Diabetes mellitus (DM) is a metabolic ailment caused by inadequate production or utilization of insulin, featured by excessive amounts of glucose in the blood. The incidence of diabetes has been rising globally. Diabetes is often diagnosed late, after the insulin-producing pancreatic ?-cells have already undergone severe damage, leading to multi-organ complications. To avoid these complications, prevention, early diagnosis, timely intervention, and development of new drugs are necessary. Animal models, particularly rodents, have been established for the study of diabetes and diabetic complications, and have helped to identify drug targets and test new drugs for DM. However, rodent physiology vastly differs from human; hence, rodent models cannot precisely mimic human diabetes. Conversely, physiology and glucose metabolism of the pig closely resemble that of the human, proving the pig a suitable model system to study human diabetes, particularly prediabetic metabolic changes. Early detection of these metabolic changes is vital for timely interventions. The two most common diabetes, type 1 and 2 diabetes (T1D and T2D), are caused by different environmental insults and multiple gene defects that are difficult to simulate in animal models. Opportunely, single gene defects of diabetes that are relatively easier to model in animals exist. For example, a recently reported point mutation in the sirtuin 1 (SIRT1) gene causes autoimmune-mediated pancreatic ?-cell destruction characteristic of T1D in human patients. To that end, the overall goal of this project is to create genome-edited swine with point mutations in the SIRT1 gene that will simulate important features of human T1D, including severe insulin deficiency and autoimmune-mediated pancreatic ?-cell destruction. This goal will be achieved by engineering SIRT1L100P mutations using CRISPR mediated homology-dependent repair (HDR) in Ossabaw fibroblasts, from which founder pigs with the specific single gene mutation will be generated through cloning. These gene-edited founder pigs will be characterized by clinical and pathological examinations. Introduction of SIRT1L100P mutation into the Ossabaw genome will exhibit typical clinicopathological features of T1D, particularly its hallmark autoimmune destruction of pancreatic ?-cells, unlike any current animal model of human diabetes. Owing to the natural propensity of Ossabaw pigs to develop T2D, the edited-pig also enables modeling large animal model of double diabetes (DD), after diet induction. The SIRT1L100P model of T1D will open new avenues for testing and developing breakthrough therapies for human use and may shift the existing clinical paradigm from frequent exogenous insulin injection towards novel corrective, or even curative, immune therapies.
This project will fund the development of monogenic autoimmune-mediated swine model of type 1 diabetes (T1D). We use gene-editing technology to introduce SIRT1L100P mutation into swine genome, which cause T1D in human. We propose that this unique large animal model mimics the human condition, and help accelerate the development of new diagnostic biomarkers and novel therapeutic targets for T1D that may lead to cure.
