Murine islet cryo preservation and corticosteroids: Functional studies

Abstract

Knowledge of protective effects of corticosteroids on traumatized cells prompted us to test the potential benefit of islet cryopreservation in the presence of hydrocortisone. Neonatal murine islets were isolated by collagenase, followed by 2- to 3-day tissue culture. Precryopreservation glucose-stimulated (50–500 mg/dl) insulin release was 25–388% above basal (mean = 113%) in 18 20 fresh islet preparations. Subsequent freezing was done in RPMI 1640 medium plus 10% ( v v ) heat-inactivated fetal calf serum and 10% ( v v ) Me 2SO with or without 1 mg/ml hydrocortisone at 0.25 °C per minute in a programmed freezing system, to −80 °C, and stored for >60 days at −196 °C. Thawing, by transfer to room air, was followed by dilution, 4 × ( v v ), in 4 °C RPMI plus 10% protein, after which glucose-stimulated insulin release was reassessed, showing 56–280% response over basal in 3 8 steroid-treated preparations and 20–220% response in 3 10 control preparations. Basal insulin release was 0.72 ng/μg protein/hr in fresh islets ( N = 20) and 0.22 ng/μg protein/hr after freeze-thawing. We conclude that functional islet survival by this method is approximately 30% and that hydrocortisone did not improve viability.