Effects of precryopreservation culture on survival of rat islets transplanted after slow cooling and rapid thawing

Abstract

Despite the frequent use of in vitro tissue culture before islet cryopreservation, no study has evaluated the ability of this procedure to improve the recovery or in vivo function of frozen-thawed islets. To evaluate this, quantities of 2500 Wistar-Furth (WF) rat islets were allocated to each of four groups ( n = 8 each): group 1, freshly isolated; group 2, 48 hr in in vitro culture; group 3, cryopreservation; group 4, cryopreservation after 48 hr in culture. Islets were frozen slowly at 0.25 °C/min and thawed rapidly at 200 °C/min. The number of islets recovered after culture or cryopreservation was determined and viability was assessed by measuring weekly indices of plasma glucose (PG), urine glucose (UG), urine volume (UV), and weight after implantation into the portal vein of streptozotocindiabetic WF recipients. Islet recovery was 97% after culture, 95% after cryopreservation, and 94% after culture-then cryopreservation. After implantation of group 1 and 2 islets, PG was <150 mg/dl at 1 week and UG and UV were normal by 1–2 weeks. Group 3 islets restored normoglycemia at 3 weeks and other indices of diabetes were reversed by 4 weeks; group 4 islets restored normoglycemia at 4 weeks and indices returned to basal after 4 weeks. At intravenous glucose tolerance testing (ivGTT), the K values (mean decline in glucose, %/min, ± SE) were group 1, 1.6 ± 0.3; group 2,1.5 ± 0.3; group 3, 0.6 ±0.1; and group 4, 0.7 ± 0.2. These data show that cryopreservation preserves freshly isolated or cultured islets that reverse the indices of diabetes after implantation. Tissue culture neither improves the recovery of rodent islets cryopreserved with these methods nor optimizes islet graft function.