Department of Rheumatology and Immunology, The Affiliated Hospital of Southwest Medical University, Luzhou, China.
Department of Clinical Research Center, Dazhou Central Hospital, Dazhou, China.
Clin Rheumatol. 2022 Sep;41(9):2765-2777. doi: 10.1007/s10067-022-06200-4. Epub 2022 May 13.
Sjögren's syndrome (SS), a systemic autoimmune disorder, is characterized by dry mouth and eyes. However, SS pathogenesis is poorly understood. We performed bioinformatics analysis to investigate the potential targets and molecular pathogenesis of SS.
Gene expression profiles (GSE157159) and methylation data (GSE110007) associated with SS patients were obtained from the Gene Expression Omnibus (GEO) database. Differentially methylated positions (DMPs) and differentially expressed genes (DEGs) were identified by the R package limma. The potential biological functions of DEGs were determined using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. Key DMPs were selected by overlap and the shrunken centroid algorithm, and corresponding genes were identified as hub genes, with their diagnostic value assessed by receiver operating characteristic (ROC) curves. The potential molecular mechanisms of hub genes were analyzed by protein-protein interaction (PPI) networks and single-gene gene set enrichment analysis (GSEA). Peripheral blood mononuclear cells (PBMCs) were collected from control and SS patients at The Affiliated Hospital of Southwest Medical University and Dazhou Central Hospital. The mRNA levels of hub genes were verified by quantitative real-time polymerase chain reaction (qRT-PCR).
We identified 788 DMPs and 2457 DEGs between the two groups. Functional enrichment analysis suggested that the DEGs were significantly enriched in T cell activation, leukocyte cell-cell adhesion, and cytokine-cytokine receptor interaction. TSS200, TSS1500, and 1stExon were identified as highly enriched areas of differentially methylated promoter CpG islands (DMCIs). In total, 61 differentially methylated genes (DMGs) were identified by the overlap of 2457 DEGs and 507 genes related to DMPs (DMPGs), of which 21 genes located near TSS200, TSS1500, and 1stExon were selected. Then, three key DMPs and the corresponding hub genes (RUNX3, HLA-DPA1, and CD6) were screened by the shrunken centroid algorithm and calculated to have areas under the ROC curve of 1.000, 0.931, and 0.986, respectively, indicating good diagnostic value. The GSEA results suggested that all three hub genes were highly associated with the immune response. Finally, positive mRNA expression of the three hub genes in clinical SS samples was verified by qRT-PCR, consistent with the GSE157159 data.
The identification of three hub genes provides novel insight into molecular mechanisms and therapeutic targets for SS. Key Points • Hub genes were screened by DNA methylation and transcriptome analyses. • The relative expression of hub genes in peripheral blood samples was verified by qRT-PCR. • HLA-DPA1 was correlated with the pathogenic mechanism of SS.
干燥综合征（SS）是一种系统性自身免疫性疾病，其特征为口干和眼干。然而，SS 的发病机制尚不清楚。我们进行了生物信息学分析，以研究 SS 的潜在靶点和分子发病机制。
从基因表达综合数据库（GEO）中获取与 SS 患者相关的基因表达谱（GSE157159）和甲基化数据（GSE110007）。使用 R 包 limma 识别差异甲基化位置（DMPs）和差异表达基因（DEGs）。通过基因本体论（GO）和京都基因与基因组百科全书（KEGG）通路分析确定 DEGs 的潜在生物学功能。通过重叠和收缩质心算法选择关键 DMPs，并确定相应的基因作为枢纽基因，通过接收器操作特征（ROC）曲线评估其诊断价值。通过蛋白质-蛋白质相互作用（PPI）网络和单基因基因集富集分析（GSEA）分析枢纽基因的潜在分子机制。从西南医科大学附属医院和达州市中心医院的对照和 SS 患者中收集外周血单核细胞（PBMCs）。通过定量实时聚合酶链反应（qRT-PCR）验证枢纽基因的 mRNA 水平。
我们在两组之间鉴定出 788 个 DMPs 和 2457 个 DEGs。功能富集分析表明，DEGs 显著富集于 T 细胞激活、白细胞细胞-细胞黏附和细胞因子-细胞因子受体相互作用。TSS200、TSS1500 和 1stExon 被鉴定为差异甲基化启动子 CpG 岛（DMCIs）的高度富集区域。总共通过 2457 个 DEGs 和 507 个与 DMPs 相关的基因（DMPGs）的重叠鉴定出 61 个差异甲基化基因（DMGs），其中 21 个基因位于 TSS200、TSS1500 和 1stExon 附近。然后，通过收缩质心算法筛选出三个关键 DMP 和相应的枢纽基因（RUNX3、HLA-DPA1 和 CD6），并计算出它们的 ROC 曲线下面积分别为 1.000、0.931 和 0.986，表明具有良好的诊断价值。GSEA 结果表明，这三个枢纽基因均与免疫反应高度相关。最后，通过 qRT-PCR 验证了临床 SS 样本中三个枢纽基因的阳性 mRNA 表达，与 GSE157159 数据一致。
三个枢纽基因的鉴定为 SS 的分子机制和治疗靶点提供了新的见解。
通过 DNA 甲基化和转录组分析筛选枢纽基因。
通过 qRT-PCR 验证外周血样本中枢纽基因的相对表达。
HLA-DPA1 与 SS 的发病机制相关。
