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Life threatening complications from chemotherapy occur frequently in cancer survivors, however little is known about genetic risk factors. We treated male normotensive rats (WKY) and strains with hypertension (SHR) and hypertension with cardiomyopathy (SHHF) with 8 weekly doses of doxorubicin (DOX) followed by 12weeks of observation to test the hypothesis that genetic cardiovascular disease would worsen delayed cardiotoxicity. Compared with WKY, SHR demonstrated weight loss, decreased systolic blood pressure, increased kidney weights, greater cardiac and renal histopathologic lesions and greater mortality. SHHF showed growth restriction, increased kidney weights and renal histopathology but no effect on systolic blood pressure or mortality. SHHF had less severe cardiac lesions than SHR. We evaluated cardiac soluble epoxide hydrolase (sEH) content and arachidonic acid metabolites after acute DOX exposure as potential mediators of genetic risk. Before DOX, SHHF and SHR had significantly greater cardiac sEH and decreased epoxyeicosatrienoic acid (EET) (4 of 4 isomers in SHHF and 2 of 4 isomers in SHR) than WKY. After DOX, sEH was unchanged in all strains, but SHHF and SHR rats increased EETs to a level similar to WKY. Leukotriene D4 increased after treatment in SHR. Genetic predisposition to heart failure superimposed on genetic hypertension failed to generate greater toxicity compared with hypertension alone. The relative resistance of DOX-treated SHHF males to the cardiotoxic effects of DOX in the delayed phase despite progression of genetic disease was unexpected and a key finding. Strain differences in arachidonic acid metabolism may contribute to variation in response to DOX toxicity., (© 2013 Elsevier Inc. All rights reserved.)

Reactive oxygen species (ROS) have been associated with many human diseases, and glutathione (GSH)-dependent processes are pivotal in limiting tissue damage. To test the hypothesis that Gr1(a1Neu) (Neu) mice, which do not express glutathione reductase (GR), would be more susceptible than are wild-type mice to ROS-mediated injury, we studied the effects of diquat, a redox cycling toxicant. Neu mice exhibited modest, dose- and time-dependent elevations in plasma alanine aminotransferase (ALT) activities, 126+/-36 U/l at 2 h after 5 micromol/kg of diquat, but no ALT elevations were observed in diquat-treated C3H/HeN mice for up to 6 h after 50 micromol/kg of diquat. Histology indicated little or no hepatic necrosis in diquat-treated mice of either strain, but substantial renal injury was observed in diquat-treated Neu mice, characterized by brush border sloughing in the proximal tubules by 1 h and tubular necrosis by 2 h after doses of 7.5 micromol/kg. Decreases in renal GSH levels were observed in the Neu mice by 2 h post dose (3.4+/-0.4 vs 0.2+/-0.0 micromol/g tissue at 0 and 50 micromol/kg, respectively), and increases in renal GSSG levels were observed in the Neu mice as early as 0.5 h after 7.5 micromol/kg (105.5+/-44.1 vs 27.9+/-4.8 nmol/g tissue). Blood urea nitrogen levels were elevated by 2 h in Neu mice after doses of 7.5 micromol/kg (Neu vs C3H, 32.8+/-4.1 vs 17.9+/-0.3 mg/dl). Diquat-induced renal injury in the GR-deficient Neu mice offers a useful model for studies of ROS-induced renal necrosis and of the contributions of GR in defense against oxidant-mediated injuries in vivo.

Hyperoxic exposure affects the levels and activities of some hepatic proteins. We tested the hypothesis that hyperoxic exposure would result in greater hepatic .NO concentrations. C3H/HeN mice were exposed to >95% O(2) for 72 or 96 h and compared to room air-breathing controls. In contrast to our working hypothesis, exposure to >95% O(2) for 96 h decreased hepatic nitrite/nitrate NO(X) concentrations (10.9 +/- 2.2 nmol/g liver versus 19.3 +/- 2.4 nmol/g liver in room air, P < 0.05). The hepatic levels of endothelial NO synthase (eNOS) and inducible NOS (iNOS) proteins were not different among the groups. The arginases, which convert L-arginine to urea and L-ornithine, may affect hepatic NOS activities by decreasing L-arginine bioavailability. Hepatic ornithine concentrations were greater in hyperoxic animals than in controls (318 +/- 18 nmol/g liver in room air, and 539 +/- 64, and 475 +/- 40 at 72 and 96 h of hyperoxia, respectively, P < 0.01). Hepatic arginase I protein levels were greater in hyperoxic animals than in controls. Hepatic carbamoyl phosphate synthetase (CPS) protein levels and activities were not different among groups. These results indicate that increases in hepatic levels of arginase I in mice exposed to hyperoxia may diminish .NO production, as reflected by lower liver levels of NO(X). The resultant greater hepatic ornithine concentrations may represent a mechanism to facilitate tissue repair, by favoring the production of polyamines and/or proline.

The primary mechanisms proposed for acetaminophen-induced hepatic necrosis should deplete protein thiols, either by covalent binding and thioether formation or by oxidative reactions such as S-thiolations. However, in previous studies we did not detect significant losses of protein thiol contents in response to administration of hepatotoxic doses of acetaminophen in vivo. In the present study we employed derivatization with the thiol-specific agent monobromobimane and separation of proteins by SDS-PAGE to investigate the possible loss of specific protein thiols during the course of acetaminophen-induced hepatic necrosis. Fasted adult male mice were given acetaminophen, and protein thiol status was examined subsequently in subcellular fractions isolated by differential centrifugation. No decreases in protein thiol contents were indicated, with the exception of a marked decrease in the fluorescent intensity, but not of protein content, as indicated by staining with Coomassie blue, of a single band of approximately 130 kDa in the mitochondrial fractions of acetaminophen-treated mice. This protein was identified by isolation and N-terminal sequence analysis as carbamyl phosphate synthetase-I (CPS-I) (EC 6.3.4.16). Hepatic CPS-I activities were decreased in mice given hepatotoxic doses of acetaminophen. In addition, hepatic glutamine synthetase activities were lower, and plasma ammonia levels were elevated in mice given hepatotoxic doses of acetaminophen. The observed hyperammonemia may contribute to the adverse effects of toxic doses of acetaminophen, and elucidation of the specific mechanisms responsible for the hyperammonemia may prove to be useful clinically. However, the preferential depletion of protein thiol content of a mitochondrial protein by chemically reactive metabolites generated in the endoplasmic reticulum presents a challenging and potentially informative mechanistic question., (Copyright 1997 Academic Press.)

Administration of hepatotoxic doses of diquat to male Fischer-344 rats increases biliary excretion of nonheme iron, whereas comparably hepatotoxic doses of acetaminophen decrease biliary export of iron. The effects of acetaminophen and diquat on the activities in bile of representative lysosomal enzymes, beta-N-acetylglucosaminidase (beta-NAG) and beta-glucuronidase (beta-GLUC) were examined as a means of assessing the possible role of lysosomal exocytosis in the effects of these hepatotoxins on biliary excretion of iron. In pentobarbital-anesthetized male Fischer-344 rats, diquat at 0.1 mmol/kg increased the biliary export of biliary beta-NAG and beta-GLUC, in conjunction with similar increases in iron. Sprague-Dawley rats, which are resistant to diquat-induced hepatic necrosis despite showing marked oxidant stress responses, showed no increases in biliary efflux of iron, beta-NAG, or beta-GLUC in response to diquat. Conversely, acetaminophen at doses of 400 or 1500 mg/kg markedly decreased biliary concentrations and efflux rates of beta-NAG and beta-GLUC in Fischer-344 rats in parallel with decreases in biliary iron, suggesting that the hepatotoxin-induced effects on biliary iron excretion may be mediated through effects on lysosomal exocytosis. Both acetaminophen and diquat increased total protein content of bile in both strains of rats; however, the proteins excreted after administration of diquat to Fischer-344 rats showed marked increases in contents of protein carbonyls, as assayed with 2,4-dinitrophenylhydrazine, whereas biliary proteins in acetaminophen-treated animals were not more oxidized than in controls. Sprague-Dawley rats given diquat showed no increase in the biliary excretion of protein carbonyls, despite the increased excretion of glutathione disulfide observed in these animals. The significant increases in biliary excretion of protein carbonyls by the diquat-treated Fischer-344 rats suggest oxidation of cellular proteins catalyzed by chemically reactive iron chelates and the excretion of at least some of the oxidized proteins to the bile, possibly through lysosomal exocytosis. The effects of acetaminophen on biliary protein excretion do not appear to involve oxidation.