Studies of chemical toxicity to fresh and cryopreserved rat hepatocytes

Abstract

Isolated hepatocytes are useful for studying the metabolism and mechanisms of hepatic toxicity of foreign chemicals. A problem with using human hepatocytes is the limited and irregular availability of normal human liver. Cryopreservation could provide a useful way of storing hepatocytes until they are needed. As a preliminary step to using human hepatocytes we have compared the toxic response to chemical toxicants of primary cultures of fresh rat hepatocytes and rat hepatocytes cryopreserved as previously described ( G. Powis, K. S. Santone, D. C. Melder, L. Thomas, D. J. Moore, and T. J. Wilke, 1987. Drug Metab. Dispos. 15, 826). After 24 hr in culture the cryopreserved hepatocytes had a plating efficiency 75% that of noncryopreserved hepatocytes. The cultured cryopreserved hepatocytes showed a small increase in spontaneous lactate dehydrogenase release compared to that of cultured noncryopreserved hepatocytes. A similar toxic chemical-induced increase in lactate dehydrogenase release occurred in the cultured cryopreserved as in the noncryopreserved hepatocytes. The 50% effective concentrations (EC50) for lactate dehydrogenase release (±SE, n = 3 preparations) from cultured cryopreserved and noncryopreserved hepatocytes for chlorpromazine were 235 ± 20 and 215 ± 30 μ m, for cadmium chloride 200 ± 5 and 272 ± 23 μ m, and for menadione (2-methyl-1,4-naphthoquinone) 24 ± 7 and 44 ± 8 μ m, respectively. The EC50 values for intracellular glutathione depletion in cultured cryopreserved and noncryopreserved hepatocytes were for chlopromazine 200 ± 8 and 235 ± 8 μ m, for cadmium chloride 242 ± 19 and 213 ± 7 μ m, and for menadione 22 ± 2 and 21 ± 3 μ m, respectively. The results show that cryopreservation offers a practical way of storing rat hepatocytes for studies of chemical toxicity.