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ABSTRACT: While progesterone plays multiple roles in the process of breast development and differentiation, its role in breast cancer is less understood. We have shown previously that progestins stimulate breast cancer cell migration and invasion because of the activation of rapid signaling cascades leading to modifications in the actin cytoskeleton and cell membrane that are required for cell movement. In this study, we have investigated the effects of progesterone on the formation of focal adhesion (FA) complexes, which provide anchoring sites for cell attachment to the extracellular matrix during cell movement and invasion. In T47-D breast cancer cells, progesterone rapidly enhances FA kinase (FAK) phosphorylation at Tyr(397) in a time- and concentration-dependent manner. As a result, exposure to progesterone leads to increased formation of FA complexes within specialized cell membrane protrusions. The cascade of events required for this phenomenon involves progesterone receptor interaction with the tyrosine kinase c-Src, which activates the phosphatidylinositol-3-kinase/Akt pathway and the small GTPase RhoA/Rho-associated kinase complex. In the presence of progesterone, T47-D breast cancer cells display enhanced horizontal migration and invasion of three-dimensional matrices, which is reversed by small interfering RNAs abrogating FAK. In conclusion, progesterone promotes breast cancer cell movement and invasion by facilitating the formation of FA complexes via the rapid regulation of FAK. These results provide novel mechanistic views on the effects of progesterone on breast cancer progression, and may in the future be helpful to develop new strategies for the treatment of endocrine-sensitive breast cancers.
Endocrine Related Cancer 03/2010; 17(2):431-43. · 4.36 Impact Factor
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ABSTRACT: Sex steroids are important for brain function and protection. However, growing evidence suggests that these actions might depend on the timing of exposure to steroids. We have studied the effects of steroid administration on the survival of neural cells and we have partially characterized the possible mechanisms. The effect of a 24h pre-treatment with 17beta-estradiol or 17beta-estradiol plus progesterone or medroxyprogesterone acetate on the toxic action of l-glutamate was used to test the experimental hypothesis. Pre-exposure to either steroid combinations turned in enhanced cell survival. Instead, addition of sex steroids together with l-glutamate, in the absence of a pre-exposure had no protective effect. Pre-treatment with the steroid combinations resulted in increased neural NOS expression and activity and blockade of NOS abolished the cytoprotective effects of steroids. These results suggest that NOS induction might be involved in sex steroid-induced neuroprotection. Furthermore, these data supports the hypothesis that prolonged and continued exposure to oestrogen and progesterone, leading to changes in gene expression, is necessary to obtain neuroprotection induced by sex steroids.
Steroids 09/2009; 74(8):650-6. · 2.83 Impact Factor
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ABSTRACT: Estrogens are important regulators of neuronal cell morphology, and this is thought to be critical for gender-specific differences in brain function and dysfunction. Dendritic spine formation is dependent on actin remodeling by the WASP-family verprolin homologous (WAVE1) protein, which controls actin polymerization through the actin-related protein (Arp)-2/3 complex. Emerging evidence indicates that estrogens are effective regulators of the actin cytoskeleton in various cell types via rapid, extranuclear signaling mechanisms. We here show that 17beta-estradiol (E2) administration to rat cortical neurons leads to phosphorylation of WAVE1 on the serine residues 310, 397, and 441 and to WAVE1 redistribution toward the cell membrane at sites of dendritic spine formation. WAVE1 phosphorylation is found to be triggered by a Galpha(i)/Gbeta protein-dependent, rapid extranuclear signaling of estrogen receptor alpha to c-Src and to the small GTPase Rac1. Rac1 recruits the cyclin-dependent kinase (Cdk5) that directly phosphorylates WAVE1 on the three serine residues. After WAVE1 phosphorylation by E2, the Arp-2/3 complex concentrates at sites of spine formation, where it triggers the local reorganization of actin fibers. In parallel, E2 recruits a Galpha(13)-dependent pathway to RhoA and ROCK-2, leading to activation of actin remodeling via the actin-binding protein, moesin. Silencing of WAVE1 or of moesin abrogates the increase in dendritic spines induced by E2 in cortical neurons. In conclusion, our findings indicate that the control of actin polymerization and branching via moesin or WAVE1 is a key function of estrogen receptor alpha in neurons, which may be particularly relevant for the regulation of dendritic spines.
Molecular Endocrinology 06/2009; 23(8):1193-202. · 4.54 Impact Factor
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ABSTRACT: Cell movement is required in relevant physiological processes such as embryonic development, tissue and organ differentiation, inflammation, immune response and wound healing, along with pathological phenomena, such as cancer metastatic spread. Cell motility is tightly controlled by a complex and often redundant array of intracellular signaling pathways largely devoted to the dynamic regulation of the actin cytoskeletal network and of its relationship with the cell membrane and the extracellular matrix. Sex steroids, particularly estrogen and progesterone, are effective regulators of cell migration and tissue organization, and recent evidence indicates that this is in part obtained through the regulation of the cytoskeleton. Intriguingly, many of these regulatory actions related to cell movement are achieved through rapid, non-classical signaling of sex steroid receptors to kinase cascades, independently from nuclear alteration of gene expression or protein synthesis. The identification of the mechanistic basis for these rapid actions on cell cytoskeleton and cell movement has special relevance for the characterization of the effects of sex steroids in physiological conditions, such their role in the control of inflammation, brain or vascular cell remodelling, angiogenesis or wound healing, as well as in the context of pathological conditions such as steroid-sensitive cancer cell invasion and metastasis. This review highlights the physiological and clinical conditions where the regulatory effects on the cytoskeleton and cell movement of sex steroids might have a special importance, as well as the recent advances in the characterization of the mechanisms, providing insights and working hypotheses on possible clinical applications for the modulation of these pathways.
Steroids 11/2008; 73(9-10):895-900. · 2.83 Impact Factor
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ABSTRACT: Raloxifene (RAL) is a selective oestrogen receptor modulator (SERM) approved for the prevention and treatment of osteoporosis and for the prevention of breast cancer in postmenopausal women. However, little is known on the effects of this SERM on breast cancer cell metastasis, which is the main cause of morbidity and death. Cell movement is critical for local progression and distant metastasis of cancer cells. These processes rely on the dynamic control of the actin cytoskeleton and of cell membrane morphology. The aim of the present study was to characterize the effects of RAL or of 17beta-estradiol (E2) plus RAL on oestrogen receptor (ER) positive T47-D breast cancer cell cytoskeletal remodelling, migration and invasion. Our findings show that, when given alone, RAL induces a weak actin cytoskeleton remodelling in breast cancer cells, with the formation of specialized cell membrane structures implicated in cell motility. However, in the presence of physiological amounts of estradiol, which potently drives breast cancer cell cytoskeletal remodelling and motility, RAL displays a powerful inhibitory effect on oestrogen-promoted cell migration and invasion. These actions are plaid through an interference of RAL with an extra-nuclear signalling cascade involving G proteins and the RhoA-associated kinase, ROCK-2, linked to the recruitment of the cytoskeletal controller, moesin. Hence, in the presence of estradiol, RAL acts as an ER antagonist. These results highlight a novel mechanism of action of the SERM raloxifene that might be important for the interference of breast cancer progression or metastasis induced by oestrogens in postmenopausal women.
Journal of Cellular and Molecular Medicine 10/2008; 13(8B):2396-407. · 4.13 Impact Factor
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ABSTRACT: The endothelial effects of progestogens are poorly investigated. Actin remodeling and cell movement are fundamental for endothelial function and are controlled by the actin-binding protein moesin. In this work, we studied the effects of progesterone and medroxyprogesterone acetate (MPA) on actin remodeling, moesin activation and cell movement in human endothelial cells. Our findings show that progesterone and MPA trigger a rapid endothelial actin rearrangement, with the formation of cortical actin complexes, pseudopodia and membrane ruffles. Both progestogens trigger a rapid progesterone receptor (PR)-dependent moesin activation via a non-genomic signaling cascade involving G proteins, the small GTPase RhoA and the Rho-associated kinase (ROCK-2). In addition, MPA signaling also requires the recruitment of phosphatidylinositol-3 kinase (PI3K). Both progestogens enhance endothelial cell migration, which is prevented by moesin silencing or by blockade of PR, G proteins, PI3K, mitogen-activated protein kinases or ROCK-2. Progesterone and MPA potentiate 17beta-estradiol (E2) induced-moesin activation. However, they partially reduce cell migration induced by E2. In conclusion, progesterone and MPA regulate endothelial cell movement by rapidly signaling to the actin-binding protein moesin and to the actin cytoskeleton. These findings provide new information on the biological actions of progestins on human endothelial cells that are relevant for vascular function.
Molecular Human Reproduction 05/2008; 14(4):225-34. · 3.85 Impact Factor
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Maria Silvia Giretti,
Xiao-Dong Fu,
Giovanni De Rosa,
Ivana Sarotto,
Chiara Baldacci,
Silvia Garibaldi,
Paolo Mannella,
Nicoletta Biglia,
Piero Sismondi,
Andrea Riccardo Genazzani,
Tommaso Simoncini
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ABSTRACT: Estrogen is an established enhancer of breast cancer development, but less is known on its effect on local progression or metastasis. We studied the effect of estrogen receptor recruitment on actin cytoskeleton remodeling and breast cancer cell movement and invasion. Moreover, we characterized the signaling steps through which these actions are enacted.
In estrogen receptor (ER) positive T47-D breast cancer cells ER activation with 17beta-estradiol induces rapid and dynamic actin cytoskeleton remodeling with the formation of specialized cell membrane structures like ruffles and pseudopodia. These effects depend on the rapid recruitment of the actin-binding protein moesin. Moesin activation by estradiol depends on the interaction of ER alpha with the G protein G alpha(13), which results in the recruitment of the small GTPase RhoA and in the subsequent activation of its downstream effector Rho-associated kinase-2 (ROCK-2). ROCK-2 is responsible for moesin phosphorylation. The G alpha(13)/RhoA/ROCK/moesin cascade is necessary for the cytoskeletal remodeling and for the enhancement of breast cancer cell horizontal migration and invasion of three-dimensional matrices induced by estrogen. In addition, human samples of normal breast tissue, fibroadenomas and invasive ductal carcinomas show that the expression of wild-type moesin as well as of its active form is deranged in cancers, with increased protein amounts and a loss of association with the cell membrane.
These results provide an original mechanism through which estrogen can facilitate breast cancer local and distant progression, identifying the extra-nuclear G alpha(13)/RhoA/ROCK/moesin signaling cascade as a target of ER alpha in breast cancer cells. This information helps to understand the effects of estrogen on breast cancer metastasis and may provide new targets for therapeutic interventions.
PLoS ONE 02/2008; 3(5):e2238. · 4.09 Impact Factor
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ABSTRACT: Limited information is available on the effects of progestins on breast cancer progression and metastasis. Cell migration and invasion are central for these processes, and require dynamic cytoskeletal and cell membrane rearrangements for cell motility to be enacted.
We investigated the effects of progesterone (P), medroxyprogesterone acetate (MPA), drospirenone (DRSP) and nestorone (NES) alone or with 17beta-estradiol (E2) on T47-D breast cancer cell migration and invasion and we linked some of these actions to the regulation of the actin-regulatory protein, moesin and to cytoskeletal remodeling.
Breast cancer cell horizontal migration and invasion of three-dimensional matrices are enhanced by all the progestins, but differences are found in terms of potency, with MPA being the most effective and DRSP being the least. This is related to the differential ability of the progestins to activate the actin-binding protein moesin, leading to distinct effects on actin cytoskeleton remodeling and on the formation of cell membrane structures that mediate cell movement. E2 also induces actin remodeling through moesin activation. However, the addition of some progestins partially offsets the action of estradiol on cell migration and invasion of breast cancer cells.
These results imply that P, MPA, DRSP and NES alone or in combination with E2 enhance the ability of breast cancer cells to move in the surrounding environment. However, these progestins show different potencies and to some extent use distinct intracellular intermediates to drive moesin activation and actin remodeling. These findings support the concept that each progestin acts differently on breast cancer cells, which may have relevant clinical implications.
BMC Cancer 02/2008; 8:166. · 3.01 Impact Factor
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ABSTRACT: Recent clinical trials indicate that synthetic progestins may be unexpectedly relevant for the development of cardiovascular disease. The aim of this study was to establish whether nomegestrol acetate induces signaling events in human endothelial cells that differ from those of other progestins, such as natural progesterone or medroxyprogesterone acetate.
We used human endothelial cells to study the action of nomegestrol acetate (either alone or in the presence of estradiol [E2]) on the synthesis of nitric oxide (NO) and on the activity or expression of endothelial nitric oxide synthase (eNOS). We compared the effects of nomegestrol acetate with those of progesterone or medroxyprogesterone acetate. In addition, we characterized the signaling events recruited by these compounds.
Progesterone and nomegestrol acetate increase NO synthesis by transcriptional and nontranscriptional mechanisms, whereas medroxyprogesterone acetate lacks such effects. When used together with physiological E2 concentrations, progesterone and nomegestrol acetate do not interfere with (or even enhance) E2 effects, whereas medroxyprogesterone acetate impairs E2 signaling. A marked difference in the recruitment of mitogen-activated protein kinase and phosphatidylinositol-3 kinase explains the divergent effects of the three gestagens.
Our findings show significant differences in the signal transduction pathways recruited by progesterone, nomegestrol acetate, and medroxyprogesterone acetate in human endothelial cells that may have relevant clinical implications.
Obstetrics and Gynecology 11/2006; 108(4):969-78. · 4.73 Impact Factor
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ABSTRACT: To investigate the effects of P, medroxyprogesterone acetate (MPA), and dydrogesterone (DYD) and its metabolite, 20-alpha-dihydrodydrogesterone (DHD) on endothelial synthesis of nitric oxide (NO) and characterize the signaling events recruited by these compounds. The Women's Health Initiative trial reports an excess of heart disease in postmenopausal women receiving MPA.
Cell culture.
Research laboratory.
Human endothelial cells from umbilical vein.
Treatments with P, MPA, DYD, or DHD.
Measure of NO release, endothelial nitric oxide synthase (eNOS) activity and expression, and activation of ERK 1/2 and Akt.
The administration of DYD alone or in combination with estrogen to endothelial cells results in neutral effects on NO synthesis and on the activity and expression of eNOS. In parallel, the stable metabolite DHD acts similarly to natural P, enhancing the expression of eNOS and inducing rapid activation of the enzyme through the regulation of the ERK 1/2 mitogen-activated protein kinase cascade. 20-Alpha-dihydrodydrogesterone and P also potentiate eNOS induction by E2. On the contrary, MPA does not trigger eNOS enzymatic activation and decreases the extent of eNOS induction by E2.
These findings support the concept that synthetic progestins act differently on vascular cells and that hormonal preparations may differ as to their cardiovascular effects.
Fertility and sterility 11/2006; 86(4 Suppl):1235-42. · 3.97 Impact Factor
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ABSTRACT: We evaluated the effects of tibolone oral administration on neuroendocrine function by investigating the modulation exerted by tibolone administration on allopregnanolone and central and peripheral beta-endorphin (beta-EP) levels in ovariectomized rats.
Female Wistar rats (N = 64) were included: 48 rats were ovariectomized, 8 cycling rats were included as controls, and 8 cycling rats were treated with placebo. The ovariectomized animals were divided into six groups: untreated rats and those that received 14-day oral treatment with either placebo, estradiol valerate (E2V) 0.05 mg/kg/d, or tibolone (0.1, 0.5, or 2 mg/kg/d. beta-EP levels were assessed in the frontal lobe, parietal lobe, hippocampus, hypothalamus, anterior pituitary, neurointermediate pituitary, and plasma, whereas allopregnanolone levels were measured in the frontal lobe, parietal lobe, hippocampus, hypothalamus, anterior pituitary, adrenal glands, and serum.
The administration of tibolone (0.5 and 2 mg/kg/d) in ovariectomized rats induces a significant increase of allopregnanolone in the frontal lobe, parietal lobe, hippocampus, hypothalamus, whereas in serum a significant increase of allopregnanolone occurs only with the dose of 2 mg/kg/d, a significant decrease in allopregnanolone levels occurs in the adrenal glands. No changes occurred in the anterior pituitary. Tibolone doses of 0.5 and 2 mg/kg/d induced a significant increase in beta-EP content in the frontal lobe, hypothalamus, and neurointermediate lobe; and, at doses of 2 mg/kg/d, in the parietal lobe, anterior pituitary, and plasma, without changes in the hippocampus. Compared with E2V, 0.5 mg/kg/d tibolone showed a similar effect on allopregnanolone and beta-EP in most brain regions.
Tibolone administration affects beta-EP and allopregnanolone levels, playing a role as a neuroendocrine modulator.
Menopause 12/2005; 13(1):57-64. · 3.76 Impact Factor
