MOLECULES AND GENES INVOLVED IN TYPE I DIABETES

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The largest gap in our understanding of the immunology of autoimmune diseases is defining the nature of the antigen involved in their pathogenesis. Although antibodies and T cell clones able to react against self determinants have been characterized in different autoimmune diseases, including IDDM. It has been difficult thus far to identify a single prevailing target antigen for each pathologic condition.The ultimate aim of the research is to determine the most important environmental factors involved in the pathogenesis of IDDM. Our recent studies on the repertoire of the T cells infiltrating the pancreata of diabetic patients at the onset of IDDM provide compelling evidence for a "superantigen" as a likely candidate for the critical environmental factor involved in the pathogenesis of IDDM. This new evidence unifies a number of apparent inconsistencies in the data generated from such diverse fields as virology, genetics, immunology and epidemiology. The investigators are in a quite favorable position to further investigate this hypothesis since isolated human pancreatic islet cells from patients who died as a result of a dramatic diabetic onset are available to them in a relatively large quantity. Further, they have already generated 61 independent T cell clones derived from the infiltrating lymphocytes from such islets and have begun their characterization. Finally, the variety of expertise available among the members of the team in the areas of T cell cloning, TCR Valpha and Vbeta repertoire testing, viral RNA detection and characterization, human and mouse anti-islet monoclonal antibody production, generation and screening of cDNA expression libraries with specific antibodies, and analysis of gene products obtained from the Baculovirus system, guarantees that this project will quickly proceed to the fulfillment of the following specific aims: a) To characterize the T cells infiltrating the pancreatic islets of patients who died at onset of IDDM in order to more precisely determine the role they play in the immunologic process that results in the destruction of the pancreatic beta cells. This characterization will include an analysis of cytokine expression in total mRNA from freshly isolated islets containing infiltrating T cells as well as in mRNA from the T cell clones derived from these islets. b) To determine the presence of viral transcripts or viral products in the cells of the pancreatic islets from the IDDM patients using molecular strategies, including R-U5 PCR and differential display of mRNAs by PCR. c) To identify and biochemically isolate proteins from human pancreatic islet cell membrane extracts that specifically activate any of the T cell clones. Finally, d) To screen cDNA expression libraries generated from the isolated pancreatic islets from those IDDM patients in order to identify and characterize the relevant antigen-encoding genes. These studies will allow a more comprehensive understanding of the etiology of IDDM which will, in turn, provide the basis for new preventive or immunotherapeutic approaches to the treatment of this disease.