ABSTRACT
This chapter deals with a technique for isolating intact islets of Langerhans from the pig pancreas based on our experience performing approximately 750 isolations. The procedure we describe involves identification of an optimal donor pancreas, purification and in vitro culture of islets, diabetes induction in recipients, and transplantation of islets and their immunomodulation. Besides the sophistication of the technical equipment employed, the major factors influencing the isolation outcome are the pig breed, the number and morphology of the islets in the donor pancreas, the quality of the collagenase/neutral protease, and the skill of the team members.
Subject(s)
Cell Separation/methods , Islets of Langerhans Transplantation/methods , Islets of Langerhans , Transplantation, Heterologous/methods , Animals , Cell Survival , Diabetes Mellitus, Experimental/therapy , Female , Humans , Islets of Langerhans/immunology , Mice , Rats , Swine , Tissue Culture Techniques , Tissue and Organ Harvesting/methodsABSTRACT
PURPOSE: Preclinical experiments on large animals are indispensable for evaluating the effectiveness of diabetes therapies. Miniature swine are well suited for such studies due to their physiological and pathophysiological responses. METHODS: We compare two methods for inducing diabetes in Goettingen minipigs (GMP), in five with the beta cell toxin streptozotocin (STZ) and in five other GMP by total pancreatectomy (PE). Glucose homeostasis was assessed with the intravenous glucose-tolerance test (IVGTT) and continual monitoring of interstitial glucose levels. At conclusion of the observation period, the pancreata were examined histologically. Three non-diabetic GMP served as control group. RESULTS: The IVGTT revealed markedly diabetic profiles in both GMP groups. STZ-GMP were found to harbor residual C-peptides and scattered insulin-positive cells in the pancreas. PE-GMP survived the total pancreatectomy only with intensive postoperative care. CONCLUSIONS: Although both methods reliably induced diabetes in GMP, the PE-GMP clearly had more health problems and required a greater expenditure of time and resources. The PE-GMP model, however, was better at eliminating endogenous insulin and C-peptide than the STZ-GMP model.
