Margaret St. John

Frequent p53 mutations and occasional loss of chromosome 4 in invasive bladder carcinoma induced by N-butyl-N-(4-hydroxybutyl)nitrosamine in B6D2F1 mice.

B6D2F1 mice (45/group) were treated with N‐butyl‐N‐(4‐hydroxybutyl)nitrosamine (BBN) or uracil as follows: Group 1 received 0.05% BBN in drinking water for the entire experiment, Group 2 received 5 mg of BBN by gastric gavage in 0.1 mL of 20% ethanol twice per week for 10 wk, Group 3 received a 2.5% uracil–containing diet for the entire experiment, and Group 4 was controls (received 0.1 mL of 20% ethanol by gavage twice per week for 10 wk). The surviving mice in Group 1 were killed after week 26 and those in the other groups after week 30. By week 15, three of 11 Group 1 and one of 15 Group 2 mice had bladder carcinoma. By 26 and 30 wk, respectively, invasive carcinomas were observed in 33 of 34 and six of 21 mice in Groups 1 and 2 and renal pelvic carcinomas in 11 of 34 and three of 21 mice in Groups 1 and 2. Four of 19 uracil‐treated mice had bladder nodular hyperplasia. By polymerase chain reaction–single‐strand conformation polymorphism and sequence analyses, 16 of 20 and two of five bladder carcinomas from Groups 1 and 2, respectively, showed mutations in the p53 gene. Ha‐ras mutation was present in one case. Loss of heterozygosity analysis with simple‐sequence length polymorphism markers for chromosome 4 showed that 10 of 21, two of 15, and nine of 13 mice in Groups 1–3, respectively, had heterozygous or homozygous deletions. B6D2F1 mice are therefore susceptible to the urothelial carcinogenic effects of BBN and develop frequent p53 mutations and chromosome 4 deletions. Chromosome 4 deletions were also seen with uracil. Mol. Carcinog. 21:70–79, 1998.

Comparison of uroplakin expression during urothelial carcinogenesis induced by N-butyl-N-(4-hydroxybutyl)nitrosamine in rats and mice.

The expression of uroplakins, the tissue-specific and differentiation-dependent membrane proteins of the urothelium, was analyzed immunohistochemically in N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN)-treated rats and mice during bladder carcinogenesis. Male Fischer 344 rats were treated with 0.05% BBN in the drinking water for 10 wk and were euthanatized at week 20 of the experiment. BBN was administered to male B6D2F1 mice; it was either provided at a rate of 0.05% in the drinking water (for 26 wk) or 5 mg BBN was administered by intragastric gavage twice weekly for 10 wk, followed by 20 wk without treatment. In rats, BBN-induced, noninvasive, low-grade, papillary, transitional cell carcinoma (TCC) showed decreased uroplakin-staining of cells lining the lumen but showed increased expression in some nonluminal cells. In mice, nonpapillary, high-grade dysplasia, carcinoma in situ, and invasive carcinoma were induced. There was a marked decrease in the number of uroplakin-positive cells lining the lumen and in nonluminal cells. This occurred in normal-appearing urothelium in BBN-treated mice and in dysplasic urothelium, in carcinoma in situ, and in invasive TCC. The percentage of uroplakin-positive nonluminal cells was higher in control mice than in rats, but it was lower in the mouse than in the rat after BBN treatment. Uroplakin expression was disorderly and focal in BBN-treated urothelium in both species. These results indicate that BBN treatment changed the expression of uroplakins during bladder carcinogenesis, with differences in rats and mice being related to degree of tumor differentiation.

Tributyl Phosphate Effects on Urine and Bladder Epithelium in Male Sprague–Dawley Rats☆

Tributyl phosphate (TBP) produces tumors of the bladder urothelium in rats at high doses (700 and 3000 ppm), with greater effects in males than in females. TBP does not produce tumors in mice and it is nongenotoxic. The dose response of TBP effects on urine and urothelium was evaluated in male Sprague-Dawley rats at 0, 200, 700, and 3000 ppm of the diet, 10 rats per group, for 10 weeks. Another group received 3000 ppm TBP plus 12,300 ppm NH4Cl to evaluate the effect of urinary acidification. An additional group of 10 rats received 12,300 ppm NH4Cl. A high-dose recovery group (10 weeks 3000 ppm TBP, then 10 weeks control diet) was included to evaluate reversibility. Urine chemistries for control and TBP-treated animals were similar except for a slight decrease in osmolality and creatinine at the highest dose. Scanning electron microscopic examination of the urine of TBP-treated rats showed no increased or abnormal crystalluria, urinary precipitate, or calculi. The urothelial effects were seen at the two highest doses, but were most severe at 3000 ppm TBP, with ulceration and hemorrhage into the bladder lumen and consequent diffuse papillary and nodular hyperplasia. Dietary NH4Cl acidified the urine but did not prevent the urothelial toxicity and regeneration. The bladder epithelial changes were reversible, but the ulcer repair process was accompanied by submucosal fibrosis. TBP at high doses appears to produce urothelial cytotoxicity with marked regenerative hyperplasia which is reversible upon withdrawal of treatment. The cytotoxicity is likely due to the direct effect of TBP or its metabolites rather than an indirect consequence of urinary changes.

A comparison of the effects of sodium saccharin in NBR rats and in intact and castrated male F344 rats.

High doses of sodium saccharin (NaSac) increase proliferation in the bladder of the rat, with a male preponderance. The possibility that alpha 2u-globulin is involved in its mechanism of action was evaluated by feeding it at 7.5% of the diet to NCI-Black-Reiter (NBR) male rats, which do not synthesize liver-derived alpha 2u-globulin. NaSac affected urinary parameters similarly in F344 and NBR male rats, but NBR rats consumed more water leading to greater urinary volume. NaSac produced less proliferation in NBR than in intact F344 rats, with intermediate changes in castrated F344 males, which had intermediate urinary alpha 2u-globulin levels.

Effect of aspirin on N-[4-(5-nitro-2-furyl)-2-thiazolyl]-formamide-induced epithelial proliferation in the urinary bladder and forestomach of the rat

The co-administration of aspirin with N-[4-(5-nitro-2-furyl)-2-thiazolyl]-formamide (FANFT) to rats resulted in a reduced incidence of FANFT-induced bladder carcinomas but a concomitant induction of forestomach tumors. An autoradiographic study was performed on male F-344 rats fed diet containing FANFT at a level of 0.2% and/or aspirin at a level of 0.5% to evaluate the effect of aspirin on the increased cell proliferation induced by FANFT in the forestomach and bladder. FANFT-induced cell proliferation in the bladder was significantly suppressed by aspirin co-administration after 4 weeks but not after 12 weeks. In the forestomach, and also in the liver, aspirin did not affect the FANFT-induced increase in labeling index. The present results are consistent with the carcinogenicity experiment suggesting that different mechanisms are involved in FANFT carcinogenesis in the bladder and forestomach, and that aspirins effect on FANFT in the forestomach is not due to an irritant effect associated with increased cell proliferation. Also, there appears to be an adaptation by the rats to the chronic ingestion of aspirin.

Effect of Sulfosulfuron on the Urine and Urothelium of Male Rats

Sulfosulfuron, developed as a herbicide, caused increased microcrystalluria and the formation of urinary tract calculi when fed to male and female rats in a chronic 2-year study at doses of 5,000 ppm and 20,000 ppm. Hyperplasia was also seen in urinary bladders at 5,000 ppm and 20,000 ppm, almost exclusively in the presence of observable calculi/microcalculi. Urinary bladder tumors were found in 2 females in the 5000 ppm group, both in the presence of calculi. No increased microcrystalluria, calculi, or tumors were found at doses of 500 ppm and lower. In the current study, 5 groups of male Sprague—Dawley rats were fed sulfosulfuron at doses of 50, 500, 5,000, and 20,000 ppm for 10 weeks. Ten animals were co-administered 5,000 ppm sulfosulfuron with 12,300 ppm NH 4Cl to determine if inhibition of the formation of calculi would prevent any urothelial effects of treatment with sulfosulfuron. Ten animals in the control group and in the high-dose sulfosulfuron group were fed only basal diet for an additional 10 weeks to determine if the effects of sulfosulfuron on the bladder epithelium were reversible. There was an increased incidence of microcrystalluria observed at 5,000 and 20,000 ppm. There was no increase in microcrystalluria observed in the urine of rats co-administered sulfosulfuron and NH4Cl. Urinary bladder calculi were found in the bladder of 1 animal fed 20,000 ppm. Examination by light microscopy showed diffuse papillary/nodular hyperplasi a of the bladder epithelium in this animal. No increased microcrystalluria was observed after withdrawal of the chemical from the diet and the bladder epithelium was normal by light microscopy. The hyperplastic effects associated with the feeding of high doses of sulfosulfuron occur only with the appearance of urinary tract calculi. Based on these results and anatomical differences between rats and humans, it may be concluded that the hyperplastic and carcinogenic effects of sulfosulfuron in rats are high-dose, threshold phenomena that are not likely to occur in humans under environmentally relevant exposures.

Effect of sodium selenite upon bromobenzene toxicity in rats: I. Hepatotoxicity

The effects of sodium selenite on bromobenzene hepatotoxicity were examined in male rats. Rats pretreated with sodium selenite (12.5 or 30 mumol/kg, ip) 72 hr prior to injection of bromobenzene (7.5 mmol/kg, ip) showed a marked reduction in bromobenzene-induced liver injury as evidenced by decreased plasma alanine and aspartate transaminase values, sorbitol dehydrogenase activity, and reduced histologic damage. Administration of bromobenzene did not affect the selenium content of blood or liver. At 72 hr after treatment with selenite, hepatic reduced (GSH) and oxidized (GSSG) glutathione values or GSH synthetic and degradation enzyme activities were not altered. However, from 3 to 12 hr following bromobenzene administration, hepatic GSH and cysteine amounts declined less rapidly in selenite-treated rats compared to control. Thus, acute selenite treatment ameliorated bromobenzene hepatotoxicity in a manner suggesting facilitation of hepatic GSH production by selenite for use in bromobenzene detoxication.

Long term dose response study of N-[4-(5-nitro-2-furyl)-2-thiazolyl] formamide-induced urinary bladder carcinogenesis.

The effect of dose was evaluated for the bladder carcinogenicity of N-[4-(5-nitro-2-furyl)-2-thiazolyl] formamide (FANFT). The chemical was fed to male weanling F344 rats for 30 weeks followed by 74 weeks of control diet. The incidences of bladder carcinoma were 100, 100, 87, 0, 0 and 0% for doses of 0.2, 0.1, 0.05, 0.01, 0.005 and 0.001%, respectively. There was an inverse relationship between dose and latency period. There was no increased incidence of tumors of other tissues.

Evaluation of Cell Proliferative Activity in the Rat Urinary Bladder After Feeding High Doses of Cacodylic Acid

Publisher Summary Cacodylic acid fed at relatively high doses in the diet (100 ppm) produces an increased incidence of bladder tumors in rats, with the effect greater in females than in males. No similar urothelial changes are seen in mice. At similar doses, cacodylic acid also enhances bladder tumor formation following prior administration of N-butyl-N-(4-hydroxybutyl) nitrosamine. The weight of the evidence strongly suggests that cacodylic acid does not produce these changes by direct interactions with DNA. Possible increased cell proliferative effects secondary to the dietary administration of cacodylic acid fed for 10 weeks were evaluated in female F344 rats. Proliferative activity was evaluated by light and scanning electron microscopy and by bromodeoxyuridine labeling index. Hyperplasia and significantly increased labeling index occurred at doses of 40 and 100 ppm but not at 2 or 10 ppm. Significant changes of necrosis and proliferation were detectable by scanning electron microscopy at the doses of 40 and 100 ppm. Urinary changes included increased volume with decreased osmolality and creatinine, but urinary calcium was increased. There was no urinary precipitate, microcrystalluria, or calculi detected related to the administration of cacodylic acid. Increased calcification occurred in the kidneys. These studies show that orally administered high doses of cacodylic acid produce urothelial toxicity and regeneration in female rats, which contribute to the ultimate development of a low incidence of bladder tumors.