Dimethylarsinic acid (DMA(V)) is carcinogenic to the rat urinary bladder when administered at high doses in the diet or drinking water. At a dietary dose of 100 ppm (microg/g), it produces cytotoxicity within 6 h and increased proliferation (hyperplasia) by 7 days of administration. We hypothesize that formation of the reactive organic intermediate dimethylarsinous acid (DMA(III)) is involved in the induction of the cytotoxicity. To evaluate the possibility that DMA(V) administration produces urothelial toxicity and regeneration by the formation of trivalent arsenicals, 2,3-dimercaptopropane-1-sulfonic acid (DMPS, 5600 ppm), a chelator of trivalent arsenicals, was co-administered with DMA(V) (100 ppm) for 2 weeks to groups of female Fischer F344 rats. Based on light and scanning electron microscopy, and bromodeoxyuridine labeling index, DMA(V) produced cytotoxicity and regenerative hyperplasia of the urothelium which was inhibited by co-administration with DMPS. The major forms of arsenic in the 24-h urine of rats administered DMA(V) were high concentrations of DMA(V) (66.4 +/- 2.7 microM) itself and the pentavalent organic arsenical trimethylarsine oxide (TMAO) (73.2 +/- 9.5 microM). Co-administration with DMPS led to an increase in DMA(V) (507 +/- 31 microM) with a decrease in TMAO (2.8 +/- 0.4 microM) excretion. The formation of TMAO from DMA(V) mechanistically suggests formation of the intermediate trivalent metabolite, DMA(III). In a second experiment evaluating fresh void urines collected on study days 1, 71, and 175, we detected DMA(III) in the urine of DMA(V) and DMA(V) plus DMPS-treated rats at approximately micromolar concentrations. Using rat (MYP3) and human (1T1) urothelial cells, cytotoxicity for trivalent arsenicals, sodium arsenite, monomethylarsonous acid (MMA(III)), and DMA(III) was demonstrated at 0.4-4.8 microM concentrations, whereas MMA(V), DMA(V), and TMAO were cytotoxic at millimolar concentrations. The presence of DMA(III) at micromolar concentrations in the urine of rats fed 100 ppm DMA(V) suggests that DMA(III) produced in vivo may be involved in the toxic effects in the rat urinary bladder after dietary administration of DMA(V).

Muraglitazar, a PPARα/γ agonist, dose-dependently increased urinary bladder tumors in male Harlan Sprague-Dawley (HSD) rats administered 5, 30, or 50 mg/kg/day for up to 2 years. To determine the mode of tumor development, male HSD rats were treated daily for up to 21 months at doses of 0, 1, or 50 mg/kg while being fed either a normal or 1% NH 4 Cl-acidified diet. Muraglitazar-associated, time-dependent changes in urine composition, urothelial mitogenesis and apoptosis, and urothelial morphology were assessed. In control and treated rats fed a normal diet, urine pH was generally ≥ 6.5, which facilitates formation of calcium-and magnesium-containing solids, particularly in the presence of other prolithogenic changes in rat urine. Urinary citrate, an inhibitor of lithogenesis, and soluble calcium concentrations were dose dependently decreased in association with increased calcium phosphate precipitate, crystals and/or microcalculi; magnesium ammonium phosphate crystals and aggregates; and calcium oxalate-containing thin, rod-like crystals. Morphologically, sustained urothelial cytotoxicity and proliferation with a ventral bladder predilection were noted in treated rats by month 1 and urinary carcinomas with a similar distribution occurred by month 9. Urothelial apoptotic rates were unaffected by muraglitazar treatment or diet. In muraglitazar-treated rats fed an acidified diet, urine pH was invariably < 6.5, which inhibited formation of calcium-and magnesium-containing solids. Moreover, dietary acidification prevented the urothelial cytotoxic, proliferative, and tumorigenic responses. Collectively, these data support an indirect pharmacologic mode of urinary bladder tumor development involving alterations in urine composition that predispose to urolithiasis and associated decreases in urine-soluble calcium concentrations.

Inorganic arsenic is a known human carcinogen of the skin and respiratory tract. Epidemiologic evidence indicates that it is also carcinogenic to the urinary bladder and other internal organs. Lack of an animal model has limited progress on understanding the mechanism of arsenic carcinogenesis. It was recently reported that high doses of an organic arsenical, dimethylarsinic acid (DMA), increased urinary bladder tumors in rats when administered in the diet or in the drinking water for 2 years, with the female being more sensitive than the male. We previously showed that high doses of DMA (40 or 100 ppm of the diet) fed for 10 weeks increased urothelial cell proliferation in the rat. Treatment with DMA also increased renal calcification and increased urinary calcium concentration. In 2 experiments, we examined the urothelial proliferative effects of treatment with 100 ppm DMA in the diet in female F344 rats for 2 and 10 weeks and for 6 and 24 h, and 3, 7, and 14 days. Cytotoxic changes in the urothelium were evident by SEM as early as 6 h after treatment was begun. Foci of cellular necrosis were detected after 3 days of treatment, followed by widespread necrosis of the urothelium after 7 days of treatment. The bromodeoxyuridine (BrdU) labeling index was not increased until after 7 days of treatment, suggesting that administration of DMA results in cytotoxicity with necrosis, followed by regenerative hyperplasia of the bladder epithelium. Although the rat provides an animal model to study the urothelial effects of DMA, the relevance of this finding to inorganic arsenic carcinogenesis in humans must be extrapolated cautiously, due to the high doses of DMA necessary to produce these changes in the rat and the differences in metabolism of arsenicals in rodents, especially rats, compared to humans.

Dimethylarsinic acid (DMA), fed to rats for 2 years, produced bladder hyperplasia and tumors at doses of 40 and 100 p.p.m., more in females than males. No urothelial proliferation was seen in mice. Our objectives were to investigate the mode of action of bladder tumor formation, evaluate the dose-response and the role of diet and to determine if the urothelial effects were reversible. The study included groups of female F344 rats fed DMA in Purina 5002 diet at doses of 0, 2, 10, 40 or 100 p.p.m. for 10 weeks; two groups of females fed DMA (0 and 100 p.p.m.) in Altromin 1321 for 10 weeks; two groups of males fed DMA (0 and 100 p.p.m.) in Purina 5002 for 10 weeks; a female high-dose recovery group (100 p.p.m. in Purina 5002 diet for 10 weeks followed by control diet for 10 weeks); and two female groups (0 and 100 p.p.m.) in Purina diet for 20 weeks. Urothelial toxicity and hyperplasia were detected by light and scanning electron microscopy (SEM), and the bromodeoxyuridine labeling index was increased in the female 40 and 100 p.p.m. groups. The effects were less in males, but were similar in females fed DMA in Altromin 1321. SEM detected no abnormal urinary solids related to treatment in any group. Urinary calcium was increased in the females fed 40 and 100 p.p.m. in Purina diet, despite overall urinary dilution. Calcification was increased in kidneys of female rats fed Purina diet. The urothelial effects of DMA were reversible. The findings support a non-DNA reactive mechanism for DMA rat bladder carcinogenicity related to urothelial toxicity and regeneration. The toxicity is probably not due to urinary solids. The toxicity and regeneration are produced in a dose-responsive manner in female rats, are greater in female than in male rats, and are reversible.