Chronic beryllium disease (CBD) is a pulmonary granulomatosis due to a cell-mediated immune response to beryllium. This class of immune response is typically triggered by an interaction involving an antigen, an HLA molecule and specific T cell receptors. The goal of this project is to define molecularly the structures of the T cell receptor (TcR) and the HLA Class II molecules involved in the immune response to beryllium. This project involves 4 major goals. 1. Is the T cell immune response to beryllium restricted to specific subfamilies of T cell receptor (TcR) alpha and beta chains? Using lung lavage lymphocytes and lung T cell lines that are reactive to beryllium, specific TcR subfamilies will be identified using primers specific for TcR alpha and beta chains and amplification using PCR. TcR alpha and beta chain cDNAs will be captured into a plasmid vector and cDNA libraries will be created. The specific TcR alpha and beta chains will be cloned and sequenced. The common variable regions of the alpha and beta chains will be identified. 2. Can polyclonal or monoclonal antibodies to these TcR subfamilies block the lymphocyte proliferative response to beryllium? The cDNA sequences of the TcR alpha and beta chains that are of interest will be cloned into expression vectors and the protein used to produce polyclonal and monoclonal antibodies. These antibodies will be tested for their ability to identify beryllium sensitive T cell lines and their ability to inhibit the lymphocyte proliferative response of beryllium sensitive T cell lines. 3. Is there a relationship between HLA Class II molecules and chronic beryllium disease? Patients with chronic beryllium disease will be serologically typed for DR and molecularly typed for DP. Preliminary studies suggest an association of HLA-DP with CBD. Using PCR, the specific HLA-DP alleles of patients with CBD will be correlated with their T cell subfamily usage. 4. Do specific HLA Class II molecules present beryllium to T Cells? Monoclonal antibodies against specific HLA Class II molecules will be used to block lymphocyte proliferative responses to beryllium. Stable L cell transfectants will be generated that will express the relevant HLA molecules identified in Section 3. The ability of these transfectants to present beryllium to beryllium sensitive T cell lines will be tested. These studies should help to define the specific epitopes on HLA Class II molecules that are involved in the presentation of beryllium-complexes to TcR. To further understand the interaction of beryllium, HLA Class II molecules, and TcR we will examine whether the beryllium-macromolecule complex requires intracellular processing or whether beryllium simply binds extracellularly. A detailed understanding of the structures of the TcR and the HLA Class II molecules involved in chronic beryllium disease will be the groundwork for the development of new methods for the diagnosis and treatment of this disorder and may add new insights into the pathogenesis and treatment of sarcoidosis.
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