Acute hepatotoxicity of adriamycin-oxidized dextran (ADM-OXD), a newly developed antineoplastic macromolecule, was compared with that of adriamycin (ADM). Biochemical parameters of hepatic function and morphological changes were monitored up to day 3 after intravenous administration of ADM-OXD or ADM to male Wistar rats at a single dose of 45 mg/kg, measured by ADM content. From day 1 after ADM-OXD administration, lipid peroxide (LPO) level in the liver tended to increase and serum transaminase activity showed pronounced elevation from day 2. Morphologically, hyaline degeneration and single cell necrosis of hepatocyte were noted from day 0 (8 hours) to day 1 and resulted in diffuse centrilobular degeneration and necrosis on day 2. ADM administration did not induce hepatotoxicity, though it did induce acute cardiotoxicity. ADM-OXD-induced acute hepatotoxicity was amplified by intermittent treatment with glutathione depleting agent after drug administration and abated by antioxidative agent. These results indicate that oxidative stress is involved in the onset of hepatotoxicity induced by ADM-OXD. Quantitative determination by HPLC after ADM administration revealed immediate excretion of ADM from the liver. After ADM-OXD administration, on the other hand, ADM levels increased gradually and peaked on days 1 and 2. Hepatotoxicity specific to ADM-OXD is essentially attributable to this long-term retention of ADM. Furthermore, observation by fluorescent microscope revealed that ADM was distributed not only to hepatocytes but also to sinusoidal lining cells after ADM-OXD administration.

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Adriamycin-oxidized dextran (ADM-OXD), a newly developed antineoplastic macromolecule, has demonstrated greater antitumor action in tumor-bearing animals than ADM. However, excess dosing of ADM-OXD readily induces specific hepatotoxicity in vivo. In this study, the toxicity of ADM-OXD to cultured hepatocytes and Kupffer cells was compared with that of ADM.
With primary cultured hepatocytes, the results obtained were completely opposite to those obtained previously in vivo. Judging from the activity of leaked hepatic enzyme and cell viability measured by MTT assay, ADM-OXD had no toxic action toward hepatocytes. ADM, on the other hand, showed severe toxicity. Morphological observation agreed with the results of biological monitoring. Cell desquamation and increased cell necrosis induced by addition of ADM were not observed after addition of ADM-OXD to the medium. With primary cultured Kupffer cells, ADM-OXD did show toxicity, but to a lesser degree than ADM. Particularly as measured by neutral red (NR) assay, which evaluates lysosomal function through NR uptake capacity, ADM-OXD as well as ADM showed severe toxicity toward Kupffer cells. Toxicity of ADM-OXD to hepatocytes, however, was induced only when hepatocytes were co-cultured with Kupffer cells. From these results it was concluded that the presence of Kupffer cells is necessary for the appearance of ADM-OXD-induced toxicity in hepatocytes. It seems likely that ADM-OXD is incorporated not into hepatocytes but into Kupffer cells, and that ADM dissociated in Kupffer cells transfers into hepatocytes, where it eventually exhibits toxic action.

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Zinostatin stimalamer (ZS) derived from an anticancer antibiotic, neocarzinostatin (NCS), was administered intravenously to F344 rats and pathological changes of the kidneys were characterized. Animals given a single injection of ZS at 0.65 mg/kg showed no changes until 3 weeks after dosing. At 4 weeks or later, proteinuria appeared, and expansion of mesangial areas with increased fibronectin was observed in the glomeruli. There were increased urea nitrogen and decreased total protein in blood plasma at 5 weeks or later. At 15 weeks after dosing, renal sclerosis was prominent. These glomerular lesions, characterized by expansion of mesangial areas, were different from those induced by other antitumor drugs mainly affecting the glomerular epithelial cells.

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In an experiment primarily designed to evaluate the long-term dose dependence of N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) administration, attention was concentrated on its influence on the renal pelvis. BBN was given at dosage levels of 50, 10, 5, 1, and 0 ppm in the drinking water to male F344 rats for 112 weeks. Even this life-span exposure did not cause any proliferative or neoplastic lesions of the renal pelvis/papilla despite papilloma and carcinoma induction by concentrations of 5 ppm and above in the urinary bladder. In a separate experiment, treatment with BBN or N-ethyl-N-(4-hydroxybutyl)nitrosamine (EHBN) at the dose of 500 ppm in the drinking water for 4 weeks was also found not to increase DNA synthesis in the renal pelvis/papilla epithelia. The fact that the bladder carcinogen BBN did not induce tumor development or an increment of DNA synthesis in the renal pelvis/papilla, which is lined like the bladder by transitional epithelium, strongly indicates that its target organ specificity is due to long urine stasis leading to prolonged metabolite (ultimate carcinogen) exposure only in the urinary bladder.

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The effects of testosterone and 17β-estradiol (E₂) on the prostate of castrated rats were investigated by histopathological and immunohistochemical procedures. Male Sprague-Dawley rats were divided into four experimental groups. Group 1 consisted of intact controls. The other animals were castrated. In group 2, rats were sacrificed 2 days after castration. The castrated animals were treated for 6 weeks with 1) testosterone 1 mg/head (Group 3) and 2) testosterone 1 mg/head plus E₂ 10 μg/head (Group 4). A significant increase in prostatic weight occurred after 6 weeks treatment with testosterone alone (Group 3) and in combination with E₂ (Group 4). The greatest increase in prostatic weight were noted in Group 4. Histopathologically, glandular hyperplasia of the prostate was clearly observed, and the number of bromo-deoxyuridine (BrdU)-positive cells showed a significant increase over that induced by testosterone alone. Immunohistochemical localization of glutathione-peroxidase (GSH-PO) which effectively reduces the lipid peroxides, was predominantly observed in the glandular epithelial cells of the prostate. The intensity of GSH-PO staining in the glandular epithelial cells was remarkably decreased after castration (Group 2), and that it was clearly recovered by testosterone (Group 3)-or testosterone plus E₂ (Group 4)- treatment to the castrated rats. In addition, androgen receptor (AR) was mainly localized in nuclei, but not in cytoplasm. Furthermore, immunodetectable AR rapidly declined after androgen withdrawal (Group 2) and returned to intact levels of staining intensity after androgen replacement (Groups 3 and 4). Thus, GSH-PO in the glandular epithelial cells suggest the very close relationship to the status of testosterone action to the cells. Therefore, it was strongly suggested that GSH-PO in the glandular epithelial cells of the rat prostate was testosterone-dependent.

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We reported the dose-response relationship between the accumulation of mercury and its toxic effect with thimerosal (ethylmercuric thiosalicylic acid, sodium salt) in mice in a previous paper. In this experiment, the toxic effects of thimerosal on the peripheral nerve were studied by light and electron microscopy since the involvement of the peripheral nerve in organic mercurial intoxication has been frequently reported in rodents. The accumulation of mercury was also examined in the cerebrum, cerebellum, liver, and kidney.
A thimerosal solution of 0.2 ml at 0.1% was injected twice into the intraperitoneal cavity of 5 week-old female BALB/c mice. The minimum dose injected in the mice was 33 μg Hg/g and the maximum was 116 μg Hg/g body weight.
The sciatic nerves were affected with the minimum dose of 33 μg Hg/g body weight. Electron microscopically, there were whorl formation and deformations of the myelin sheaths, which were irregularly swollen and thickened. The axons showed atrophy with high electron density and degeneration of Schwann cells was also noted.
The cerebrum, cerebellum, liver, and kidney demonstrated strong positive reactions to mercury-histochemistry with photoemulsion method. An assay of the total and inorganic mercury content in these organs was also performed since the ratio of inorganic mercury to total mercury in the brain increased with time because of biotransformation of organic to inorganic mercury. No definite relationship was noted between the mercury dose injected into the mice and the accumulation of mercury in the organs of mice. Thimerosal was suspected to be poorly absorbed through the peritoneal cavity because multiple intraperitoneal injections caused severe peritonitis.

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In order to examine the variation of pathologic modifications with endogenous endotoxin, galactosamine (GalN) 400 mg/kg was administered intraperitoneally to male rats combined with lactulose (LTS) 6,000 mg/kg/day × 7 days, po, cyclophosphamide (CPM) 100 mg/kg, ip, or ricinoleic acid (RA) 2,000 mg/body, po. LTS was used for alteration of intestinal flora, CPM for depression of local intestinal immune system, and RA for disruption of intestinal epithelial barrier. In the GalN alone group, hepatic changes such as morphologic activation of Kupffer cells, and degeneration and necrosis of hepatocytes in the liver were observed microscopically. However, there were no remarkable changes in the liver in the GalN/LTS group. In the GalN/CPM and GalN/RA groups, although damage of the liver parenchymal cells was milder than those of the GalN alone group, activation of Kupffer cells and mononuclear cell aggregation in the GalN/RA group were more prominent than the GalN alone group. From the results, as to occurrence of pathologic changes due to endogenous endotoxin, we suggest that the situation of the intestinal flora and the intestinal epithelial barrier, and the balance between quantity of endotoxin and the RES function are important. We concluded that, according to toxicity of a test compound, it is necessary to consider the influence of endogenous endotoxin for assessment and judgement of the toxicity study's results.

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Light microscopic, histochemical, and ultrastructural observations were conducted to examine a spontaneous malignant atriocaval tumor of the heart in an old Sprague-Dawley rat among the untreated controls in a carcinogenicity study. The tumor, located from the right atrioventricular septum to the atrium, was composed of numerous tubular structures lined by columnar or cuboidal cells of distinctly epithelial appearance, but had no continuum to the mesothelium of the epicardium or the pleura. Metastatic small foci and emboli were noted in the bilateral pulmonary tissues. Hyaluronic acid was detected in the tumor cells of epithelial appearance. There were no argyrophilic granules of neuroendocrine cells in the tumor tissue. Ultrastructural examination revealed long slender microvilli on the surface and junctional complexes but no filament bundles or secretory granules in the tumor cells. Thus the tumor in question closely resembled the congenital atriocaval mesotheliomas described in a special strain of NZR/Gd inbred rats. In searching the literature no reports of malignant atriocaval tumor of the heart were found as having occurred in Sprague-Dawley rats.

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