Identification of hnRNP-A2 (RA33) as a major B-cell and T-cell autoantigen in pristane–induced arthritis

Pristane-induced arthritis (PIA) in rats is considered an excellent model for rheumatoid arthritis (RA) since it fulfils the criteria for RA including a chronic relapsing disease course and is not dependent on immunization with exogenous antigen. Although the adjuvant pristane is not immunogenic, the disease is MHC associated and dependent on the activation of (autoreactive) T cells. However, so far it has not been possible to link the immune response to joint antigens or other endogenous components. HnRNP A2, the RA33 autoantigen (A2/RA33), is a multi-functional RNA binding protein involved in splicing and other aspects of post-transcriptional regulation of gene expression. Autoantibodies as well as autoreactive T cells against A2/RA33 have been found in patients with RA but the pathogenetic role of these autoimmune responses is unresolved [1]. It was therefore the aim of this study to elucidate a potential involvement of A2/RA33 in PIA.

Autoantibodies against A2/RA33 were determined by immunoblotting, and MHC association of the anti-A2/RA33 immune response was specified by the presence of autoantibodies, delayed-type hypersensitivity reactions and T-cell cytokine secretion in DA rats of different haplotypes. Interferon gamma and tumour necrosis factor secretion by T cells isolated from draining lymph nodes 10 days after pristane injection and restimulation with A2/RA33 in vitro was determined. Expression of A2/RA33 in joints and organs was analysed by immunohistochemistry and western blotting. Nasal vaccinations were performed with A2/RA33 7 days prior to pristane injection.

Although anti-A2/RA33 autoantibodies were detected in all four rat strains investigated, the immune response appeared to be particularly linked to the F and U MHC haplotypes. Autoantibodies to A2/RA33 were found in 60% of DA1.F sera, and T cells of all DA1.F rats tested produced intermediate to high levels of interferon gamma upon stimulation with A2/RA33. The reaction seemed to be entirely produced by CD4⁺ cells showing a Th1 phenotype. Furthermore, nasal vaccination with A2/RA33 significantly delayed the onset and decreased severity of arthritis in DA1.F rats. Finally, immunohistochemical and western blot analyses revealed pronounced overexpression of A2/RA33 in joints of rats suffering from acute PIA, but not in healthy joints or in joints from animals with chronic PIA.

The A2/RA33 autoantigen is targeted by autoantibodies and Th1 cells in rats with PIA shortly after pristane injection. The presence of autoreactive Th1 cells in conjunction with synovial overexpression of A2/RA33 strongly suggests involvement of this autoantigen in the pathogenesis of PIA. This is further bolstered by the observed alleviation and delay of onset of PIA following nasal vaccination with A2/RA33. Thus, A2/RA33 seems to be one of the primary autoantigens in PIA that in conjunction with previous observations on the presence of autoreactive T cells in RA patients may argue for a pathogenetic role also in human RA.