Although the thymus is not generally considered to be a site of B cell activity, in myasthenia gravis (MG) it is a scene of prominent B cell responses. This activity, which is highlighted by production of anti-AChR antibody (anti-AChR), likely reflects intrathymic immune events that are important in disease pathogenesis. Several reasons could account for the prominent albeit incongruous thymic B cell responses. They include: 1) thymic B cells are a particularly reactive population, one which can be distinguished on the basis of phenotypic properties from blood B cells, 2) the thymic microenvironment is particularly supportive of B cell differentiation and 3) thymic B cells have undergone some degree of differentiation in vivo perhaps as a result of locally encountered stimuli. The experiments described in this proposal will help to 1) fully characterize the phenotypic properties of these unusual thymic B cells; 2) correlate in vitro function of these cells with their phenotypic properties; 3) determine the phenotype of antigen specific thymic helper T cells and thus elucidate the developmental stage at which helper activity is acquired in the thymus; 4) determine if MG thymus cells constitutively produce soluble factors required to support B cell differentiation and 5) characterize the thymic populations expressing AChR prior to determining if such receptor cells provide a stimulus for anti-AChR production. Whenever possible, comparisons will be made between 1) MG thymic cells and autologous blood cells, 2) thymic cells of control subjects. Technical approaches to accomplish the above objectives include: cell subset analysis with monoclonal antibodies along with automated flow cytometry and immunofluorescence microscopy, subset fractionation by affinity chromatography and FACS sorting for use in several in vitro cell culture systems, and ELISA and radioimmunoassay for measurement of Ig secreted into culture supernatants. Increased knowledge of the mechanisms underlying the anomalous thymic B cell responses in MG will likely enhance our understanding of aberrant local intrathymic immune events and ultimately may help to clarify mechanisms responsible for initiation, potentiation or perpetuation of disease.
