Michiko Shibata

Promoting effects of various chemicals in rat urinary bladder carcinigenesis initiated by N-nitroso-n-butyl-(4-hydroxybutyl)amine

We studied the capacity of various chemicals to promote urinary bladder cancer in male F344 rats after initiation by N-nitroso-n-butyl-(4-hydroxybutyl)amine (BBN). The rats were given initially 0.01% BBN in the drinking-water for 4 wk and then the test compound in the diet for 34 wk. Effects were judged by measuring the formation of preneoplastic lesions papillary or nodular hyperplasia (PN hyperplasia) of the urinary bladder. Administration of 5%, but not 0.5% (w/w) sodium saccharin in the diet significantly increased the incidence and extent of PN hyperplasia. This finding could be related to the induction of cancers in the rat urinary bladder by high levels of saccharin. Sodium ascorbate (5%). DL-tryptophan (5%) and allopurinol (0.02%) also significantly increased the extent of PN hyperplasia in the affected animals, but other test chemicals, such as acetazolamide (0.35%) and quercetin (5%) did not. The results with sodium saccharin and DL-tryptophan were consistent with previous findings and suggest that sodium ascorbate and allopurinol have promoting activities in urinary bladder carcinogenesis in rats. No correlation was found between the extent of crystalluria and promotion of preneoplastic lesions.

Chronic toxicity of butylated hydroxytoluene in Wistar rats

Abstract Groups of 57 Wistar rats of each sex were maintained on diet containing 0.25 or 1% butylated hydroxytoluene (BHT) for 104 wk; control groups comprised 36 rats of each sex. Treated rats of both sexes showed reduced body-weight gain, relative spleen weight and white-blood-cell count while in the males there was also a reduction in serum triglyceride. BHT-treated animals of both sexes showed increased relative liver weight and total blood cholesterol but increases in red-blood-cell count could be seen only in females and only the males showed increased γ-glutamyltransferase. No significant histological changes were observed in the liver or haematopoietic system to explain these haematological and biochemical changes. Tumours were found in the liver, pancreas, mammary glands, uterus, pituitary gland, adrenal glands and in some other organs of some of the treated rats, but their incidence was not significantly different from that in controls. This experiment showed no carcinogenic effect of BHT on rats.

Lack of carcinogenicity of butylated hydroxytoluene on long-term administration to b6c3f1 mice

Groups of approximately 50 male and 50 female B6C3F1 mice were given butylated hydroxytoluene (BHT) at concentrations of 200, 1000 or 5000 ppm in their diet for 96 wk followed by a basal diet for 8 wk and were then killed. Similar groups of male and female controls were given basal diet throughout the 104 wk. Females given 1000 or 5000 ppm BHT and males given 5000 ppm showed reduced weight gain. Neither survival rates nor food consumption differed between experimental and control groups. No significant changes attributable to BHT treatment were found in the haematological examinations or serum and urine analyses. Tumours were found in many organs, especially the lungs, liver, lymph nodes and spleen, in both the experimental and control groups, but none were related to BHT treatment. Thus this experiment provided no evidence of BHT carcinogenicity in mice.

Participation of urinary Na+, K+, pH, and L-ascorbic acid in the proliferative response of the bladder epithelium after the oral administration of various salts and/or ascorbic acid to rats

Changes in urinary parameters (particularly electrolyte levels and pH), and DNA synthesis and the morphology of the bladder epithelium were investigated in rats that were fed for 4 or 8 wk on diets containing various Na, K, Mg or Ca carbonate salts, with or without L-ascorbic acid (AsA). [The carbonate salts were fed at a level of 3% in the diet, and AsA or AsA-Na was administered at 5% in the diet. NH4Cl was at 1% in the diet.] The effects of treatment with NH4Cl (used as a urine acidifier), and of combined treatment with sodium ascorbate (AsA-Na) and NH4Cl were also investigated. Urinary pH was significantly elevated in groups given NaHCO3, K2CO3, AsA + NaHCO3, AsA + K2CO3 and AsA-Na, whereas treatment with AsA or NH4Cl alone caused a significant drop in urinary pH. An increase in urinary electrolytes or ascorbic acid was associated with the corresponding dosing regimen. DNA synthesis in the bladder epithelium was increased in groups given NaHCO3, K2CO3, AsA + NaHCO3, AsA + K2CO3 or AsA-Na. Furthermore, all treatments that induced an elevation of DNA synthesis also induced some morphological alterations in the bladder epithelium. The administration of AsA in conjunction with NaHCO3 or K2CO3 induced levels of change greater than those with either salt alone. In contrast, the degree of response in the bladder epithelium of rats given AsA-Na was reduced by the simultaneous administration of NH4Cl. These results suggest that the degree of DNA synthesis and/or morphological alteration in the rat-bladder epithelium after treatment with various bases may depend on changes in urinary concentrations of Na+ or K+ ions and/or pH, and the presence of ascorbic acid in the urine. The results are discussed in relation to the possible promotion by various treatment regimens (salts +/- AsA) of urinary bladder carcinogenesis.

Intrahepatic bile duct proliferation induced by 4,4′-Diaminodiphenylmethane in rats

Prolonged intoxication with 1000 ppm of 4,4′-diaminodiphenylmethane (DDPM) led to hyperplasia of bile ducts and proliferation of oval cells. Hyperplastic areas gradually regressed and were replaced by fibrous tissue after cessation of DDPM administration. Elevation of γ -glutamic transpeptidase and alkaline phosphatase in serum accompanied the proliferation of bile ductular cells.

Promoting effect of sodium chloride in 2-stage urinary bladder carcinogenesis in rats initiated by N-Butyl-N-(4-hydroxybutyl)-nitrosamine

SummaryThe promoting effect of sodium chloride (NaCl) in 2-stage urinary bladder carcinogenesis in F344 rats initiated by 2 doses of N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) was investigated. The incidences of PN hyperplasia were significantly higher in rats initiated with 0.01 or 0.05% BBN when they were given diet containing 10% NaCl for 32 weeks than when they were given control diet. The incidence of papilloma in rats given 0.05% BBN followed by diet containing 10% or 5% NaCl tended to be higher than that in control rats. The urine of rats given diet containing NaCl was larger in volume and had lower osmolality than that of controls. The total urinary sodium and chloride contents were also increased, whereas those of potassium and phosphorus were decreased. No calculus formation or crystalluria was observed. These data suggest that excess intake of sodium as NaCl has a weak promoting effect in 2-stage urinary bladder carcinogenesis.

Long-term toxicity and carcinogenicity study of sodium o-phenylphenate in B6C3F1 mice.

Sodium o-phenylphenate (OPP-Na) was given at dietary levels of O (control), 0.5, 1.0 and 2.0% to groups of 50 male and 50 female mice for 96 wk, and all the animals were maintained without OPP-Na for a further 8 wk. Both sexes given 2% OPP-Na and females given 0.5% and 1% OPP-Na showed growth retardation. Serum alkaline phosphatase activity in OPP-Na treated females was significantly increased in a dose-related manner. There were no treatment-related effects on clinical signs, mortality, urinalyses, haematology or organ weights. The incidences of several non-neoplastic and neoplastic lesions achieved statistical significance but none was considered to be related to treatment. There were increased incidences of haemangiosarcomas of the liver in males given 1% and of hepatocellular carcinomas in 1 and 2% males, and haemangiomas and leiomyomas of the uterus, present in the controls, were absent or decreased in all treated females. Therefore, this study did not demonstrate any clear carcinogenic effect of OPP-Na on mice at dietary levels of up to 2%.

Long-term toxicity and carcinogenicity test of ammonia-process caramel colouring given to B6C3F1 mice in the drinking-water.

Caramel colouring (ammonia process) was given at levels of 0 (control), 1.25 and 5.0% in the drinking-water to groups of 50 male and 50 female mice for 96 wk, and then all all the animals were maintained without caramel for a further 8 wk. Males given 5.0% caramel showed increased cumulative mortality from wk 100 to the end of the experiment. The white blood cell count in treated males was significantly elevated in a dose-related manner. However, these changes were not considered to be biologically significant. There were no treatment-related effects on clinical signs, body or organ weights, results of urine analyses, or histological features. Therefore, this study did not demonstrate any carcinogenic effect of caramel on mice at levels of up to 5.0% in the drinking-water.

Lack of carcinogenicity of magnesium chloride in a long-term feeding study in B6C3F1 mice

Groups of 50 male and 50 female B6C3F1 mice were given magnesium chloride (MgCl2.6H2O) at dose levels of 0 (control), 0.5 and 2% in the diet for 96 wk, after which all animals received the control diet for 8 wk and were then necropsied. In females of the high-dose group a decrease in body weight was observed. However, survival rates did not differ between the treatment and control groups for males or females. Furthermore, clinical signs and urinary, haematological or serum clinical chemistry parameters showed no treatment-related effects. On histological examination, tumours were mainly found in the skin/subcutis, liver and lymphatic system. However, with the exception of a significant decrease in the incidence of liver tumours among males of the high-dose group, no differences were noted in the tumour incidence between the treated and control animals. Thus, the study described here clearly shows a lack of carcinogenicity of MgCl2.6H2O given to B6C3F1 mice in the diet.

Lack of carcinogenicity of monosodium l-glutamate in Fischer 344 rats

Monosodium L-glutamate (MSG) was administered to groups of 50 male and 50 female Fischer 344 rats in the diet at levels of 0, 0.6, 1.25, 2.5 and 5.0% for a 2-yr period. Survival of dosed and control rats of both sexes was comparable, although animals receiving 5.0% MSG showed a trend towards (or significant) growth retardation. Urinary biochemistry values indicated increased pH and Na+ levels and decreased K+ in rats of both sexes given 2.5 or 5.0%. However, no significant increases in proliferative or neoplastic lesion development in the urinary tract were observed in any MSG-treated animals. In addition, occurrences of neoplastic lesions in other organs did not differ between treated and control groups. The present study thus did not demonstrate any long-term carcinogenic effect of MSG administered to rats at levels up to 5.0% in the diet.