Prevention of oxidative injury in islet isolation

Islet transplantation has emerged as a plausible therapy for the treatment of type 1 diabetes. However, islets are faced with various types of stress related to the isolation and transplantation procedure. Variable periods of ischemia and oxidative stress can affect the number and function of islets recovered, frequently having to pool islets from more than one donor to achieve normoglycemia in one recipient.; Our rodent model of pancreatic warm ischemia showed that ischemia during preservation and islet isolation increased oxidative stress, damaging islets and affecting isolation outcomes. Our studies further revealed that by adding an antioxidant precursor (L-Glutamine) to the standard intraductal pancreas perfusate together with the collagenase enzyme, we were able to increase the levels of Glutathione in islets. This showed improvement in islet yields, viability and function after isolation and transplantation, compared to the standard intraductal flush. Furthermore, when applying this therapy in the clinical setting, isolation outcomes were improved by glutamine treatment and oxidative stress was decreased in glutamine-treated islets leading to lower oxidative injury and apoptosis than the control. As in the rodent studies, in vivo islet function was also improved after transplantation into diabetic athymic mice, rendering a higher percentage of transplanted mice normoglycemic in a shorter period of time than the control.; Ischemia produces damage of mitochondrial structures and precedes the occurrence of oxidative stress by the introduction of O2, an entity known as reperfusion injury. We tested the effect of pancreatic intraductal administration of a hemoglobin-based O2 carrier (PolyHeme) loaded with O2, after 30 min warm ischemia. Viability, in vitro and in vivo function of islets were significantly improved by PolyHeme pretreatment. Mitochondrial integrity and function were also superior in treated islets shown by improved mitochondrial membrane potential and increased fluorescent intensity evenly distributed around the nuclei. In our study, the introduction of O2 by PolyHeme did not increase oxidative stress or damage to the islets.; The intraductal administration of Glutamine and Hemoglobin O2 carriers (PolyHeme) as a simple addition to current standard isolation techniques provides protection of pancreatic islets from oxidative stress and ischemia, improving islet isolation outcomes and islet function after transplantation.