Effect of protein supplement level on the productive and reproductive parameters of replacement heifers managed in intensive grazing systems.

Evaluations of replacement heifers in intensively managed grazing systems in tropical conditions are warranted. Thus, we aimed to evaluate performance, muscle and mammary gland development, oocyte quality, and in vitro production of embryos of crossbred heifers grazing an intensively managed pasture and supplemented with high or low protein concentrates. Eighteen pubertal crossbred heifers (Holstein x Gyr) with an initial weight of 350 ± 8.0 kg were used in a 60-day trial. Two supplement types, 12% crude protein (CP) (S12CP) or 24% CP (S24CP), and a control treatment (mineral mixture, CON) were randomly distributed to the heifers. Throughout the experiment, four digestibility trials were performed over four consecutive days. Four ovarium pick-ups were performed to evaluate oocyte quality and in vitro embryo production. Lastly, ultrasounds of carcasses and mammary glands were performed. The intakes of dry matter (DM), digestible energy (DE), and CP were greater for supplemented (SUP) compared with CON heifers. The SUP heifers had a greater average daily gain (ADG) (645 versus 390 g/d) and rib eye area (58.78 versus 53.32 cm2) than the CON heifers. Oocyte recovery, quality, and follicle features were not affected by supplementation strategy. However, the cleavage rate (47.17% versus 30.31%) and blastocyst rate (27.91% versus 10.12%) were negatively affected by supplementation. The S12CP presented a blastocyst rate much lower than the S24CP (3.02% versus 17.23%). Carcass ultrasonography indicated a trend for greater rib eye area for S24CP and mammary ultrasonography indicated no effects of supplementation on mammary gland development. In summary, supplementation seems to be an appropriate strategy for satisfactory performance, with greater muscle deposition and no negative impacts on mammary gland development. However, in vitro embryo production was impaired when the animals received the supplementation with 12% CP.

Folate Deficiency Inhibits Development of the Mammary Gland and its Associated Lymphatics in FVB Mice.

BACKGROUND: Folate is essential for DNA synthesis, DNA repair, cell proliferation, development, and morphogenesis. Folic acid (FA) is a nutritional supplement used to fortify human diets. OBJECTIVES: We investigated the effects of dietary FA on early mammary gland (MG) development and hyperplasia. METHODS: Study 1: nulliparous female FVB wild-type (WT) mice were fed control (Con; 2 mg FA/kg), deficient (Def; 0 mg FA/kg), excess (Ex; 5 mg FA/kg), or super excess (S-Ex; 20 mg FA/kg) diets for 8 wk before mating to WT or heterozygous FVB/N-Tg[mouse mammary tumor virus long terminal repeat (MMTV)-polyomavirus middle T antigen (PyVT)]634Mul/J (MMTV-PyMT+/-) transgenic males. Dams were fed these diets until they weaned WT or MMTV-PyMT+/- pups, which were fed the dam's diet from postnatal day (PND) 21 to 42. Tissues were collected from female progeny at PNDs 1, 21, and 42. Study 2: Con or Def diets were fed to WT intact females and males from PND 21 to 56, or to ovariectomized females from PND 21 to 77; tissues were collected at PND 56 or 77. Growth of all offspring, development of MGs, MG hyperplasia, supramammary lymph nodes, thymus and spleen, cell proliferation, and expression of MG growth factors were measured. RESULTS: Study 1: Ex or S-Ex did not affect postnatal MG development or hyperplasia. The rate of isometric MG growth (PND 1-21) was reduced by 69% in Def female progeny (P < 0.0001). Similarly, hyperplastic growth in MGs of Def MMTV-PyMT+/- offspring was 18% of Con (P < 0.05). The Def diet reduced supramammary lymph node size by 20% (P < 0.0001) and increased MG insulin-like growth factor 2 mRNA by 200% (P < 0.05) and protein by 130%-150% (P < 0.05). Study 2: the Def diet did not affect MG growth, but it did reduce supramammary lymph node size (P < 0.05), spleen weight (P < 0.001), and thymic medulla area (P < 0.05). CONCLUSIONS: In utero and postnatal folate deficiency reduced the isometric development of the MGs and early MG hyperplasia. Postnatal folate deficiency reduced the development of lymphatic tissues.

Valine supplementation during late pregnancy in gilts increases colostral protein synthesis through stimulating mTOR signaling pathway in mammary cells.

Mammary gland development during late pregnancy in sows is a major factor affecting the composition of colostrum and milk and the pre-weaning growth of piglets, while valine is essential for protein and nitrogen metabolism in mammary gland of sow. However, the effects of valine and its underlying mechanism on mammary gland development during late pregnancy are still unclear. Here, we hypothesized that dosage of dietary valine during late pregnancy will affect protein synthesis of colostrum in gilts. The results showed that supplementation of valine during late pregnancy significantly increased content of protein (P < 0.01), fat (P = 0.02) and solids-non-fat (P = 0.04) in colostrum. Our in vitro study also confirmed that valine supplementation increased protein synthesis and cell proliferation in porcine mammary epithelial cells (PMEC). Furthermore, these changes were associated with elevated phosphorylation levels of mammalian target of rapamycin (mTOR), and ribosomal protein S6 kinase (S6) and eukaryotic initiation factor 4E-binding protein-1 (4EBP1) in valine-supplemented cells, which could be effectively blocked by the antagonists of mTOR. These findings indicated that valine enhanced mammary gland development and protein synthesis in colostrum via the mTOR signaling pathway. These results, using an in vivo and in vitro model, helped to understand the beneficial effects of dietary valine supplementation on gilts.

Review: Mammary gland development in swine: embryo to early lactation.

Milk production by the sow is a major factor limiting the growth and survival of her litter. Understanding the process of morphogenesis of the sow's mammary gland and the factors that regulate mammary development are important for designing successful management tools that may enhance milk production. Primordia of the mammary glands are first observable in the porcine embryo at approximately 23 days of gestation. The glands then progress through a series of morphologically distinct developmental stages such that, at birth, each mammary gland is composed of the teat, an organized fat pad and two separate lactiferous ducts each with a few ducts branching into the fat pad. The glands continue to grow slowly until about 90 days of age when the rate of growth increases significantly. The increased rate of mammary gland growth coincides with the appearance of large ovarian follicles and an increase in circulating estrogen. After puberty, the continued growth of the gland and elongation and branching of the duct system into the fat pad takes place in response to the elevated levels of estrogen occurring as part of the estrous cycles. After conception, parenchymal mass of each gland increases slowly during early pregnancy and then grows increasingly rapidly during the final trimester. This growth is in response to estrogen, progesterone, prolactin and relaxin. Lobuloalveolar development occurs primarily during late pregnancy. By parturition, the fat pad of the mammary gland has been replaced by colostrum-secreting epithelial cells that line the lumen of the alveoli, lobules and small ducts. All mammary glands develop during pregnancy, however, the extent of development is dependent on the location of the mammary gland on the sow's underline. The mammary glands undergo significant functional differentiation immediately before and after farrowing with the formation of colostrum and the transition through the stages of lactogenesis. Further growth of the glands during lactation is stimulated by milk removal. Individual glands may grow or transiently regress in response to the intensity of suckling during the initial days postpartum. Attempts to enhance milk production by manipulation of mammary development at stages before lactation generally have met with limited success. A more in depth understanding of the processes regulating porcine mammary gland morphogenesis at all stages of development is needed to make further progress.

Prenatal Exposure to Unconventional Oil and Gas Operation Chemical Mixtures Altered Mammary Gland Development in Adult Female Mice.

Unconventional oil and gas (UOG) operations, which combine hydraulic fracturing (fracking) and directional drilling, involve the use of hundreds of chemicals, including many with endocrine-disrupting properties. Two previous studies examined mice exposed during early development to a 23-chemical mixture of UOG compounds (UOG-MIX) commonly used or produced in the process. Both male and female offspring exposed prenatally to one or more doses of UOG-MIX displayed alterations to endocrine organ function and serum hormone concentrations. We hypothesized that prenatal UOG-MIX exposure would similarly disrupt development of the mouse mammary gland. Female C57Bl/6 mice were exposed to ~3, ~30, ~ 300, or ~3000 µg/kg/d UOG-MIX from gestational day 11 to birth. Although no effects were observed on the mammary glands of these females before puberty, in early adulthood, females exposed to 300 or 3000 µg/kg/d UOG-MIX developed more dense mammary epithelial ducts; females exposed to 3 µg/kg/d UOG-MIX had an altered ratio of apoptosis to proliferation in the mammary epithelium. Furthermore, adult females from all UOG-MIX-treated groups developed intraductal hyperplasia that resembled terminal end buds (i.e., highly proliferative structures typically seen at puberty). These results suggest that the mammary gland is sensitive to mixtures of chemicals used in UOG production at exposure levels that are environmentally relevant. The effect of these findings on the long-term health of the mammary gland, including its lactational capacity and its risk of cancer, should be evaluated in future studies.

Stearic acid suppresses mammary gland development by inhibiting PI3K/Akt signaling pathway through GPR120 in pubertal mice.

It has been demonstrated that dietary high fat diet negatively affects the pubertal mammary gland development. The aim of the present study was to investigate the effects of stearic acid (SA), an 18-carbon chain saturated fatty acid, on mammary gland development in pubertal mice and to explore the underlying mechanism. Our results demonstrated that dietary supplementation of 2% SA suppressed mammary duct development, with significant reduction of terminal end bud (TEB) number and ductal branch. In accord, the expression of proliferative marker Cyclin D1 was markedly decreased by dietary SA. Furthermore, dietary SA led to increase of G protein-coupled receptor 120 (GPR120) expression and inhibition of PI3K/Akt signaling pathway in mammary gland of pubertal mice. In good agreement with the in vivo findings, the in vitro results showed that 40 µM SA significantly suppressed proliferation of mouse mammary epithelial cell HC11 by regulating mRNA and/or protein expression of proliferative markers such as Cyclin D1/3, p21, and PCNA. Meanwhile, SA activated GPR120 and inhibited PI3K/Akt signaling pathway in a GPR120-dependent manner. In addition, SA-induced inhibition of PI3K/Akt signaling pathway, suppression of HC11 proliferation, and alteration of proliferative markers expression were abolished by knockdown of GPR120 with siRNA. Collectively, these findings showed that SA suppressed mammary gland development of pubertal mice, which was coincident with the SA-inhibited HC11 proliferation, and was associated with inhibition of PI3K/Akt signaling pathway through activation of GPR120. These data provided new insights into the regulation of mammary gland development by dietary fatty acids.

Detection and comparison of microRNAs in the caprine mammary gland tissues of colostrum and common milk stages.

BACKGROUND: MicroRNAs (miRNAs) have a great influence on various physiological functions. A lot of high-throughput sequencing (HTS) research on miRNAs has been executed in the caprine mammary gland at different lactation periods (common milk lactation and dry period), but little is known about differentially expressed miRNAs in the caprine mammary gland of colostrum and peak lactation periods. RESULT: This study identified 131 differentially expressed miRNAs (P < 0.05 and log2 colostrum normalized expression (NE)/peak lactation NE > 1 or log2 colostrum NE/peak lactation NE < -1), including 57 known miRNAs and 74 potential novel miRNAs in the colostrum and peak lactation libraries. In addition, compared with differentially expressed miRNAs in the peak lactation period, 45 miRNAs in the colostrum lactation period were remarkably upregulated, whereas 86 miRNAs were markedly downregulated (P < 0.05 and log2 colostrum NE/peak lactation NE > 1 or log2 colostrum NE/peak lactation NE < -1). The expressions of 10 randomly selected miRNAs was analyzed through stem-loop real-time quantitative PCR (RT-qPCR). Their expression patterns were the same with Solexa sequencing results. Pathway analysis suggested that oestrogen, endocrine, adipocytokine, oxytocin and MAPK signalling pathways act on the development of mammary gland and milk secretion importantly. In addition, the miRNA-target-network showed that the bta-miR-574 could influence the development of mammary gland and lactation by leptin receptor (LEPR), which was in the adipocytokine signalling pathway. Chr5_3880_mature regulated mammary gland development and lactation through Serine/threonine-protein phosphatase (PPP1CA), which was in the oxytocin signalling pathway. CONCLUSIONS: Our finding suggested that the profiles of miRNAs were related to the physiological functions of mammary gland in the colostrum and peak lactation periods. The biological features of these miRNAs may help to clarify the molecular mechanisms of lactation and the development of caprine mammary gland.

Trans-Fatty Acid-Stimulated Mammary Gland Growth in Ovariectomized Mice is Fatty Acid Type and Isomer Specific.

We previously reported that the trans-18:2 fatty acid trans-10, cis-12 conjugated linoleic acid (t10,c12-CLA) stimulates mammary gland development independent of estrogen and its receptor. Given the negative consequences of dietary trans-fatty acids on various aspects of human health, we sought to establish whether other trans-fatty acids could similarly induce ovary-independent mammary gland growth in mice. Prepubertal BALB/cJ mice were ovariectomized at 21 days of age then were fed diets enriched with cis-9, trans-11 CLA (c9,t11-CLA), or mixtures of trans-18:1 fatty acids supplied by partially hydrogenated sunflower, safflower, or linseed oil. The resultant mammary phenotype was evaluated 3 weeks later and compared to the growth response elicited by t10,c12-CLA, or the defined control diet. Whereas partially hydrogenated safflower oil increased mammary gland weight, none of the partially hydrogenated vegetable oils promoted mammary ductal growth. Similarly, the c9,t11-CLA supplemented diet was without effect on mammary development. Taken together, our data emphasize a unique effect of t10,c12-CLA in stimulating estrogen-independent mammary gland growth manifest as increased mammary ductal area and elongation that was not recapitulated by c9,t11-CLA or the partially hydrogenated vegetable oil diets.

Paternal selenium deficiency but not supplementation during preconception alters mammary gland development and 7,12-dimethylbenz[a]anthracene-induced mammary carcinogenesis in female rat offspring.

Breast cancer is a global public health problem and accumulating evidence indicates early-life exposures as relevant factors in the disease risk determination. Recent studies have shown that paternal nutrition can influence offspring health including breast cancer risk. Selenium is a micronutrient with essential role in central aspects of embryogenesis, male fertility and cancer and that has been extensively studied as a chemopreventive agent in several breast cancer experimental models. Thus, we designed an animal study to evaluate whether paternal selenium deficiency or supplementation during preconception could affect the female offspring mammary gland development and breast cancer susceptibility. Male Sprague-Dawley rats were fed AIN93-G diet containing 0.15 ppm (control diet), 0.05 ppm (deficient diet) or 1 ppm (supplemented diet) of selenium for 9 weeks and mated with control female rats. Mammary carcinogenesis was induced with 7,12-dimethylbenz[a]anthracene (DMBA) in their female offspring. Paternal selenium deficiency increased the number of terminal end buds, epithelial elongation and cell proliferation in the mammary gland of the female rat offspring and these effects were associated with higher susceptibility to DMBA-induced mammary tumors (increased incidence and higher grade tumors). On the other hand, paternal selenium supplementation did not influence any of these parameters. These results highlight the importance of father's nutrition including selenium status as a relevant factor affecting daughter's breast cancer risk and paternal preconception as a potential developmental stage to start disease preventive strategies.

Exogenous genistein in late gestation: effects on fetal development and sow and piglet performance.

Due to their functional similarity to estradiol, phytoestrogens could prove to be beneficial in late gestating sows. The goal of this study was to determine the impact of providing the phytoestrogen genistein during late pregnancy on the performance of sows and their litters. In total, 56 gilts were equally divided into the two following groups on day 90 of gestation: (1) controls (CTL); and (2) two daily i.m. injections of 220 mg of genistein (GEN). Treatments were carried out until farrowing. Jugular blood samples were collected from 16 gilts/treatment on days 89 and 110 of gestation, and on days 3 and 21 of lactation. Milk samples were also obtained from those sows on day 3 of lactation. A male piglet from 16 CTL and 15 GEN litters was slaughtered at 24 h postpartum and a blood sample was obtained. The liver, heart and visceral organs were weighed and the semitendinosus (ST) muscle was collected and carcass composition was determined. The treatment increased (P0.1) on weight or backfat loss of sows during lactation, milk composition or weights of piglets. The pre-weaning mortality rate of piglets was very low (0.1). However, carcasses from GEN litters contained more fat than those from CTL litters (9.63% v. 8.34%, P0.1). In conclusion, injecting gilts with 440 mg/day of genistein in late gestation increased IGF1 concentrations in gilts and carcass fat in neonatal piglets, but had minimal effect on muscle development of piglets at birth and on the performance of lactating sows and their litters.

Estrogens in the wrong place at the wrong time: Fetal BPA exposure and mammary cancer.

Iatrogenic gestational exposure to diethylstilbestrol (DES) induced alterations of the genital tract and predisposed individuals to develop clear cell carcinoma of the vagina as well as breast cancer later in life. Gestational exposure of rodents to a related compound, the xenoestrogen bisphenol-A (BPA) increases the propensity to develop mammary cancer during adulthood, long after cessation of exposure. Exposure to BPA during gestation induces morphological alterations in both the stroma and the epithelium of the fetal mammary gland at 18 days of age. We postulate that the primary target of BPA is the fetal stroma, the only mammary tissue expressing estrogen receptors during fetal life. BPA would then alter the reciprocal stroma-epithelial interactions that mediate mammogenesis. In addition to this direct effect on the mammary gland, BPA is postulated to affect the hypothalamus and thus in turn affect the regulation of mammotropic hormones at puberty and beyond.

Effects of the plant extract silymarin on prolactin concentrations, mammary gland development, and oxidative stress in gestating gilts.

The impacts of supplementing the diet of gestating gilts twice daily with 4 g of the plant extract silymarin on circulating hormonal concentrations, oxidative status, mammary development, and mammary gene expression at the end of gestation were determined. Gilts were fed conventional diets during gestation and on d 90 they were assigned as controls (CTL; n = 16) or treated (TRT; n = 17) animals. Treatment consisted of providing 4 g of silymarin twice daily until d 110, at which time all gilts were slaughtered to collect mammary tissue for compositional analyses and measures of gene expression and oxidative status, and liver and corpora lutea for measures of oxidative stress variables. Blood samples for hormonal assays and evaluation of oxidative stress biomarkers were obtained on d 89, 94, and 109 of gestation. Silymarin increased (P = 0.05) circulating concentrations of prolactin over all samples in the repeated in time analysis. In separate analyses for each sampling time, prolactin concentrations in TRT gilts tended (P < 0.10) to be greater than in CTL gilts on d 94 of gestation. Repeated in time analysis also revealed that silymarin reduced (P ≤ 0.05) plasmatic accumulation of biomarkers of oxidative damage to protein (protein carbonyls) between d 89 and 109. There was no effect (P > 0.10) of treatment on progesterone, estradiol, leptin, or 8-hydroxy-2'-deoxyguanosine concentrations. Percent fat in mammary parenchyma was greater (P ≤ 0.05), percent protein was lesser (P ≤ 0.05), and concentrations of both RNA (P ≤ 0.01) and DNA (P < 0.05) were lesser in TRT than CTL gilts. Mammary parenchyma from TRT gilts had lower (P ≤ 0.05) mRNA abundance for STAT5A and leptin and tended to have lower (P ≤ 0.10) abundance for STAT5B than CTL gilts. Silymarin reduced (P ≤ 0.001) protein carbonyls concentrations in liver of TRT gilts. No effect of treatment was observed on antioxidant gene expression and enzymatic activities in liver samples while total superoxide dismutase activity tended to be higher (P ≤ 0.10) in the corpora lutea of TRT animals when compared with CTL. This is the first demonstration that, in female pigs, silymarin can increase prolactin concentrations and protect against oxidative stress, yet the increase in prolactin was not enough to have beneficial effects on mammary gland development in late gestation.

Postweaning dietary genistein exposure advances puberty without significantly affecting early pregnancy in C57BL/6J female mice.

An epidemiological study indicates higher plasma level of genistein in girls with earlier puberty. This study tests the hypothesis in C57BL/6J mice that postweaning (peripubertal) dietary genistein exposure could result in earlier puberty in females assessed by vaginal opening, estrous cyclicity, corpus luteum and mammary gland development. Newly weaned female mice were fed with 0, 5, 100, or 500 ppm genistein diets. Decreased age at vaginal opening, increased length on estrus stage, and accelerated mammary gland development were detected in 100 and 500 ppm genistein-treated groups. Increased presence of corpus luteum was found in 5 ppm genistein-treated group at 6 weeks old only. Increased expression of epithelial-specific genes but not that of ERα or ERß was detected in 500 ppm genistein-treated mammary glands at 5 weeks old. No significant adverse effect on embryo implantation was observed. These data demonstrate causal effect of dietary genistein on earlier puberty in female mice.

Feeding soy protein isolate and treatment with estradiol have different effects on mammary gland morphology and gene expression in weanling male and female rats.

Isoflavones are phytochemical components of soy diets that bind weakly to estrogen receptors (ERs). To study potential estrogen-like actions of soy in the mammary gland during early development, we fed weanling male and female Sprague-Dawley rats a semipurified diet with casein as the sole protein source from postnatal day 21 to 33, the same diet substituting soy protein isolate (SPI) for casein, or the casein diet supplemented with estradiol (E2) at 10 µg/kg/day. In contrast to E2, the SPI diet induced no significant change in mammary morphology. In males, there were 34 genes for which expression was changed ≥2-fold in the SPI group vs. 509 changed significantly by E2, and 8 vs. 174 genes in females. Nearly half of SPI-responsive genes in males were also E2 responsive, including adipogenic genes. Serum insulin was found to be decreased by the SPI diet in males. SPI and E2 both downregulated the expression of ERα (Esr1) in males and females, and ERß (Esr2) only in males. Chromatin immunoprecipitation revealed an increased binding of ERα to the promoter of the progesterone receptor (Pgr) and Esr1 in both SPI- and E2-treated males compared with the casein group but differential recruitment of ERß. ER promoter binding did not correlate with differences in Pgr mRNA expression. This suggests that SPI fails to recruit appropriate co-activators at E2-inducible genes. Our results indicate that SPI behaves like a selective estrogen receptor modulator rather than a weak estrogen in the developing mammary gland.

Endocrine disrupting effects in rats perinatally exposed to a dietary relevant mixture of phytoestrogens.

Dietary phytoestrogens may prevent certain human diseases, but endocrine activity has been reported in animal studies. Sprague-Dawley rats were exposed perinatally to a 1-, 10- or 100-fold "high human dietary intake" mixture of 12 phytoestrogens consisting of mainly the lignan secoisolarici resinol and the isoflavones genistein and daidzein. This mixture induced persistent adverse effects, as adult male mammary glands showed hypertrophic growth. A reduced anogenital distance in newborn males indicated an anti-androgenic mode of action. Testosterone levels, testis and prostate weights, and expression of selected genes in testis and prostate were unaffected. Decreased serum estradiol was seen in genistein-exposed dams. This study indicated adverse effects at high intake levels in rats, but does not provide evidence for risk of phytoestrogen-mediated endocrine disruption at normal human dietary consumption levels. Further studies are warranted to increase the knowledge upon which risk assessment on dietary phytoestrogen exposure during pregnancy and infancy is based.

Endocrine disruptors and the breast: early life effects and later life disease.

Breast cancer risk has both heritable and environment/lifestyle components. The heritable component is a small contribution (5-27 %), leaving the majority of risk to environment (e.g., applied chemicals, food residues, occupational hazards, pharmaceuticals, stress) and lifestyle (e.g., physical activity, cosmetics, water source, alcohol, smoking). However, these factors are not well-defined, primarily due to the enormous number of factors to be considered. In both humans and rodent models, environmental factors that act as endocrine disrupting compounds (EDCs) have been shown to disrupt normal mammary development and lead to adverse lifelong consequences, especially when exposures occur during early life. EDCs can act directly or indirectly on mammary tissue to increase sensitivity to chemical carcinogens or enhance development of hyperplasia, beaded ducts, or tumors. Protective effects have also been reported. The mechanisms for these changes are not well understood. Environmental agents may also act as carcinogens in adult rodent models, directly causing or promoting tumor development, typically in more than one organ. Many of the environmental agents that act as EDCs and are known to affect the breast are discussed. Understanding the mechanism(s) of action for these compounds will be critical to prevent their effects on the breast in the future.

Diet-induced metabolic change induces estrogen-independent allometric mammary growth.

Lifetime breast cancer risk reflects an unresolved combination of early life factors including diet, body mass index, metabolic syndrome, obesity, and age at first menses. In parallel, the onset of allometric growth by the mammary glands around puberty is widely held to be estrogen (E)-dependent. Here we report that several physiological changes associated with metabolic syndrome in response to a diet supplemented with the trans-10, cis-12 isomer of conjugated linoleic acid lead to ovary-independent allometric growth of the mammary ducts. The E-independence of this diet-induced growth was highlighted by the fact that it occurred both in male mice and with pharmacological inhibition of either E receptor function or E biosynthesis. Reversal of the metabolic phenotype with the peroxisome proliferator-activated receptor-γ agonist rosiglitazone abrogated diet-induced mammary growth. A role for hyperinsulinemia and increased insulin-like growth factor-I receptor (IGF-IR) expression during mammary growth induced by the trans-10, cis-12 isomer of conjugated linoleic acid was confirmed by its reversal upon pharmacological inhibition of IGF-IR function. Diet-stimulated ductal growth also increased mammary tumorigenesis in ovariectomized polyomavirus middle T-antigen mice. Our data demonstrate that diet-induced metabolic dysregulation, independently of ovarian function, stimulates allometric growth within the mammary glands via an IGF-IR-dependent mechanism.

Hormonally active doses of isoflavone aglycones promote mammary and endometrial carcinogenesis and alter the molecular tumor environment in Donryu rats.

Our research is focused on modifying effects of an isoflavone aglycones (IAs)-rich extract at a hormonally active dose of 150 mg/kg body weight/day on mammary and endometrial carcinogenesis in female Donryu rats. IA administered for 2 weeks in a phytoestrogen-low diet exerted estrogenic activity and induced cell proliferation in the uterus of ovariectomized rats. Furthermore, administration for 4 weeks resulted in elevation of cell proliferation in the mammary glands of 7,12-dimethylbenz[a]anthracene (DMBA)-treated animals. Forty weeks of postpubertal administration of IA to 5-week-old rats after initiation of mammary and endometrial carcinogenesis with DMBA and N-ethyl-N'-nitro-N-nitrosoguanidine (ENNG) caused significant increase of incidence and multiplicity of mammary adenocarcinoma, multiplicities of endometrial atypical hyperplasia, adenomatous polyps, and an increased trend of uterine adenocarcinomas. Liquid chromatography with tandem mass spectrometry and immunohistochemical analyses revealed significant elevation of tumorigenesis-related proteins such as S100 calcium-binding protein A8, kininogen 1, and annexins 1 and 2 in mammary adenocarcinomas and cadherin EGF LAG seven-pass G-type receptor 2, DEAD box polypeptide 1, and cysteine- and glycine-rich protein 1 in uterine proliferative lesions of IA-treated animals. Those changes are likely to be related to modulation of estrogen receptor (ER), AP1, nuclear factor-kappa B, and actin signaling pathways. Our results indicate that the postpubertal exposure of Donryu rats to IA at an estrogenic dose results in promotion of mammary and uterine carcinogenesis induced by DMBA and ENNG, which might be related to the activation of ER-dependent signaling and alteration of the molecular tumor environment in the mammary gland and endometrium.

Recent insights into the effect of natural and environmental estrogens on mammary development and carcinogenesis.

The present work reviews recent findings related to the action of steroidal (physiological) estrogens on normal mammary gland development and carcinogenesis, as well as effects of related environmental mediators (phyto- and xeno-estrogens), the role of which remains controversial. Orchestration by estrogen receptors (i.e. ERα and ERß) and coregulators of growth, apoptosis and differentiation of epithelial cells, directed our analysis. The bidirectional coordination between epithelium and stroma in parallel with maintenance of stemness are also investigated. The relevance of nuclear and extranuclear localization of ERs and other eventual estrogen binding sites, mediating differential actions in regard to these various topics, is critically addressed to delineate the importance of direct and indirect activation procedures and delicate feedback loops (ligand-induced or/and cross-talk activation, respectively). The inclusion of the outlined regulatory concepts in drug design programs for the prevention and treatment of breast cancer may have potent effects.

Abnormal peripubertal development of the rat mammary gland following exposure in utero and during lactation to a mixture of genistein and the food contaminant vinclozolin.

The impact of early exposure to endocrine disruptor mixtures on mammary gland development is poorly known. Here, we identify the effects of a conception to weaning exposure of rats to the phytoestrogen genistein (G) and/or the antiandrogen vinclozolin (V) at 1mg/kg-d, alone or in association. Using several approaches, we found that G- and GV-exposed rats displayed significantly greater epithelial branching and proliferation, wider terminal end buds than controls at PND35, as well as ductal hyperplasia and periductal fibrosis. Focal branching defects were present in V-exposed rats. An increased ER and AR expression was observed in G- and GV- as compared to V-exposed rats at PND35. Surprisingly, a significant number of GV- and to a lesser extent, V-exposed animals displayed abnormal hyperplasic alveolar structures at PND50. Thus, gestational and lactational exposure to low doses of genistein plus vinclozolin may seriously affect peripubertal development of the rat mammary gland.