Optimizing Temperature and Oxygen Supports Long-term Culture of Human Islets

摘要

Background: Islet transplantation is a promising treatment for type-1 diabetes; however, donor shortage is a concern. Even when a pancreas is available, low islet yield limits the success of transplantation. Islet culture enables pooling of multiple low-yield isolations into an effective islet mass, but isolated islets rapidly deteriorate under conventional culture conditions. Oxygen (O-2) depletion in the islet core, which leads to central necrosis and volume loss, is one of the major reasons for this deterioration. Methods. To promote long-term culture of human islets in PIM-R medium (used for islet research), we adjusted temperature (12 degrees C, 22 degrees C, and 37 degrees C) and O-2 concentration (21% and 50%). We simulated the O-2 distribution in islets based on islet O-2 consumption rate and dissolved O-2 in the medium. We determined the optimal conditions for O-2 distribution and volume maintenance in a 2-week culture and assessed viability and insulin secretion compared to noncultured islets. In vivo islet engraftment was assessed by transplantation into diabetic nonobese diabetic-severe combined immunodeficiency mouse kidneys. We validated our results using CMRL 1066 medium (used for clinical islet transplantation). Results. Simulation revealed that 12 degrees C of 50% O-2 PIM-R culture supplied O-2 effectively into the islet core. This condition maintained islet volume at greater than 90% for 2 weeks. There were no significant differences in viability and function in vitro or diabetic reversal rate in vivo between 2-week cultured and noncultured islets. Similar results were obtained using CMRL 1066. Conclusions. By optimizing temperature and O-2 concentration, we cultured human islets for 2 weeks with minimal loss of volume and function.