Although B-cells are not necessary to initiate the inflammatory reactions of RA, these cells are involved in their perpetuation. Central to this perpetuation is the activation of subsets of B-cells by unknown (auto)antigens. Once activated, these cells proliferate in the synovium and elaborate Igs, only some of which are RF. Since the cells in the synovium eventually form aggregates, it is presumed that they create a lymphoid follicle that allows mutation of Ig V genes and the generation of new antigen-combining sites. Because RF have a major, defined autoantigenic target that is important clinically, they have been extensively studied. However, dilemmas relating to their combining sites remain. Much less attention has been paid to the antigenic reactivities of the nonRF, even though these are more plentiful than RF. The investigators propose to focus on these issues by addressing three Specific Aims. First, they will determine whether the synovial tissue in RA can sustain the diversification and selection of antigen-activated B-cells. This will involve analyzing by immunofluorescent and immunohistochemical techniques, the lymphoid and non-lymphoid cells in the synovium. They will specifically attempt to identify B-cells that resemble either follicular mantle, marginal zone, or germinal center lymphocytes, appropriate T-cells, follicular dendritic cells, and apoptotic B-cells. DNA sequencing data generated by RT in situ PCR will directly address whether somatic mutation can occur in RA synovium. Next, after production of B-cell lines from the various B-cell subsets, the investigators will determine the molecular genetic and serologic characteristics of the RF-producing B-cell lines derived from them to resolve several apparent structure/function incongruities. Finally, the investigators will attempt to determine the antigenic reactivities of the nonRF antibodies produced by plasma cells in the synovium in hopes that they will help to identify the initiating or perpetuating (auto)antigens of this disease. If new auto- or exo-antigenic specificities are determined, synovial fluid and sera from a series of RA patients, collected at different stages of their disease, will be screened for the presence of such antibodies.
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