Mechanisms of Drug-Induced Liver Disease

今年，我们提供了更完整的超微结构证据，证明氟烷的三氟乙酰化蛋白加合物(TFAPA)可能导致肝脏损伤。所采取的方法是用肝毒性剂量的氟烷治疗雌性Balb/CJ小鼠。12小时后，这是肝脏毒性的早期时间点，从氟烷处理和未处理的小鼠身上取出肝脏碎片。对切片进行化学固定，用常规的透射电子显微镜(TEM)观察氟烷剂量对亚细胞超微结构的影响。透射电子显微镜图像显示许多肝细胞的超微结构改变。与对照组相比，氟烷处理组小鼠肝细胞含有更多的脂滴，更多的线粒体受损和更多的扩张的平滑内质网。各种形态变化表明线粒体受损。包括基质中的小的透光区、明显的肿胀、致密的絮状体、内外膜的分离和线粒体的凝聚。将部分肝组织固定、冷冻、冷冻，用TFAPA特异性抗血清对肝细胞进行TFAPA标记。氟烷治疗后形态正常的肝细胞在整个细胞上有稀疏的免疫金标记物。相反，许多肝细胞在内质网、细胞质和较小程度的细胞核上被大量标记。这些肝细胞表现为脂滴增多，部分线粒体受损。然而，这些肝细胞中的大多数线粒体上的标记是稀疏的，包括一些形态受损的线粒体。一些浓缩的线粒体以及过氧化物体和自溶酶体被标记为与周围细胞质相似的标记。我们的结果表明，氟烷处理12h后，TFAPA在受损的肝细胞中集中在几个隔室中，但在未受损和部分受损的线粒体中的浓度要低得多。这表明，尽管线粒体损伤是肝毒性的早期迹象，但氟烷引起的线粒体损伤可能是通过作用于其他细胞质细胞器间接发生的。 结论：氟烷诱导的小鼠肝损伤模型继续为药物蛋白加合物引起肝损伤的亚细胞部位提供重要信息。

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.