Background RA associated immunity develops years before disease development and might be initiated at extra-articular sites, such as the lungs.
Objectives To investigate molecular lung changes in patients with rheumatoid arthritis (RA) at disease onset and to address the contribution of these changes to disease initiation.
Methods 24 RA patients with patient-reported symptom duration less than 1 year and naive to DMARD treatment and 9 healthy individuals were subjected to bronchoscopy and mucosal large bronchial biopsies were retrieved. For comparison 16 large bronchial biopsies and 79 BAL samples from healthy volunteers were available. Histological analysis for identification of inducible bronchia associated lymphoid tissues (iBALT) and lymphocyte infiltration were performed. Further immunohistochemical analysis was performed in RA biopsies to detect PAD enzymes, CD3, HLA-DQ and HLA-DR and to identify citrullinated targets. Mass spectrometry was used for identification of citrullinated epitopes in 6 of the lung RA biopsies and additional 8 synovial RA biopsies. One candidate peptide was synthesized and used to investigate presence of antibodies by ELISA in RA patients (n=393), healthy controls (n=152) and disease controls (n=236). HLA-DRB1 shared epitope (SE) alleles were detected in RA patients.
Results Bronchial lymphocyte infiltration and iBALT formation was observed at baseline in half of the ACPA+ RA patients but only 1 out of 6 ACPA- patients (17%) and 1 out of 9 healthy volunteers (10%). Higher expression of HLA-DR and citrullinated targets was observed in bronchial biopsies of ACPA+ as compare to ACPA- RA (p<0.05). CD3 and HLA-DQ expression also showed a tendency to higher expression in the ACPA+ as compared to ACPA- RA patients. ACPA+ RA patients had significantly higher number of lymphocytes and neutrophils as compared to healthy controls (p<0.05). Markers of T cell activation (CD69 and CD103) were expressed by significantly more CD4+ BAL T cells in ACPA+ RA patients as compared to healthy controls. BAL fluids of ACPA+ RA patients were enriched in both IgG and IgA ACPA as compared to paired serum samples. Mass spectometry identified 5 proteins in the synovium (in total 8 sites) and 4 in the lungs (in total 6 sites) containing citrullinated residues. Two vimentin derived citrullinated peptides were present in a majority of both synovial and lung biopsies with slightly higher citrullinated/unmodified peptides ratios in the smokers as compared to non-smokers. Antibodies against a synthesized cit-vim peptide candidate (covering both cit-vim peptides identified in vivo) were present in 15% of RA patients and 3.4% of disease controls and associated with presence of the SE alleles in RA.
Conclusions Signs of inflammation, accumulation of highly activated and differentiated BAL CD4+ T cells and local ACPA enrichment are present early in bronchial tissues of ACPA+ RA patients. Shared citrullinated targets in the lung and joints as well as systemic reactivity against these targets are present in RA patients. Our findings support the notion that early inflammatory events in the lungs may represent a critical initiating factor in the development of ACPA+ RA.
Disclosure of Interest None declared