Effects of azathioprine, 6-mercaptopurine, and 6-thioguanine on canine primary hepatocytes

Abstract

OBJECTIVE To investigate the cytotoxic effects of azathioprine, 6-mercaptopurine, and 6-thioguanine on canine hepatocytes. SAMPLE Commercially available cryopreserved canine primary hepatocytes. PROCEDURES The study consisted of 2 trials. In trial 1, hepatocytes were incubated with azathioprine, 6-mercaptopurine, or 6-thioguanine at 1 of 6 concentrations (0.468, 0.937, 1.875, 3.750, 7.500, or 15.000 µmol/L) for 24, 48, or 72 hours. At each time, cell viability and lactate dehydrogenase (LDH) activity were determined for each thiopurine-concentration combination, and alanine aminotransferase (ALT) activity was determined for cells incubated with each thiopurine at a concentration of 15 µmol/L. In trial 2, hepatocytes were incubated with azathioprine, 6-mercaptopurine, or 6-thioguanine at 1 of 3 concentrations (18.75, 37.50, or 75.00 µmol/L) for 24 hours, after which the free glutathione concentration was determined for each thiopurineconcentration combination and compared with that for hepatocytes incubated without a thiopurine (control). RESULTS Incubation of hepatocytes with each of the 3 thiopurines adversely affected cell viability in a time- and concentration-dependent manner; however, this decrease in cell viability was not accompanied by a concurrent increase in LDH or ALT activity. Likewise, free glutathione concentration for hepatocytes incubated for 24 hours with supratherapeutic thiopurine concentrations (> 18.75 µmol/L) did not differ significantly from that of control cells. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that thiopurines adversely affected the viability of canine hepatocytes in a time- and concentration-dependent manner but had a nonsignificant effect on the LDH and ALT activities and free glutathione depletion of those hepatocytes.