Kanz M F, Gunasena G H, Kaphalia L, Hammond D K, Syed Y A
Department of Pathology, University of Texas Medical Branch, Galveston 77555-0609, USA.
Toxicol Appl Pharmacol. 1998 Jun;150(2):414-26. doi: 10.1006/taap.1998.8382.
The threshold for hepatotoxicity and cholestasis induced by methylene dianiline (DAPM) in rats is between 25 and 75 mg/kg (Bailie et al., Environ. Health Perspect., 124, 25-30, 1993). Our objectives were to determine if a minimally toxic dose of DAPM provided a model system for studies of selective injury to biliary epithelial cells (BEC) in vivo. Thus, we examined the effects of 50 mg DAPM/kg on (1) biliary constituents, (2) liver constituents likely involved in DAPM biotransformation/detoxification, and (3) early morphological and histochemical changes in the liver. Male Sprague Dawley rats had biliary cannulas positioned under pentobarbital anesthesia. After 1 h of control bile collection, rats received 50 mg DAPM/kg po in 35% ethanol or 35% ethanol only. Bile was collected for another 6 h. Histochemical, ultrastructural, and biochemical liver alterations were assessed at 3 h or at 3 and 6 h. DAPM had minimal effects on biliary bile salt and bilirubin excretion over 6 h. Biliary glucose and protein excretion were increased approximately 2-fold starting in Hour 1, while inorganic phosphate excretion was not increased until Hour 2. Biliary glutathione excretion initially increased (Hour 1) but then declined steadily for 5 h. Microsomal cytochrome P-450 activities were transiently decreased at 3 h but had returned to control values by 6 h. Liver glutathione (GSH and GSSG) was not affected by DAPM at 3 or 6 h. Necrosis of intrahepatic bile ducts was severe at 6 h with moderate injury in smaller bile ducts. Ultrastructural alterations were observed in BEC mitochondria and microvilli at 3 h with no apparent alterations in hepatocyte mitochondria or tight junctions between cells. In addition, histochemical staining of liver sections and assays of mitochondrial enzyme activities in vitro at 3 h revealed no loss of mitochondrial function in hepatocytes. These results provide strong evidence for defining DAPM as a selective bile duct toxicant.
大鼠体内亚甲基二苯胺（DAPM）诱导肝毒性和胆汁淤积的阈值在25至75毫克/千克之间（Bailie等人，《环境健康展望》，第124卷，第25 - 30页，1993年）。我们的目标是确定最小毒性剂量的DAPM是否能为体内胆管上皮细胞（BEC）选择性损伤研究提供一个模型系统。因此，我们研究了50毫克DAPM/千克对（1）胆汁成分、（2）可能参与DAPM生物转化/解毒的肝脏成分以及（3）肝脏早期形态学和组织化学变化的影响。雄性Sprague Dawley大鼠在戊巴比妥麻醉下放置胆管插管。在收集1小时对照胆汁后，大鼠经口给予50毫克DAPM/千克，溶于35%乙醇中，或仅给予35%乙醇。再收集6小时胆汁。在3小时或3和6小时评估肝脏的组织化学、超微结构和生化改变。DAPM在6小时内对胆汁胆盐和胆红素排泄影响极小。胆汁葡萄糖和蛋白质排泄从第1小时开始增加约2倍，而无机磷酸盐排泄直到第2小时才增加。胆汁谷胱甘肽排泄最初增加（第1小时），但随后5小时稳步下降。微粒体细胞色素P - 450活性在3小时短暂降低，但到6小时已恢复至对照值。3小时或6小时时，肝脏谷胱甘肽（GSH和GSSG）不受DAPM影响。6小时时肝内胆管坏死严重，较小胆管有中度损伤。3小时时在BEC线粒体和微绒毛中观察到超微结构改变，肝细胞线粒体或细胞间紧密连接无明显改变。此外，3小时时肝脏切片的组织化学染色和体外线粒体酶活性测定显示肝细胞线粒体功能未丧失。这些结果为将DAPM定义为选择性胆管毒物提供了有力证据。
