Abstract
BACKGROUND: A burgeoning body of evidence has substantiated the association between alterations in the composition of the gut microbiota and rheumatoid arthritis (RA). Nevertheless, our understanding of the intricate mechanisms underpinning this association is limited. METHODS: To investigate whether the gut microbiota influences the pathogenesis of RA through metabolism or immunity, we performed rigorous synthesis analyses using aggregated statistics from published genome-wide association studies (GWAS) using two-sample Mendelian randomization (MR) and mediated MR techniques, including two-step MR and multivariate MR analyses. Subsequently, we conducted in vitro cellular validation of the analyzed Microbial-Cytokine-RA pathway. We determined the optimal culture conditions through co-culture experiments involving concentration and time. Cell Counting Kit-8 (CCK-8) assays were employed to assess cellular viability, and enzyme-linked immunosorbent assays (ELISA) were performed to assess tumor necrosis factor-inducible gene 6 protein (TSG-6) and tumor necrosis factor-α (TNF-α) levels. RESULTS: Our univariable MR results confirmed 15 microbial traits, 7 metabolites and 2 cytokines that may be causally associated with RA (P (FDR) < 0.05). Mediation analysis revealed that microbial traits influence the risk of RA through metabolite or cytokine (proportion mediated: 7.75% - 58.22%). In vitro experiments demonstrated that TSG-6 was highly expressed in the Subdoligranulum variabile treatment group and was correlated with decreased RA severity (reduced TNF-α expression). Silencing the TSG-6 gene significantly increased TNF-α expression, regardless of treatment with S. variabile. Additionally, S. variabile-secreted exosomes exhibited the same effect. CONCLUSION: The results of this study suggest that S. variabile has the potential to promote TSG-6 secretion, thereby reducing RA inflammation.