell senescence is one of the key components of immunosenescence,closely associated with the occurrence and development of various age-related diseases and tumors.However,there is a lack of research exploring the molecular mechanisms of T cell senescence based on transcriptomics.Objective To establish a T cell senescence model and perform transcriptome sequencing,then use bioinformatics methods to analyze potential molecular mechanisms and identify key genes during T cell senescence.Methods T cells were isolated from human peripheral blood mononuclear cells,after a brief period of in vitro culture for expansion,the cells were randomly divided into senescent T cell group,cultured in a medium containing 20mg/mL D-galactose,and normal control group,cultured in a medium without D-galactose.After 48 hours of culture,changes in telomerase activity,telomere length,senescence-associated secretory phenotype,andβ-galactosidase activity were assessed in T cells before and after cell senescence.mRNA transcriptome sequencing of the cell models was performed,and bioinformatics methods were used to analyze significantly differentially expressed genes(DEGs).Enrichment analysis and the construction of protein-protein interaction(PPI)networks were conducted on the DEGs to analyze the functional status of DEGs during T cell senescence and to screen for key genes.Results After 48 hours of D-galactose treatment,T cells showed impaired telomere function,increased expression of p21 gene and protein,increased secretion of cytokines,enhancedβ-galactosidase activity,and increased intracellular lipid synthesis.Enrichment analysis revealed that senescent T cells showed enhanced activity in cytokine-cytokine receptor interaction,p53,PI3K/Akt/mTOR,NF-kappaB signaling pathways.PPI network analysis suggested that ISGs and RPs gene families might play a crucial role in T cell senescence.Conclusion Senescent T cells exhibit telomere dysfunction,lipid metabolism disorder,enhanced pro-inflammatory capacity, and increased activity of cell senescence-related signaling pathways. Moreover, the ISGs and RPs gene families may play significant roles in the process of T cell senescence.