Toxicity of low dose azathioprine and 6-mercaptopurine in rat hepatocytes. Roles of xanthine oxidase and mitochondrial injury

• Published in: Journal of hepatology 2004-03, Vol.40 (3), p.454-463
• Main Authors: Tapner, Michael J, Jones, Brett E, Wu, Wan M, Farrell, Geoffrey C
• Format: Article
• Language: English
• Subjects: 6-Mercaptopurine; Adenosine Triphosphate - metabolism; Allopurinol - pharmacology; Animals; Antioxidants - pharmacology; Azathioprine; Azathioprine - administration & dosage; Azathioprine - poisoning; Biological and medical sciences; Cell Differentiation; Cell Survival; Cells, Cultured; Chromans - pharmacology; Drug toxicity and drugs side effects treatment; Enzyme Inhibitors - pharmacology; Gastroenterology. Liver. Pancreas. Abdomen; Glutathione - metabolism; Glutathione Disulfide - metabolism; Hepatocytes - drug effects; Hepatocytes - enzymology; Hepatocytes - metabolism; Hepatocytes - pathology; Hepatotoxicity; Male; Medical sciences; Mercaptopurine - administration & dosage; Mercaptopurine - poisoning; Microscopy, Electron; Mitochondria, Liver - metabolism; Mitochondria, Liver - pathology; Mitochondrial injury; Necrosis; Oxidative Stress; Pharmacology. Drug treatments; Rats; Rats, Wistar; Time Factors; Toxicity: digestive system; Xanthine oxidase; Xanthine Oxidase - metabolism
• ISSN: 0168-8278; 1600-0641
• DOI: 10.1016/j.jhep.2003.11.024

To study effects of pharmacologic concentrations of azathioprine and 6-mercaptopurine (6-MP) on rat hepatocytes. Hepatocytes cultured on matrigel were incubated with azathioprine or 6-MP; effects of putative protective agents were studied. Viability (LDH leakage), reduced (GSH) and oxidized glutathione (GSSG), mitochondrial (mt) GSH, ATP and ultrastructural changes were determined. Azathioprine and 6-MP (0.5–5 μmol/l) reduced viability 5–34% at day 1 and 42–92% by day 4. Allopurinol (20 μM) (xanthine oxidase inhibitor) and 2 mM Trolox (vitamin E analog) together provided near complete protection. During culture with azathioprine, GSSG increased before cell death and there was a disproportionate reduction of mtGSH and ATP, together with ultrastructural abnormalities in mitochondria. All changes were prevented by allopurinol and trolox. Discontinuation of 1 μmol/l azathioprine restored ATP levels and arrested cell injury, while culture in glucose-enriched media augmented ATP levels and ameliorated cell death. Clinically relevant concentrations of azathioprine and 6-MP are toxic to rat hepatocyte cultures by a mechanism that involves oxidative stress, mitochondrial injury and ATP depletion. This can lead to irreversible de-energization and cell death by oncosis (necrosis).