Xiaofang Liu ^1* Bin Li ^1* Yuxi Lin ^1* Xueying Ma ^1* Yingying Liu ^1* Lili Ma ^1* Xiaomeng Ma ^1* Xia Wang ^2* Nanjing Li ^2* Xiaoyun Liu ^1* Xiaohong Chen ^1*

• ¹ Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, China
• ² Huizhou Sixth People's Hospital, Huizhou, Guangdong Province, China

Background Emerging evidence underscores the comorbidity mechanisms among autoimmune diseases (AIDs), with innovative technologies such as single-cell RNA sequencing (scRNA-seq) significantly advancing the explorations in this field. This study aimed to investigate the shared genes among three AIDs-Multiple Sclerosis (MS), Systemic Lupus Erythematosus (SLE), and Rheumatoid Arthritis (RA) using bioinformatics databases, and to identify potential biomarkers for early diagnosis.We retrieved transcriptomic data of MS, SLE, and RA patients from public databases.Weighted Gene Co-Expression Network Analysis (WGCNA) was employed to construct gene coexpression networks and identify disease-associated modules. Functional enrichment analyses and Protein-Protein Interaction (PPI) network was constructed. We used machine learning algorithms to select candidate biomarkers and evaluate their diagnostic value. The Cibersort algorithm was and scRNA-seq analysis was performed to identify key gene expression patterns and assess the infiltration of immune cells in MS patients. Finally, the biomarkers' expression was validated in human and mice experiments.Several shared genes among MS, SLE, and RA were identified, which play crucial roles in immune responses and inflammation regulation. PPI network analysis highlighted key hub genes, some of which were selected as candidate biomarkers through machine learning algorithms.Receiver Operating Characteristic (ROC) curve analysis indicated that some genes had high diagnostic value (Area Under the Curve, AUC > 0.7). Immune cell infiltration pattern analysis showed significant differences in the expression of various immune cells in MS patients. scRNAseq analysis revealed clusters of genes that were significantly upregulated in the single cells of cerebrospinal fluid in MS patients. The expression of shared genes was validated in the EAE mose model. Validation using clinical samples confirmed the expression of potential diagnostic biomarkers.This study identified shared genes among MS, SLE, and RA and proposed potential early diagnostic biomarkers. These genes are pivotal in regulating immune responses, providing new targets and theoretical basis for the early diagnosis and treatment of autoimmune diseases.