Research On Exploring The Molecular Mechanism Of Sjogren’s Syndrome Based On Multi-Omics Technology

Objective(s):Primary Sjogren’s syndrome(pSS)is one of the most common chronic immune diseases.The pathogenesis of pSS is complex and no definite conclusions have been reached.At present,the efficacy of some targeted therapeutic drugs,such as B-cell depleting agents and B-cell activation inhibitors,still a long way from clinical trials,and treatments for the disease are limited.Rapid and correct diagnosis is the key to successful treatment of the disease.Therefore,in this study,differentially expressed genes were screened by whole transcriptome sequencing.In addition,it is difficult to analyze at the single-cell level using traditional sequencing technology.The development of single-cell RNA sequencing(RNA-Seq)opens up a new approach and has been widely applied in the study of the pathogenesis of various immune diseases,as an effective means to parse the pathogenesis of pSS.Method(s):Peripheral blood mononuclear cells(PBMCs)were analyzed in three pSS patients and three healthy controls.pSS patients and healthy controls were sequenced by whole transcriptome sequencing and four types of RNA information were obtained,including mRNA,lnc RNA,circ RNA and mi RNA.Meanwhile,peripheral blood mononuclear cell samples were collected from 6 pSS patients and 6 healthy controls.Immune cells in the patient group and healthy control group were analyzed by sc RNA-seq.In addition,an in-depth analysis of the TCR immune repertoires at the single-cell level has been performed.Result(s):Compared with the healthy control group,mRNAs,lnc RNAs,circ RNAs and mi RNAs were differentially expressed in pSS patients,and the differentially expressed genes with higher and lower expression levels were screened out(p<0.05 and fold change(FC)>2 or<0.5).In the disease group,205 mRNAs were differentially expressed,of which 86 were up-regulated and 119 were down-regulated.A total of 543 Inc RNAs were differentially expressed,of which 301 were up-regulated and 242 were down-regulated.A total of 28 circ RNAs were differentially expressed,of which 17 were up-regulated and 11 down-regulated.A total of 18 mi RNAs were differentially expressed,among which 8 were up-regulated and 10were down-regulated.At the same time,we performed functional enrichment analysis of differentially expressed genes of four types of RNA and we constructed network map of Inc RNA-mi RNA-mRNA.The enrichment immune pathways may be related to the pathogenesis of pSS.Analysis showed that Inc RNA regulate mRNA expression through competitive binding of mi RNA and affect the disease course of pSS.In addition,by analyzing immune cells using the sc RNA-seq,we identified novel cell subsets between tissues,including activated CD4~+T cells,CD4~+CD8~+T cells,CD14~+mononuclear cells,early progenitor cells,macrophages,MAIT cells,memory B cells,megakaryocytes,dendritic cells,mononuclear cells,naive B cells,naive CD4~+T cells,naive CD8~+T cells,NK cells,NKT cells,plasma cells,CD8~+TEM cells and Treg cells,etc.We also determined the number of cells and gene expression level in each cell type.By analyzing signaling pathways and cell-to-cell communication,abnormal cell subsets and dysregulated signaling pathways involved in disease initiation,maintenance,and progression were identified.Finally,an in-depth analysis of deep profiling of TCR immune repertoires at the single-cell level has been performed.It was found that biased use of TRBV and TRBJ gene subtypes appeared in patient samples.Conclusion(s):Through whole transcriptome sequencing,we preliminarily obtained differentially expressed mRNAs,lnc RNAs,circ RNAs and mi RNAs.These genes may be related to the occurrence and development of diseases,but the mechanisms need to be further studied by expanding the sample size.In addition,we performed analysis of PBMCs in pSS patients and healthy controls based on sc RNA-seq,effectively identifying their distinct cell types and states.By identifying the differentially expressed genes of cell subtypes,abnormal signaling pathways and cell communication networks,as well as changes in the TCR immune repertoires,it contributes to a deeper understanding of the role of immune cells in the pathogenesis of pSS,and some findings may provide new targets for pSS therapy.