Mechanisms of Drug-Induced Liver Disease

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This year we provide more complete ultrastructural evidence of sites where trifluoroacetylated proteins adducts (TFAPA)of halothane may cause liver injury. The approach taken was to treat female Balb/cJ mice with a hepatotoxic dose of halothane. After 12 hours, a time point early in hepatotoxicity, liver pieces were removed from halothane treated and untreated mice. Pieces were chemically fixed for conventional transmission electron microscopy (TEM) to investigate any changes in subcellular ultrastructure resulting from the halothane dose. TEM images indicated ultrastructural changes in many hepatocytes. Hepatocytes from halothane treated mice contained more lipid droplets, more damaged mitochondria and more dilated smooth endoplasmic reticulum than hepatocytes from control mice. Various morphological changes indicated mitochondrial damage. These included small lucent regions in the matrix, overt swelling, dense flocculent bodies, separation of the inner and outer membrane and condensation of the mitochondrion. Some pieces of liver were fixed, cryoprotected and frozen for cryo-immunogold labeling to localize TFAPA in the hepatocytes, using TFAPA specific antiserum. Hepatocytes that appeared morphologically normal after the halothane treatment had sparse immunogold label over the entire cell. In contrast, many hepatocytes were heavily labeled with gold over the endoplasmic reticulum, cytoplasm and to a lesser extent, the nuclei. These hepatocytes showed increased lipid droplets and some damaged mitochondria. However, the labeling was sparse over most of the mitochondria in these hepatocytes, including some mitochondria with morphological damage. Some condensed mitochondria as well as peroxisomes and autolysosomes were labeled similar to surrounding cytoplasm. Our results indicate that 12 hours after treatment with halothane, TFAPA are concentrated in several compartments in the damaged hepatocytes, but are at much lower concentrations in undamaged and some damaged mitochondria. This suggests the possibility that, although mitochondrial damage is an early sign of hepatotoxicity, halothane-induced damage of mitochondria may occur indirectly through action on other cytoplasmic organelles. Conclusion: The murine model of halothane-induced liver injury continues to provide important information concerning subcellular sites where protein adducts of drugs may initiate liver injury caused not only by halothane, but also by other drugs.