Species-dependent induction of peroxisome proliferation by haloxyfop, an aryloxyphenoxy herbicide.

The potential of haloxyfop [2-(4-((3-chloro-5-(trifluoromethyl)-2- pyridinyl)oxy)phenoxy)propanoic acid; HAL] to induce the proliferation of hepatocellular peroxisomes (PP) was examined in rats, mice, dogs, and monkeys. Chemically induced PP is associated with the development of liver tumors in rodents via an apparent species-dependent, nongenotoxic mechanism of action. HAL is nongenotoxic yet has been shown to cause liver tumors in female B6C3F1 mice. Ingestion of HAL by rats and/or mice (0.1-14 mg/kg/day for 2 to 4 weeks) resulted in significant dose-related PP as evidenced by hepatocellular hypertrophy, increased peroxisome volume density (VD), and induction of peroxisomal enzymes and CYP4A1. Only a relatively weak induction of PP was noted at a carcinogenic dosage in female mice. In contrast to rodent species, ingestion of up to 20 mg/kg/day HAL by male and female Beagle dogs for 13 weeks failed to increase peroxisomal VD while causing only a slight increase in peroxisomal enzyme activity at the highest dosages. Oral administration of up to 30 mg/kg/day HAL by male and female Cynomolgus monkeys for 13 weeks failed to induce PP. While a direct relationship of PP with tumor formation, at least in mice, was not demonstrated, these data still support the concept that PP represents a potential marker of nongenotoxic tumorigenic activity, at some dosage, in rodents.

Induction of fatty acid beta-oxidation and peroxisome proliferation in the liver of rhesus monkeys by DL-040, a new hypolipidemic agent.

Many structurally unrelated hypolipidemic agents and certain phthalate-ester plasticizers induce hepatomegaly and proliferation of peroxisomes in liver parenchymal cells of rodents, but there is relatively limited evidence regarding the ability of such compounds to induce peroxisome proliferation in the livers of nonrodent species including man. The present study was designed to determine if DL-040 (4-(((1,3-benzodioxol)-5-yl)methyl)amino-benzoic acid), a newly developed hypolipidemic agent, induces peroxisome proliferation in the liver of adult rhesus monkeys. Feeding of DL-040 (300 mg/kg body wt for 1 week; and 400 mg/kg body wt for 10 weeks) caused a significant increase in peroxisome population as determined by ultrastructural and morphometric analyses. The DL-040-induced peroxisome proliferation was accompanied by increases in the levels of catalase, carnitine acetyltransferase and the peroxisomal fatty acid beta-oxidation system. As expected, DL-040 caused a significant reduction of serum cholesterol and low density lipoprotein content. These data suggest that hepatic peroxisome proliferation is inducible in nonhuman primates at dose levels that exceed therapeutic levels.

Pancreatic beta cell vacuolation in rats after oral administration of hexamethylmelamine.

The effects of hexamethylmelamine (HMM) on morphology and function of the endocrine pancreas were investigated following the induction of a diabetic-like syndrome in a reproduction study in rats. Doses of 0, 20, 40, or 80 mg/kg/day were administered once daily, by gavage, to groups of 10 male and 10 female Sprague-Dawley rats for up to 77 days. Signs of toxicity were observed at all treatment levels and were of such severity at 80 mg/kg/day that this group was terminated after 8 days of treatment. Measurement of serum glucose levels revealed a dose- and time-related hyperglycemia in rats from the 20- and 40-mg/kg/day groups with levels exceeding 700 mg/dl in some rats after 77 days of treatment. Reversal of the hyperglycemia after discontinuing treatment with HMM occurred in some rats receiving 20 mg/kg/day, whereas the effects of higher doses were still present following a 2-month recovery period. Microscopically, there were hydropic degeneration of renal tubular epithelium and vacuolation of pancreatic beta cells. Ultrastructurally, vesiculation of the Golgi-smooth membrane reticular complex and a marked reduction in the number of insulin granules were observed. It appeared that HMM exerted a disruptive effect on the production of insulin at or prior to the level of the Golgi complex. The severity and time course of morphologic changes and hyperglycemia were dependent on dose and duration of treatment.

[Comparative study of the effects of probucol, fenofibrate and clofibrate on liver ultrastructure in rats (author's transl)]. / Etude comparative de l'action du probucol, du fénofibrate et du clofibrate sur l'ultrastructure hépatique chez le rat.

The effects on rat liver of probucol (250 or 500 mg/kg/24 h), clofibrate (250 mg/kg/24 h) and fenofibrate (100 mg/kg/24 h) were compared after 28 days' treatment. Th bodyweight of animals treated with probucol 250 mg/kg/24 h has increased, as compared with a control group of untreated animals. The increase in liver weight and live weight/bodyweight ratio was greater in rats treated with clofibrate or fenofibrate than in rats treated with probucol. Cholesterol and triglyceride plasma levels were lower in clofibrate-treated animals. There were no changes in SGOT and SGPT in any of the three groups. Alkaline phosphatases were increased only in rats treated with clofibrate (these enzymes were not assayed in rats which received fenofibrate). Electron microscope study of liver sections showed the following changes in clofibrate--and fenofibrate-treated rats as compared with the control group: increase in smooth-surfaced endoplasmic reticulum, ribosome detachment, slight glycogen depletion and increase in the number of peroxysomes. In the probucol-treated group, apart from an occasional increase in smooth-surfaced endoplasmic reticulum, liver cells were identical with those of the control group; in particular, there was no increase in the number of peroxysomes.

Eight-year toxicity study in monkeys and reproduction studies in rats and rabbits treated with probucol.

Probucol [4,4'-(isopropylidenedithio)bis(2,6-di-tert-butylphenol)], a hypocholesterolemic agent, was given orally to male and female rhesus monkeys at 0, 60, 125, 250, and 500 mg/kg . d for more than 8 yr without adverse effect. Of 40 monkeys on test, 14 were killed for interim studies (wk 81 and 102), 21 were maintained for more than 8 yr, and 5 were submitted for necropsy for conditions unrelated to treatment. Monitored parameters included growth rate, demeanor, hematology, clinical chemistries, urinalyses, ophthalmoscopy, organ weights, and gross, histopathologic, and electron microscopic evaluations. Bone marrow smears at the conclusion of the test revealed no differences between control and treated animals. Electron microscopy of liver specimens from monkeys treated for 8 yr revealed comparable hepatocellular ultrastructure in control and treated monkeys. Probucol was given orally at 0, 100, 500, and 1000 mg/kg to Sprague-Dawley rats during appropriate time intervals to evaluate effects on fertility and postnatal development. The same dose levels were given during organogenesis, or in some cases prior to breeding and throughout organogenesis, to Sprague-Dawley rats and New Zealand white rabbits. No adverse effects on fertility or postnatal development were observed in rats, and no evidence of teratogenicity was observed in either species. The results indicate that probucol does not affect reproduction of rats, lacks teratogenic potential in the species studied, and is nontoxic to subhuman primates treated for more than 8 yr.

Comparative ultrastructural study of rat hepatocytes after treatment with the hypolipidemic agents probucol, clofibrate, and fenofibrate.

Experimental findings are reported for two similarly conducted studies. One was designed to compared rat liver cell ultrastructure during and after 91 d of dosing with probucol, a hypocholesterolemic agent, and clofibrate, a hypolipidemic drug known to elicit marked alteration of rat hepatocellular morphology. The second was designed to similarly assess male rats after 28 d of treatment with the hypolipidemic agent fenofibrate. Diet mixes for these studies were prepared to attain dosage levels of approximately 500 mg/kg . d for probucol, 250 mg/kg . d for clofibrate, and 100 mg/kg . d for fenofibrate. Control rats were given untreated basal ration. Weekly adjustments in dietary concentrations were made in accordance with group mean food consumption and body weight changes. Probucol and clofibrate treatments produced statistically significant reductions in mean serum cholesterol levels of both sexes after 28 and 91 d of dosing. Only male rats were given fenofibrate, and they exhibited statistically significant cholesterol reductions after 28 d. Clofibrate and fenofibrate administration resulted in marked increases in liver weight/body weight ratios. Probucol had no statistically significant effects on liver weight/body weight ratios after 28 and 91 consecutive days of treatment. Light microscopy of liver sections stained with hematoxylin and eosin revealed an abnormal amount of cytoplasmic granularity within hepatocytes from rats given clofibrate and fenofibrate. The granules were identified by electron microscopy and cytochemistry as enlarged, proliferated peroxisomes--a known rat hepatocellular response to treatment with many hypolipidemic drugs. In addition, ultrastructural cytochemistry suggested reduced amounts of catalase in individual peroxisomes after clofibrate and fenofibrate dosing. Liver tissue from rats given probucol showed no abnormal cytoplasmic granularity and, ultrastructurally, no peroxisomal changes. Liver tissues from probucol-treated rats revealed features similar to those encountered in tissues from untreated control animals. It was concluded that the hypocholesterolemic response elicited by probucol treatment does not involve significant changes in rat liver cell morphology.