m⁶A-associated GAS6 expression is associated with pathogenic activation of fibroblast-like synoviocytes in rheumatoid...

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by persistent synovial inflammation and progressive joint destruction. Although N6-methyladenosine (m⁶A) modification is increasingly recognized as an important epitranscriptomic regulatory mechanism, its role in RA remains incompletely understood. This study integrated transcriptomic analysis, m⁶A methylome profiling, and functional experiments to investigate the potential role of m⁶A-associated growth arrest-specific 6 (GAS6) in RA. Synovial tissue samples from patients with RA and healthy controls (HC) were subjected to mRNA sequencing, and integrative analyses were performed using public m⁶A epitranscriptomic datasets. Peripheral blood mononuclear cells (PBMCs) and serum samples were collected from patients with RA and HC subjects. GAS6 and MER proto-oncogene, tyrosine kinase (MERTK) expression levels were measured by quantitative real-time PCR (qRT-PCR) and western blotting, and interleukin (IL)-1β, IL-6, IL-10, and transforming growth factor-β1 (TGF-β1) levels were measured by enzyme-linked immunosorbent assay (ELISA). Small interfering RNA (siRNA) was used to silence GAS6 and MERTK in RA fibroblast-like synoviocytes (RA-FLS). Cell viability, apoptosis, migration, and protein expression were evaluated using Cell Counting Kit-8 (CCK-8), flow cytometry, wound-healing assays, immunofluorescence staining, and western blotting. Integrated multi-omics analysis identified 323 genes with both differential expression and altered m⁶A modification, with enrichment in efferocytosis-related pathways, T helper 17 cell differentiation, and cellular senescence. Among these candidates, GAS6 showed prominent m⁶A hypermethylation and significant upregulation, suggesting a potential epitranscriptomic association. GAS6 was also linked to the efferocytosis-related receptors MERTK and AXL in pathway and interaction analyses. In patients with RA, GAS6 and MERTK expression levels were significantly increased and were associated with inflammatory burden and disease activity. In vitro, silencing GAS6 or MERTK attenuated several pathogenic features of RA-FLS, including enhanced proliferation, migration, pro-inflammatory cytokine production, and resistance to apoptosis. Integrative omics analysis suggests that aberrant m⁶A modification exhibits a potential epitranscriptomic association with activation of the GAS6/MERTK signaling axis in RA. As a hypothesis-generating study, these findings provide new insights into the context-dependent role of the m⁶A-associated GAS6/MERTK axis in RA and suggest that this axis may represent a potential biomarker or therapeutic target warranting further mechanistic investigation.