Certain disease conditions can modify drug-induced toxicities, which, in turn, may cause a medication-related health crisis. Therefore, preclinical investigations into the alterations in drug-induced toxicities using appropriate disease animal models are very important. This paper reviews the reported data related to the effects of diabetes and hypertriglyceridemia, common lifestyle-related diseases in a modern society, on acetaminophen (APAP)-induced hepatotoxicity and nephrotoxicity in rats and mice. It has generally been reported that diabetes protects rats and mice from APAP-induced hepatotoxicity and there are several reports that help to speculate on the effects of diabetes on APAP-induced nephrotoxicity. In fructose-induced hypertriglyceridemic rats, hepatotoxicity of APAP becomes apparently less severe, whereas nephrotoxicity of APAP becomes significantly more severe. The mechanisms of alteration of APAP-induced hepatorenal toxicity under diabetic and hypertriglyceridemic conditions are also discussed in this paper.
Quantification of polycyclic aromatic hydrocarbons (PAH) and their metabolites within living cells and tissues in real time using fluorescence methods is complicated due to overlaping excitation and/or emission spectra of metabolites. In this study, simultaneous analysis of several metabolites of a prototype carcinogenic PAH, benzo[a]pyrene (BaP) in undifferentiated (MCF10A) and differentiated (MCF10CA1h) breast cancer cells was performed using single-cell multiphoton spectral analysis. The two cell types were selected for this study because they are known to have differences in BaP uptake and metabolism and induction of aryl hydrocarbon receptor-dependent ethoxyresorufin-O-deethylase (EROD) activity. Multiphoton microscopy spectral analysis performed in cells exposed to BaP for 24 hr identified 5 major peaks of fluorescence that were monitored within spectral bands. A comparison of the fluorescence peaks within these bands to those of BaP metabolite standards indicated that a peak in the spectral range of 393-415 nm matched benzo[a]pyrene-r-7,t-8-dihydrodiol-t-9,10-epoxide(±),(anti) (BPDE), the ultimate carcinogenic BaP metabolite. In addition, the 426-447 nm band matched the major metabolites 3-hydroxybenzo[a]pyrene (3-OH BaP) and 9-hydroxybenzo[a]pyrene (9-OH BaP); the 458-479 nm band corresponded to the secondary metabolite benzo[a]pyrene-3,6-dione (3,6 BPQ); and a peak at 490-530 nm matched the parent compound, BaP. Multiphoton spectral analysis also revealed differences in fluorescence intensities between MCF10A and MCF10CA1h cells within three spectral bands: 393-415 nm, 426-447 nm and 458-479 nm which were partially reversed with cyclosporine A suggesting differences in efflux mechanisms between cell lines. These results demonstrate the feasibility of analyzing BaP metabolism in situ by multiphoton spectral analysis and also identifying cell-type differences in BaP accumulation and metabolism.
Estimation of liver damage is important in the pathophysiological and toxicological study of liver disease. As a novel, non-invasive marker of liver damage, we studied the efficacy of urine bile acids (UBA) in a rat model of liver disease. Thioacetamide (TAA)-treated rats were used in this study. Single intraperitoneal administration of high-dose TAA induces severe damage to the liver, and thus is used as a model of acute hepatitis. Continuous administration of low-dose TAA yields mild damage to the liver, and induces cirrhosis and hepatic tumors. In this study, it was found that both acute and chronic administration of TAA was associated with a dose-dependent elevation of UBA. The elevation of UBA content correlated with the alteration of blood biochemical indicators, and UBA screening showed a remarkable ability to distinguish liver-damaged rats from healthy rats. In particular, UBA analysis was found to have high sensitivity, specificity, and positive predictive value for the screening of rats with abnormal serum alkaline phosphatase (ALP) activity due to chronic liver damage, which was confirmed to include cholestasis and subsequent cirrhosis by liver histological analysis. In conclusion, we demonstrated that measurement of UBA is a simple, non-invasive and effective method for the screening of cholestasis in TAA-treated rats. We suggest that UBA analysis may have potent applicability for monitoring the progress of liver damage in animal models of chronic liver disease, such as cirrhosis and hepatic encephalopathy.
Totally implantable catheter animal models are considered useful for pharmacological and toxicological studies. In this report, we assessed the feasibility of using an indwelling vascular access port (VAP) in rats for long-term evaluation of repeated and intermittent dose toxicity studies. In Experiment 1, the VAP devices were implanted in male and female rats and a saline solution administered intravenously via the posterior vena cava for 2 weeks (4 ml/kg, 2 ml/min, 5 times/week, 10 times total). General conditions, body weight and blood chemistry showed no toxicological changes compared with the rats in the non-implanted, non-treated group. Hematology changes such as transient increases in peripheral blood reticulocytes and eosinophils were noted post-implantation. In pathology, proliferation of the endothelium at the site of VAP implantation and perivascular inflammatory cell infiltration including eosinophils in lung were noted at the end of the treatment period. Moreover, we found that the lumbar area is more suitable for VAP implantation than the back of neck for young, still growing rats. Experiment 2 included a 1-month intravenous intermittent dose (4 ml/kg, 2 ml/min, 1 time/week, 5 times total) toxicity study in VAP-implanted rats followed by a 1-month recovery period conducted under Good Laboratory Practice (GLP) regulations. The results suggested that an animal model with implanted VAP is useful for intermittent intravenous dosing of drugs. Moreover, VAP implantation in animals is expected to be extrapolated to use VAP in humans in clinical studies.
N,N-Dimethylformamide (DMF), a ubiquitous contaminant in living and working environments, enters the human body by inhalation, as well as by oral and dermal routes of exposure. In order to provide bioassay data for carcinogenic risk assessment of humans exposed to DMF by multiple routes of exposure, hepatocarcinogenic effect of combined inhalation and oral exposures of rats to DMF was examined. A group of 50 male F344 rats, 6-week-old, was exposed by inhalation to 0 (clean air), 200, or 400 ppm (v/v) of DMF vapor-containing air for 6 hr/day and 5 days/week during a 104-week period, and each inhalation group was given ad libitum DMF-formulated drinking water at 0, 800 or 1,600 ppm (w/w) for 104 weeks. Incidences of hepatocellular adenomas and carcinomas and their combined incidences were significantly increased in the combined-exposure groups compared with the untreated control group or each of the inhalation-alone and oral-alone groups with matching concentrations. Incidences of hepatocellular adenomas and carcinomas induced by the combined exposures were greater than the sum of the two incidences of the hepatocellular adenomas and carcinomas induced by the single-route exposures through inhalation and ingestion. The combined exposures enhanced tumor malignancy. It was concluded that the combined inhalation and oral exposures markedly enhance the incidences and malignancy of hepatocellular tumors, suggesting that the hepatocarcinogenic effect of the combined exposures is greater than the effect that would be expected under the assumption that the two effects of single-route exposures through inhalation and drinking are additive.
The present study assessed a carcinogenic hazard of multi-wall carbon nanotube (MWCNT) in intact (not genetically modified) rodents. MWCNT (1 mg/kg body weight, 7 animals), crocidolite (2 mg/kg body weight, 10 animals) or vehicle (2% carboxymethyl cellulose, 5 animals) was administered to male Fischer 344 rats (12 weeks old) by a single intrascrotal injection. Rats were autopsied immediately after death, when becoming moribund or at the end of the maximal observation period scheduled to be 52 weeks. After 37-40 weeks, however, 6 MWCNT-treated animals died or became moribund due to intraperitoneally disseminated mesothelioma (6/7, 85.7%) with bloody ascites. Peritoneal mesothelium was generally hypertrophic, and numerous nodular or papillary lesions of mesothelioma and mesothelial hyperplasia were developed. While mesothelioid cells were predominant in relatively early stage tumors, advanced stage mesotheliomas were constituted by 2 portions occupied by mesothelioid cells on the surface and spindle-shaped sarcomatous cells in the depth. In the latter, the histological transition was apparently observed between these 2 portions. Mesotheliomas were invasive to adjacent organs and tissues, and frequently metastasized into the pleura. Only 1 rat survived for 52 weeks in the MWCNT-treated group, and similar findings except mesothelioma were observed. All 10 crocidolite-treated and 5 vehicle-treated rats survived for 52 weeks without any particular changes except deposition of asbestos in the former case. It is thus indicated that MWCNT possesses carcinogenicity causing mesothelioma at a high rate in intact male rats under the present experimental conditions. The present data identifies a carcinogenic hazard of MWCNT and will serve as one of the indispensable evidences to be used for the risk assessment crucial for not only protection and improvement of human health and welfare, but also safe and acceptable development and prevalence of this and similar upcoming materials.
Reserpine, a natural product extracted from Rauwolfia serpintina or Rauwolfia vomitoria, is a known dopamine depleter that inhibits several neurotransmitters. Reserpine has been used clinically to control hypertension, schizophrenia, insomnia and insanity. The use of this drug, however, has been limited because of its side effects which include oxidative damage to organs, including the liver. Green tea catechins are potent antioxidants that have the potential to counteract reserpine induced oxidative stress. This study investigated the merits of administering green tea concurrently with reserpine to prevent oxidative hepatic damage in Sprague-Dawely (SD) rats. Reserpine was found to cause hepatic damage, with elevated levels of oxidative stress markers, such as Thiobarbituric Acid Reactive Substances (TBARS), transaminases and cholesterol. Reserpine also induced hepatic ultra-structural damage in the cytoplasmic membrane, nuclear envelope, endoplasmic reticulum (rER), ribosomal stripping and mitochondria. Electron microscopy examination showed revival of liver cells as a result of green tea extract administration to experimental rats.
Class II alcohol dehydrogenase (π-ADH), encoded by alcohol dehydrogenase (ADH4), is considered to contribute to ethanol (EtOH) oxidation in the liver at high concentration. Four single nucleotide polymorphisms (SNPs) were found in the promoter region of this gene. Analysis of genotype distribution in 102 unrelated Japanese subjects revealed that four loci were in strong linkage disequilibrium and could be classified into three haplotypes. The effects of these polymorphisms on transcriptional activity were investigated in HepG2 cells. Transcriptional activity was significantly higher in cells with the -136A allele than in those with the -136C allele. To investigate whether this difference in transcriptional activity caused a difference in EtOH elimination, previous data on blood EtOH changes after 0.4 g/kg body weight alcohol ingestion were analyzed. When analyzed based on aldehyde dehydrogenase-2 gene (ALDH2) 487Glu/Lys genotype, the significantly lower level of EtOH at peak in subjects with -136C/A and -136A/A genotype compared with subjects with -136C/C genotype indicated that -136 bp was a suggestive locus for differences in EtOH oxidation. This effect was observed only in subjects with ALDH2487Glu/Glu. These results suggested that the SNP at -136bp in the ADH4 promoter had an effect on transcriptional regulation, and that the higher activity of the -136A allele compared with the -136C allele caused a lower level of blood EtOH after alcohol ingestion; that is, individuals with the -136A allele may consume more EtOH and might have a higher risk for development of alcohol dependence than those without the -136A allele.
Chemical characterization and acute and sub-acute toxicity study of Trikatu, a generic herbal formulation of Indian system of medicine, was carried out in Charles Foster (CF) rats for safety profiling. In acute toxicity experiment, Trikatu at 2,000 mg/kg body weight once orally was well tolerated by the experimental animals (both male and female) and no changes were observed in mortality, morbidity, gross pathology, gain in weight, vital organ weight, hematological (total white blood cells (WBC) and red blood cells (RBC) count), biochemical parameters such as serum creatinine, serum glutamate oxaloacetate transaminase (SGOT), serum glutamate pyruvate transaminase (SGPT), serum lipid profile and tissue biochemical parameters such as reduced glutathione and malonaldehyde content as oxidative stress markers. In sub-acute experiment, Trikatu was administered at 5, 50 and 300 mg/kg body weight once daily for 28 days in female CF rats, and non-significant changes were found in most of the parameters studied such as acute experiment except significant increase in low density lipoprotein (LDL) cholesterol level at 50 and 300 mg/kg body weight, decrease in high density lipoprotein (HDL) cholesterol level at 300 mg/kg body weight, increase in SGPT activity at 50 mg/kg body weight and decrease in WBC count at 300 mg/kg body weight on 28th day post treatment.
Dicyclanil (DC) generates reactive oxygen species (ROS) due to Cyp1a1 induction, and DNA damage caused by oxidative stress is probably involved in hepatocarcinogenesis in mice. To clarify the modifying effect of the Siraitia grosvenorii extract (SGE), which has antioxidative properties, we employed a 2-stage liver carcinogenesis model in partially hepatectomized male ICR mice. Mice maintained on diet containing DC at a concentration of 1,500 ppm for 9 weeks after a single intraperitoneal injection of diethylnitrosamine (DEN) at a dose of 30 mg/kg and they were given water containing 2,500 ppm of SGE for 11 weeks including 2 weeks as pre-administration on DC. SGE inhibited the induction of γ-glutamyltranspeptidase-positive hepatocytes, lipid peroxidation, and gene expression of Cyp1a1, all ofwhich were caused by DC. To examine whether SGE indirectly inhibits Cyp1a1 expression induced by inhibition of aryl hydrocarbon receptor (Ahr)-mediated signal transduction caused by DC, mice with high (C57BL/6J mice) and low affinities (DBA/2J mice) to Ahr were given DC-containing diet and/or SGE-containing tap water for 2 weeks. Cyp1a1 gene expression was significantly lower in C57BL/6J mice administered DC + SGE than in C57BL/6J mice administered DC alone; there was no difference in the Cyp1a1 expression between DBA/2J mice administered DC + SGE and DC alone. These results suggest that SGE suppresses the induction of Cyp1a1, leading to inhibition of ROS generation and consequently inhibited hepatocarcinogenesis, probably due to suppression of Ahr activity.
A mutagenicity test was conducted on water-soluble ZnO nanoparticles capped with tetramethylammonium hydroxide in a bacterial reverse mutation assay using Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537, and Escherichia coli strain WP2uvrA−, with and without metabolic activation by S9 mix in the preincubation method. Mutagenicity was negative in all strains.
The purpose of tumorigenicity testing, as applied not only to cell substrates used for viral vaccine manufacture but also stem cells used for cell-based therapy, is to discriminate between cells that have the capacity to form tumors and cells that do not. Therefore, tumorigenicity testing is essential in assessing the safety of these biological materials. Recently developed NOD/Shi-scid IL2Rgnull (NOG) mice have been shown to be superior to NOD/Shi-scid (SCID) mice for xenotransplantation of both normal and cancerous cells. To select a suitable mouse strain as a xenogenic host for tumorigenicity testing, we compared the susceptibility of NOG (T, B, and NK cell-defective), SCID (T and B cell-defective), and the traditionally used nude (T cell-defective) mice to tumor formation from xenotransplanted HeLa S3 cells. When 104 HeLa S3 cells were subcutaneously inoculated into the flanks of these mice, the tumor incidence on day 22 was 10/10 (100%) in NOG, 2/10 (20%) in SCID, and 0/10 (0%) in nude mice. The subcutaneous tumors formed reproducibly and semiquantitatively in a dose-dependent manner. Unexpectedly, half of the NOG mice (5/10) that had been inoculated with a mere 101 HeLa S3 cells formed progressively growing subcutaneous tumors on day 78. We confirmed that the engrafted tumors originated from inoculated HeLa S3 cells by immunohistochemical staining with anti-HLA antibodies. These data suggest that NOG mice may be the best choice as a suitable strain for testing tumorigenicity.
The effects of restricted feeding (20 g/day from gestational day (GD) 6 to 28) on pregnancy outcome and blood parameters were examined in pregnant rabbits. As compared with the group which was allowed free access to diet throughout the gestational period (NT group), the group subjected to restricted feeding (R group) showed significantly lower values in many parameters such as total protein, albumin and triglyceride on GDs 22 and 28, reflecting low nutritive conditions. In addition, there were significant changes in blood coagulation-related parameters, suggesting an imbalance between coagulation and anti-coagulation factors. Moreover, abortions occurred in about half of the animals of the R group between GDs 23 and 27. When aborted rabbits were compared with those which could maintain pregnancy under restricted feeding, total protein, albumin, platelets and antithrombin III values and especially blood progesterone concentration were significantly lower in aborted rabbits on GD 22, prior to occurrence of abortion. These results suggested that abortions due to restricted feeding might be brought about by lower nutritive conditions, an imbalance of blood coagulation-related factors and lower blood progesterone concentrations.