COLD AND CRYOPRESERVATION OF MONKEY LIVER SLICES

Both cynomolgus and rhesus monkeys are utilized in chemical toxicity screening and drug development studies by industry and government laboratories. Along with better utilization of their tissue for in vivo studies, it would be advantageous if the tissue could be cold-preserved or cryopreserved for future in vitro experimentation. Therefore, livers were excised from control monkeys and precision-cut tissue slices were prepared. The objective of this study was twofold: to compare cold-preservation solutions (V-7 and Viaspan) and to compare controlled-rate and vitrification cryopreservation protocols. Monkey liver slices were cold-stored in V-7 or Viaspan preservation solutions for 7 days. V-7 maintained slice viability for 5 days whereas Viaspan maintained slice viability for 1 day. In the controlled-rate freezing procedure, slices were exposed to 10% dimethyl sulfoxide and 90% fetal calf serum (FCS); cooled at the rate of 0.5°C, 1°C, or 12°C per min to -70°C; then placed into liquid nitrogen. Vitrification was accomplished by exposing slices stepwise to increasing concentrations of 1,2-propanediol (1.2, 2.4, and 4 M) in FCS with direct submersion into liquid nitrogen. In both protocols, slices were rewarmed quickly to 37°C and then incubated in FCS for 4 h. Three viability parameters were used to measure slice viability - retention of potassium, leakage of lactate dehydrogenase, and synthesis of protein. Liver slices cryopreserved at a rate of 0.5°C per min and vitrified successfully retained 80 to 90% of their viability. These results confirm the feasibility of functional cold preservation and cryopreservation protocols for monkey liver slices that would allow for a more efficient use of monkey tissue.