Processing and presentation of the rheumatoid arthritis candidate autoantigen aggrecan, by antigen-specific B cells

Abstract

The proteoglycan aggrecan, which is a major structural component of cartilage, has been identified as a candidate autoantigen in rheumatoid arthritis (RA). This is principally due to its degradation early in the disease pathology, its ability to induce an RA-like disease in mouse models and the presence of elevated numbers of reactive T and B cells in RA patients. Studies have also defined an essential requirement for autoantigen-specific B cells as antigen presenting cells (APC) in RA although the cellular mechanisms involved in antigen processing and presentation of joint-derived autoantigens by B cells remains unknown. To investigate the role of autoreactive B cells as APC in RA, I have used two complimentary approaches to generate B cells expressing an aggrecan-specific B cell receptor (BCR). The first was based on a modified monoclonal antibody production protocol and the second involved the transfection of B lymphoma cells with newly generated plasmids encoding an aggrecan-specific BCR. Using the second approach, I have successfully generated aggrecan-specific B cell lines (C71-4C5 and C71-5F10). I have shown that these B cells specifically bind aggrecan leading to efficient processing and the generation of the immunogenic T cell epitope 84-103 that is recognised by both aggrecan-specific T cell hybridomas and CD4+ T cells isolated from an aggrecan-specific TCR transgenic mouse. The aggrecan-specific B cells are able to present aggrecan at least 104 fold more efficiently than non-specific B cells, 102 fold more efficiently than macrophages and comparable to that seen by dendritic cells. By using a panel of inhibitors, I have also shown that the generation of the 84-103/MHC complex by aggrecan-specific B cells requires an acidic environment, proteolysis by aspartic, serine and metalloproteinases and the “classical” pathway of MHC class II biosynthesis. During this PhD, I have highlighted a novel role for aggrecan-specific B cells as important APC involved in aggrecan-presentation, as well as elucidating a role for metalloproteinases in aggrecan processing and presentation by APC.