Influence of caffeine on development of benign and carcinomatous mammary gland tumors in female rats treated with the carcinogens 7,12-dimethylbenz(a)anthracene and N-methyl-N-nitrosourea

The effect of chronic caffeine consumption (500 mg/liter of drinking water) on the initiation and promotion stages of 7,12-dimethylbenz(a)anthracene (DMBA) (a low dose, 0.5 mg/100 g body weight, i.v.) and N-methyl-N-nitrosourea (MNU) (a standard dose, 2.5 mg/100 g body weight, i.v.) induced mammary gland tumorigenesis in female Sprague-Dawley rats was determined. In the initiation studies, caffeine was administered for 30 days prior to and for 3-4 days after carcinogen treatment (carcinogens administered at 55-57 days of age); in the promotion studies, caffeine was administered beginning 3-4 days after carcinogen treatment and until experiment termination (DMBA study and MNU study, 48 and 26 weeks after carcinogen treatment, respectively). In the DMBA study, there were 62-73 rats/group, in the MNU study, 40 rats/group. Eighty-nine % of the mammary tumors induced by DMBA were benign (adenomas, fibroadenomas, often with cystic secretory activity), 11% were carcinomas (intraductal and invasive); virtually all of the MNU-induced mammary tumors were carcinomas (approximately 99%). Caffeine consumption during the initiation stage in the DMBA-treated rats resulted in a significant decrease in the mean number of mammary carcinomas per rat (50% reduction, P less than 0.01) and mean number of benign mammary tumors per rat (28% reduction, P less than 0.05); caffeine consumption during the promotion stage significantly decreased the mean number of benign mammary tumors per rat (57% reduction, P less than 0.001) while not significantly influencing mammary carcinoma number. In contrast, caffeine consumption during either the initiation or promotion stages of MNU-treated rats did not significantly influence this tumorigenic process. The influence of caffeine on urinary and fecal excretion of tritiated DMBA and on rat mammary gland development at the time of carcinogen treatment also was determined. Slightly reduced levels of tritium in 24-h urinary samples were observed in caffeine-treated animals (P = 0.06). No significant effect of caffeine on 24- to 96-h fecal or 48- to 96-h urinary excretion of the isotope was observed. No apparent effect of caffeine on rat mammary gland development (number of ducts, degree of lobuloalveolar development) was observed. That caffeine significantly suppresses the initiation stage of DMBA-induced rat mammary gland tumorigenesis, while not influencing this stage when MNU is used as a carcinogen, suggests that caffeine acts via an alteration in carcinogen (DMBA) activation. The lack of a pronounced effect of caffeine on tritiated DMBA excretion, however, does cast some doubt on this mechanism.(ABSTRACT TRUNCATED AT 400 WORDS)     

In vitro metabolism of 7,12-dimethylbenz[a]anthracene in mammary epithelial cells from rats with different susceptibilities to carcinogenesis

These in vitro metabolism studies of 7,12-dimethylbenz[a]-anthracene(DMBA) by rat mammary epithelial cells were initiated to determinewhether differences in the incidence of DMBA-induced mammarycarcinomas observed between young virgin, old virgin and parousSprague-Dawley rats can be explained through variations in theirability to metabolize DMBA. The results show that: (a) qualitatively,the metabolic profiles produced by epithelial cells from the3 groups of rats were similar; (b) the level of metabolism withcells from young virgin and parous rats was similar but was1.5–2.0 times higher than that obtained with cells fromold virgin rats; (c) the major ethyl acetate-soluble metabolitedetected by h.p.l.c. from cells of old virgin and parous ratswas the trans-8,9-dihydrodiol (50–65% of total metabolites),whereas the majority of the radioactivity following incubationwith cells from young virgin rats co-eluted with very polarmetabolites (65% of total metabolites); and (d) the amount ofphenolic metabolites produced by young virgin rat cells washigher than in old virgin or parous rat cells. The data suggestthat the susceptibility of young virgin rat mammary cells toDMBA may be due in part to the conversion of a greater percentageof DMBA to phenolic compounds and to water- and ethyl acetate-solublepolar metabolites, an event known to influence the susceptibilityof cultured cells to the cytotoxic effects of DMBA.     

Caffeine (1,3,7-trimethylxanthine), a temperate promoter of DMBA-induced rat mammary gland carcinogenesis

The administration of caffeine to the drinking water (250 mg/l and 500 mg/l) of female Sprague-Dawley rats after treatment with 7,12-dimethylbenzanthracene (DMBA) (5 mg i.g.) resulted in an increase in mammary carcinoma incidence. The confidence levels of statistical significance for the groups of rats receiving the 250-mg and 500-mg doses of caffeine were >0.70 and >0.99, respectively. This increase in mammary carcinoma incidence was observed when caffeine treatment was initiated commencing 3 days after DMBA treatment and continued for 21 weeks. This effect was also observed when caffeine treatment was initiated commencing 20 weeks after DMBA treatment and continued for 6 weeks in rats relatively refractory to carcinogen treatment (mammary tumor-free at onset of treatment) and in rats relatively sensitive to the carcinogen (mammary tumor bearing at onset of caffeine treatment). Caffeine treatment of rats prior to and during carcinogen treatment did not significantly affect mammary carcinoma incidence. Thus caffeine consumption has been shown to significantly enhance the promoting phase but is without effect on the initiating phase of this carcinogenic process.     

Refractoriness to mammary tumorigenesis in parous rats: is it caused by persistent changes in the hormonal environment or permanent biochemical alterations in the mammary epithelia?

Administration of a single i.v. injection of 50 mg N-methyl-N-nitrosourea(MNU)/kg body wt to 50- to 60-day-old virgin rats, 120-day-oldvirgin rats, and 120-day-old parous rats (Sprague-Dawley; n= 18–37) resulted in a high incidence of mammary carcinomasin the virgin animals (97.3% in 50- to 60-day-old virgin rats;75.0% in 120-day-old virgin rats), but mammary carcinomas didnot develop in the parous rats. The concentrations in serumof various mammotropic hormones were measured in identical groupsof rats at the time of MNU treatment. Growth hormone (GH) concentrationwas significantly reduced in parous rats, as compared with youngor age-matched virgin rats. The concentrations of prolactin,17ß-estradiol, progesterone, corticosterone and thyroxinewere not significantly altered in the parous rats compared tothe two groups of virgin animals. Histological examination ofthe mammary glands from the three groups of rats showed thatthe epithelia of the parous animals were in a stage of regression,whereas the mammae of the young virgin rats showed the highestdegree of lobulo-alveolar development. The levels of estrogenreceptor (ER), epidermal growth factor (EGF) receptor (EGF-R)and GH receptor (GHR) in the mammary glands of the animals werealso measured. We found a reduction in the receptor levels forboth estrogen and EGF in mammary tissues from parous animals.Receptors for GH were present in normal mammary tissues fromboth virgin and parous rats. We hypothesize that the reductionin the circulating concentration of GH caused the reduced susceptibilityof parous rats to mammary carcinogenesis possibly by decreasingthe levels of ER and/or EGF-R in the mammary gland.     

Obesity increases the incidence of 7,12-dimethylbenz(a)anthracene-induced mammary tumors in an ovariectomized Zucker rat model

Obesity is associated with increased risk for postmenopausal, but not premenopausal breast cancer. Recently, we reported that intact obese Zucker rats had increased susceptibility to DMBA-induced mammary tumors compared to lean Zucker rats. In the present study, we investigated whether excessive adipose tissue would promote mammary tumor induction in the absence of ovarian estrogen. Lean and obese rats were sham-operated or ovariectomized at 40 days old and were gavaged at 50 days old with 65 mg/kg DMBA. Rats were weighed and palpated twice weekly for detection of mammary tumors and sacrificed 135 days post-DMBA treatment. Obese sham-operated (O/S) rats had a shorter latency period (102 days) compared to lean sham-operated (L/S) (134 days) and obese ovariectomized (O/O) rats (123 days). At the end of the experiment, 36% of the O/O rats developed mammary tumors while lean ovariectomized (L/O) rats developed no mammary tumors (P<0.001), and 59% of the O/S rats developed mammary tumors compared to 30% of the L/S rats (P<0.05). In summary, obesity increases the susceptibility of ovariectomized Zucker rats to DMBA-induced mammary tumors, suggesting that adipose tissue-derived estrogen in obese animals may be sufficient to promote DMBA-induced tumors in this model. These results suggest that obesity in postmenopausal women may increase breast cancer risk due to increased breast tissue exposure to adipose tissue-derived estrogen. In conclusion, we have developed an animal model to further investigate the role of obesity in breast cancer development in postmenopausal women.     

Effect of selective ablation of proliferating mammary epithelial cells on MNU induced rat mammary tumorigenesis

Proliferating cells within the terminal end buds of the virgin female rat mammary gland are the most susceptible to chemical carcinogen induced tumorigenesis. We hypothesized that selective ablation of proliferating cells in the mammary gland would reduce mammary tumor incidence upon carcinogen challenge. Selective ablation of proliferating cells was achieved by intraductal injections of Adv-RSV-tk and gancyclovir administration. Despite efficient viral transduction of the thymidine kinase protein and the apparent elimination of >90% of the prolif-erating cells, the rats exhibited a higher incidence of MNU induced mammary tumors arising with shorter latency as compared to control animals. Several possible explanations of the puzzling relationship between elimination of cycling cells and increased tumor incidence are discussed and alternative strategies for the prevention of breast cancer are proposed.     

The prophylaxis of rat and mouse mammary gland tumorigenesis by suppression of prolactin secretion: A reappraisal

Summary The purpose of this study was to evaluate the effect of early and temporal CB-154 induced suppression of prolactin secretion on the genesis of mammary tumors in female Sprague-Dawley rats treated i.g. with 3 doses of 7,12-dimethylbenzanthracene (DMBA); doses which yield a high (20 mg), moderate (5 mg) and low (1.25 mg) mammary carcinoma incidence. In addition, the effect of prolactin suppression on the genesis of mammary tumors in strains of rodents which, when treated with maximally tolerated doses of DMBA, develop a moderate (female Lewis rats) and a low (female Long-Evans rats and female Balb/c mice) mammary carcinoma incidence was also evaluated. Daily suppression of prolactin secretion during DMBA treatment (30 days before, during and 30 days after) (series 1) or commencing 30 days after DMBA treatment (for 60–81 days) (series 2) significantly (P<0.05) reduced the number of mammary carcinomas in Sprague-Dawley rats treated with 20 and 5 mg DMBA and in Lewis rats (series 2); treatment with CB-154 did not significantly lower mammary carcinoma incidence in Sprague-Dawley rats treated with 1.25 mg DMBA, nor in Lewis rats (series 1), Long-Evans rats and Balb/c mice. These results provide evidence that the effectiveness of prolactin suppression in the prophylaxis of chemical carcinogenesis of the rodent mammary gland is enhanced when a moderate or large dose of the carcinogen is used, or a strain of rodent is used which is moderately or highly susceptible to the mammary oncogenic effects of the carcinogen. Low doses of the carcinogen or use of a rodent strain relatively resistent to the carcinogen nullifies the chemopreventive activities of early prolactin suppression. Address for reprints: Clifford W. Welsch, Ph.D., Department of Anatomy, Michigan State University, East Lansing, MI 48824.     

Suppression by pregnancy of chemically induced preneoplastic cells of the rat mammary gland

Since a previous study suggested that pregnancy either eliminated preneoplastic cells or increased their latency period in rat mammary glands, additional experiments were performed to determine the fate of these cells. Following administration of the carcinogen dimethylbenzanthracene, few mammary cancers appeared after rats completed pregnancy and lactation. Because these results are similar to those previously obtained with N-methyl-N-nitrosourea (MNU), the effect of pregnancy appears to be independent of the carcinogen used to induce cancer. For rats dosed with MNU, relatively few cancers developed either during an extended observation following pregnancy and lactation or following administration of prolactin, a growth stimulator of mammary cancers. In the latter experiment, the average number of mammary cancers per rat in the prolactin-treated virgin rats was 7.1, while prolactin-treated parous rats had only 0.8 cancers per rat; i.e., 89% fewer cancers. Thus, pregnancy appears to suppress many of the preneoplastic cells induced by carcinogens in the mammary gland of rats.     

Relationship between stages of mammary development and sensitivity to gamma-ray irradiation in mammary tumorigenesis in rats

Mature Wistar-MS rats were ovariectomized and treated with estradiol benzoate and/or progesterone. Control animals were treated with olive oil. The rats were then exposed to γ-rays and implanted with a pellet of diethylstilbestrol. The incidence of mammary tumors in rats treated with estradiol benzoate or with progesterone was significantly higher than in rats in the non-treated control group, whereas, in rats treated with both estradiol benzoate and progesterone, the incidence was not significantly different from that in the controls. Histological examination of the mammary tumors showed 2 types of neoplasm: adenocarcinoma and fibroadenoma. Interestingly, over half of all the tumors in the rats treated with estradiol benzoate were adenocarcinomas, while fibroadenomas were mainly induced in the rats treated with progesterone or with both estradiol benzoate and progesterone. The expression of estrogen and progesterone receptors in the tumor tissues showed some differences according to whether the groups were treated with estradiol benzoate or with progesterone. Morphologically, mammary glands at irradiation showed well-developed lobuloalveoli in both the estradiol-benzoate-treated rats and in those rats treated with both estradiol benzoate and progesterone. This was consistent with the higher incorporation of [³H]thymidine into the DNA in the mammary glands of rats in both of these groups. Our findings suggest that a more advanced developmental stage of the mammary glands, dependent upon ovarian hormones, is related to a higher incidence of mammary tumors induced by irradiation. © 1995 Wiley-Liss, Inc.     

Pregnancy-induced antitumor cytotoxicity of T cell-rich fraction against mammary adenocarcinoma cells in rats.

Our earlier observations indicated that splenocytes from parous rats have high cytotoxic activity against mammary tumor cells in vitro. It has also been observed that this cytotoxic activity of splenocytes from parous rats can be adoptively transferred to virgin rats of same age. The present investigation is an attempt to determine the cell type in spleens of parous rats that cause the cytotoxicity against mammary tumors. The spleens from both virgin and parous rats were removed aseptically and T and B cell-rich fractions were separated. 7,12-Dimethylbenz[a]anthracene-induced rat mammary tumor epithelial cells were used as targets and the T and B cell-rich fractions from either virgin or parous rats were used as effectors. The parous rats were divided into two groups according to the time period after parturition. Group I contained rats that were 5-13 days after delivery, while the rats in group II were 14 or more days post parturition. In vitro cytotoxicity was determined by incubating the tumor cells with spleen cells in the target/effector ratio of 1:3, 1:10 and 1:30 and using the standard 51Cr release assay. In all ratio groups of T cell-rich fractions particularly in group II, there was significantly higher cytotoxicity against mammary tumor cells. None of the B cell-rich fractions from parous rats were significantly more cytotoxic than those from virgin controls.     

Perinatal Exposure to Bisphenol A or Diethylstilbestrol Increases the Susceptibility to Develop Mammary Gland Lesions After Estrogen Replacement Therapy in Middle-Aged Rats

The development of the mammary gland is a hormone-regulated event. Several factors can dysregulate its growth and make the gland more susceptible to cellular transformation. Among these factors, perinatal exposure to xenoestrogens and hormone replacement therapy has been associated with increased risk of developing breast cancer. Here, we assessed the effects induced by estrogen replacement therapy (ERT) in ovariectomized (OVX) middle-aged rats and whether perinatal exposure to diethylstilbestrol (DES) or bisphenol A (BPA) modified these effects in the mammary gland. Pregnant rats were orally exposed to vehicle, 5 μg DES/kg/day, or 0.5 or 50 μg BPA/kg/day from gestational day 9 until weaning. Then, 12-month-old offspring were OVX and treated with 17β-estradiol for 3 months. Morphological changes and the percentage of epithelial cells that proliferated or expressed estrogen receptor alpha (ESR1) and progesterone receptor (PR) were analyzed in mammary gland samples of 15-month-old animals. ERT induced lobuloalveolar hyperplasia and ductal cysts in the mammary gland of middle-aged rats, associated with a higher proliferation index of epithelial cells. Perinatal exposure to DES followed by ERT increased the number of cysts and induced the formation of fibroadenoma and ductal carcinoma in situ, without modifying the expression of ESR1 or PR. Also, after 3 months of ERT, BPA-exposed rats had a higher incidence of ductal hyperplasia and atypical lobular hyperplasia than animals under ERT alone. In conclusion, perinatal exposure to xenoestrogens increases the susceptibility of the mammary gland to develop cysts and hyperplastic lesions when confronted with ERT later in life.     

Identification of rat mammary tumor-1 gene (RMT-1), which is highly expressed in rat mammary tumors.

Full-term pregnancy early in life results in a permanent reduction in lifetime breast cancer risk in women. Parous rats and mice are also refractory to chemical carcinogenesis. Therefore, investigation of the differences between mammary glands from virgin and parous rats would provide valuable information regarding the protective effects of early full-term pregnancy. In this report, we examined the gene expression patterns in mammary glands from virgin and parous Lewis rats. Using differential display technology, a novel 4.2 kb cDNA, designated rat mammary tumor-1 (RMT-1) was isolated. Northern blot analysis of RMT-1 showed that RMT-1 expression was higher in the pre-pubertal and pubertal stages during rat mammary gland development while it was down-regulated in mammary glands from mature virgin and parous rats. RMT-1 expression was highest in rat mammary cancers compared with either the mammary glands of virgin or parous rats. At the Northern blot sensitivity level, RMT-1 expression was found only in the mammary gland. Northern blot analysis also showed that the expression of this gene was found in 74% of N-methyl-nitrosourea (MNU)-induced mammary cancers while it was not found in MNU-induced cancers from other organs. The examination of the RMT-1 gene structure revealed that it consists of five exons spanning 5.9 kb. Using fluorescence in situ hybridization, the gene was localized on rat chromosome 1 band q 43-51. The present data show that there is a correlation between high RMT-1 expression and rat mammary carcinogenesis or decreased RMT-1 expression and parity associated refractoriness to chemically induced mammary carcinogenesis. However, whether or not RMT-1 gene has a functional role in these processes remains to be investigated.     

Relationship between previous reproductive history and chemically induced mammary cancer in rats

The relationship between a previous history of multiple pregnancies and/or lactations and 7,12-dimethylbenz(a)anthracene-induced mammary carcinogenesis was studied in groups of Sprague-Dawley rats. All rats received the carcinogen at approximately 6 1/2 months of age. Virgin control rats which did not receive the carcinogen did not develop mammary cancer within 13 months although carcinogen-fed virgin rats exhibited a mammary cancer incidence of 38.5%. A single pregnancy prior to feeding the carcinogen resulted in a decrease in the incidence and an increase in the mean time of appearance of the first palpable tumor. A single pregnancy followed by a subsequent nursing period reduced the percentage incidence and increased the mean time of appearance of the first palpable tumor when compared to the carcinogen-fed virgin rats. A further increase in the number of pregnancies and/or lactations did not greatly influence the incidence of mammary cancer nor the number of cancers per rat. The mean time of tumor appearance was, however, increased. These data indicated that the relationship between party and carcinogen-induced mammary cancer in the rat is similar to that existing in the human female but opposite to that found for the mouse.     

Mammary tumors induced by human adenovirus type 9: A role for the viral early region 4 gene

Summary Human adenovirus type 9 (Ad9) elicits exclusively estrogen-dependent mammary tumors when injected into female rats. Three different histological types of mammary tumor (benign fibroadenomas, phyllodes tumors, and malignant solid sarcomas) have been described in Ad9-infected animals, with benign fibroadenomas being seen most frequently. Interestingly, in contrast to other adenoviruses, in which oncogenic viral functions are entirely encoded within the E1 region, Ad9 requires an E4 region transforming protein (ORF1) for its unique mammary oncogenicity. Studies of Ad9-induced rat mammary tumors may lead to a detailed molecular understanding for the development of fibroadenoma, a common human breast tumor.     

Refractoriness to mammary carcinogenesis in the parous mouse is reversible by hormonal stimulation induced by pituitary isografts

We have previously reported that mouse mammary epithelial cells transformed in vitro yield tumors which vary qualitatively and quantitatively as a function of the mitogenic environment in which the cells are propagated at the time of carcinogen treatment. One milieu supportive of transformation in vitro was medium supplemented with progesterone and prolactin as the mitogens. We have performed parallel studies in which virgin mice were isografted with pituitaries resulting in elevated serum titers of progesterone and prolactin. After carcinogen treatment, these mice developed mammary tumors which included those identical genotypically and phenotypically to tumors induced in vitro in cells grown in progesterone and prolactin during carcinogen exposure. Our current working hypothesis is that the mitogenic environment around the time of carcinogen administration can modulate the incidence and phenotype of the resultant tumors. To further test this hypothesis, we have evaluated the susceptibility of hormonally-stimulated parous mice to chemically induced mammary carcinogenesis since parity is known to significantly reduce the susceptibility of the mouse mammary gland to carcinogenesis. Virgin or multiparous BALB/c mice were isografted with two pituitaries. Five weeks after surgery, the mice were injected with N-methyl-N-nitrosourea (MNU; 50 μg/g i.v.). Mammary carcinomas arose in 85% (11/13) with a median latency of 22.8 weeks and 1.9 tumors per virgin mouse and 80% (24/30) with a median latency of 22.1 weeks at a frequency of 1.9 tumors per parous mouse. Only 14% (2/14) of the non-isografted, age-matched parous controls developed tumors when injected with MNU. Fourteen parous mice receiving only pituitary isografts (no MNU), did not develop mammary carcinomas within the 7-month period of the study. These results demonstrate that parous BALB/c mice are refractory to MNU-induced mammary carcinogenesis and that this refractoriness is not permanent, but can be overcome by hormonal stimulation mediated by pituitary isografts.     

The parity-related protection against breast cancer is compromised by cigarette smoke during rat pregnancy: observations on tumorigenesis and immunological defenses of the neonate

Early pregnancy is a powerful negative risk factor for breast cancer (BCa) in women. Pregnancy also protects rats against induction of BCa by carcinogens such as N-methyl-N-nitrosourea (MNU), making the parous rat a useful model for studying this phenomenon. Smoking during early pregnancy may lead to an increased risk of BCa in later life, possibly attributable to carcinogens in cigarette smoke (CS), or to reversal of the parity-related protection against BCa. To investigate these possibilities, 50-day-old timed first-pregnancy rats were exposed to standardized mainstream CS (particle concentration = 50 mg/m3) or to filtered air (FA) 4 h/day, Day 2-20 of gestation. Age-matched virgin rats were similarly exposed to CS or FA. At age 100 days, the CS or FA-exposed, parous and virgin rats were injected s.c. with MNU (50 mg/kg body wt), or with MNU vehicle. Mammary tumors (MTs) first appeared in virgin rats 9 weeks post-MNU injection. While no MTs were detected in FA-exposed parous rats until 18 weeks post-MNU, MTs appeared in the CS-exposed parous rats as early as 10 wks (P < 0.02). As no MTs developed in CS-exposed rats not injected with MNU, CS did not act as a direct mammary carcinogen. Serum prolactin concentration on Day 19 of pregnancy in CS-exposed dams was reduced by 50% compared with FA-exposed dams (P < 0.005). CS exposure during a pregnancy may thus 'deprotect' rats, enhancing their vulnerability to MNU-induced BCa. Prenatal CS exposure had no detectable effect on the immune responses of the pups examined at 3, 8 or 19 weeks of age. However, prolactin concentration in stomach contents (milk) of 3-day-old pups suckled by CS-exposed dams was decreased when compared with that of FA-exposed dams (P < 0.032). As milk-borne prolactin modulates development of the central nervous and immune systems of neonatal rats, CS exposure of the dams could adversely affect later maturation of these systems by reducing milk prolactin.     

Comparative effect of chlormadinone acetate and diethylstilbestrol as promoters in mammary tumorigenesis of rats irradiated with γ-rays during lactation

The purpose of this study was to determine the promotional role of estrogen and progestin in the development of radiation-induced mammary tumors. To eliminate the effects of endogenous ovarian hormones on tumor promotion, all rats were ovariectomized immediately after the initiation by irradiation with 2.6 Gy -rays at day 21 of lactation, and were divided into 3 groups. For the control experiment, rats were implanted with a cholesterol pellet 1 month after the irradiation. Only one rat developed a fibroadenoma (4.3% mammary tumor incidence) during the 1 year period of the implantation. In the other two groups, chlormadinone acetate (CMA) to increase progestin level or diethylstilbestrol (DES) to increase estrogenic activity were administered, respectively, as tumor promoters for 1 year. Treatment with CMA did not significantly increase the incidence of mammary tumors as compared with the controls. However, administration of DES resulted in a significantly higher mammary tumor incidence (79.3%) than control treatment. Compared with cholesterol administration, DES treatment caused an increase in prolactin concentration in serum (5-fold), and reduction of estradiol-17 concentration (22% of control). These results suggest that DES ia a potent effective promoter for tumorigenesis of radiation-initiated mammary cells, but CMA is not. DES may act directly on the irradiated mammary cells by binding to ER, and indirectly by stimulating prolactin secretion from the pituitary glands.     

Prevention of age-related spontaneous mammary tumors in outbred rats by late ovariectomy

Background: Breast cancer prevention trials have shown that the antiestrogen tamoxifen inhibits development of estrogen receptor (ER)-positive tumors. In Sprague-Dawley rats, removal of ovarian function in young animals can reduce the incidence of spontaneous age-dependent mammary tumors. However, it is not known whether removal of ovaries late in life, before middle age onset, can still prevent mammary tumor development. Methods: In this study we used Hsd:Sprague-Dawley^® SD^® (Hsd) rats to determine the effect of late ovariectomy on mammary tumor development. Intact, sham-ovariectomized and ovariectomized rats were followed until 110 weeks of age, or over their life span. In some experiments, palpable tumors were surgically removed upon presentation. Results: Removal of ovaries before middle age onset (∼5-7 months) inhibited development of spontaneous mammary tumors by 95%. Only one mammary tumor was observed in 19 late ovariectomized animals while 47 total tumors developed in 42 non-ovariectomized animals. Tumor incidence was reduced from 73.8 to 5.3% (relative risk = 0.05, 95% CI = 0.0072-0.354). The frequency of mammary carcinomas in non-ovariectomized virgin female rats was one in eight rats. Spontaneous rat carcinomas expressed ER and other biomarkers, such as cyclin D1. When palpable tumors were removed by surgical excision, tumor multiplicity increased from 0.76 to 1.61 tumors per rat. Surprisingly, ovariectomy increased the 110-week survival rate and maximum life span of Hsd rats. Conclusion: Late ovariectomy prevents spontaneous mammary tumor development in Hsd rats. This animal model may be useful for evaluating novel interventions in breast cancer prevention.     

Immuno-neutralization of circulating relaxin does not alter the breast cancer-protective action of parity in MNU-treated rats

Early pregnancy and childbirth protects women against future development of breast cancer by an unknown mechanism. Parity likewise reduces mammary cancer incidence in rats exposed to the carcinogen, N-methyl-N-nitrosourea (MNU), providing a model for the human phenomenon. We hypothesized that relaxin, a 6KD luteal mammotropic hormone of pregnancy, might be the anti-cancer pregnancy factor, and that induced relaxin deficiency during rat gestation would restore carcinogen sensitivity. Forty-one pregnant (age 50 days) and 25 age-matched virgin Sprague-Dawley rats were used. Relaxin deficiency was induced by injecting mouse monoclonal anti-rat relaxin antibody (MCA1) days 12-18 of gestation. Pregnant controls were injected with vehicle or mouse IgG on the same schedule. Because MCA1 disrupts parturition, all rats underwent cesarean section on day 22. At age 100 days, all rats were injected i.v. with MNU (50mg/Kg) and examined daily for tumors until euthanized at age 240 days.Mammary tumor incidence and frequency were significantly (p<0.01) reduced and tumor latency was increased (p<0.001) in primiparous as compared with virgin rats. However, tumor incidence, type, size and latency were similar in MCA1-treated and control primiparous rats. Thus, luteal relaxin does not appear to be the factor responsible for resistance to breast cancer.