Evaluation of the effect of the antihistaminic preparations dimebon and diprazin on embryogenesis in rats

A single administration of dimebon (800 mg/kg) to rats per os (2/3 of the LD50 isoeffective for females) raises the preimplantation death, whereas pipolphen increases the intrauterine lethality and inhibits the development of fetuses. Pipolphen in a dose of 175 mg/kg and dimebon in doses of 300-150 mg/kg (exceeding 25- and 300-150-fold, respectively, the therapeutic dose for man) do not exert any specific embryotropic action.     

Saturation toxicokinetics of thioacetamide: role in initiation of liver injury.

Thioacetamide (TA), a potent centrilobular hepatotoxicant, undergoes a two-step bioactivation mediated by microsomal CYP2E1 to TA sulfoxide (TASO), and further to TA-S,S-dioxide (TASO2), a reactive metabolite that initiates cellular necrosis. Our earlier studies showed that bioactivation-mediated liver injury of TA is not dose-proportional. The objective of this study was to examine whether increasing doses of TA lead to enzyme saturation, thereby resulting in lack of dose-response for injury: bioactivation of TA --> TASO --> TASO2 may follow zero-order kinetics. A 12-fold dose range of TA (50, 300, and 600 mg/kg i.p.) was injected into male Sprague-Dawley rats. TA and TASO were quantified in plasma, liver, and urine by high-performance liquid chromatography. With increasing doses, the apparent elimination half-lives of TA and TASO increased linearly, indicating that TA bioactivation exhibits saturation kinetics. Increasing TA dose resulted in greater-than-proportional increases in plasma TA and TASO levels. The TASO/TA ratio was inversely proportional to the dose of TA. Covalent binding of 14C-TA-derived radiolabel to liver macromolecules showed a less-than-dose-proportionate increase with a 12-fold higher dose. Less than dose-proportional covalent binding was confirmed in liver microsomal incubations with 14C-TA. Three-fold higher excretion of TASO was seen in urine at the highest dose (600 mg/kg) compared with the lowest dose (50 mg TA/kg). Incubation of TA with rat liver microsomes and purified baculovirus-expressed rat and human CYP2E1 Supersomes, over a concentration range of 0.01 to 10 mM, revealed saturation of TA conversion to TASO at and above 0.05 mM TA concentration, comparable to in vivo plasma and liver levels achieved upon administration of higher doses. Calculated K(m) values for TA (0.1 mM) and TASO (0.6 mM) suggest that the second step of TA bioactivation is 6-fold less efficient. Collectively, the findings indicate saturation of CYP2E1 at the first (TA to TASO) and second (TASO to TASO2) steps of TA bioactivation.     

Role of CYP2E1 and saturation kinetics in the bioactivation of thioacetamide: Effects of diet restriction and phenobarbital

Thioacetamide (TA) undergoes saturation toxicokinetics in ad libitum (AL) fed rats. Diet restriction (DR) protects rats from lethal dose of TA despite increased bioactivation-mediated liver injury via CYP2E1 induction. While a low dose (50 mg TA/kg) produces 6-fold higher initial injury, a 12-fold higher dose produces delayed and mere 2.5-fold higher injury. The primary objective was to determine if this less-than-expected increase in injury is due to saturation toxicokinetics. Rats on AL and DR for 21 days received either 50 or 600 mg TA/kg i.p. T(1/2) and AUCs for TA and TA-S-oxide were consistent with saturable kinetics. Covalent binding of (14)C-TA-derived-radiolabel to liver macromolecules after low dose was 2-fold higher in DR than AL rats. However, following lethal dose, no differences were found between AL and DR. This lack of dose-dependent response appears to be due to saturation of bioactivation at the higher dose. The second objective was to investigate the effect of phenobarbital pretreatment (PB) on TA-initiated injury following a sub-lethal dose (500 mg/kg). PB induced CYP2B1/2 approximately 350-fold, but did not increase covalent binding of (14)C-TA, TA-induced liver injury and mortality, suggesting that CYP2B1/2 has no major role in TA bioactivation. The third objective was to investigate the role of CYP2E1 using cyp2e1 knockout mice (KO). Injury was assessed over time (0-48 h) in wild type (WT) and KO mice after LD(100) dose (500 mg/kg) in WT. While WT mice exhibited robust injury which progressed to death, KO mice exhibited neither initiation nor progression of injury. These findings confirm that CYP2E1 is responsible for TA bioactivation.     

Toxicological profile of carboxymethyl inulin.

Carboxymethylinulin (CMI), formed by carboxylation of a natural carbohydrate obtained from the chicory plant, is particularly effective in sequestration of hard water cations, and thus serves as a unique anti-scalant which could find uses in food processing. A series of toxicological studies has been performed to investigate its toxiciologic properties following repeated exposure, possible sensitization, and its potential to elicit genotoxic activity; all studies conformed to internationally accepted safety test guidelines currently in force. Subacute (4-week) oral toxicity was investigated in groups of rats exposed via gavage to 0, 50, 150 and 1000 mg/kg/day CMI. No treatment-related effects were observed in body weight, food consumption, mortality, hematology, clinical blood chemistry, organ weights or gross or microscopic pathology up to the highest dose (1000 mg/kg/day) tested. Motor activity, as observed in a functional observation battery, was elevated in high-dose females, and is not considered of significance toxicologically. Lack of adverse toxicity seen with CMI at this dosage is consistent with a similar lack of significant toxicity exhibited by other dietary carbohydrates (sorbitol, sucrose, glucose), oligofructoses (inulin/FOS) and carboxylated cellulose in repeated-dose rat studies at approximately the same dosage. No evidence of dermal sensitization was observed in groups of guinea pigs following CMI testing by the Magnusson-Kligman maximization test methodology. No mutagenic activity was observed when CMI was tested in four Salmonella strains-TA1535, TA1537, TA98 and TA100-or in Escherichia coli WP2uvrA bacterial point mutation assays or in an in vitro Chinese hamster ovary cell chromosomal aberration assay. The results obtained in the present study with CMI are consistent with similar data derived on numerous dietary carbohydrate fibers generally recognized as safe in the human diet.     

Toxicology and carcinogenesis studies of bis(2-chloroethoxy)methane (CAS No. 111-91-1) in F344/N rats and B6C3F1 mice (dermal studies)

Bis(2-chloroethoxy)methane is used as a solvent and the starting agent in the production of fungicides and polysulfide polymers. Bis(2-chloroethoxy)methane was nominated for study by the National Institute of Environmental Health Sciences because of its widespread use as a starting material to produce polysulfide elastomers, and because there were no 2-year carcinogenicity studies reported in the literature. Male and female F344/N rats and B6C3F1 mice received dermal applications of bis(2-chloroethoxy)-methane in ethanol (greater than 98% pure) for 2 weeks, 3 months, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium and Escherichia coli, rat bone marrow cells, and mouse peripheral blood erythrocytes. 2-WEEK STUDY IN RATS: Groups of five male and five female rats were dermally administered 0, 12.5, 25, 50, 100, or 200 mg bis(2-chloroethoxy)methane/kg body weight in ethanol, 5 days per week for 16 days. All rats survived to the end of the study. Mean body weights of dosed rats were similar to those of the vehicle control groups. There were no histopathologic lesions related to bis(2-chloroethoxy)methane administration. 2-WEEK STUDY IN MICE: Groups of five male and five female mice were dermally administered 0, 12.5, 25, 50, 100, or 200 mg bis(2-chloroethoxy)methane/kg body weight in ethanol, 5 days per week for 17 days. All mice survived to the end of the study. Mean body weights of dosed mice were similar to those of the vehicle control groups. There were no histopathologic lesions related to bis(2-chloroethoxy)methane administration. 3-MONTH STUDY IN RATS: Groups of 10 male and 10 female rats were dermally administered 0, 50, 100, 200, 400, or 600 mg bis(2-chloroethoxy)methane/kg body weight in ethanol, 5 days per week for 14 weeks. Additional clinical pathology groups of 10 male and 10 female rats were administered the same doses for 23 days. All core study 600 mg/kg males and females and two 400 mg/kg females died before the end of the study. The cause of death was considered to be related to the cardiotoxic effect of bis(2-chloroethoxy)methane. There were no significant differences between final mean body weights of dosed rats and those of the vehicle control groups; the mean body weight gain of 400 mg/kg males was significantly less than that of the vehicle controls. Clinical findings included prostration and ataxia in 600 mg/kg rats during the first week of the study and nasal/eye discharge, lethargy, ataxia, and abnormal breathing in 400 and 600 mg/kg females beginning week 5. An enlarged heart was noted in one 100 mg/kg female rat. Relative kidney weights of 100, 200, and 400 mg/kg males were significantly greater than that of the vehicle control group. Increased incidences and severities of myofiber cytoplasmic vacuolization and interstitial mononuclear cell infiltration in the heart occurred in 400 and 600 mg/kg male and female rats and in 200 mg/kg females. Increased incidences and severities of myofiber necrosis occurred in 600 mg/kg males and females; one female each in the 200 and 400 mg/kg groups also had this lesion. Three 600 mg/kg males had atrial thrombosis. 3-MONTH STUDY IN MICE: Groups of 10 male and 10 female mice were dermally administered 0, 50, 100, 200, 400, or 600 mg bis(2-chloroethoxy)methane/kg body weight in ethanol, 5 days per week for 14 weeks. Except for three 600 mg/kg females, all mice survived to the end of the study. Mean body weights of dosed and vehicle control mice were similar. One 600 mg/kg female that died early exhibited lethargy, abnormal breathing, and tremors, and one animal had clonic seizures. One 600 mg/kg female that died early had focal erosion of the glandular stomach and a focus in the duodenum found to consist of acute suppurative inflammation and thrombosis. Absolute and relative kidney weights of 400 and 600 mg/kg males and 600 mg/kg females were significantly greater than those of the vehicle control groups. Absolute liver weights of 400 and 600 mg/kg females were also significantly increased. Significantly increased incidences of myofiber cytoplasmic vacuolization occurred in 400 and 600 mg/kg females. 2-YEAR STUDY IN RATS: Groups of 50 male and 50 female rats were dermally administered 0, 75, 150, or 300 mg bis(2-chloroethoxy)methane/kg body weight in ethanol, 5 days per week for 105 weeks. Survival of all dosed groups of rats was generally similar to that of the vehicle controls. Mean body weights of dosed rats were similar to those of the vehicle controls throughout the study. Clinical findings in 300 mg/kg females that died during the first year of the study included abnormal breathing, lethargy, thinness, nasal discharge, and ataxia. Significantly increased incidences of degeneration of the olfactory epithelium in the nose occurred in all dosed groups of males and in 150 and 300 mg/kg females. The incidences of inflammation of the forestomach were significantly increased in 150 and 300 mg/kg males, and the incidence of ulcers was significantly increased in 300 mg/kg males. Increased incidences of cystic degeneration of the liver occurred in 150 and 300 mg/kg male rats; the incidence was significantly increased in the 300 mg/kg group. 2-YEAR STUDY IN MICE: Groups of 50 male mice were dermally administered 0, 150, 300, or 600 mg bis(2-chloroethoxy)methane/kg body weight in ethanol, 5 days per week for 105 weeks. Groups of 50 female mice were dermally administered 0, 100, 200, or 400 mg/kg in ethanol, 5 days per week for 104 weeks. Survival of 600 mg/kg male mice was significantly less than that of the vehicle control group. Mean body weights of dosed mice were generally similar to those of the vehicle controls throughout the study. Clinical findings observed in 600 mg/kg male mice that died during the first year of the study included lethargy and thinness. Myocardial heart changes were recorded according to the characteristic lesions of cardiomyopathy syndrome (necrosis, mononuclear cell infiltration, myocardial cell vacuolization, and interstitial fibrosis) separately, and in addition, where appropriate, they were also categorized as cardiomyopathy. Increased incidences of cardiomyopathy and mononuclear cell infiltration occurred in 600 mg/kg males and 400 mg/kg females; the incidences were significantly increased in 600 mg/kg males compared to the vehicle controls. Significantly increased incidences of cardiomyocyte vacuolization and interstitial fibrosis occurred in 600 mg/kg males. A few early deaths in the 600 mg/kg males were considered to be due, at least in part and probably exclusively, to bis(2-chloroethoxy)methane-induced cardiotoxicity. The incidence of ulceration of the forestomach was significantly increased in 600 mg/kg males. Significantly increased incidences of dermal inflammation and fibrosis and epidermal hyperplasia at the site of application occurred in 600 mg/kg male mice. GENETIC TOXICOLOGY: Bis(2-chloroethoxy)methane was mutagenic in S. typhimurium strains TA100 and TA1535 in the presence of exogenous metabolic activation enzymes (S9) in one study; results from a second bacterial mutagenicity test were judged to be equivocal based on responses observed in TA100 and in E. coli strain WP2 uvrA/pKM101 in the presence of S9. No mutagenicity was observed in other tester strains or in the absence of S9. Bis(2-chloroethoxy)methane did not increase the frequency of micronucleated reticulocytes in bone marrow of male F344/N rats following three daily treatments by gavage or micronucleated erythrocytes in peripheral blood of male or female mice after 3 months of dermal exposure. CONCLUSIONS: Under the conditions of these 2-year dermal studies, there was no evidence of carcinogenic activity of bis(2-chloroethoxy)methane in male or female F344/N rats administered 75, 150, or 300 mg/kg. There was no evidence of carcinogenic activity of bis(2-chloroethoxy)methane in male B6C3F1 mice administered 150, 300, or 600 mg/kg or in female B6C3F1 mice administered 100, 200, or 400 mg/kg. The administration of bis(2-chloroethoxy)methane for 2 years resulted in increased incidences of nonneoplastic lesions in the nose of male and female rats, the forestomach of male rats, the heart of male and female mice, and the forestomach and skin of male mice.     

Disrupted G1 to S phase clearance via cyclin signaling impairs liver tissue repair in thioacetamide-treated type 1 diabetic rats

Previously we reported that a nonlethal dose of thioacetamide (TA, 300 mg/kg) causes 90% mortality in type 1 diabetic (DB) rats because of irreversible acute liver injury owing to inhibited hepatic tissue repair, primarily due to blockage of G(0) to S phase progression of cell division cycle. On the other hand, DB rats receiving 30 mg TA/kg exhibited equal initial liver injury and delayed tissue repair compared to nondiabetic (NDB) rats receiving 300 mg TA/kg, resulting in a delay in recovery from liver injury and survival. The objective of the present study was to test the hypothesis that impaired cyclin-regulated progression of G(1) to S phase of the cell cycle may explain inhibited liver tissue repair, hepatic failure, and death, contrasted with delayed liver tissue repair but survival observed in the DB rats receiving 300 in contrast to 30 mg TA/kg. In the TA-treated NDB rats sustained MAPKs and cyclin expression resulted in higher phosphorylation of retinoblastoma (pRb), explaining prompt tissue repair and survival. In contrast, DB rats receiving the same dose of TA (300 mg/kg) exhibited suppressed MAPKs and cyclin expression that led to inhibition of pRb, inhibited tissue repair, and death. On the other hand, DB rats receiving 30 mg TA/kg exhibited delayed up regulation of MAPK signaling that delayed the expression of CD1 and pRb, explaining delayed stimulation of tissue repair observed in this group. In conclusion, the hepatotoxicant TA has a dose-dependent adverse effect on cyclin-regulated pRb signaling: the lower dose causes a recoverable delay, whereas the higher dose inhibits it with corresponding effect on the ultimate outcomes on hepatic tissue repair; this dose-dependent adverse effect is substantially shifted to the left of the dose response curve in diabetes.     

Developmental toxicity in the pregnant rabbit by the class III antiarrhythmic drug sotalol

The purpose of this study was to investigate the potential of sotalol to cause developmental toxicity in the pregnant rabbit. Sotalol is a beta-adrenoceptor blocking drug which also has class III antiarrhythmic properties via Ikr channel blockade. EXPERIMENT 1: Nine pregnant New Zealand White rabbits were given doses of either 300, 225, or 150 mg/kg of sotalol during gestational days, called Days, 13-16 which resulted in total litter loss. EXPERIMENT 2: A single dose of sotalol, 100 or 150 mg/kg was administered during Days 8-17 to 15 rabbits. Dosing on Day 8, 9, or 10 resulted in a slightly higher incidence of embryonic death compared to historical controls. There was marked increased embryonic death of 55-90% (four does with total litter loss), decreased number of live foetuses per litter, and elevated mean foetal weight after dosing during Days 12-16. EXPERIMENT 3: 16 pregnant rabbits were administered single doses of sotalol of either 100, 85, 75, 60 or 50 mg/kg on Day 14. The main finding was increased embryonic death, which ranged from total litter loss to approximately 30% at 50 mg/kg. At 50 mg/kg, the maternal Cmax, AUC(1-24 hr), and t1/2 were approximately 45 microM, 340 micromol x hr/l, and 6 hr, respectively. In conclusion, sotalol treatment resulted in embryonic death in the rabbit in early pregnancy in the same way as has been seen for other drugs with Ikr blocking properties (class III antiarrhythmics) in rodents. The observed developmental toxicity in the rabbit is most likely secondary to embryonic arrhythmia as has been shown in rodent studies. The results may indicate that Ikr blocking agents are developmental toxicants across species including man.     

Toxicity of Reldan (Chlorpyrifos-methyl) in Nubian goats

Eight Nubian goats were given single oral doses of 1200, 600, 300 or 150 mg/kg of Reldan. The 2 goats receiving the highest dose died after 1 and 16 hr whereas other animals survived the 7-day observation period. Twelve Nubian goats were given Reldan orally at daily dosages of 600, 300, 150 or 75 mg/kg. The goats died or were killed in extremis between 2 and 25 days. Pathology, clinical chemistry and hematology findings are reported.     

Enhanced hepatotoxicity and toxic outcome of thioacetamide in streptozotocin-induced diabetic rats

Diabetes is known to potentiate thioacetamide (TA)-induced liver injury via enhanced bioactivation. Little attention has been given to the role of compensatory tissue repair on ultimate outcome of hepatic injury in diabetes. The objective of this study was to investigate the effect of diabetes on TA-induced liver injury and lethality and to investigate the underlying mechanisms. We hypothesized that hepatotoxicity of TA in diabetic rats would increase due to enhanced bioactivation-mediated liver injury and also due to compromised compensatory tissue repair, consequently making a nonlethal dose of TA lethal. On day 0, male Sprague-Dawley rats (250-300 g) were injected with streptozotocin (STZ, 60 mg/kg ip) to induce diabetes. On day 10 the STZ-induced diabetic rats and the nondiabetic rats received a single dose of TA (300 mg/kg ip). This normally nonlethal dose of TA caused 90% mortality in the STZ-induced diabetic rats. At various times (0-60 h) after TA administration, liver injury was assessed by plasma alanine aminotransferase (ALT), sorbitol dehydrogenase (SDH), and liver histopathology. Liver function was evaluated by plasma bilirubin. Cell proliferation and tissue repair were evaluated by [(3)H]thymidine ((3)H-T) incorporation and proliferating cell nuclear antigen (PCNA) assays. In the nondiabetic rat, liver necrosis peaked at 24 h and declined thereafter toward normal by 60 h. In the STZ-induced diabetic rat, however, liver necrosis was significantly increased from 12 h onward and progressed, culminating in liver failure and death. Liver tissue repair studies showed that, in the liver of nondiabetic rats, S-phase DNA synthesis was increased at 36 h and peaked at 48 h following TA administration. However, DNA synthesis was approximately 50% inhibited in the liver of diabetic rats. PCNA study showed a corresponding decrease of cell-cycle progression, indicating that the compensatory tissue repair was sluggish in the diabetic rats. Further investigation of tissue repair by employing equitoxic doses (300 mg TA/kg in the non-diabetic rats; 30 mg TA/kg in the diabetic rats) revealed that, despite equal injury up to 24 h following injection, the tissue repair response in the diabetic rats was much delayed. The compromised tissue repair prolonged liver injury in the diabetic rats. These studies suggest that the increased TA hepatotoxicity in the diabetic rat is due to combined effects of increased bioactivation-mediated liver injury of TA and compromised compensatory tissue repair.     

26-week intravenous toxicity test of cefpiramide in cynomolgus monkeys

A chronic toxicity of cefpiramide (CPM) was studied in Cynomolgus monkeys. Groups of 4 males and 4 females were given daily doses of 100, 300 mg/kg or 600 mg/kg by intravenous administration for 26 weeks. Another group of 4 males and 4 females was given physiological saline and served as the control. In CPM groups, diarrhea or soft feces was observed after the commencement of administration at every doses. The frequency of these signs decreased as the study progressed and animals at lower doses returned to normal earlier. There were no treatment-related changes in body weights, fecal occult blood and electrocardiograms. A female in the 600 mg/kg group died after 13 weeks of treatment. The cause of death was assumed to be an acute myocardial necrosis resulting from thromboarteritis. It was not considered to be treatment-related. Ophthalmological examination revealed no abnormalities attributable to the treatment. Erythrocyte counts, hemoglobin and hematocrit values decreased in 3 animals receiving 600 mg/kg of the drug. The changes, however, were transient at least in 2 of these animals. There were no effects on plasma biochemical and urinalysis parameters. Small yellow spotes were noted in the renal cortex of 2 monkeys in the 600 mg/kg group at necropsy. Liver and kidneys were slightly heavier in animals receiving 600 mg/kg. Histopathological examinations revealed focal nonsuppurative interstitial nephritis in 1 animal given 300 mg/kg and 5 animals given 600 mg/kg. No treatment-related changes were observed in other organs and tissues. From these results, the maximum non-effective dose level of CPM was considered to be 100 mg/kg.     

Temporal Changes in Tissue Repair Permit Survival of Diet-Restricted Rats from an Acute Lethal Dose of Thioacetamide

Although, diet restriction (DR) has been shown to substantiallyincrease longevity while reducing or delaying the onset of agerelateddiseases, little is known about the mechanisms underlying thebeneficial effects of DR on acute toxic outcomes. An earlierstudy (S. K. Ramaiah et al., 1998, Toxicol. Appl. Pharmacol.150, 12–21) revealed that a 35% DR compared to ad libitum(AL) feeding leads to a substantial increase in liver injuryof thioacetamide (TA) at a low dose (50 mg/kg, ip). Higher liverinjury was accompanied by enhanced survival. A prompt and enhancedtissue repair response in DR rats at the low dose (sixfold higherliver injury) occurred, whereas at equitoxic doses (50 mg/kgin DR and 600 mg/kg in AL rats) tissue repair in AL rats wassubstantially diminished and delayed. The extent of liver injurydid not appear to be closely related to the extent of stimulatedtissue repair response. The purpose of the present study wasto investigate the time course (0–120 h) of liver injuryand liver tissue repair at the high dose (600 mg TA/kg, ip,lethal in AL rats) in AL and DR rats. Male Sprague-Dawley rats(225–275 g) were 35% diet restricted compared to theirAL cohorts for 21 days and on day 22 they received a singledose of TA (600 mg/kg, ip). Liver injury was assessed by plasmaALT and by histopathological examination of liver sections.Tissue repair was assessed by [³H]thymidine incorporation intohepatonuclear DNA and proliferating cell nuclear antigen (PCNA)immunohistochemistry during 0–120 h after TA injection.In AL-fed rats hepatic necrosis was evident at 12 h, peakedat 60 h, and persisted thereafter until mortality (3 to 6 days).Peak liver injury was approximately twofold higher in DR ratscompared to that seen in AL rats. Hepatic necrosis was evidentat 36 h, peaked at 48 h, persisted until 96 h, and returnedto normal by 120 h. Light microscopy of liver sections revealedprogression of hepatic injury in AL rats whereas injury regressedcompletely leading to recovery of DR rats by 120 h. Progressionof injury led to 90% mortality in AL rats vs 30% mortality inDR group. In the surviving AL rats, S-phase DNA synthesis wasevident at 60 h, peaked at 72 h, and declined to base levelby 120 h, whereas in DR rats S-phase DNA synthesis was evidentat 36 h and was consistently higher until 96 h reaching controllevels by 120 h. PCNA studies showed a corresponding increasein cells in S and M phase in the AL and DR groups. DR resultedin abolition of the delay in tissue repair associated with thelethal dose of TA in ad libitum rats. Temporal changes and highertissue repair response in DR rats (earlier and prolonged) arethe conduits that allow a significant number of diet restrictedrats to escape lethal consequence.     

Developmental Toxicity in the Pregnant Rabbit by theClass III Antiarrhythmic Drug Sotalol

The purpose of this study was to investigate the potential of sotalol to cause developmental toxicity in the pregnant rabbit. Sotalol is a β-adrenoceptor blocking drug which also has class III antiarrhythmic properties via Ikr channel blockade. Experiment 1: Nine pregnant New Zealand White rabbits were given doses of either 300, 225, or 150 mg/kg of sotalol during gestational days, called Days, 13–16 which resulted in total litter loss. Experiment 2: A single dose of sotalol, 100 or 150 mg/kg was administered during Days 8–17 to 15 rabbits. Dosing on Day 8, 9, or 10 resulted in a slightly higher incidence of embryonic death compared to historical controls. There was marked increased embryonic death of 55–90% (four does with total litter loss), decreased number of live foetuses per litter, and elevated mean foetal weight after dosing during Days 12–16. Experiment 3: 16 pregnant rabbits were administered single doses of sotalol of either 100, 85, 75, 60 or 50 mg/kg on Day 14. The main finding was increased embryonic death, which ranged from total litter loss to ∼30% at 50 mg/kg. At 50 mg/kg, the maternal C_max, AUC_{1–24 hr}, and t_1/2 were approximately 45 μM, 340 μmol×hr/l, and 6 hr, respectively. In conclusion, sotalol treatment resulted in embryonic death in the rabbit in early pregnancy in the same way as has been seen for other drugs with Ikr blocking properties (class III antiarrhythmics) in rodents. The observed developmental toxicity in the rabbit is most likely secondary to embryonic arrhythmia as has been shown in rodent studies. The results may indicate that Ikr blocking agents are developmental toxicants across species including man.     

The combined effect of dursban and Reldan on nubian goats.

Symptoms, lesions and changes in clinical chemistry and hematology were examined in goats orally dosed with dursban (150 mg/kg), reldan (150 mg/kg) and their mixture (300 mg/kg) in certain proportions. More rapid death and severe changes occurred in goats receiving reldan and dursban at 50% of the combined dose than either of the individual compounds.     

Toxicology and carcinogenesis studies of isoeugenol (CAS No. 97-54-1) in F344/N rats and B6C3F1 mice (gavage studies)

Isoeugenol is one of several structurally similar phenylpropenoid compounds produced by plants. It has been extracted from calamus, savory, basil, ylang-ylang, clove, tuberose, jonquil, nutmeg, tobacco, sandalwood, dill seed, mace, gardenia, petunia, and other flowers. Isoeugenol can also be produced by isomerization of eugenol, which occurs naturally in clove, pimento, bay leaf, and cinnamon. As a fragrance with a spicy, carnation-like odor, isoeugenol is incorporated into numerous household and personal hygiene products, including perfumes, cream lotions, soaps, and detergents. As a flavoring agent, isoeugenol is added to nonalcoholic drinks, baked foods, and chewing gums. Isoeugenol was nominated by the National Cancer Institute and was selected for carcinogenicity testing because of widespread human exposure through its use as a flavoring and fragrance agent and because of its structural similarity to phenylpropenoids such as safrole, isosafrole, eugenol, methyleugenol, estragole, and anethole, most of which are known rodent carcinogens. Male and female F344/N rats and B6C3F1 mice were administered isoeugenol (99% or greater pure) in corn oil by gavage for 3 months or 2 years. Genetic toxicity tests were conducted in Salmonella typhimurium, Escherichia coli, cultured Chinese hamster ovary cells, and mouse peripheral blood erythrocytes. 3-MONTH STUDY IN RATS: Groups of 10 male and 10 female rats were exposed to isoeugenol in corn oil by gavage at doses of 0, 37.5, 75, 150, 300, or 600 mg/kg, 5 days per week for 14 weeks. All rats survived to the end of the study except one 600 mg/kg male and one 37.5 mg/kg female that were killed in dosing accidents. Mean body weights of all exposed groups of males were significantly less than that of the vehicle control group; however, only the decrease for the 600 mg/kg group exceeded 10% and was considered related to isoeugenol exposure. Liver weights were significantly increased in 300 and 600 mg/kg females. The incidences of minimal atrophy of the olfactory epithelium of the nose were significantly increased in 150 mg/kg or greater males and in 300 or 600 mg/kg females. The incidence of atrophy of olfactory nerve bundles was significantly increased in 600 mg/kg females. Minimal to mild periportal hepatocellular cytoplasmic alteration occurred in all 300 or 600 mg/kg females. 3-MONTH STUDY IN MICE: Groups of 10 male and 10 female mice were exposed to isoeugenol in corn oil by gavage at doses of 0, 37.5, 75, 150, 300, or 600 mg/kg, 5 days per week for 14 weeks. All mice survived to the end of the study. The mean body weight of 600 mg/kg males was significantly less (12%) than that of the vehicle controls. Liver weights of 300 and 600 mg/kg males were significantly greater than those of the vehicle controls. Minimal to moderate atrophy of olfactory epithelial tissue and nerve bundles was observed in 600 mg/kg males and females. 2-YEAR STUDY IN RATS: Groups of 50 male and 50 female rats were exposed to isoeugenol in corn oil by gavage at doses of 0, 75, 150, or 300 mg/kg, 5 days per week for 105 weeks. Survival rates of exposed male and female rats were similar to those of vehicle controls. Mean body weights of 300 mg/kg male rats were 9% greater than the vehicle controls at the end of the study. The general lack of toxicity and nonneoplastic lesions indicates that rats might have been able to tolerate higher doses. Two male rats in the 300 mg/kg group had rare benign or malignant thymomas, while two other males in this group had rare mammary gland carcinomas. Low incidences of minimal atrophy and minimal to mild respiratory metaplasia of the olfactory epithelium were increased in 150 mg/kg males and 300 mg/kg males and females. Similar incidences of minimal to mild olfactory epithelial degeneration in 300 mg/kg males were also increased. Incidences of keratoacanthoma of the skin were decreased in 150 and 300 mg/kg males. 2-YEAR STUDY IN MICE: Groups of 50 male and 50 female mice were exposed to isoeugenol in corn oil by gavage at doses of 0, 75, 150, or 300 mg/kg, 5 days per week for 104 (females) or 105 (males) weeks. Survival of 300 mg/kg males was significantly decreased compared to the vehicle controls. Mean body weights of 300 mg/kg male and female groups were less than those of vehicle controls at the end of the study, 10% and 15% less, respectively. In all groups of exposed males, the incidences of hepatocellular adenoma, hepatocellular carcinoma, and hepatocellular adenoma or carcinoma (combined) were significantly greater than those in the vehicle control group; incidences of multiple hepatocellular adenoma were also significantly increased. Incidences of clear cell focus were significantly increased in 75 and 150 mg/kg male mice. There was a significant positive trend in the incidences of histiocytic sarcoma in females, and this neoplasm occurred in multiple tissues. Incidences of respiratory metaplasia in olfactory epithelium in all exposed groups and of atrophy and hyaline droplet accumulation in all exposed groups except 75 mg/kg females were significantly greater than those in corresponding vehicle control groups. Incidences of minimal to marked hyperplasia of Bowman's gland were increased significantly in all exposed groups. Incidences of minimal to mild necrosis of renal papilla and mild to moderate necrosis of renal tubules were increased significantly in 300 mg/kg females. Incidences of forestomach squamous hyperplasia, inflammation, and ulceration (males only) increased with exposure and were significant in the 300 mg/kg groups. The incidence of glandular stomach ulcers was low but significantly increased in the 300 mg/kg groups. GENETIC TOXICOLOGY: Isoeugenol was not mutagenic in two independent assays in bacteria (S. typhimurium and E. coli) conducted with and without exogenous metabolic activation (S9 liver enzymes). Neither did it induce chromosomal aberrations in cultured Chinese hamster ovary cells, with or without S9 activation. Frequencies of micronucleated erythrocytes were not increased in peripheral blood of male mice exposed to isoeugenol by gavage for 3 months; however, an increasing trend and a threefold increase in the 600 mg/kg group indicate a positive result for this test in female mice. CONCLUSIONS: Under the conditions of these 2-year gavage studies, there was equivocal evidence of carcinogenic activity of isoeugenol in male F344/N rats based on increased incidences of rarely occurring thymoma and mammary gland carcinoma. There was no evidence of carcinogenic activity of isoeugenol in female F344/N rats administered 75, 150, or 300 mg/kg. There was clear evidence of carcinogenic activity of isoeugenol in male B6C3F1 mice based on increased incidences of hepatocellular adenoma, hepatocellular carcinoma, and hepatocellular adenoma or carcinoma (combined). There was equivocal evidence of carcinogenic activity of isoeugenol in female B6C3F1 mice based on increased incidences of histiocytic sarcoma. Exposure to isoeugenol resulted in nonneoplastic lesions of the nose in male and female rats; of the nose, forestomach, and glandular stomach in male and female mice; and of the kidney in female mice.     

Mechanisms of increased liver tissue repair and survival in diet-restricted rats treated with equitoxic doses of thioacetamide.

Moderate dietary or caloric restriction (DR) modulates animal physiology in a beneficial fashion. Previously, we have reported an equitoxic dose experiment where liver injury in DR male Sprague-Dawley rats exposed to a low dose of thioacetamide (TA, 50 mg/kg) was similar to that observed in ad libitum fed (AL) rats exposed to a 12-fold higher dose (600 mg/kg). Paradoxically, the AL rats experienced 90% mortality while all of the DR rats, with the same amount of initial bioactivation-mediated liver injury, survived. The protection observed in the DR rats was due to efficient compensatory liver tissue repair, which was delayed and attenuated in the AL rats, leading to progression of liver injury. The objective of the present study was to investigate the molecular mechanisms of the enhanced tissue repair in the DR rats upon equitoxic challenge with TA. Promitogenic mechanisms and mediators such as proinflammatory cytokines (TNF-alpha and IL-6), growth factors (TGF-alpha and HGF), and inducible nitric oxide synthase (iNOS) were estimated over a time course after equitoxic challenge (50 mg/kg to DR vs. 600 mg/kg to AL rats). Except for TNF-alpha, all other molecules were expressed earlier and in greater amount in the DR rats. IL-6 was 10-fold greater and peaked 12 h earlier; HGF also peaked 12 h sooner in the DR rats, when it was 2.5-fold greater than the value in the AL rats. TGF-alpha expression in livers of DR rats increased after TA administration and peaked at 24 h. In the AL rats, it was lower and peaked at 36 h. Diet restriction alone induced iNOS 2-fold in the DR rats and remained elevated until 12 h after TA administration, then declined thereafter. The lower iNOS activity in the AL rats further decreased after TA injection. DR rats exhibited higher apoptosis after thioacetamide administration, which further increased the efficiency of tissue repair. Taken together, these data indicate that even though the liver injury is near equal in AL and DR rats, sluggish signal transduction leads to delayed liver regeneration, progression of liver injury, and death in the AL rats. The equitoxic dose experiment indicates that stimulation of tissue repair is independent of the extent of initial liver injury and is governed by physiology of diet restriction. DR stimulates promitogenic signaling leading to a quick and timely response upon liver injury, arrest of progressive injury on one hand, and recovery from injury on the other, paving the way for survival of the DR rats.     

Toxicokinetics and toxicity of thioacetamide sulfoxide: a metabolite of thioacetamide

Thioacetamide (TA) is bioactivated by CYP2E1 to TA sulfoxide (TASO), and to the highly reactive sulfdioxide (TASO₂), which initiates hepatic necrosis by covalent binding. Previously, we have established that TA exhibits saturation toxicokinetics over a 12-fold dose range, which explains the lack of dose–response for bioactivation-based liver injury. In vivo and in vitro studies indicated that the second step (TASO → TASO₂) of TA bioactivation is less efficient than the first one (TA → TASO). The objective of the present study was to specifically test the saturation of the second step of TA bioactivation by directly administering TASO, which obviates the contribution from first step, i.e. TA → TASO. Male SD rats were injected with low (50 mg/kg, ip), medium (100 mg/kg) and high (LD₇₀, 200 mg/kg) doses of TASO. Bioactivation-mediated liver injury that occurs in the initial time points (6 and 12 h), estimated by plasma ALT, AST and liver histopathology over a time course, was not dose-proportional. Escalation of liver injury thereafter was dose dependent: low dose injury subsided; medium dose injury escalated upto 36 h before declining; high dose injury escalated from 24 h leading to 70% mortality. TASO was quantified in plasma by HPLC at various time points after administration of the three doses. With increasing dose (i.e., from 50 to 200 mg/kg), area under the curve (AUC) and C_max increased more than dose proportionately, indicating that TASO bioactivation exhibits saturable kinetics. Toxicokinetics and initiation of liver injury of TASO are similar to that of TA, although TASO-initiated injury occurs at lower doses. These findings indicate that bioactivation of TASO to its reactive metabolite is saturable in the rat as suggested by previous studies with TA.     

Pharmacokinetics of lopromide Liposomes in Rabbits

Purpose. The dose-proportionality of pharmacokinetics of an iodinated contrast medium, iopromide, encapsulated into liposomes was investigated. Methods. Following single intravenous administration of 150 mg iodine/kg (potential diagnostic dose) and a five-fold higher dose in rabbits the pattern of elimination was studied until 7 d and the blood concentrations were monitored up to 72 h after administration. The iodine concentration in the liver was calculated on the basis of the blood concentration and related to the concentration measured in the rabbit liver. Results. The dose-normalized blood concentration-time profiles of the encapsulated iodine were not superimposable. Contrary to the low dose a steady-state concentration of 2.8 mg iodine/mL was observed in blood for 60 min after the high dose administration indicating a saturation of the liposomal liver uptake. For both doses the elimination of iodine occurred predominantly via the kidneys and was complete 7 d after administration. The dose-normalized amounts of iodine excreted with the urine were similar for both dose groups. From the blood data it was calculated that doses up to about 300 mg iodine/kg should result in a dose-proportional increase of liposomal liver uptake before saturation occurs. This was confirmed by the measured iodine liver concentrations after increasing the doses stepwise from 150 to 750 mg iodine/kg. Conclusions. In rabbits for the dose range 150 to 750 mg iodine/kg iopromide liposomes reveal dose-dependent pharmacokinetics due to a saturation in liver uptake which occurs for doses of 300 mg iodine/kg corresponding to 300 mg lipid/kg onwards.     

Teratogenesis of Calcium Valproate in Rats

Teratogenesis of Calcium Valproate in Rats. Ong, Lina L., Schardein,James L., Petrere, Judith A., Sakowski, Raymond, Jordan, Hollis,Humphrey, Ronald R., Fitzgerald James E., and de la Iglesia,Felix A. (1983). Fundam. Appl. Toxicol. 3:121-126. Studies wereconducted to determine the teratogenic potential of the calciumsalt of valproic acid in rats when given orally at doses of600, 150, and 50 mg/kg on days 6–15 of gestation. Thesodium salt of valproic acid was used as a reference agent ata dose level of 600 mg/kg. The administration of 600 mg/kg/dayof either calcium or sodium valproate resulted in transient,severe sedation in the dams. Four dams receiving 600 mg/kg ofeither salt died during the experiment, with deaths occurringbetween day 7 and 11 of gestation. Food consumption and bodyweight gain were significantly reduced during the dosing periodwith both salts at dose levels of 600 mg/kg. Embryotoxicityat the high doses(600 mg/kg) with either salt was manifestedby increases in fetal resorption, reduced body weights, andsignificantly increased Incidence of supernumerary ribs andbifid vertebral centra among the surviving fetuses. A teratogeniceffect was evident at 600 mg/kg with either salt of valproicacid. Seven of 16 fetuses from dams given the calcium salt wereabnormal. Findings included one with omphalocele and six otherswith skeletal malformations. Eleven of 24 fetuses from damsgiven the sodium salt were abnormal: three littermates had bilateralectrodactyly of the rear feet and malformed vertebral centraand eight others had skeletal malformations. No teratogeniceffect was evident among the fetuses from dams given 150 mg/kgcalcium salt. Embryotoxicity was demonstrated by a significantincrease in the incidence of supernumerary ribs. No adverseeffect was observed among the fetuses from dams given 50 mg/kgof the calcium salt.     

The Development Toxicity of 2-Ethylhexanoic Acid in Wistar Rats

The developmental toxicity of 2-ethylhexanoic acid (2-EHA),a wood preservative and a mammalian metabolite of di-(2-ethylhexyl)phthalatewas examined in Wistar rats (20–21 pregnant females/dose).Mated animals were exposed to 2-EHA in their drinking waterat doses of 100, 300, or 600 mg/kg/day on Days 6–19 ofgestation. Control animals received vehicle water. The fetuseswere examined (on Gestational Day 20) for external, visceral,and skeletal malformations and variations. 2-EHA was marginallytoxic to the dams at 600 mg/kg, but not at lower doses, sincethe mean near term body weight was reduced by 11%. This doselevel was also slightly fetotoxic as indicated by a 5 to 8%decrease in the mean fetal body weight both in males and females.No treatment-related effects were observed in the number ofimplantations or live fetuses. At doses of 100 mg/kg and above,2-EHA caused skeletal malformations (clubfoot, absence of fibula,polydactyly), while the development of visceral tissues wasless affected. The number of affected fetuses increased in adose-dependent way (4.9, 8.9, and 15.3% of treated offspringat 100, 300, and 600 mg/kg/day, respectively, vs 2.4% control).These results indicate that 2-EHA is teratogenic in rats alreadyat doses which are not yet maternally toxic. The skeleton appearsto be the main target of 2-EHA in developing rats.     

烟酰胺单核苷酸对内毒素休克小鼠死亡率的影响

目的 探究烟酰胺单核苷酸(NMN)对脂多糖(LPS)诱导的内毒素休克小鼠死亡率的影响.方法 将10周龄C57BL/6J雄性小鼠随机分组,均腹腔注射LPS(10 mg/kg)造模.NMN腹腔注射给药,分为3种方式:①造模后0.5 h给药,剂量为10、30、100、300 mg/kg;②造模前0.5 h给药,剂量为30、1...