Specific inhibitory effect of dietary eicosapentaenoic acid on N-nitroso-N-methylurea-induced mammary carcinogenesis in female Sprague-Dawley rats

We have investigated the effects of two qualitatively different types of unsaturated fatty acids on N-nitroso-N-methylurea (NMU)-induced mammary carcinogenesis in female Sprague-Dawley rats. Semipurified diets containing 4.7% eicosapentaenoic acid (EPA) plus 0.3% linoleic acid or 5% linoleic acid were prepared. Animals maintained on these diets were given an i.v. injection of NMU (50 mg/kg body wt) at 50 days of age and killed 20 weeks later. Both tumor incidence and tumor number per rat were significantly lower in the EPA diet group (60.0% and 2.3 +/- 2.5 versus 93.3% and 5.1 +/- 4.5 respectively) for the 5% linoleic acid diet. Furthermore, the average weight of tumor material (total) per rat was significantly lower in the EPA as compared to linoleic acid diet group (2.9 +/- 4.2 g and 11.4 +/- 12.2 g respectively). Analysis of phospholipid fatty acids in the mammary tumors in the EPA diet group showed a higher proportion of C16:0, C18:2, omega-3 fatty acids C20:5 and C22:6 and a lower proportion of C20:4. Furthermore, mammary tumors in rats fed the EPA diet demonstrated significant reduction in prostaglandins. The results thus suggest that inhibition by EPA of NMU-induced mammary carcinogenesis may be mediated via the modulation of lipid metabolism and associated reduction in prostaglandin synthesis.     

Low zinc intake suppressed N-methyl-N-nitrosourea-induced mammary tumorigenesis in Sprague-Dawley rats

Zinc has been shown to be accumulated in N-methyl-N-nitrosourea (MNU)-induced rat mammary tumors. Zinc is required for cell proliferation and tumorigenesis is characterized by dysregulation of cell proliferation. An accumulation of zinc in mammary tumors perhaps indicates a reliance on zinc to sustain tumor growth. Limiting zinc supply by means such as reduced zinc intake should negatively modulate mammary tumorigenesis. Our objective was to determine the effects of zinc status on MNU-induced mammary tumorigenesis in sexually mature female rats. Twenty-one-day-old Sprague-Dawley rats were assigned to low-zinc (3 mg zinc/kg diet) or adequate-zinc (12 mg zinc/kg diet) ad libitum or pair-fed control group (n = 25-33 rats/group). On day 50 of age, all rats were intraperitoneally injected with MNU (50 mg/kg body wt) to induce mammary tumorigenesis. Rats were further maintained on their assigned diet until 14 weeks post-MNU injection. Total food intake and overall body weight gain were lower in low-zinc rats than in adequate-zinc ad libitum control rats, but were similar to adequate-zinc pair-fed control rats. Plasma zinc concentration was lower in low-zinc rats than in adequate-zinc ad libitum and pair-fed control rats, confirming moderately low-zinc status in low-zinc rats. Tumor incidence (46 versus 84 and 80%; P < 0.05) and tumor multiplicity (0.8 versus 5.0 and 2.6 tumors/rat; P < 0.05) and tumor number (28 versus 123 and 66 tumors) were reduced in low-zinc rats compared with that in adequate-zinc ad libitum and pair-fed control rats, respectively. Tumor latency in low-zinc and adequate-zinc pair-fed control rats was not significantly different, but was longer than in adequate-zinc ad libitum control rats (P < 0.05), suggesting that reduced food intake associated with low-zinc intake prolonged tumor latency. Tumor burden and size were not affected by zinc intake. Overall, our observations showed that moderately low-zinc status suppressed MNU-induced rat mammary tumorigenesis.     

Growth arrest of DMBA-induced mammary carcinogenesis with ibuprofen treatment in female Sprague-Dawley rats

We examined effects of ibuprofen on the growth and development of DMBA-induced mammary cancers in mature female Sprague-Dawley rats. Ibuprofen was added to the standard diet at approximately 1,000 mg/kg rodent chow, resulting in an average daily dose of 25 mg per day per 0.25 kg rat. After five weeks of ibuprofen treatment, there was a 37% reduction in tumor volume compared to a 260% increase in the volume of tumors in untreated rats (p<0.001). These results suggest that ibuprofen may have potential in the chemoprevention and treatment of breast cancer.     

Dietary folate deficiency suppresses N-methyl-N-nitrosourea-induced mammary tumorigenesis in rats

Epidemiologic studies have suggested that dietary folate intake is inversely related to breast cancer risk. However, epidemiologic evidence has not been consistent nor has it provided unequivocal support for this purported inverse relationship. This study investigated the effect of dietary folate on N-methyl-N-nitrosourea (MNU)-induced mammary tumorigenesis in rats. Weanling, female Sprague-Dawley rats were fed diets containing either 0 (deficient; n = 22), 2 (basal dietary requirement, control; n = 20) or 8 mg (supplemented; n = 20) folate/kg diet for 30 weeks. At 50 days of age, rats received an i.p. injection of MNU (50 mg/kg body wt). At necropsy, all macroscopic mammary tumors were identified and examined microscopically. The effect of dietary folate on genomic DNA methylation in mammary tumorigenesis was determined by the in vitro methyl acceptance assay. The incidence of mammary adenoma and adenocarcinoma in the folate-deficient group was lower than that of the control and folate-supplemented groups (55 versus 90 and 75%, respectively, P = 0.043). Kaplan-Meier analyses also demonstrated a similar trend in the rates of appearance of either adenoma or adenocarcinoma (P = 0.06). In contrast, folate supplementation did not significantly modulate mammary tumorigenesis compared with the control group. Although mammary tumors were significantly hypomethylated compared with non-neoplastic mammary tissues in each dietary group (P < 0.03), folate status did not significantly affect the extent of DNA methylation. The data suggest that dietary folate deficiency of a moderate degree suppresses, whereas folate supplementation at four times the basal dietary requirement does not significantly modulate, mammary tumorigenesis in this model. The role of folate in mammary tumorigenesis needs to be clarified for safe and effective prevention of breast cancer.     

Protective effects of pregnancy and lactation against N-methyl-N-nitrosourea-induced mammary carcinomas in female Lewis rats

The role of parity before and after N-methyl-N-nitrosourea (MNU) treatment in protection against mammary carcinogenesis was investigated. The effect of lactation on reduction in the incidence of mammary carcinoma was also examined. Parous rats were compared with respective age-matched virgins (AMVs). Pregnancy and lactation prior to MNU exposure significantly reduced both the incidence of mammary carcinoma (22 versus 72%) and the average number of mammary carcinomas per rat (0.22 versus 0.86) and significantly prolonged the latency of the carcinomas (247 versus 215 days). Pregnancy and lactation following MNU exposure also significantly reduced both the incidence of mammary carcinoma (25 versus 94%) and the average number of mammary carcinomas per rat (0.25 versus 1.50) and significantly prolonged the latency (240 versus 155 days). Lactation showed an additive effect on the reduction in mammary cancer. Pregnancy suppressed the number of estrogen receptor (ER)- and progesterone receptor (PgR)-positive cells and lowered the cell proliferation rate in the non-tumoral mammary glands. Since the majority (>76%) of the mammary carcinomas was hormone dependent in both the parous and AMV rats, pregnancy and lactation appear to decrease the ER- and/or PgR-positive cells presumed to be the progenitors of hormone-dependent carcinomas and they lowered the cell turnover necessary for tumor promotion in parous rats, resulting in a lower mammary carcinoma yield.     

Site-dependent modulating effects of conjugated fatty acids from safflower oil in a rat two-stage carcinogenesis model in female Sprague-Dawley rats

Modifying effects of dietary administration of conjugated fatty acids from safflower oil (CFA-S), rich in conjugated linoleic acid, on major organs were examined in the post-initiation stage of a two-stage carcinogenesis model in female rats. Groups of 21 or 22 F344 female rats were treated sequentially with 2,2'-dihydroxy-di-n-propylnitosamine (intragastrically, i.g.), 7,12-dimethylbenz[a]anthracene (i.g.), 1,2-dimethylhydrazine (subcutaneously) and N-butyl-N-(4-hydroxybutyl)nitrosamine (in drinking water) during the first 3 weeks for initiation, and then administered diet containing 1 or 0.1% CFA-S for 33 weeks. Further groups of animals were treated with carcinogens or 1% CFA-S alone, or maintained as non-treated controls. All surviving animals were killed at week 36, and major organs were examined histopathologically for development of pre-neoplastic and neoplastic lesions. The 1 and 0.1% CFA-S treatment significantly decreased the incidence and multiplicity of mammary carcinomas, though a clear dose response was not observed. In the urinary bladder, the incidence of papillary or nodular hyperplasia but not tumors was significantly increased in the 1% CFA-S-treated group. The results indicate that low dose CFA-S may find application as a potent chemopreventor of mammary carcinogenesis.     

Promotion of dimethylbenz[a]anthracene-initiated mammary carcinogenesis by iron in female Sprague-Dawley rats

Iron body-stores and iron dietary intake have been sporadically reported to increase the risk of cancer in humans. To investigate the effect of iron on the development of mammary tumors, female Sprague- Dawley rats were given dimethylbenz[a]anthracene (DMBA) (5 mg/kg, i.g., 1x) at 55 days of age. Eight days later, rats received iron(II) sulfate s.c. (50 micromol/kg, 2x/week) for 53 weeks. Mammary tumors started to appear 6-8 weeks after DMBA initiation. At 20 weeks after DMBA treatment, iron(II) increased mammary tumor frequency twofold (11/30 versus 5/30 with DMBA alone). Tumor frequency increased with time and was significantly higher in iron-promoted rats after 40 weeks of treatment (24/30 versus 11/30, P = 0.001). Also, mammary tumors in iron- promoted rats were significantly larger than in DMBA-only rats at 20 weeks after initiation (P = 0.04) and this difference remained significant through the observation time point at 40 weeks. Iron could be detected histochemically in the stromal connective tissue, but not in the epithelial cells of mammary carcinomas. Mammary tumors in the DMBA-only group were mostly adenomas and adenocarcinomas, while those promoted by iron sulfate included fibroadenomas, adenomas and adenocarcinomas. Thus, iron(II) administered s.c. subsequent to DMBA initiation, greatly accelerated mammary carcinogenesis, implying its promoting activity for mammary tissue of female rats.      

Dietary fat and DMBA mammary carcinogenesis in rats

While there is no convincing direct evidence of an influence of specific dietary factors on breast cancer in women, the overall geographic correlation between risk of breast cancer and food consumption patterns suggests a positive link. Epidemiologic studies have correlated breast cancer rates with fat consumption. Increased dietary fat, through intestinal microflora production of estrogens, might expose breast tissue to chronic, excessive stimulation and increase cancer risk. Laboratory animal studies have shown that dietary fat affects response to DMBA carcinogenesis. Studies in our own laboratory have shown that 20% corn oil or lard increased DMBA mammary tumorigenesis, compared to rats fed 5% fat; 20% corn oil accelerated sexual maturation, but 20% lard did not. The mechanisms of tumorigenesis under such circumstances are unknown.     

N-methyl-N-nitrosourea-induced mammary carcinogenesis is promoted by short-term treatment with estrogen and progesterone mimicking pregnancy in aged female Lewis rats

We examined the effects of short-term estrogen and progesterone treatment mimicking pregnancy in aged female Lewis rats on the development of N-methyl-N-nitrosourea (MNU)-induced mammary carcinoma. Rats were administered a single intraperitoneal injection of 20 mg/kg MNU at 7 weeks of age and half of those rats were administered a subcutaneously implanted 21-day release pellet containing 0.5 mg 17beta-estradiol and 32.5 mg progesterone (E/P) at 24 weeks of age. The rats were then monitored for the occurrence of mammary tumors. Rats were sacrificed when the largest mammary tumor became > or = 1 cm in diameter, or when the rat reached 48 weeks of age. Development of MNU-induced mammary carcinomas was accelerated after short-term E/P treatment, compared with E/P-untreated rats: the incidence of > or = 1-cm mammary carcinomas tended to increase (60 vs. 44%); the latency tended to shorten (28.7 vs. 34.6 weeks); and cancer multiplicity (number of all-sized carcinomas per rat) significantly increased (1.8 vs. 0.8). In E/P-treated rats, comedo necrosis was frequently seen and the incidence of estrogen receptor and/or progesterone receptor-negative mammary carcinomas was significantly increased. Early age at full-term pregnancy or short-term hormone treatment mimicking pregnancy may suppress the risk of breast cancer, but the age of hormone exposure is a crucial factor, because hormone exposure mimicking pregnancy in aged individuals may exert effects opposite of those exerted in younger individuals.     

Prevention of N-methyl-N-nitrosourea-induced mammary carcinogenesis in rats by 1alpha-hydroxyvitamin D(5)

BACKGROUND: Although the active form of vitamin D, i.e., 1,25-dihydroxyvitamin D(3), is a potent cell-differentiating agent, its use in cancer prevention or therapy is precluded because it induces excessive blood calcium levels (hypercalcemia). However, less calcemic or noncalcemic synthetic analogues of vitamin D(3) are poorly effective against mammary carcinogenesis. We synthesized an analogue of vitamin D(5), 1alpha-hydroxy-24-ethylcholecalciferol (1alpha-hydroxyvitamin D(5)), which was less calcemic than 1,25-dihydroxyvitamin D(3) and prevented the development of precancerous lesions in mammary glands. Here, we evaluate its efficacy in an experimental rat mammary carcinogenesis model. METHODS: Sprague-Dawley rats were treated with 1alpha-hydroxyvitamin D(5) beginning 2 weeks before carcinogen treatment. Animals received an intravenous injection of N-methyl-N-nitrosourea at 80 days of age and continued to receive dietary 1alpha-hydroxyvitamin D(5) for an additional 105 days. Tumor incidence and multiplicity were determined, and plasma concentrations of calcium and phosphorus were measured. The efficacy of 1alpha-hydroxyvitamin D(5) at different stages of carcinogenesis was determined in mouse mammary gland organ culture. All statistical tests were two-sided. RESULTS: The tumor incidence was reduced from 80% (95% confidence interval [CI] = 51.9%-95.7%) in control rats to 53.3% (95% CI = 26.6%-78.8%) and 46.6% (95% CI = 21.3%-73.4%) in rats treated with 1alpha-hydroxyvitamin D(5) at 25 microg/kg diet and 50 microg/kg diet, respectively. The tumor multiplicity was reduced from 1.6 tumors per rat to 1.2 (95% CI for the difference = -0.45 to 1.25; P=.34) and 0.8 (95% CI for the difference = 0.14-1.46; P =.02), respectively. There was no statistically significant increase in the plasma calcium or phosphorus concentration at either dose level. The vitamin D(5) analogue was effective during both the initiation and the promotion stages of mammary lesion formation in organ culture. CONCLUSION: Our findings indicate that 1alpha-hydroxyvitamin D(5) reduces the incidence of mammary carcinogenesis in vivo. This analogue appears to be a good candidate for further development as a chemopreventive agent.     

Dietary vitamin E intake and mammary carcinogenesis in rats

Using the dimethylbenz[a]anthracene-induced mammary tumor model,the present study demonstrated that a low vitamin E intake (7.5mg/kg of diet) had minimal effect on carcinoma development inrats fed a 5% stripped corn oil diet, but resulted in a markedenhancement in tumor incidence and yield in those rats fed a25% stripped corn oil ration. Control animals in this experimentreceived an adequate supply of vitamin E (30 mg/kg as DL--tocopherylacetate). Thus, the effect of vitamin E deficiency on mammarycarcinogenesis was accentuated in rats maintained on a highpolyunsaturated fat diet, an observation similar to that ofselenium deficiency which was reported by the author in a previouspublication. In view of the biochemical interaction betweenvitamin E and selenium as endogenous antioxidants, another experimentwas conducted to determine whether supranutritional supplementationof vitamin E (1000 mg/kg) was able to block the enhancementin mammary tumorigenesis due to selenium deprivation. Resultsof this experiment indicated that vitamin E excess failed toovercome the augmented tumor yield in selenium-deficient rats,nor did it provide any protection in rats that received an adequatesupply of selenium. In summary, vitamin E deficiency may increasethe risk of neoplastic development, especially when coupledwith a high polyunsaturated fat intake; however, a high vitaminE supplementation does not seem to have any prophylactic effecton tumorigenesis by Itself.     

A combination of resveratrol and melatonin exerts chemopreventive effects in N-methyl-N-nitrosourea-induced rat mammary carcinogenesis

The neurohormone melatonin is primarily involved in the regulation of circadian rhythms, but also acts as an antioxidant and anticarcinogenic agent, especially in breast cancer. Resveratrol (3,5,4'-trihydroxy-trans-stilbene) is a widely known polyphenolic agent from red wine, which has been shown to exert antioxidant, anti-inflammatory and anticarcinogenic effects. The objective of this study was therefore to investigate the effects of melatonin in combination with resveratrol in a rat model of experimental mammary carcinogenesis. Female Sprague-Dawley rats aged 31 days were used in the experiment. Mammary carcinogenesis was induced by N-methyl-N-nitrosourea (NMU), which was administered in two intraperitoneal doses (50 mg/kg of body weight). Chemoprevention with resveratrol and melatonin started 2 weeks before the first dose of NMU and lasted until the end of the experiment. The basic parameters evaluated were: tumour incidence, latency period, tumour frequency per group and tumour volume. In addition, oestrogen receptors ERα and ER?, melatonin receptor MT1, proliferating cell nuclear antigen and vascular endothelial growth factor were determined by immunohistochemical staining. The combination of resveratrol and melatonin reduced tumour incidence by approximately 17% and significantly decreased the quantity of invasive and in-situ carcinomas. Food intake declined in the second and seventh weeks after the administration of carcinogen. Resveratrol in combination with melatonin returned food intake to the level of intact controls. Resveratrol in combination with melatonin has some protective effects on NMU-induced rodent breast cancer. Further studies are necessary to confirm these effects of this promising combination.     

Chemoprevention of mammary carcinogenesis in female rats by rofecoxib

Epidemiological and experimental studies indicate that non-steroidal anti-inflammatory drugs play role in the prevention of human neoplasia including mammary gland cancer. In this study, tumour suppressive effects of a selective inhibitor of cyclooxygenase-2 (COX-2) rofecoxib in the prevention of N-methyl-N-nitrosourea (NMU)-induced mammary carcinogenesis in female Sprague-Dawley rats were evaluated. Rofecoxib was dietary administered in two concentrations-0.01 mg/1 g (ROFE 0.001%), or 0.05 mg/1 g (ROFE 0.005%). Rofecoxib decreased the incidence by 40% (ROFE 0.001%) and by 42.5% (ROFE 0.005%) (P=0.043) when compared to the control group. Similarly, tumour frequency and tumour volume decreased depending on an increasing rofecoxib dose by 18.5 or 40% (tumour frequency) and by 35 or 43% (tumour volume). Rofecoxib shifted the rate of fibroadenomas from 15% (control group) to 32% (ROFE 0.001%), or 38% (ROFE 0.005%), respectively. The present study points to pronounced dose-dependent tumour suppressive effects of rofecoxib in mammary carcinogenesis.     

Protective effect of dietary brussels sprouts against mammary carcinogenesis in Sprague-Dawley rats

The effect of dietary brussels sprouts (Brassica oleracea, L.) on mammary carcinogenesis induced by 7,12-dimethylbenz[a]anthracene (DMBA) was studied in Sprague-Dawley female rats. Rats fed a 20% brussels sprouts diet only during the initiation period of carcinogenesis had a palpable mammary tumor incidence of 13%, while those fed a casein-cornstarch semi-purified diet during this initiation period had a tumor incidence of 77% after 15 weeks post DMBA dose. When the rats were switched from the semi-purified diet to the 20% brussels sprouts diet at this time, there appeared to be a regression of small mammary tumors after 6 weeks on this dietary treatment. This regression was transitory since during the final 10 weeks of this 1 year study, 100% of this group of rats developed tumors. The rats fed the 20% brussels sprouts diet during tumor initiation exhibited a 67% incidence of fibroadenomas. The rats fed the semi-purified diet during initiation, but switched later to the brussels sprouts diet, showed over a 90% incidence of adenocarcinomas.     

Citrus auraptene suppresses cyclin D1 and significantly delays N-methyl nitrosourea induced mammary carcinogenesis in female Sprague-Dawley rats

Background  

Breast cancer is a major problem in the United States leading to tens of thousands of deaths each year. Although citrus auraptene suppresses cancer in numerous rodent models, its role in breast cancer prevention previously has not been reported. Thus, our goal was to determine the anticarcinogenic effects of auraptene against breast cancer.     

Metabolism of retinol and retinoic acid in N-methyl-N-nitrosourea-induced mammary carcinomas in rats

This study was conducted to examine the in vivo uptake and metabolism of natural retinoids by N-methyl-N-nitrosourea-induced mammary carcinomas. In this study, endogenous retinol and retinyl esters were present in normal mammary epithelial cells, but were undetectable in N-methyl-N-nitrosourea-induced mammary carcinomas in rats as determined by high-pressure liquid chromatography. No differences were found in plasma levels of retinol, in liver retinyl esters, or total content of vitamin A between tumor-bearing and control animals. Administered labeled retinol was taken up and esterified by normal mammary epithelial cells. Tumor-bearing rats were given injections i.p. of either [3H]retinol or [3H]retinoic acid. Radioactivity increased progressively with time in liver and other tissues except in breast tumor, where the uptake fluctuated over the 8 days after the injection of [3H]retinol; in mammary tumors practically no metabolism of [3H]retinol occurred, while in other tissues extensive esterification was detectable. In contrast, in animals given injections of [3H]retinoic acid, the uptake and metabolism of the label in the breast tumors paralleled with those found in other tissues. Neither the activity of acyl coenzyme A:retinol acyl transferase nor the activity of retinyl ester hydrolase was altered in the mammary tumor compared to the normal mammary gland. On the other hand, a significant decrease in the retinal oxidase activity was found in tumor tissue compared to normal mammary tissue. Since no esterification of [3H]retinol occurred in vivo despite the presence of acyl coenzyme A:retinol acyl transferase activity, it is possible that a specific defect in the cellular uptake of retinol may exist in N-methyl-N-nitrosourea-induced mammary carcinomas.     

Effects of arctiin on PhIP-induced mammary, colon and pancreatic carcinogenesis in female Sprague-Dawley rats and MeIQx-induced hepatocarcinogenesis in male F344 rats

Chemopreventive effects of arctiin, a lignan isolated from Arctium lappa (burdock) seeds, on the initiation or post initiation period of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) induced mammary carcinogenesis in female rats and on 2-amino-3, 8-dimethylimidazo[4,5-f]quinoxaline (MeIQx)-associated hepatocarcinogenesis in male rats were examined. In experiment 1, female Sprague-Dawley (SD) rats were given intragastric doses of 100 mg/kg body wt of PhIP once a week for 8 weeks as initiation. Groups of 20 rats each were treated with 0.2 or 0.02% arctiin during or after PhIP initiation. Control rats were fed 0.2 or 0.02% arctiin, or basal diet alone during the experimental period. Animals were killed at the end of week 48. Although the incidence of mammary carcinomas did not significantly differ among the PhIP-treated groups, multiplicity was significantly decreased in rats given 0.2 (0.7+/-0.7, P<0.05) or 0.02% (1.0+/-1.1, P<0.05) arctiin after PhIP initiation as compared with the PhIP alone controls (2.1+/-2.5). The average number of colon aberrant crypt foci was also significantly decreased in these two groups. Pancreas acidophilic foci were induced in PhIP treated animals with slight decrease in the multiplicity with arctiin during the initiation phase. For liver carcinogenesis, groups of 15 male F344 rats were given a single intraperitoneal injection of diethylnitrosamine (DEN) and starting 2 weeks later, they were administered 0.03% MeIQx in the diet, MeIQx together with 0.5% arctiin, 0.1% arctiin or basal diet for 6 weeks. They were subjected to two-third partial hepatectomy 3 weeks after DEN initiation and killed at the end of week 8 for glutathione S-transferase placental form (GST-P) immunohistochemistry. The numbers and areas of preneoplastic GST-P positive foci were elevated by the treatment with MeIQx, and further increased by the simultaneous treatment with arctiin. These results indicate that arctiin has a protective effect on PhIP-induced carcinogenesis particularly in the mammary gland in the promotion period. On the other hand, it may have a weak co-carcinogenic influence on MeIQx-induced hepatocarcinogenesis. In addition, the results suggested that PhIP is a weak pancreatic carcinogen in female SD rats, targeting acinar cells.     

Adrenal regulation of mammary tumorigenesis in female Sprague-Dawley rats: histopathology of mammary tumors

Mammary tumors induced in female Sprague-Dawley rats by feeding 7,12-dimethylbenz(a)anthracene (DMBA; 20 mg/100 g body weight) were classified according to histological criteria of tissue differentiation, cellular atypia, and evidence of invasion. The 549 tumors could be placed in three categories, nodular hyperplasia, nodular hyperplasia with atypia, and carcinoma, and combinations of all three. Although tumors classified histologically as carcinomas did not metastasize, upon transplantation to the kidney capsule, a tumor classified as a carcinoma grew for eight generations and metastasized. Tumor heterogeneity was a common finding in DMBA-initiated tumors. Carcinomas were an early lesion. As the length of time between DMBA treatment and sacrifice increased, more tumors with areas of carcinoma were found. Therefore, DMBA-initiated tumors progressed to carcinomas either soon after initiation or later by development within nodular hyperplasias. In 4 separate groups of animals (74 adrenalectomized rats and 90 intact rats), postinitiation adrenalectomy increased the numbers of carcinomas compared to intact animals. This effect was consistently seen in the cervical and thoracic mammary glands. We propose that the mechanism for enhancement of progression to greater malignancy by adrenalectomy may be inhibition of differentiation of initiated cells in the absence of glucocorticoids.     

Effect of molybdenum and tungsten on mammary carcinogenesis in Sprague-Dawley (SD) rats

Virgin female rats of SD strain were given ad libitum a nutritionally adequate semipurified diet containing 0.026 ppm molybdenum and deionized water (groups 1-3) or the same diet with 150 ppm tungsten and the drinking water (group 4). Group 1 was used as control. After 15 days, all the animals in groups 2-4 received an intravenous injection of N-nitroso-N-methylurea (NMU) 5 mg/100 g body weight. One week after administration of carcinogen, 10 ppm Mo was added to the drinking water in group 3. After 125 days, the mammary cancer incidence in group 4 (79.2%) was significantly higher than that in group 2 (50.0%) or group 3 (45.5%) (P less than 0.05). After 198 days, the average number of mammary cancer in each animal and mammary cancer incidence in group 3 (1.5 and 50.0%) were obviously lower than those in group 2 (2.0 and 90.5%) or group 4 (2.6 and 95.7%). The first palpable mammary tumor was found in the W-supplemented group only 56 days after the injection of NMU, whereas in the W-unsupplemented and Mo-supplemented groups, the first mammary tumor was observed 71 and 85 days after NMU treatment. Of these 181 mammary tumors, 177 (97.8%) were adenocarcinoma or papillary carcinoma, only 4 (2.2%) fibroadenocarcinoma. The results of this study show, for the first time, the inhibitory effect of Mo on the mammary carcinogenesis and promoting effect of Tungsten, an antagonist of molybdenum, on the tumor growth.     

Dietary seaweed (Laminaria) and mammary carcinogenesis in rats

To test the potential in vivo antitumor effect of dietary seaweed, we induced mammary tumors in female Sprague-Dawley rats with the carcinogen 7,12-dimethylbenz(a)anthracene. Twenty-one-day-old rats (n = 108) were divided into two groups. Controls were fed a standard semipurified diet, and experimental rats received the control diet with 5% Laminaria, a brown seaweed, replacing 5% alphacel . At 55 days of age, each rat received 5 mg 7,12-dimethylbenz(a)anthracene intragastrically. Rats were palpated for mammary tumors and weighed weekly for 26 weeks. Complete autopsies were then done on all rats. The seaweed diet did not alter weight gain or weights of body organs at autopsy. Experimental rats had a significant delay in the time to tumor (p = 0.007); median time until tumor was 19 weeks in experimental rats and 11 weeks in control animals. Among mammary adenocarcinoma tumor-bearing animals, experimental rats had fewer adenocarcinomas/individual (p less than 0.05). There was also an overall 13% reduction in the number of experimental rats with histologically confirmed adenocarcinomas (76% among the control rats compared to 63% among the experimental rats). Components of Laminaria which might account for the observed difference in mammary tumor growth are varied and include the sulfated polysaccharide fucoidan . Rats in the top row of cages had a significant (p = 0.01) delay in time to tumor compared to rats in the lower four rows. In each row, the seaweed-fed rats had a longer time to tumor than did the control rats.