In this grant, attempts will be made to identify a putative arthritogenic epitope of B. burgdorferi using molecular biologic techniques to map T and B cell epitopes of the spirochete. The postulate is that in patients who are genetically susceptible to chronic Lyme arthritis, one antigenic fragment of B. burgdorferi combines with a distinct conformation encoded by the DR4 haplotype. Preliminary work suggests that this epitope may be located on the OspA or B proteins. The interaction of this MHC-peptide complex with CD4-positive T cells may trigger an untoward immune response with autoreactive features leading to chronic arthritis. In an effort to identify such an epitope, fusion proteins will be generated in an E. coli expression vector using restriction fragments of the 49 kb plasmid encoding for the OspA and B proteins. To begin the process of mapping B cell epitopes, the fusion proteins will be tested in an ELISA for their reactivity with the sera of DR4-positive of negative patients who have the antibody responses associated with either brief or chronic Lyme arthritis. To begin the process of T cell mapping, the fusion proteins will be tested in a proliferative assay for their reactivity with peripheral blood and/or synovial fluid lymphocytes from the same groups of patients. B. burgdorferi-reactive T cell clones will be generated and those that react with the OspA and B proteins and with the fusion proteins will be identified. The sequences of the immunoreactive fusion proteins will be deduced from the known sequences of the OSPA and B proteins, and overlapping synthetic peptides will be made and tested against patients' sera and cells in an effort to identify the putative arthritogenic peptide sequence. This work is likely to influence the treatment of patients with chronic Lyme arthritis; it will provide guidelines for mapping other T and B cell epitopes of B. burgdorferi, and it may have implications for vaccine development since immunity to the Osp proteins may play a role in protection, but may also trigger an autoimmune response in genetically susceptible individuals.
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