Modifying effects of simultaneous treatment with butylated hydroxyanisole (BHA) on rat tumor induction by 3,2'-dimethyl-4-aminobiphenyl, 2,2'-dihydroxy-di-n-propylnitrosamine and N-methylnitrosourea

The modifying effects of concurrent treatment with high or low doses of butylated hydroxyanisole (BHA) on wide-spectrum carcinogen-induced carcinogenesis were studied in male F344 rats. Groups of 20 animals were treated with 2 or 0.04% BHA for 24 weeks. Starting 2 weeks after the commencement of BHA treatment, they were given s.c. injection of 50 mg/kg body weight 3,2'-dimethyl-4-aminobiphenyl (DMAB) once a week, i.g. administrations of 200 mg/kg body weight 2,2'-dihydroxy-di-n-propylnitrosamine (DHPN) once every 2 weeks, or i.p. injection of 15 mg/kg body weight N-methylnitrosourea (MNU) once every 2 weeks for 22 weeks. Further groups of rats were treated with DMAB, DHPN, MNU, or 2 or 0.04% BHA alone. All surviving animals were killed 24 weeks after the beginning of the experiment and the target organs examined histopathologically. The BHA treatment dose-dependently decreased the incidence of DMAB-induced liver preneoplastic lesions but was associated with significant tumor induction in the forestomach (papillomas, 40%, P less than 0.01) and urinary bladder (papillomas, 53%, P less than 0.001; carcinomas, 80%, P less than 0.001), where no lesions were observed in the group given only DMAB. Concurrent administration of 2% BHA also significantly inhibited the development of alveolar hyperplasia (P less than 0.001) of the lung in DHPN-treated animals, while enhancing induction of forestomach papillomas (P less than 0.05) and simple hyperplasia in the urinary bladder. Neither MNU nor 2% BHA alone induced forestomach carcinoma or papillary or nodular hyperplasia (PN hyperplasia) in the urinary bladder. However, these lesions were observed in 100% (P less than 0.001) and 55% (P less than 0.001) of animals respectively, receiving the two compounds in combination. These results demonstrated that concurrent treatment with BHA not only inhibits but can also strongly enhance carcinogenesis depending on the organ, irrespective of whether the carcinogens act directly or require metabolism. The finding that BHA potently modified carcinogenesis at 0.04% in diet, 1/50 of the carcinogenic dose, suggests that actual dietary levels close to the human situation might play a significant role in tumor development in man.     

Selective in vivo inhibition of rat mammary 7,12-dimethylbenz[a]anthracene-DNA adduct formation by dietary butylated hydroxytoluene

The in vivo formation of specific 7,12-dimethylbenz[a]-anthracene (DMBA)-DNA adducts in the mammary gland of the female Sprague-Dawley rat was studied in response to dietary butylated hydroxytoluene (BHT). Dietary BHT concentrations of 0.4 and 0.8% significantly inhibited total DMBA-DNA binding by 41.5 and 35.6% respectively, as compared to controls. However, the decrease in total binding associated with intake of BHT was not due to a uniform inhibition in the formation of all individual adducts. The formation of two adducts resulting from the binding of the anti-dihydrodiolepoxide of DMBA to deoxyguanosine (anti-dGuo) was significantly decreased by a combined average of 51.5% for rats fed BHT-supplemented diets as compared to controls. However, syn-derived DMBA-DNA adducts were not consistently inhibited by dietary BHT. Adduct formation resulting from the binding of the syn-dihydrodiolepoxide of DMBA to deoxyadenosine (syn-dAdo) was significantly inhibited only for rats fed a diet supplemented with 0.4% BHT. The formation of the syn-dGuo adduct was not affected by the feeding of BHT-supplemented diets. These results suggest that in vivo inhibition by BHT of mammary DNA adducts formed from the anti-diastereomer of DMBA may be an important contribution to the inhibitory effect of BHT on the initiation stage of DMBA-induced mammary carcinogenesis.     

Dose-dependent effects of butylated hydroxyanisole, butylated hydroxytoluene and ethoxyquin for promotion of bladder carcinogenesis in N-butyl-N-(4-hydroxybutyl)nitrosamine-initiated, unilaterally ureter-ligated rats

Dose-dependent effects of 3 antioxidants, butylated hydroxyanisole (BHA, 2.0, 1.0 and 0.5%), butylated hydroxytoluene (BHT, 1, 0.5 and 0.25%) and ethoxyquin (0.5, 0.25 and 0.125%) on the development of preneoplastic lesions in the bladder of N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN)-treated rats were investigated. Feeding of the antioxidants after pretreatment of 0.05% BBN commenced and unilateral ureteric ligation was combined at week 3 of the experiment. Surviving rats were killed at the end of week 24. BHA and BHT, but not ethoxyquin increased dose-dependently the incidence and number of preneoplastic lesions, papillary or nodular hyperplasia of the urinary bladder in rats treated with BBN. Particularly, the incidence and number of PN hyperplasia in rats treated with 2.0% BHA and 1.0% BHT were significantly higher than those of the control group. Thus, promoting activities of BHA and BHT, but not ethoxyquin for the urinary bladder were confirmed in this system of BBN-initiated, unilaterally ureter-ligated rats.     

Organ dependent enhancement of rat 3,2'-dimethyl-4-aminobiphenyl (DMAB) carcinogenesis by 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP): positive effects on the intestine but not the prostate

In order to evaluate tumor enhancing effects of the heterocyclic carcinogen, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), doses of 100 and 300 p.p.m. PhIP were given for 40 weeks to male F344 rats, which initially received 3,2'-dimethyl-4-aminobiphenyl (DMAB). DMAB shows a similar carcinogenic organ spectrum to that of PhIP, including the prostate and colon. PhIP alone at a dose of 300 p.p.m. resulted in the development of prostate and intestine cancers. Furthermore, among the DMAB-treated group, enhancement of intestinal carcinogenesis by 300 p.p.m. PhIP was observed. However, no prostate enhancement was demonstrated in the DMAB + PhIP group. Since PhIP-DNA adduct formation in the prostate epithelial cells in a satellite experiment was not affected by pre-treatment with DMAB, it is speculated that the contradictory findings between the intestine and prostate may be due to the specific biological effects of PhIP. Taking into account previous data, that PhIP clearly enhanced rat 1,2-dimethylhydrazine-initiated colon tumorigenesis, the potential of PhIP to enhance colon carcinogenesis may be initiator dependent.     

Increased expression of cyclooxygenase-2 protein in rat urinary bladder tumors induced by N-butyl-N-(4-hydroxybutyl) nitrosamine

The anti-inflammatory drugs, aspirin and piroxicam, are known to possess chemopreventive potential against rat superficial urinary bladder carcinogenesis induced by N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN). Recently, we found similar inhibitory effects with a selective cyclooxygenase (COX)-2 inhibitor, nimesulide. In order to clarify the inhibitory mechanisms, we have further studied the expression of COX-2 protein in urinary bladder tumors induced by BBN in Fischer 344 male rats. For comparison, papillomatosis caused by uracil-induced urolithiasis, and normal epithelial cells, were also investigated. Western blot analysis revealed COX-2 protein to be barely expressed in the normal epithelial cells, whereas it was increased 13-22-fold in varying sizes of urinary bladder tumors and 7-fold in papillomatosis. Immunohistochemically, COX-2 protein was diffusely expressed in transitional cell carcinomas and nodulo-papillary hyperplasia but weakly expressed only in basal cells in simple hyperplasia and normal-looking surrounding epithelia. In papillomatosis, it was moderately expressed only in endothelial cells in stroma. These results indicate that COX-2 plays important roles in the development of preneoplastic and neoplastic lesions in the rat urinary bladder, and therefore could be a good target for chemoprevention of superficial lesions.     

Inhibition of N-butyl-N-(4-hydroxybutyl)nitrosamine-induced rat urinary bladder carcinogenesis by alpha-difluoromethylornithine

The effect of oral administration of alpha-difluoromethylornithine (DFMO), an irreversible ornithine decarboxylase inhibitor, on N-butyl-N-(4-hydroxybutyl)nitrosamine (BHBN)-induced rat urinary bladder carcinogenesis was investigated. Four-wk-old male Fischer 344 rats, 30-38 per group, were divided into 3 groups; each group was divided into 3 subgroups. In Group A, 6-wk treatment with 0.05% BHBN in drinking water was followed by either 0.5% (A1), 0.2% (A2), or 0% (A3) DFMO in drinking water for 34 wk. In Group B, coadministration in drinking water of 0.01% BHBN and either 0.5% (B1), 0.2% (B2), or 0% (B3) DFMO was continued for 30 wk. Group C consisted of animals receiving 0.5%, 0.2%, or 0% DFMO in drinking water for 34 wk without prior or cocarcinogen treatment. Bladder tumorigenesis was clearly inhibited by DFMO; tumor incidence was 14 of 37 (38%) in A1, 16 of 38 (42%) in A2, and 31 of 35 (89%) in A3, and 7 of 35 (20%) in B1, 14 of 35 (40%) in B2, and 28 of 35 (80%) in B3 (P less than 0.01, DFMO groups as compared to the respective control A3 or B3). The average tumor volume was strikingly reduced in Group A rats given DFMO (3.0 mm3 in A1, 5.0 in A2, and 38.6 in A3). Significant suppression of tumor multiplicity (number of tumors/tumor-bearing bladder) was observed in DFMO-treated subgroups in Group B (1.1 in B1, 1.3 in B2, and 1.8 in B3). In both Groups A and B, however, DFMO failed to suppress hyperplastic changes (simple hyperplasia) or preneoplastic lesions (nodulopapillary hyperplasia). Systematic examination of all pertinent organs excluding the brain showed no adverse effects attributable to DFMO treatment except for decrease in body weight (less than 7%), which was consistently observed in the groups receiving 0.5% DFMO, and reduction in the combined weight of the prostate and seminal vesicles (less than 20%), which was noted in Group B in which exposure to DFMO was started at a younger age. These results indicate that oral administration of DFMO is quite effective in suppressing (or retarding) BHBN-induced carcinogenesis with minimal untoward effects and confirm the similar inhibitory effects demonstrated earlier with the heterotopically transplanted rat urinary bladder system.     

Loss of heterozygosity in (LewisxF344)F1 rat urinary bladder tumors induced with N-butyl-N-(4-hydroxybutyl)nitrosamine followed by dimethylarsinic acid or sodium L-ascorbate.

Dimethylarsinic acid (DMA), a main metabolite of arsenicals which are carcinogenic in man, exerts tumor-promoting activity on rat urinary bladder carcinogenesis initiated with N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN). Sodium L-ascorbate (Na-AsA) is also a strong tumor promoter in this animal model. In this study, we used (LewisxF344)F, rats to compare molecular alterations in urinary bladder tumors caused by BBN followed by DMA or Na-AsA. Male, 6-week-old rats were given 0.05% BBN in their drinking water for 4 weeks, and then the rats in group 1 were maintained with no further treatment for 40 weeks. The animals of groups 2 and 3 were administered 0.01% DMA in their drinking water (group 2) or 5% Na-AsA in the powder diet (group 3) after the BBN treatment. Group 4 rats were given 0.05% BBN continuously for 36 weeks. At weeks 12, 20, 36 and 44, subgroups of rats were killed. Histopathological examination revealed promoting activity for DMA and, to a greater extent, Na-AsA on urinary bladder carcinogenesis. Loss of heterozygosity (LOH), detected with the polymerase chain reaction using 36 microsatellite markers, was found to be present in 2 of 9 (22%) urinary bladder tumors after treatment with DMA and 3 of 22 (14%) induced by continuous administration with BBN. No LOH was, however, detected in urinary bladder tumors after treatment with Na-AsA. The results thus suggest that the mechanisms of action of these two promoters, DMA and Na-AsA, may differ in rat urinary bladder carcinogenesis.     

Effect of butylated hydroxyanisole on the metabolism of N-nitrosodi-n-butylamine and N-nitrosobutyl(4-hydroxybutyl)amine by rat hepatic S9 preparations in vitro

N-Nitrosodi-n-butylamine (NDBA), the NADPH generating system and various concentrations of butylated hydroxyanisole (BHA) added to rat hepatic S9 fractions resulted in a significant drop (30-50%) in N-nitrosobutyl(4-hydroxybutyl)amine (NBHBA) formation and a consequent rise in the amount of substrate recovered unchanged. When NBHBA and NAD+ were incubated with BHA and S9 fractions, the amount of N-nitrosobutyl(3-carboxypropyl)amine (NBCPA) was decreased by 20-40%, and the amount of unmetabolized NBHBA increased.     

Absence of ras oncogene activation in rat urinary bladder carcinomas induced by N-methyl-N-nitrosourea or N-butyl-N-(4-hydroxybutyl)nitrosamine.

Previously, we demonstrated point mutations of the H-ras gene in N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide (FANFT)-induced rat urinary bladder carcinomas. In this study, ras oncogene activation was examined in urinary bladder carcinomas induced by N-(4-hydroxybutyl)nitrosamine (BBN) or N-methyl-N-nitrosourea (MNU) administration followed by uracil treatment. In the first experiment, MNU (20 mg/kg body wt) was i.p. injected into 11 male F344 rats twice a week for 4 weeks, followed by feeding 3% uracil for 20 weeks (MNU/uracil group). Ten rats were given only 3% uracil without MNU pretreatment. In the second experiment, 20 male F344 rats were given 0.05% BBN in the drinking water for 4 weeks, then fed 3% uracil for 20 weeks (BBN/uracil group). Another 20 rats were fed 3% uracil without the BBN pretreatment. Transitional cell carcinomas were induced in the urinary bladder of all rats in the MNU/uracil and BBN/uracil groups. Papillomas and hyperplasias were present in the rats given uracil without prior BBN or MNU. DNA and protein were extracted from the tumors (MNU/uracil or BBN/uracil groups) or from the scraped bladder epithelium (uracil alone groups). Sequences around codons 12, 13 and 61 of H-, K- and N-ras genes were examined by direct sequencing after polymerase chain reaction, and p21 was examined by Western blotting. No mutation was found within the examined sequences and p21 showed no changes in mobility. There was no difference in the level of p21 expression between rats treated with MNU/uracil or BBN/uracil compared to corresponding uracil alone groups. These results indicate that the ras oncogene was not activated in urinary bladder carcinomas induced by BBN or MNU in combination with uracil treatment, in contrast to previous findings with FANFT.     

DNA methylation adduct formation and H-ras gene mutations in progression of N-butyl-N-(4-hydroxybutyl)nitrosamine-induced bladder tumors caused by a single exposure to N-methyl-N-nitrosourea

After receiving 500 p.p.m. N-butyl-N-(4-hydroxybutyl) nitrosamine(BBN) in their drinking water for an initial 10 weeks, ratswere given a single i.p. injection of N-methyl-N-nitrosourea(MNU) at a dose of 50 mg/kg body wt at week 20 (at a stage whenbladder tumor development had already occurred), and then maintaineduntil they were killed at week 40. Three and six hours afterthe MNU injection, the DNA methylation adducts, O⁶-methyldeoxyguanine(O⁶-medG) and 7-methyldeoxyguanine (7-medG), were immunohistochemicallyrevealed to be markedly more frequent in urothelial preneoplasiasor neoplasias than in normal cells. These adducts were rapidlyrepaired, and although 7-dmeG in tumor cells still persistedafter 72 h, they appeared essentially to have returned to normallevels. At the termination, conversion of transitional cellcarcinomas (TCC) to squamous cell carcinomas (SCC) of the urinarybladder was significantly increased in the BBN + MNU group.The extent of invasion was also significantly greater with theadditional MNU treatment Expression of p21 protein, detectedby immunohistochemistry, was comparable between the groups.Mutations in the H-ras gene were observed in one case each ofthe BBN and BBN + MNU groups, and both cases showed a G:C toA:T transition at codon 12. The present study thus suggestedthat while an additional single treatment with MNU of rats bearingBBN-induced bladder neoplasias is associated with significant,possibly mutation-dependent tumor progression, H-ras mutationsare not necessary events.     

Combined effects of L-ascorbic acid, citric acid or their sodium salts on tumor induction by N-butyl-N-(4-hydroxybutyl)nitrosamine or N-ethyl-N-(4-hydroxybutyl)nitrosamine in the rat urinary bladder

L-Ascorbic acid, citric acid or their sodium salts (at levels equivalent to 5% sodium L-ascorbate) were fed in the diet simultaneously with N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) or N-ethyl-N-(4-hydroxybutyl)nitrosamine (EHBN) (0.025% BBN or 0.021% EHBN) in the drinking water to male F344 rats for 20 weeks to determine whether urinary pH changes affect the carcinogenicity of BBN or EHBN. In the urine, pH was decreased in rats fed the acidic chemicals and increased in rats fed their corresponding sodium salts. Histopathologically, the incidences and numbers of preneoplastic and neoplastic lesions in groups treated with each test chemical were not different from those in control groups except for sodium citrate-treated groups in which induction of carcinomas was higher, resulting from increased intake of either carcinogen and also from increased urinary excretion of main carcinogenic metabolites. These results show that the test chemicals do not affect the carcinogenicity of BBN or EHBN on the rat urinary bladder when simultaneously administered despite significant differences in urinary pH.     

Effect of UFT on urinary bladder tumors in rats induced by N-butyl-N-(4-hydroxybutyl)nitrosamine

The effect of combined tegafur and uracil (UFT) on the development of rat urinary bladder tumors induced by N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) was studied. Two hundred F344 male rats were divided into 10 groups. Groups 1 to 5 were given 0.05% BBN in drinking water for the initial 8 weeks of the experiment, and Groups 6 to 10 were the controls of the prior 5 groups treated with BBN. UFT in commercial diet was administered at daily doses of 100mg (30.9 mg as FT-207) per kg of body weight in Groups 2 and 4 and 200 mg (61.7 mg as FT-207) per kg of body weight in Groups 3 and 5. Groups 2 and 3 received UFT throughout the period of the experiment, and Groups 4 and 5 for 12 weeks after 8 week treatment with BBN. All animals were sacrificed at 20 weeks, and studied histopathologically. In Groups 1 to 5, urinary bladder tumors developed in 20 of 20, 13 of 20, 6 of 20, 14 of 20 and 6 of 20, respectively. Incidences of tumors in the 4 groups treated with UFT were significantly lower than that in Group 1 treated with BBN alone. This result shows that UFT inhibits the development of urinary bladder tumors in rats induced by BBN.     

Initiating potential of 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide (AF-2), butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT) and 3,3',4',5,7-pentahydroxyflavone (quercetin) in two-stage mouse skin carcinogenesis

Tumor initiating activity of 3,3',4',5,7-pentahydroxyflavone (quercetin), butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT) and 2-(2-furyl)-3-(5-nitro-2-furyl) (AF-2) acrylamide) were tested in a two-stage mouse skin carcinogenesis model using 12-O-tetradecanoyl phorbol-13-acetate (TPA) as a promoter. These compounds dissolved in dimethyl sulfoxide were topically applied twice weekly for 5 weeks on the dorsal skin, and then followed by TPA for 47 weeks. The total initiating dose was 100 mg for each compound. 7,12-Dimethylbenz(a)anthracene (DMBA) at a total dose of 100 micrograms was used as a positive control compound. AF-2 induced skin tumors in 35% of the mice (average of 0.4 tumors/mouse), HBA in 15% in (0.2/mouse), BHT in 13% (0.13/mouse) and quercetin in 5% (0.1/mouse). No tumors appeared in the groups treated with either test chemicals alone or TPA alone. Statistical analysis according to either Fisher's exact test or Peto's trend test revealed significant differences for tumor appearance in the AF-2/TPA and BHA/TPA followed by TPA groups as compared to in the DMSO/TPA group. The results indicate that AF-2 and BHA have weak tumor initiating activity on mouse skin, but such effects are not apparent for BHT or quercetin.     

Aberrant expression of p27(Kip1) is associated with malignant transformation of the rat urinary bladder epithelium

Alteration in cell cycle regulators is considered to play an important role in carcinogenesis. In order to cast light on changes in reversible hyperplastic and irreversible tumorigenic lesions in the rat urinary bladder, expression of p27(Kip1), cyclin D1 and cyclin E proteins was sequentially compared. In the first study, 3% uracil was fed for 4 weeks to cause urinary calculi and consequent hyperplasia and papillomatosis, both regressing after withdrawal of the insult. Compared with normal bladder epithelium, in papillomatosis at week 4, the BrdU index and immunohistochemical positivities for cyclin D1 and cyclin E were significantly elevated, whereas values for p27(Kip1) tended to be reduced. One week after withdrawal of uracil, the BrdU index and positivities for cyclin D1 and cyclin E were decreased to below the control levels, while positivity for p27(Kip1) was dramatically increased, with a strong staining intensity. In a second study, rats were initiated with a bladder carcinogen, N-butyl-N-(4-hydroxybutyl)nitrosamine for 4 weeks, then fed 3% uracil for 8 weeks. During this latter period, expression of cyclin D1, cyclin E and p27(Kip1) in hyperplastic urothelium were comparable with those in the first study. One week after withdrawal of uracil, most urothelial lesions regressed, showing high p27(Kip1) and low cyclin D1 and cyclin E staining. Two weeks after uracil withdrawal, transitional cell carcinomas, with a low p27(Kip1) and high cyclin D1 and cyclin E staining pattern, could be easily distinguished from surrounding regressing epithelium. These data indicate that during regression of papillomatosis after cessation of a proliferative stimulus, expression of p27(Kip1)is elevated, accompanied by a lowering of cyclin D1 and cyclin E. In irreversible tumorous bladder lesions, on the other hand, persistent low expression of p27(Kip1) and elevated cyclin D1 and cyclin E are characteristic.     

Activation of H-ras oncogene in rat bladder tumors induced by N-butyl-N-(4-hydroxybutyl)nitrosamine

Ras-oncogene activation was investigated in the bladder tumors of F344 male rats given N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) in drinking water. DNA from one of the nine transitional cell carcinomas contained an H-ras oncogene detectable by the NIH/3T3 transfection assay. Analysis of p21 ras proteins suggested that the activating mutation resided within codon 61 of the H-ras gene and that such activating mutations were not present in other tumors. In contrast to mutational activation of ras genes, enhanced expression of p21 was observed in all tumors examined by immunohistochemical techniques with the use of Formalin-fixed paraffin-embedded tissue sections and an anti-ras p21 antibody, RAP-5. Further histochemical analysis of bladder tissues at various stages of the BBN-induced carcinogenic process indicated that the enhanced expression of p21 appeared early; the reactivity with RAP-5 was observed in diffuse hyperplastic epithelia after 5 weeks of exposure to BBN. The frequency of ras oncogenes, activated either by point mutations or overexpression of p21, in BBN-induced rat bladder carcinomas has thus been shown to be similar to that observed in human bladder carcinomas.     

Induction of dorsolateral prostate adenocarcinomas and other accessory sex gland lesions in male Wistar rats by a single administration of N-methyl-N-nitrosourea, 7,12-dimethylbenz(a)anthracene, and 3,2'-dimethyl-4-aminobiphenyl after sequential treatment with cyproterone acetate and testosterone propionate

Groups of 20-25 male Wistar rats (Cpb:WU), nine groups of 4-week-old rats, and nine groups of 8-week-old rats, were given cyproterone acetate (CA) s.c. or by gavage daily for 18 days at a dose of 50 mg/kg/day. Directly following CA treatment, the rats received 3 daily s.c. injections with testosterone propionate (TP) at a dose of 100 mg/kg/day. On the day after the last TP administration, a single dose of one of the following carcinogens was given to 3 groups: N-methyl-N-nitrosourea (MNU), 50 mg/kg i.v.; 7,12-dimethylbenz(a)anthracene, 30 mg/kg i.v.; 3,2'-dimethyl-4-aminobiphenyl, 250 mg/kg s.c. Three other groups received the same carcinogen treatments after 7 days of recovery from the CA administration. The last 3 groups received carcinogen without TP treatment, but immediately after CA pretreatment was stopped. A 25% incidence of invasively growing, metastasizing adenocarcinomas was found in the dorsolateral prostate region of 8-week-old rats that had received MNU after treatment with CA plus TP. In addition, this group had a 5% incidence of carcinoma in situ and a 5% incidence of atypical hyperplasia in the dorsolateral prostate. Lower incidences of adenocarcinoma of the dorsolateral prostate region and of carcinoma in situ and atypical hyperplasia of the dorsolateral prostate were found in other groups that were treated with MNU or 7,12-dimethylbenz(a)anthracene after pretreatment with CA, followed by TP or recovery, but never in rats that had been treated with CA only. In the groups treated with 3,2'-dimethyl-4-aminobiphenyl, which is slowly metabolized, these lesions were also found in groups that were pretreated with only CA. The carcinomas seemed to originate from the dorsolateral prostate and their average latency time was approximately 61 weeks. The 8-week-old rat given a MNU injection after sequential treatment with CA and TP may provide a relevant animal model for human prostatic cancer.     

Agglutination of isolated rat bladder cells by lectins after administration of N-butyl-N-(4-hydroxybutyl) nitrosamine

Lectins, concanavalin A (Con A), wheat germ agglutinin (WGA), and Ricinus communis agglutinin (RCA) were used for agglutination of isolated rat bladder cells after administration of N-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN). The results showed that WGA and RCA, as well as Con A, agglutinated rat bladder cells during the early stage of bladder carcinogenesis. The agglutinations by the 3 kinds of lectins were different; WGA and Con A agglutinated only BBN-treated cells but RCA agglutinated both BBN-treated and untreated cells. These findings may be useful in analyzing the membrane alterations during the early phase of bladder carcinogenesis.     

Characterization of adenocarcinomas of the dorsolateral prostate induced in Wistar rats by N-methyl-N-nitrosourea, 7,12-dimethylbenz(a)anthracene, and 3,2'-dimethyl-4-aminobiphenyl, following sequential treatment with cyproterone acetate and testosterone propionate

Carcinomas of the rat prostate induced by a single injection of N-methyl-N-nitrosourea, 7,12-dimethylbenz(a)anthracene, and 3,2'-dimethyl-4-aminobiphenyl, after sequential treatment with cyproterone acetate and testosterone propionate, were evaluated as potential animal models for prostatic cancer. All ten carcinomas examined were located in the dorsolateral prostate region and did not involve the distal parts of the seminal vesicles and coagulating glands. The incidence of urinary obstruction leading to the animals' death was 6 of 10 rats, and metastases in the lung, abdominal lymph nodes, and/or liver also occurred in 6 of 10 rats. The tumors were invasive adenocarcinomas, showing frequent perineural invasion and a variable degree of differentiation. There were ultrastructural similarities with human prostatic carcinomas, such as intracellular lumina. Plasma acid phosphatase was increased. Enzyme histochemical analysis revealed similarities with the Dunning R3327H and -HI prostatic carcinomas but was not helpful in determining the site of origin of the tumors. The gross and microscopic appearance of the tumors and the observation of preneoplastic lesions exclusively located in the dorsolateral prostate suggest this lobe as site of origin of the carcinomas. Preneoplastic lesions (n = 9) included atypical hyperplasias (n = 5) and lesions with all histological characteristics of carcinoma except for local invasion and metastases, which were classified as carcinoma in situ (n = 4). Although androgen sensitivity could not be assessed, the observed characteristics of the tumors [their long latency time (46-80 weeks), the presence of preneoplastic lesions, and the short duration of the treatment, leaving the animals intact] all indicate that the present approach is a valid animal model for the study of prostatic carcinogenesis.