Background/Purpose:  It has been proposed that immunological triggers at mucosal sites, such as the gut microbiota, may promote autoimmunity affecting joints in patients with rheumatoid arthritis (RA). We recently reported evidence for immune relevance of Prevotella copri, a gut microbe, in a subgroup of new-onset and chronic RA patients. However, it has been unclear how immune responses to gut microbes may be linked to autoimmune joint pathology.

Methods:  To identify disease-relevant microbial and self antigens involved in the pathogenesis of RA, we used a novel proteomic approach to identify HLA-DR-presented peptides (T cell epitopes) in patients’ samples by tandem mass spectrometry. Immunoreactive peptides or their source proteins were then tested for T cell reactivity by IFN-γ ELISpot assay and for antibody responses by ELISA in our large cohort of RA patients or control subjects. Serum samples were also analyzed for innate, Th1, and Th17 mediators by Luminex. Immunoreactive epitopes were searched for microbial sequence homology using BLASTp, and homologous peptides were tested for T cell reactivity. All RA patients met the 2010 ACR/EULAR criteria for RA.

Results: From proteomic analyses, we identified two novel autoantigens, N-acetylglucosamine-6-sulfatase (GNS) and filamin A (FLNA), as targets of T and B cell responses in about half of RA patients. These autoantibody responses were found primarily in patients with antibodies to P. copri, and both IgG and IgA responses to P. copri correlated with the autoantibody levels. These microbial and self immune responses occurred specifically in RA patients, and not in those with other rheumatic diseases or in healthy subjects. Both self proteins were highly expressed in synovia. GNS (but not FLNA) appeared to be citrullinated, and antibody to the citrullinated GNS correlated with anti-citrullinated-protein antibody levels. Anti-GNS and anti-FLNA autoantibodies also correlated with inflammatory cytokines IFN-γ, IL-12, IL17-F, and IL-22, indicative of Th1 and Th17 adaptive immune responses. In a search for T cell epitope mimicry, the HLA-DR-presented GNS peptide was found to have marked sequence homology with epitopes from sulfatase proteins of the gut microbes of the Prevotella and Parabacteroides sp., whereas the HLA-DR-presented FLNA peptide had homology with epitopes from proteins of of Prevotella and Butyricimonas sp., another gut commensal. Patients with T cell reactivity with each self-peptide had responses to the corresponding microbial peptides, and the levels correlated directly. These responses were more common in patients with shared-epitope alleles.

Conclusion: These findings provide evidence for immune relevance of a related order of gut commensals in a subgroup of RA patients, and suggest that gut dysbiosis may compromise the mucosal barrier resulting in leakage of microbes. Moreover, the identification of T cell epitope mimicry between microbial and self epitopes as well as significant correlations in patients’ immune responses to these antigens provide a mechanism linking mucosal immunity and joint autoimmunity in these patients. Finally, the specificity of these responses may advance diagnostic testing in RA.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/sequence-homology-and-immune-reactivity-between-t-cell-epitopes-of-related-gut-microbes-and-two-novel-autoantigens-provide-a-link-between-microbial-and-host-immunity-in-patients-with-rheumat/