Interaction of Cooling Rate, Warming Rate, and Extent of Permeation of Cryoprotectant in Determining Survival of Isolated Rat Islets of Langerhans During Cryopreservation

Cryopreservation of islets of Langerhans offers a number of important benefits for attempts to cure diabetes by transplantation. In the published literature, a variety of cooling rates, ranging from 0.25 to 75°C/min, in conjunction with warming rates of 4–200°C/min have been proposed to give optimal preservation of islets. In view of the general importance of rates of temperature change in determining survival and because of the possibility of modulating tissue immunogenicity by freezing and thawing, we have studied the interaction of cooling rate and warming rate for isolated rat islets that had been either fully or partially equilibrated with 2 M dimethyl sulfoxide (DMSO). Batches of islets were stored at −196°C after cooling at 0.3, 3.0, 10, 30, 60, 150, or 1000°C/min and then warmed at either 10 or 50°C/min. Survival was assessed by measuring the secretion of insulin during static incubation in alternating nonstimulatory and stimulatory media. Cooling rates extending over three orders of magnitude proved not to be a major determinant of survival when the islets were equilibrated with 2 M DMSO: >50% survival was achieved at all cooling rates studied when the warming rate was at 50°C/min. Peak survival (83%) was attained at a cooling rate of 0.3°C/min, but only slightly lower recoveries were obtained at 60 and >1000°C/min. However, in islets only partially equilibrated with cryoprotectants, functional recovery was highly dependent on the cooling and warming rates, with peak survivals after slow cooling and rapid warming. Full permeation of the tissue with cryoprotectant offered maximal recovery of function.