Using a proteomic strategy, we recently identified a new class of post-translational protein modification that occurs in pancreatic beta cells. This modification is a result of insulin fragments that become covalently linked to other protein fragments that form in beta cells. The resulting hybrid insulin peptides (HIPs) contain non-germline encoded amino acid sequences and provide a plausible explanation on how tolerance to self-antigens may be lost leading to the development of type 1 diabetes (T1D). We demonstrated that autoreactive CD4 T cells, isolated from the residual islets of T1D organ donors, target various HIPs. Additionally, diabetes triggering CD4 T cells from non-obese diabetic (NOD) mice, a major animal model for T1D, were shown to be HIP reactive. Using mass spectrometric analyses on cell extracts of beta cell tumors, isolated from NOD-RIPTAg mice, we confirmed the presence of HIPs in those cells. Key questions that arise from these observations are whether HIP formation is a process that takes place in human beta cells, and whether individuals that do not develop diabetes also form HIPs. BALB/c mice do not develop spontaneous diabetes and here we analyzed pancreatic islets from these mice by mass spectrometry. We verified the presence of various HIPs in those islets. Data indicate that the quantity of HIPs formed in NOD islets is equivalent to the quantity formed in BALB/c islets. However, additional quantification steps will be required to validate these observations. It is difficult to obtain sufficient amounts of residual islets from T1D organ donors that allow a detailed proteomic analysis, but our findings now set the stage to look at more accessible islets of non-T1D organ donors to verify the formation of HIPs in humans.


T. Wiles: None. K.M. Haskins: None. T. Delong: None.

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