PMID: 8937857Nov 1, 1996Paper

Cryopreservation of rat and mouse hepatocytes. II. Assessment of metabolic capacity using testosterone metabolism

Drug Metabolism and Disposition : the Biological Fate of Chemicals
N J SwalesJ Caldwell

Abstract

Cryopreservation of hepatocytes is widely used, but to validate the use of cryopreserved (CP) hepatocytes in metabolic studies, CP cells must compare favorably with fresh cell activities. We have assessed the metabolic capacity of fresh and CP rat and mouse hepatocytes in primary culture. Total cytochrome P450 (P450) contents and metabolism of testosterone were measured up to 72 hr in culture. At 0 hr, total P approximately 450 in CP rat hepatocytes was 102.5 +/- 32.8 pmol/10(6) cells, compared with fresh rat hepatocytes that had 148.2 +/- 75.7 pmol/10(6) cells. The P450 contents of mouse hepatocytes were also unaltered by cryopreservation (176.7 +/- 56.0 pmol/ 10(6) fresh cells; 196.4 +/- 59.9 pmol/10(6) CP cells). There were no significant differences in the total P450 contents of fresh and CP rat and mouse cell cultures with time over 72 hr in culture. The overall metabolism of testosterone was lower in CP suspensions than in freshly isolated hepatocytes. When CP hepatocyte suspensions were permeabilized (with digitonin) and incubated with NADPH and ATP, testosterone metabolism was significantly increased. Testosterone hydroxylase activities (16 alpha-, 6 beta-, 2 alpha-, and 7 alpha-hydroxylase) were equivalent in fresh and...Continue Reading

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