RESUMEN
OBJECTIVE: Increased adolescent obesity rates in the United States are a significant public health concern. Obesity or increased adiposity during puberty in girls, an important period of breast development and a window of exposure sensitivity, may influence breast development and cancer risk. The purpose of this study was to investigate the impact of a high fat diet (HFD) on mammary gland development in obesity-susceptible C57BL/6 and obesity-resistant BALB/c mice. DESIGN: Pubertal or adult C57BL/6 and BALB/c mice were fed an HFD or control diet (CD) from 3 to 7 weeks of age or from 10 to 14 weeks of age, respectively. The effects of HFD diet on body weight, adiposity, mammary gland development, and mammary gland response to estrogen were evaluated. RESULTS: Pubertal C57BL/6 mice fed the HFD had a significant increase in body weight and adiposity, and this was accompanied by stunted mammary duct elongation and reduced mammary epithelial cell proliferation. Ovariectomy and estrogen (17-ß-estradiol, E) treatment of pubertal HFD-fed C57BL/6 mice showed decreased mammary gland stimulation by E. Amphiregulin, a downstream mediator of pubertal E action, was reduced in mammary glands of HFD-fed C57BL/6 mice. Weight loss and reduced adiposity initiated by switching C57BL/6 mice from HFD to CD restored ductal elongation. Pubertal BALB/c mice fed the HFD did not exhibit a significant increase in body weight or adiposity; HFD caused increased mammary epithelial cell proliferation and had no effect on response to E. HFD had no effect on body weight or the mammary glands of adult mice. CONCLUSIONS: HFD during puberty had a profound strain-specific effect on murine mammary gland development. Obesity and increased adiposity were associated with reduced responsiveness to estrogen and stunted ductal growth. Importantly, the effect of diet and adiposity on the mammary gland was specific to the pubertal period of development.
Asunto(s)
Grasas de la Dieta/administración & dosificación , Estrógenos/fisiología , Glándulas Mamarias Animales/fisiología , Obesidad/fisiopatología , Maduración Sexual/fisiología , Animales , Estrógenos/metabolismo , Femenino , Glándulas Mamarias Animales/crecimiento & desarrollo , Glándulas Mamarias Animales/metabolismo , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Obesidad/metabolismoRESUMEN
Estrogen receptor positive (ER+), estrogen (E) responsive MCF-7 breast cancer cells cultured on the extracellular matrix (ECM) protein laminin (LM), exhibit significantly reduced E-induced proliferation compared with cells cultured on collagen I (Col I) that is not due to a loss of ER. Based on reported differences in integrin-activated pathways on Col I vs. LM, we investigated the potential role of Rac1/c-jun-N-terminal kinase (JNK) activation and downstream regulation of cyclin D1 by E on Col I vs.LM. E-induced proliferation was increased on LM in MCF-7 cells expressing constitutively active Rac1 (CA Rac1) and decreased in dominant negative Rac1-(DN Rac1) expressing cells on Col I. siRNA knockdown established the specificity and requirement for Rac1 activation for E-induced regulation of cyclin D1. More robust c-Jun activation occurred on Col I than on LM and E-induced proliferation was abolished after treatment with a JNK inhibitor. These results provide evidence that Rac1/JNK/c-Jun activation promotes E-induced proliferation on Col I and reduced Rac1/JNK/c-Jun activation on LM contributes significantly to reduced E-induced proliferation in MCF-7 cells on LM. These results identify a novel role for extracellular matrix (ECM)-integrin regulation of Rac1-JNK pathway in E-regulated proliferation in ER+ breast cancer cells. These findings suggest that tumor stromal environment, i.e., ECM composition, may contribute to loss of E regulation in ER+ breast cancers. Defining molecular markers for early identification of ER+ tumors that are ER+ but antiestrogen resistant would allow the design and use of alternative therapies to inhibit tumor growth and improve survival.
Asunto(s)
Estrógenos/metabolismo , Matriz Extracelular/metabolismo , Proteína de Unión al GTP rac1/metabolismo , Línea Celular Tumoral , Proliferación Celular , Colágeno/metabolismo , Ciclina D1/metabolismo , Humanos , Laminina/metabolismo , Modelos Biológicos , ARN Interferente Pequeño/metabolismo , Receptores de Estrógenos/metabolismo , Transducción de Señal , Proteína de Unión al GTP cdc42/metabolismo , Proteínas de Unión al GTP rac/metabolismoRESUMEN
Despite the critical importance of ovarian steroids in the treatment of breast cancer, little is known about the acquisition or loss of estrogen and progesterone responsiveness in either the normal or neoplastic mammary gland. This review focuses on the interactions among mammary stroma-derived extracellular matrix (ECM) proteins, integrins and ovarian hormone-dependent proliferation in normal and neoplastic mammary cells both in vivo and in vitro. In vitro studies show that fibronectin is required for progesterone-induced proliferation of normal mammary epithelial cells and that specific ECM proteins also regulate interactions between growth factors and ovarian hormones. Studies with human breast cancer cell lines have shown that laminin inhibits estrogen-induced proliferation and estrogen-response-element-mediated transcription in vitro and also inhibits estrogen-induced proliferation in vivo. Reciprocally, ovarian steroids regulate the expression of ECM proteins and their cellular receptors, integrins, during mammary gland development in vivo. The fibronectin-specific integrin, alpha5beta1 is regulated by ovarian steroids and its expression is positively correlated with developmental stages of peak proliferation. These studies suggest that the coordinated regulation of ovarian hormone responsiveness and ECM/integrin expression may be critical to normal mammary gland development and breast cancer growth and progression.
Asunto(s)
Mama/metabolismo , Estrógenos/farmacología , Proteínas de la Matriz Extracelular/farmacología , Glándulas Mamarias Animales/metabolismo , Progesterona/farmacología , Animales , Mama/citología , Neoplasias de la Mama/metabolismo , Femenino , Fibronectinas/metabolismo , Humanos , Glándulas Mamarias Animales/citología , Ratones , Neoplasias Hormono-Dependientes/metabolismoRESUMEN
Extracellular matrix (ECM) proteins have been shown to regulate mammary epithelial cell proliferation, differentiation, and apoptosis in vitro. However, little is known about the hormonal regulation and functional role of ECM proteins and integrins during mammary gland development in vivo. We examined the temporal and spatial localization and hormone regulation of collagen I, collagen IV, laminin, and fibronectin. Among these ECM proteins only fibronectin changed appreciably. Fibronectin levels increased 3-fold between the onset of puberty and sexual maturity, remaining high during pregnancy and lactation. This increase occurred specifically in the epithelial basement membrane. Fibronectin was decreased 70% by ovariectomy and increased 1.5- and 2-fold by estrogen or estrogen plus progesterone treatment, respectively. The fibronectin-specific integrin, alpha(5)beta(1), was localized in myoepithelial cells; it increased 2.2-fold between puberty and sexual maturity and decreased in late pregnancy and lactation. The basal localization of alpha(5)beta(1) was notably increased in pubertal and adult virgin mice. alpha(5)beta(1) concentrations decreased 40-50% after ovariectomy in pubertal and adult mice, which was reversed by estrogen plus progesterone treatment in adult mice. The high basal expression of alpha(5)beta(1) during active proliferation and the low expression in nonproliferating and lactating glands indicate that fibronectin signaling may be required for hormone-dependent proliferation in the mammary gland.
Asunto(s)
Envejecimiento/metabolismo , Estrógenos/fisiología , Fibronectinas/metabolismo , Glándulas Mamarias Animales/metabolismo , Progesterona/fisiología , Receptores de Fibronectina/metabolismo , Animales , Western Blotting , Proteínas de la Matriz Extracelular/metabolismo , Femenino , Inmunohistoquímica , Lactancia/fisiología , Glándulas Mamarias Animales/citología , Ratones , Ratones Endogámicos BALB C , Ovariectomía , Embarazo , Valores de Referencia , Distribución TisularRESUMEN
Hormone replacement therapy (HRT) with ovarian hormones is an important therapeutic modality for postmenopausal women. However, a negative side effect of HRT is an increased risk of breast cancer. Surgical induction of menopause by ovariectomy (OVX) in mice is an experimental model that may provide insights into the effects of hormone replacement therapy on the human breast. We have developed a mouse model of early and late postmenopausal states to investigate the effects of HRT on the normal mammary gland. The purpose of this study was to determine if HRT-induced proliferation was due to the direct action of the hormones on the mammary gland, or mediated systemically by hormones or growth factors produced elsewhere in the body. Estrogen (E) or E plus the synthetic progestin, R5020, were implanted directly into the mammary glands of early (1 week post OVX) and late (5 week post OVX) postmenopausal mice instead of administration by injection. We report that responses of early and late postmenopausal mice to implanted hormones were the same as those observed previously with systemically administered hormones. Implanted E conferred an enhanced proliferative response in the late postmenopausal gland characterized morphologically by enlarged duct ends. E+R5020 implants induced similar degrees of cell proliferation in both postmenopausal states but the morphological responses differed. Ductal sidebranching was observed in early postmenopausal mice, whereas duct end enlargement was observed in late postmenopausal mice. The differences in morphological response to E+R5020 in 5 week post OVX were associated with an inability of E to induce progesterone receptors (PR) in the late postmenopausal gland. The responses of the late postmenopausal glands to E and E+P were very similar to that observed previously in immature pubertal glands in ovary-intact mice. In pubertal mice, PR cannot be induced by E unless the mammary gland is pre-treated with EGF-containing implants. Similarly, herein pre-treatment of the late postmenopausal mammary gland with EGF-containing implants restored PR induction by E. Thus, EGF may determine the sensitivity of the mammary gland to E and E+P in late postmenopause and at puberty.
Asunto(s)
Mama/efectos de los fármacos , Estrógenos/farmacología , Posmenopausia/efectos de los fármacos , Congéneres de la Progesterona/farmacología , Promegestona/farmacología , Animales , Mama/citología , Mama/fisiología , División Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Implantes de Medicamentos , Factor de Crecimiento Epidérmico/farmacología , Estrógenos/administración & dosificación , Femenino , Ratones , Ratones Endogámicos BALB C , Modelos Animales , Ovariectomía , Congéneres de la Progesterona/administración & dosificación , Promegestona/administración & dosificación , Receptores de Progesterona/metabolismo , Factores de TiempoRESUMEN
OBJECTIVE: The aim of the study was to analyze the proliferative response of the normal mammary gland to combination hormone replacement therapy with estrogen and progesterone in a murine model of early versus late postmenopausal states. STUDY DESIGN: Ovariectomized mice were injected daily for up to 56 days with estrogen plus progesterone, starting at either 1 or 5 weeks after ovariectomy to simulate early and late menopausal periods, respectively. At various times after treatment, proliferation was analyzed by deoxyribonucleic acid histoautoradiography and whole-mount preparations. The induction of progesterone receptor by estrogen was also analyzed. To distinguish between estrogen- and progesterone-specific responses, we tested the effects of the antiprogesterone mifepristone (RU 486) and the antiestrogen ICI 182,780. RESULTS: The acute response to estrogen-progesterone therapy in the early postmenopausal period resulted in duct-end enlargement, ductal side branching, alveolar bud formation, and a 100-fold increase in epithelial cell proliferation. This was caused by the dominant effect of progesterone acting through the progesterone receptor. In the late postmenopausal period the acute response produced only duct-end enlargement; the 100-fold increase in epithelial cell proliferation resulted from the dominant effect of estrogen. After long-term treatment, both early and late postmenopausal glands exhibited similar morphologic features and a 9-fold higher steady-state proliferation rate than was found in control-treated groups. CONCLUSIONS: Starting combined estrogen and progesterone hormone replacement therapy in either early or late postmenopause produced a persistent, steady-state 9-fold increase in epithelial cell proliferation, which could be a contributing factor to increased breast cancer risk. The acute response in the late postmenopausal period mimics the hormonal response of the pubertal mammary gland, which in rodents is the stage most susceptible to carcinogen-induced mammary tumorigenesis. These observations raise questions about increased susceptibility of the late postmenopausal gland to carcinogenesis and a role for hormone replacement therapy in the promotion of tumorigenesis.
Asunto(s)
Estradiol/análogos & derivados , Estradiol/farmacología , Terapia de Reemplazo de Hormonas , Glándulas Mamarias Animales/efectos de los fármacos , Progesterona/farmacología , Animales , División Celular/efectos de los fármacos , Antagonistas de Estrógenos/farmacología , Femenino , Fulvestrant , Antagonistas de Hormonas/farmacología , Procesamiento de Imagen Asistido por Computador , Inmunohistoquímica , Glándulas Mamarias Animales/citología , Glándulas Mamarias Animales/diagnóstico por imagen , Glándulas Mamarias Animales/fisiología , Ratones , Ratones Endogámicos BALB C , Mifepristona/farmacología , Modelos Animales , Ovariectomía , Posmenopausia/fisiología , Cintigrafía , Receptores de Progesterona/metabolismo , Timidina/administración & dosificación , Tritio/administración & dosificaciónRESUMEN
The purpose of the present study was to investigate the role of extracellular matrix proteins (ECMs; collagens I and IV, fibronectin, and laminin) in modulating proliferative responses of normal mammary epithelial cells in serum-free culture to epidermal growth factor (EGF) and insulin-like growth factor I (IGF-I). As EGF and IGF-I can alter steroid responses, the interactions among growth factors, estrogen, and R5020 were also investigated. We report the novel finding that all ECMs tested, but not a nonspecific attachment factor, poly-L-lysine (PL), promoted a highly synergistic proliferative response to EGF plus IGF-I. EGF receptors were significantly increased with culture time on all ECMs, but not on PL. IGF receptor expression was significantly 2- to 4-fold higher on all ECMs compared with PL. EGF decreased IGF-binding protein-2 (IGFBP-2) and IGFBP-3 by more than 50% in the presence of IGF-I on PL or collagen I. These results indicate that ECM-specific IGF-I/EGF synergism occurs in response to ECM up-regulation of growth factor receptors and EGF down-regulation of inhibitory IGFBPs. Growth factors did not synergize with estrogen and/or R5020. Instead, estrogen plus R5020 decreased EGF-plus IGF-I-induced proliferation in an ECM-dependent manner. These studies demonstrate that proliferation of normal mammary epithelial cells involves complex interactions among steroids, growth factors, binding proteins, and ECMs.
Asunto(s)
Glándulas Mamarias Animales/citología , Animales , División Celular/efectos de los fármacos , Células Cultivadas , Sinergismo Farmacológico , Factor de Crecimiento Epidérmico/farmacología , Células Epiteliales/citología , Células Epiteliales/metabolismo , Matriz Extracelular/metabolismo , Proteínas de la Matriz Extracelular/farmacología , Femenino , Hormonas/metabolismo , Hormonas/farmacología , Proteínas de Unión a Factor de Crecimiento Similar a la Insulina/metabolismo , Factor I del Crecimiento Similar a la Insulina/farmacología , Glándulas Mamarias Animales/metabolismo , Ratones , Ovario/metabolismo , Receptores de Estrógenos/metabolismo , Receptores de Progesterona/metabolismoRESUMEN
Breast tumors that lack estrogen responsiveness have a poor prognosis. Despite the critical importance to breast cancer treatment, little is known about the loss of estrogen responsiveness and the development of antiestrogen resistance. We have examined the regulation of estrogen-induced proliferation, estrogen regulation of progesterone receptor (PR) expression, and estrogen signaling pathways in estrogen receptor (ER) positive (MCF-7 and T47D) breast cancer cell lines by specific extracellular matrix proteins (ECM) under serum-free conditions. Estrogen, supplemented with submaximal concentrations of insulin-like growth factor I (IGF-I) and epidermal growth factor (EGF), stimulated DNA synthesis of MCF-7 cells 7- to 10-fold and T47D cells 2-fold on collagen I or fibronectin. However, estrogen-induced proliferation was greatly reduced on laminin. In contrast, IGF-I or EGF, alone, stimulated proliferation of MCF-7 and T47D cells on all ECM. Thus, ER+ breast cancer cells were not refractory to mitogens when cultured on laminin. Similarly, estrogen induction of PR occurred on fibronectin or collagen I, but not on laminin. While ER content was similar on all ECM, estrogen stimulation of estrogen response element (ERE)-luciferase activity was significantly lower in MCF-7 cells cultured on laminin. Therefore, changes in ECM composition that occur in breast cancer may alter estrogen-responsiveness and the effectiveness of antiestrogen therapies in ER+ breast cancer cells.
Asunto(s)
Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Antagonistas de Estrógenos/farmacología , Estrógenos/farmacología , Laminina/farmacología , Adhesión Celular , División Celular/efectos de los fármacos , Medios de Cultivo , Medio de Cultivo Libre de Suero , ADN/biosíntesis , Factor de Crecimiento Epidérmico/farmacología , Matriz Extracelular/metabolismo , Proteínas de la Matriz Extracelular/metabolismo , Humanos , Factor I del Crecimiento Similar a la Insulina/farmacología , Receptores de Estrógenos/análisis , Receptores de Progesterona/metabolismo , Elementos de Respuesta , Transducción de Señal/efectos de los fármacos , Células Tumorales CultivadasRESUMEN
The relative effects of postmenopausal hormone replacement therapy (HRT) with estrogen alone vs. estrogen+progestin on breast cell proliferation and on breast cancer risk are controversial. A cross-sectional observational study was carried out to examine the proliferative effects of HRT with estrogen or estrogen plus the progestin, medroxyprogesterone acetate, in breast tissue of postmenopausal women. Benign breast biopsies from 86 postmenopausal women were analyzed with antiproliferating cell nuclear antigen (anti-PCNA) and Ki67 antibodies to measure relative levels of cell proliferation. Epithelial density and estrogen and progesterone receptor status were also determined. The women were categorized either as users of: 1) estrogen (E) alone; 2) estrogen+medroxyprogesterone acetate (E+P); or 3) no HRT. Compared with no HRT, the breast epithelium of women who had received either E+P or E alone had significantly higher PCNA proliferation indices, and treatment with E+P had a significantly higher index (PCNA and Ki67) than treatment with E alone. Breast epithelial density was significantly greater in postmenopausal women treated with E and E+P, compared with no HRT. Thus, the present study shows that postmenopausal HRT with E+P was associated with greater breast epithelial cell proliferation and breast epithelial cell density than E alone or no HRT. Furthermore, with E+P, breast proliferation was localized to the terminal duct-lobular unit of the breast, which is the site of development of most breast cancers. Further studies are needed to assess the possible association between the mitogenic activity of progestins and breast cancer risk.
Asunto(s)
Mama/citología , División Celular , Terapia de Reemplazo de Estrógeno , Acetato de Medroxiprogesterona/uso terapéutico , Posmenopausia , Adulto , Anciano , Anciano de 80 o más Años , Biopsia , Mama/química , Neoplasias de la Mama/inducido químicamente , Estudios Transversales , Células Epiteliales/citología , Terapia de Reemplazo de Estrógeno/efectos adversos , Femenino , Humanos , Antígeno Ki-67/análisis , Acetato de Medroxiprogesterona/efectos adversos , Persona de Mediana Edad , Antígeno Nuclear de Célula en Proliferación/análisis , Receptores de Estrógenos/análisis , Receptores de Progesterona/análisis , Factores de RiesgoRESUMEN
Hormone replacement therapy (HRT) with estrogen alleviates menopausal symptoms and is effective in reducing osteoporosis and cardiovascular disease when taken in early postmenopause. Older, late postmenopausal women who never previously received HRT are also believed to benefit from estrogen treatment. On the other hand, increased lifetime exposure of the mammary gland to estrogen may increase the risk of breast cancer. The development of suitable experimental animal model systems can advance our understanding of the effects of estrogen and the timing of HRT on the postmenopausal breast. Toward this end, early and late postmenopausal states were induced in mice by short vs. long term ovariectomy (1 vs. 5 weeks), and the effects of 17beta-estradiol (E) on mammary gland morphology, cell proliferation, and progesterone receptor (PR) levels were investigated. We report that in late postmenopausal mice, E caused a pronounced enlargement of duct ends and 6.5- and 4-fold greater mitogenic responses in the duct end epithelium and adjacent stromal cells, respectively, compared with the response in early postmenopausal mice. Furthermore, after long term, daily treatment with E, steady state levels of proliferation remained 2-fold higher than those of similarly treated, early postmenopausal mice. E failed to increase mammary PR levels in late postmenopausal, but not in early postmenopausal mice. Stimulation of duct ends by E and lack of PR inducibility are characteristics of the immature pubertal mammary gland and indicate that the late postmenopausal mammary gland resembled the immature state. In contrast, minimal E-induced proliferation and increased PR inducibility, characteristics of the adult, sexually mature mammary gland, were retained in early postmenopausal mice. The lack of difference in the numbers of estrogen receptor-positive epithelial or stromal cells or in estrogen receptor cellular concentration after short vs. long term ovariectomy indicates that the observed greater efficacy of E is mediated at a step beyond receptor-ligand binding. This mouse model of experimentally induced early vs. late postmenopausal states should prove useful in better understanding alterations in hormone responsiveness and their implications for timing of HRT on the human breast.
Asunto(s)
Terapia de Reemplazo de Estrógeno , Estrógenos/farmacología , Glándulas Mamarias Animales/efectos de los fármacos , Menopausia , Factores de Edad , Animales , División Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Glándulas Mamarias Animales/patología , Ratones , Ratones Endogámicos BALB C , Ovariectomía , Receptores de Progesterona/análisis , Factores de TiempoRESUMEN
Genistein, found in soy products, is a phytochemical with several biological activities. In the current study, our research focused on the estrogenic and proliferation-inducing activity of genistein. We have demonstrated that genistein enhanced the proliferation of estrogen-dependent human breast cancer (MCF-7) cells in vitro at concentrations as low as 10 nM, with a concentration of 100 nM achieving proliferative effects similar to those of 1 nM estradiol. Expression of the estrogen-responsive gene pS2 was also induced in MCF-7 cells in response to treatment with a concentration of genistein as low as 1 microM. At higher concentrations (above 20 microM), genistein inhibits MCF-7 cell growth. In vivo, we have shown that dietary treatment with genistein (750 ppm) for 5 days enhanced mammary gland growth in 28-day-old ovariectomized athymic mice, indicating that genistein acts as an estrogen in normal mammary tissue. To evaluate whether the estrogenic effects observed in vitro with MCF-7 cells could be reproduced in vivo, MCF-7 cells were implanted s.c. in ovariectomized athymic mice, and the growth of the estrogen-dependent tumors was measured weekly. Negative control animals received the American Institute of Nutrition (AIN)-93G diet, the positive control group received a new s.c. estradiol (2 mg) pellet plus the AIN-93G diet, and the third group received genistein at 750 ppm in the AIN-93G diet. Tumors were larger in the genistein (750 ppm)-treated group than they were in the negative control group, demonstrating that dietary genistein was able to enhance the growth of MCF-7 cell tumors in vivo. Increased uterine weights were also observed in the genistein-treated groups. In summary, genistein can act as an estrogen agonist in vivo and in vitro, resulting in the proliferation of cultured human breast cancer cells (MCF-7) and the induction of pS2 gene expression. Here we present new information that dietary genistein stimulates mammary gland growth and enhances the growth of MCF-7 cell tumors in ovariectomized athymic mice.
Asunto(s)
Neoplasias de la Mama/patología , Estrógenos/farmacología , Genisteína/farmacología , Receptores de Estrógenos/análisis , Animales , Neoplasias de la Mama/química , División Celular/efectos de los fármacos , Femenino , Genisteína/sangre , Humanos , Glándulas Mamarias Animales/efectos de los fármacos , Ratones , Ratones Desnudos , Ovariectomía , Proteínas/genética , ARN Mensajero/análisis , Factor Trefoil-1 , Células Tumorales Cultivadas , Proteínas Supresoras de TumorRESUMEN
The purpose of the present studies was to investigate the role of epidermal growth factor (EGF) in the acquisition of estrogen (E) and progestin (P) responsiveness in the mouse mammary gland in vivo. Using the Elvax 40P implant technique to introduce bioactive molecules directly into the mammary gland to produce a localized effect, we have made the novel observation that EGF implanted into glands of pubertal mice followed by E treatment resulted in the precocious acquisition of E-inducible progesterone receptors (PR). In sexually mature mice, EGF implants alone were able to increase PR. A neutralizing antibody specific for EGF blocked E-dependent stimulation of end-bud development and PR induction. Furthermore, the antiestrogen ICI 182,780 blocked the EGF-induced stimulation end-buds and PR induction, indicating that these EGF effects are mediated via estrogen receptors (ER). Immunohistochemical analysis showed that the endogenous EGF content of mammary glands of mature mice was higher than pubertal mice, that E implants caused a localized increase in mammary gland EGF content in both pubertal and mature mice, and that in mature mice E caused an increase in stromal cell EGF content. We have previously shown that the acquisition of E-inducible PR can be modulated by mammary stroma, and the present results indicate that mammary stroma could modulate hormonal responsiveness through control of local growth factor concentration. Taken together, these results provide evidence that E-dependent responses of mouse mammary gland in vivo, such as end-bud proliferation and PR regulation, may be mediated by EGF through an ER-dependent mechanism.
Asunto(s)
Factor de Crecimiento Epidérmico/metabolismo , Estrógenos/farmacología , Glándulas Mamarias Animales/metabolismo , Progestinas/farmacología , Animales , Células Cultivadas , Femenino , Inmunohistoquímica , Ratones , Ovario/metabolismoRESUMEN
Ovarian steroid hormones play a critical role in regulating mammary gland growth and development. The mammary gland sequentially acquires and cyclically exhibits proliferative responses to estrogen and/or progesterone from birth to postmenopause. The focus of this review is to present our current understanding of estrogen and progesterone receptor distribution in epithelial and stromal cells and their functions in relation to mammary gland development. Insights gained from the study of the normal mammary gland are relevant to our understanding of the conditions which may predispose women to the development of breast cancer as well as to alterations in hormonal regulation that occur in breast cancer.
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Mama/fisiología , Estrógenos/fisiología , Glándulas Mamarias Animales/fisiología , Progesterona/fisiología , Receptores de Estrógenos/fisiología , Receptores de Progesterona/fisiología , Animales , Mama/citología , Femenino , Humanos , Glándulas Mamarias Animales/citología , Ratones , Ovario/fisiología , PosmenopausiaRESUMEN
Postnatal mammary gland development is highly dependent on the ovarian steroids, estrogen and progesterone. However, evidence from both in vitro and in vivo studies indicates that steroid-induced development occurs indirectly, requiring stromal cooperation in epithelial proliferation and morphogenesis. Stromal cells appear to influence epithelial cell behavior by secretion of growth factors and/or by altering the composition of the extracellular matrix in which epithelial cells reside. This review will discuss the requirement for stromal tissue in modulating proliferative responses to ovarian hormones during postnatal development and the potential role of the EGF, IGF, HGF and FGF3 growth factor families. Additionally, the roles of extracellular matrix proteins, including fibronectin, collagens and laminin, will be summarized.
Asunto(s)
Mama/fisiología , Estrógenos/fisiología , Glándulas Mamarias Animales/fisiología , Ovario/fisiología , Progesterona/fisiología , Células del Estroma/fisiología , Animales , Mama/citología , División Celular , Femenino , Sustancias de Crecimiento/fisiología , Humanos , Glándulas Mamarias Animales/citología , Células del Estroma/citologíaRESUMEN
Mammary stromal cells can modulate steroid hormone responsiveness both in vivo and in vitro. One of the mechanisms by which stromal cells can influence epithelial cell behavior is by modifying the composition of the extracellular matrix (ECM). In this report, we have investigated the effects of five ECM molecules on control of epithelial cell proliferation by estrogen (E2) and progestin (R5020) under serum-free culture conditions. To assess the contribution of mammary gland differentiation in determining epithelial cell interactions with ECM, the behavior of mammary epithelial cells derived from nulliparous and pregnant mice was compared. We report the novel finding that the proliferative responses of mammary epithelial cells to progestin is influenced by specific ECM molecules. However, the primary determinant of hormonal responsiveness is the developmental state of the gland from which the epithelial cells were derived. Nulliparous-derived epithelial cells, proliferated in response to R5020 only on fibronectin (FN) and collagen IV (Col IV). The more highly differentiated, pregnancy-derived epithelial cells were not responsive to E2 or R5020 on any ECM. To determine if steroid hormone receptors were targets of ECM-mediated effects, ER and PR levels were analyzed. In both nulliparous and pregnancy-derived cultures, PR binding levels were maintained at similar levels on all ECMs. However, ER levels were not maintained in nulliparous-derived cultures, and this may have contributed to the lack of a significant response to E2. Alternatively or in addition, E2-induced responses may require additional signals or growth factors that are provided by stromal cells in vivo or by serum supplementation in vitro. These results demonstrate the ECM molecules, fibronectin and collagen IV, can modulate responsiveness of mammary epithelial cells to R5020 in vitro, and may be the mediators of stromal influences on hormone responsiveness in vivo. However, the specific effects of ECM and hormones are also determined by the developmental state of the mammary gland from which the cells are derived. Thus, mammary gland differentiation, ovarian hormones, and ECM composition may act in concert to determine the outcome of hormone treatment on cell proliferation.
Asunto(s)
Proteínas de la Matriz Extracelular/farmacología , Matriz Extracelular/fisiología , Glándulas Mamarias Animales/citología , Receptores de Estrógenos/metabolismo , Receptores de Progesterona/metabolismo , Animales , Caseínas/análisis , Caseínas/biosíntesis , Diferenciación Celular , División Celular , Células Cultivadas , Colágeno/farmacología , Medio de Cultivo Libre de Suero , ADN/biosíntesis , Células Epiteliales , Epitelio/efectos de los fármacos , Epitelio/fisiología , Estradiol/farmacología , Femenino , Fibronectinas/farmacología , Glándulas Mamarias Animales/efectos de los fármacos , Glándulas Mamarias Animales/fisiología , Ratones , Ratones Endogámicos BALB C , Embarazo , Promegestona/farmacología , Timidina/metabolismoRESUMEN
The first known member of the Polyomavirus family, murine Polyomavirus (MPyV), was discovered because of its oncogenic properties. The genetic simplicity of MPyV (shared with all members of the Py family), the wide spectrum of tumors induced by MPyV, and the convenient properties of its natural host, the mouse, make it a particularly interesting model system to study oncogenesis. This paper briefly reviews the virus infectious cycle and our current understanding of the viral proteins that are involved in oncogenesis, and focuses on recent studies on oncogenesis of the mammary gland. Mammary gland ductal adenocarcinomas develop at high frequency and with short latency in infected immunoincompetent adult female or normal neonatal mice or in transgenic mice expressing the viral oncogene, middle T. These tumors provide excellent model systems for the study of human breast cancer.
Asunto(s)
Carcinoma Ductal de Mama/virología , Modelos Animales de Enfermedad , Neoplasias Mamarias Experimentales/virología , Infecciones por Polyomavirus/virología , Poliomavirus , Infecciones Tumorales por Virus/virología , Animales , Animales Recién Nacidos , Carcinoma Ductal de Mama/genética , Transformación Celular Neoplásica/genética , Femenino , Humanos , Neoplasias Mamarias Experimentales/genética , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Ratones Transgénicos , Poliomavirus/fisiología , Infecciones por Polyomavirus/genética , Infecciones Tumorales por Virus/genética , Replicación ViralRESUMEN
Polyomavirus infection of adolescent athymic female mice causes a high incidence of mammary adenocarcinomas. We have examined the role of ovarian hormones, age and mammary gland developmental stage at infection on subsequent tumor induction, viral replication and gene expression. Ovariectomy (OVX) of adolescent mice 1 week before infection decreased mammary tumor incidence and number, and significantly increased tumor incidence and number, and significantly increased tumor latency. Reduction in tumorigenesis was observed to a lesser degree if mice were OVX at the time of or after infection, indicating that ovarian hormones are mainly required for tumor initiation. Tumor incidence was also reduced with increasing age; OVX prior to infection at older ages drastically reduced tumor development. Treatment of OVX adult mice with estrogen + progesterone for 1-3 weeks prior to infection was unable to restore tumorigenesis to the level observed in intact mice. Thus, in contrast to adolescent mice, the continued presence of ovarian hormones after infection was required for maximal tumorigenesis in adult mice. The decreased tumorigenesis observed in older animals is not likely due to increased differentiation since late pregnant mice with well differentiated mammary glands remained highly susceptible to tumorigenesis. At 10 days post infection, the levels of viral genomes were moderately high and similar in all experimental groups. Early viral protein and middle T-associated kinase levels were undetectable in infected tissues in all experimental conditions. However, high levels were found in tumors, perhaps reflecting a high dosage requirement for oncogenesis.
Asunto(s)
Estradiol/farmacología , Neoplasias Mamarias Experimentales/fisiopatología , Neoplasias Mamarias Experimentales/virología , Ovario/fisiología , Infecciones por Polyomavirus/fisiopatología , Poliomavirus , Progesterona/farmacología , Infecciones Tumorales por Virus/fisiopatología , Envejecimiento/fisiología , Animales , Neoplasias Óseas/fisiopatología , Neoplasias Óseas/virología , Diferenciación Celular , Femenino , Genoma Viral , Glándulas Mamarias Animales/citología , Glándulas Mamarias Animales/efectos de los fármacos , Glándulas Mamarias Animales/crecimiento & desarrollo , Ratones , Ratones Desnudos , Ovariectomía , Ovario/crecimiento & desarrollo , Poliomavirus/aislamiento & purificación , Poliomavirus/fisiología , Embarazo , Neoplasias Cutáneas/fisiopatología , Neoplasias Cutáneas/virología , Factores de Tiempo , Replicación ViralRESUMEN
Mice lacking cyclin D1 have been generated by gene targeting in embryonic stem cells. Cyclin D1-deficient animals develop to term but show reduced body size, reduced viability, and symptoms of neurological impairment. Their retinas display a striking reduction in cell number due to proliferative failure during embryonic development. In situ hybridization studies of normal mouse embryos revealed an extremely high level of cyclin D1 in the retina, suggesting a special dependence of this tissue on cyclin D1. In adult mutant females, the breast epithelial compartment fails to undergo the massive proliferative changes associated with pregnancy despite normal levels of ovarian steroid hormones. Thus, steroid-induced proliferation of mammary epithelium during pregnancy may be driven through cyclin D1.
Asunto(s)
Ciclinas/fisiología , Glándulas Mamarias Animales/embriología , Proteínas Oncogénicas/fisiología , Retina/embriología , Animales , Diferenciación Celular , División Celular , Ciclina D1 , Ciclinas/genética , Femenino , Regulación del Desarrollo de la Expresión Génica , Marcación de Gen , Masculino , Glándulas Mamarias Animales/citología , Glándulas Mamarias Animales/metabolismo , Ratones , Ratones Mutantes , Neoplasias Experimentales/etiología , Proteínas Oncogénicas/genética , Fenotipo , Embarazo , Receptores de Estrógenos/metabolismo , Retina/citología , Retina/metabolismo , Células Madre/citología , Esteroides/metabolismoRESUMEN
An in vitro serum-free culture system provides an important approach to the understanding of local hormonal regulation of mammary epithelial and fibroblast cells, avoiding the complexity of the in vivo environment and the influence of undefined serum factors. The substratum conditions and medium components have been examined for the basal growth of epithelial cells, fibroblasts, and combined epithelial and fibroblast cells in monolayer cultures. Epithelial cells and mixed cells exhibit good attachment and maintenance on a collagen-coated surface in a minimal medium supplemented with fetuin and insulin. In contrast, fibroblast-enriched cultures require a plastic substratum and a medium supplemented with insulin, fetuin, and hydrocortisone. In mixed cell culture, fibroblasts are maintained well in the minimal media which supports the maintenance of epithelial cells. These results indicate that the presence of epithelial cells in mixed cell cultures can influence fibroblast function. The media developed in the present study can be used in future studies of fibroblast and epithelial cell interactions with regard to hormone and growth factor regulation of their growth and differentiation.
Asunto(s)
Glándulas Mamarias Animales/citología , Células del Estroma/citología , Animales , Sangre , División Celular/efectos de los fármacos , Células Cultivadas , Medios de Cultivo , Células Epiteliales , Femenino , Fibroblastos/citología , Insulina/farmacología , Ratones , Ratones Endogámicos BALB C , Embarazo , Albúmina Sérica Bovina/farmacología , Transferrina/farmacología , alfa-Fetoproteínas/farmacologíaRESUMEN
Estrogen (E), progesterone (P), and epidermal growth factor (EGF) are involved in the growth and development of the normal mammary gland. While studies have been carried out to investigate the in vivo effects of EGF in the immature mammary gland, nothing is known about the growth effects of EGF or its potential interactions with E and/or P in the adult mammary gland. The present studies were undertaken to investigate the effects of EGF, E, and P on mammary cell proliferation in immature, peripubertal vs. adult, sexually mature mice. We have found that EGF promotes epithelial and stromal cell proliferation in both the immature and adult mammary glands. In the immature gland, the end bud epithelium is most responsive to the proliferative effects of EGF and there is no apparent interaction between EGF, E, and/or P. In contrast, in the mature gland EGF adds to the proliferative effects of E+P in the ductal epithelium resulting in more extensive ductal sidebranching. Thus these results demonstrate that the developmental state of the mammary gland determines the nature and extent of the interactions between EGF, E, and P in growth and development.