The long term objective of this research proposal is to determine the mechanisms leading to immunologic destruction of the beta cells in the Islets of Langerhans in type I insulin-dependent diabetes in the NOD mouse. Specifically, we plan to clone and sequence cDNA copies of the I-A alpha and beta chains from the non-obese non-diabetic (NON) mouse and compare these to the sequences of the same genes from the NOD mouse in order to assess the role of specific allelic sequence in conferring susceptibility to IDDM in the NOD mouse. A separate set of experiments will attempt to produce NOD mice which are more than 95% B cell depleted, to determine whether these mice also develop diabetes with the same incidence as in the control NOD mouse line. This would indicate that the immunologic destruction of the beta cells is not primarily mediated by humoral antibody mechanisms. Parallel experiments will attempt to isolate T cell clones specific for beta cells from diabetic NOD mice. In order to isolate these T cell clones, we will establish an insulinoma cell line from mice which are H-2 identical with the NOD mouse. This will be done by crossing the NOD mouse strain with a transgenic mouse strain which has a high incidence of spontaneous islet cell tumors due to the previous introduction of the SV40 T antigen gene coupled to the insulin promoter. T cell lines isolated by stimulation with H-2 identical insulinoma cells will be characterized for MHC restriction (Class I vs. Class II), antigenic specificity, and for the ability to transfer diabetes to histocompatible, young male NOD mice (which normally do not develop diabetes). Finally, attempts will be made to isolate islet cell specific monoclonal antibodies from the NOD mouse using lymphocytes obtained from diabetic NOD mice and from NOD mice immunized with islet cells. Such monoclonal antibodies, if isolated, will be used to characterize cell surface molecules unique to beta cells.
