RESUMO
In female mammals, the cessation of ovarian functions is associated with significant metabolic alterations, weight gain, and increased susceptibility to a number of pathologies associated with ageing. The molecular mechanisms triggering these systemic events are unknown because most tissues are responsive to lowered circulating sex steroids. As it has been demonstrated that isoform alpha of the estrogen receptor (ERα) may be activated by both estrogens and amino acids, we test the metabolic effects of a diet enriched in specific amino acids in ovariectomized (OVX) mice. This diet is able to block the OVX-induced weight gain and fat deposition in the liver. The use of liver-specific ERα KO mice demonstrates that the hepatic ERα, through the control of liver lipid metabolism, has a key role in the systemic response to OVX. The study suggests that the liver ERα might be a valuable target for dietary treatments for the post-menopause.
Assuntos
Aminoácidos Essenciais/farmacologia , Receptor alfa de Estrogênio/metabolismo , Fígado/efeitos dos fármacos , Ovariectomia/efeitos adversos , Aminoácidos de Cadeia Ramificada/farmacologia , Aminoácidos de Cadeia Ramificada/uso terapêutico , Aminoácidos Essenciais/uso terapêutico , Animais , Dietoterapia , Receptor alfa de Estrogênio/deficiência , Feminino , Metabolismo dos Lipídeos/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Knockout , Caracteres Sexuais , Transcriptoma/efeitos dos fármacos , Aumento de Peso/efeitos dos fármacosRESUMO
Adjuvant tamoxifen therapy improves survival in breast cancer patients. Unfortunately, long-term treatment comes with side effects that impact health and quality of life, including hot flashes, changes in bone density, and fatigue. Partly due to a lack of proven animal models, the tissues and cells that mediate these negative side effects are unclear. Here, we show that mice undergoing tamoxifen treatment experience changes in temperature, bone, and movement. Single-cell RNA sequencing reveals that tamoxifen treatment induces widespread gene expression changes in the hypothalamus and preoptic area (hypothalamus-POA). These expression changes are dependent on estrogen receptor alpha (ERα), as conditional knockout of ERα in the hypothalamus-POA ablates or reverses tamoxifen-induced gene expression. Accordingly, ERα-deficient mice do not exhibit tamoxifen-induced changes in temperature, bone, or movement. These findings provide mechanistic insight into the effects of tamoxifen on the hypothalamus-POA and indicate that ERα mediates several physiological effects of tamoxifen treatment in mice.
Estrogen is a hormone often known for its role in female development and reproduction. Yet, it also has an impact on many biological processes such as immunity and the health of bones, the heart, or the brain. It usually works by attaching to receptor proteins in specific cells. For instance, estrogen-responsive cells are present in the hypothalamus, the brain area that controls energy levels as well as the body's temperature and internal clock. Breast cancer cells are also often sensitive to estrogen, with the hormone fuelling the growth of tumors. The drug tamoxifen blocks estrogen receptors, stopping cells from responding to the hormone. As such, it is often used to reduce the likelihood that estrogen-dependent breast cancer will come back after treatment. However, its use can induce hot flashes, changes in bone density, fatigue and other life-altering side effects. Here, Zhang et al. investigated how estrogen receptors in the hypothalamus and a related region known as the preoptic area could be responsible for these side effects in mice. When the rodents were given tamoxifen for 28 days, they experienced changes in temperature, bone density and movement similar to those found in humans. In fact, genetic analyses revealed that the drug altered the way genes were turned on and off in certain cells types in the hypothalamus. Crucially, mice whose hypothalamus and preoptic area lacked estrogen receptors did not experience these behavioral and biological alterations. The findings by Zhang et al. help to understand how the side effects of tamoxifen emerge, singling out estrogen receptors in particular brain regions. This result could help to develop new therapies so that breast cancer can be treated with a better quality of life.
Assuntos
Antineoplásicos Hormonais/farmacologia , Hipotálamo/metabolismo , Área Pré-Óptica/metabolismo , Tamoxifeno/farmacologia , Animais , Temperatura Corporal/efeitos dos fármacos , Densidade Óssea/efeitos dos fármacos , Receptor alfa de Estrogênio/deficiência , Feminino , Regulação da Expressão Gênica , Camundongos , Movimento/efeitos dos fármacosRESUMO
Estrogen receptors are involved in regulating the proliferation and invasion process of neuroblastoma. As a kind of estrogen-like environmental endocrine disruptors (EEDs), whether mono-2-ethylhexyl phthalate (MEHP) can affect the proliferation and invasion of neuroblastoma cells via ERs is unknown. The present study aimed to explore the role of ERα in MEHP-induced proliferation, migration, and invasion of SH-SY5Y cells. SH-SY5Y cells were cultured in DMEM with 10 % FBS. Wild-type SH-SY5Y cells and ERα-knockdown SH-SY5Y cells were treated with MEHP (0, 10, 50, and 250 µM) for 12 h and 24 h. The viability of SH-SY5Y cells was detected with a CCK8 kit and cell cycle was measured by flow cytometry. Cell migration was measured using a scratch assay, and cell invasion was tested using a Transwell migration assay. The expression levels of proliferating cell nuclear antigen (PCNA), matrix metalloproteinase 2 (MMP-2), matrix metalloproteinase 9 (MMP-9), tissue inhibitor of matrix metalloproteinase 2 (TIMP-2), ERα, and ERß were detected with real-time qPCR and western blotting. MEHP promoted the proliferation of SH-SY5Y cells. The results also showed that MEHP significantly increased the relative migration distance of wild-type SH-SY5Y cells. Conversely, MEHP treatment did not increase the relative migration distance of ERα-knockdown SH-SY5Y cells, suggesting that MEHP promotes the migration of neuroblastoma through ERα. Similarly, MEHP significantly increased the relative number of invaded wild-type SH-SY5Y cells, while the MEHP-induced invasion effect was significantly decreased in ERα-knockdown SH-SY5Y cells. Moreover, the expression levels of PCNA, MMP-2, MMP-9, and ERα cells were upregulated by MEHP in wild-type SH-SY5Y, and the expression level of its tissue inhibitor TIMP-2 was downregulated. In contrast, the expression of PCNA, MMP-2, MMP-9, and ERα was significantly downregulated in ERα-knockdown SH-SY5Y cells, while the expression of TIMP-2 was significantly upregulated. In conclusion, MEHP can upregulate PCNA, MMP-2, and MMP-9, and downregulate TIMP-2, further promoting proliferation, migration, and invasion of neuroblastoma through ERα.
Assuntos
Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Dietilexilftalato/análogos & derivados , Receptor alfa de Estrogênio/fisiologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/fisiologia , Dietilexilftalato/toxicidade , Relação Dose-Resposta a Droga , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Técnicas de Silenciamento de Genes/métodos , Humanos , Invasividade Neoplásica/patologiaRESUMO
Women with pulmonary arterial hypertension (PAH) exhibit better right ventricular (RV) function and survival than men; however, the underlying mechanisms are unknown. We hypothesized that 17ß-estradiol (E2), through estrogen receptor α (ER-α), attenuates PAH-induced RV failure (RVF) by upregulating the procontractile and prosurvival peptide apelin via a BMPR2-dependent mechanism. We found that ER-α and apelin expression were decreased in RV homogenates from patients with RVF and from rats with maladaptive (but not adaptive) RV remodeling. RV cardiomyocyte apelin abundance increased in vivo or in vitro after treatment with E2 or ER-α agonist. Studies employing ER-α-null or ER-ß-null mice, ER-α loss-of-function mutant rats, or siRNA demonstrated that ER-α is necessary for E2 to upregulate RV apelin. E2 and ER-α increased BMPR2 in pulmonary hypertension RVs and in isolated RV cardiomyocytes, associated with ER-α binding to the Bmpr2 promoter. BMPR2 is required for E2-mediated increases in apelin abundance, and both BMPR2 and apelin are necessary for E2 to exert RV-protective effects. E2 or ER-α agonist rescued monocrotaline pulmonary hypertension and restored RV apelin and BMPR2. We identified what we believe to be a novel cardioprotective E2/ER-α/BMPR2/apelin axis in the RV. Harnessing this axis may lead to novel RV-targeted therapies for PAH patients of either sex.
Assuntos
Apelina/metabolismo , Receptores de Proteínas Morfogenéticas Ósseas Tipo II/metabolismo , Estradiol/metabolismo , Receptor alfa de Estrogênio/metabolismo , Hipertensão Pulmonar/fisiopatologia , Função Ventricular Direita/fisiologia , Animais , Cardiotônicos/metabolismo , Modelos Animais de Doenças , Células Endoteliais/metabolismo , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Feminino , Humanos , Masculino , Camundongos , Camundongos Knockout , Modelos Cardiovasculares , Miócitos Cardíacos/metabolismo , Ratos , Ratos MutantesRESUMO
Estrogen receptor α (ERα) modulates gene expression by interacting with chromatin regions that are frequently distal from the promoters of estrogen-regulated genes. Active chromatin-enriched "super-enhancer" (SE) regions, mainly observed in in vitro culture systems, often control production of key cell type-determining transcription factors. Here, we defined super-enhancers that bind to ERα in vivo within hormone-responsive uterine tissue in mice. We found that SEs are already formed prior to estrogen exposure at the onset of puberty. The genes at SEs encoded critical developmental factors, including retinoic acid receptor α (RARA) and homeobox D (HOXD). Using high-throughput chromosome conformation capture (Hi-C) along with DNA sequence analysis, we demonstrate that most SEs are located at a chromatin loop end and that most uterine genes in loop ends associated with these SEs are regulated by estrogen. Although the SEs were formed before puberty, SE-associated genes acquired optimal ERα-dependent expression after reproductive maturity, indicating that pubertal processes that occur after SE assembly and ERα binding are needed for gene responses. Genes associated with these SEs affected key estrogen-mediated uterine functions, including transforming growth factor ß (TGFß) and LIF interleukin-6 family cytokine (LIF) signaling pathways. To the best of our knowledge, this is the first identification of SE interactions that underlie hormonal regulation of genes in uterine tissue and optimal development of estrogen responses in this tissue.
Assuntos
Cromatina/metabolismo , Receptor alfa de Estrogênio/metabolismo , Útero/metabolismo , Animais , Sítios de Ligação , Cromatina/química , Estradiol/farmacologia , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Feminino , Histonas/metabolismo , Proteínas de Homeodomínio/metabolismo , Fator Inibidor de Leucemia/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ligação Proteica , Receptor alfa de Ácido Retinoico/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Útero/efeitos dos fármacosRESUMO
Sexually dimorphic bone structure emerges largely during puberty. Sex steroids are critical for peak bone mass acquisition in both genders. In particular, the biphasic effects of estrogens mediate the skeletal sexual dimorphism. However, so far the stimulatory vs inhibitory actions of estrogens on bone mass are not fully explained by direct effects on bone cells. Recently, it has become evident that there is possible neuroendocrine action of estrogen receptor alpha (ERα) on the skeleton. Based on these considerations, we hypothesized that neuronal ERα-signaling may contribute to the skeletal growth during puberty. Here, we generated mice with tamoxifen-inducible Thy1-Cre mediated ERα inactivation during late puberty specifically in extrahypothalamic neurons (N-ERαKO). Inactivation of neuronal ERα did not alter the body weight in males, whereas N-ERαKO females exhibited a higher body weight and increased body and bone length compared to their control littermates at 16 weeks of age. Ex vivo microCT analysis showed increased radial bone expansion of the midshaft femur in female N-ERαKO along with higher serum levels of insulin-like growth factor (IGF)-1 as well as IGF-binding protein (IGFBP)-3. Furthermore, the 3-point bending test revealed increased bone strength in female N-ERαKO. In contrast, inactivation of neuronal ERα had no major effect on bone growth in males. In conclusion, we demonstrate that central ERα-signaling limits longitudinal bone growth and radial bone expansion specifically in females potentially by interacting with the GH/IGF-1 axis.
Assuntos
Desenvolvimento Ósseo/fisiologia , Receptor alfa de Estrogênio/metabolismo , Neurônios/metabolismo , Maturidade Sexual/fisiologia , Animais , Fenômenos Biomecânicos , Densidade Óssea/genética , Densidade Óssea/fisiologia , Desenvolvimento Ósseo/genética , Osso e Ossos/anatomia & histologia , Osso e Ossos/diagnóstico por imagem , Osso e Ossos/fisiologia , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Caracteres Sexuais , Maturidade Sexual/genética , Transdução de Sinais , Microtomografia por Raio-XRESUMO
Estrogen contributes to females' strong antibody response to microbial vaccines and proneness to autoimmunity, particularly antibody-mediated systemic autoimmunity, in females. We have hypothesized that this is due to estrogen-mediated potentiation of class switch DNA recombination (CSR) and somatic hypermutation (SHM). As we have shown, estrogen boosts AID expression, which is critical for both CSR and SHM, through upregulation of HoxC4, which together with NF-κB critically mediates Aicda (AID gene) promoter activation. We contend here that additional regulation of Aicda expression by estrogen occurs through epigenetic mechanisms. As we have shown, histone deacetylase inhibitors (HDIs) short-chain fatty acid (SCFA) butyrate and propionate as well as the pharmacologic HDI valproic acid upregulate miRNAs that silence AID expression, thereby modulating specific antibody responses in C57BL/6 mice and autoantibody responses in lupus-prone MRL/Faslpr/lpr mice. Here, using constitutive knockout Esr1-/- mice and B cells as well as conditional knockout Aicdacre/creEsr1flox/flox mice and B cells, we showed that the HDI-mediated downregulation of Aicda expression as well as the maturation of antibody and autoantibody responses is reversed by estrogen and enhanced by deletion of ERα or E2 inhibition. Estrogen's reversion of HDI-mediated inhibition of Aicda and CSR in antibody and autoantibody responses occurred through downregulation of B cell miR-26a, which, as we showed, targets Aicda mRNA 3'UTR. miR-26a was significantly upregulated by HDIs. Accordingly, enforced expression of miR-26a reduced Aicda expression and CSR, while miR-26a-sponges (competitive inhibitors of miR-26a) increased Aicda expression and CSR. Thus, our findings show that estrogen reverses the HDI-mediated downregulation of AID expression and CSR through selective modulation of miR-26a. They also provide mechanistic insights into the immunomodulatory activity of this hormone and a proof-of-principle for using combined ER inhibitor-HDI as a potential therapeutic approach.
Assuntos
Autoanticorpos/biossíntese , Linfócitos B/efeitos dos fármacos , Butiratos/farmacologia , Citidina Desaminase/biossíntese , Estradiol/farmacologia , Inibidores de Histona Desacetilases/farmacologia , Switching de Imunoglobulina/efeitos dos fármacos , Isoanticorpos/biossíntese , MicroRNAs/biossíntese , Propionatos/farmacologia , Ácido Valproico/farmacologia , Regiões 3' não Traduzidas , Animais , Autoanticorpos/imunologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Sequência de Bases , Ligação Competitiva , Citidina Desaminase/deficiência , Citidina Desaminase/genética , Regulação para Baixo/efeitos dos fármacos , Receptor alfa de Estrogênio/deficiência , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Switching de Imunoglobulina/genética , Isoanticorpos/imunologia , Lúpus Eritematoso Sistêmico/tratamento farmacológico , Lúpus Eritematoso Sistêmico/imunologia , Camundongos , Camundongos Endogâmicos MRL lpr , Camundongos Knockout , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Fosfoproteínas Fosfatases/genética , Monoéster Fosfórico Hidrolases/genética , Regiões Promotoras Genéticas , Estudo de Prova de Conceito , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Caracteres Sexuais , Transdução GenéticaRESUMO
OBJECTIVE: Estrogen receptor-α (ERα) is a nuclear receptor family member thought to substantially contribute to the metabolic regulation of skeletal muscle. However, previous mouse models utilized to assess the necessity of ERα signaling in skeletal muscle were confounded by altered developmental programming and/or influenced by secondary effects, making it difficult to assign a causal role for ERα. The objective of this study was to determine the role of skeletal muscle ERα in regulating metabolism in the absence of confounding factors of development. METHODS: A novel mouse model was developed allowing for induced deletion of ERα in adult female skeletal muscle (ERαKOism). ERαshRNA was also used to knockdown ERα (ERαKD) in human myotubes cultured from primary human skeletal muscle cells isolated from muscle biopsies from healthy and obese insulin-resistant women. RESULTS: Twelve weeks of HFD exposure had no differential effects on body composition, VO2, VCO2, RER, energy expenditure, and activity counts across genotypes. Although ERαKOism mice exhibited greater glucose intolerance than wild-type (WT) mice after chronic HFD, ex vivo skeletal muscle glucose uptake was not impaired in the ERαKOism mice. Expression of pro-inflammatory genes was altered in the skeletal muscle of the ERαKOism, but the concentrations of these inflammatory markers in the systemic circulation were either lower or remained similar to the WT mice. Finally, skeletal muscle mitochondrial respiratory capacity, oxidative phosphorylation efficiency, and H2O2 emission potential was not affected in the ERαKOism mice. ERαKD in human skeletal muscle cells neither altered differentiation capacity nor caused severe deficits in mitochondrial respiratory capacity. CONCLUSIONS: Collectively, these results suggest that ERα function is superfluous in protecting against HFD-induced skeletal muscle metabolic derangements after postnatal development is complete.
Assuntos
Receptor alfa de Estrogênio/metabolismo , Insulina/metabolismo , Mitocôndrias/metabolismo , Músculo Esquelético/metabolismo , Animais , Receptor alfa de Estrogênio/deficiência , Feminino , Humanos , Camundongos , Camundongos Knockout , Músculo Esquelético/citologiaRESUMO
Background The cardiovascular protective effects of estrogens in premenopausal women depend mainly on estrogen receptor α (ERα). ERα activates nuclear gene transcription regulation and membrane-initiated signaling. The latter plays a key role in estrogen-dependent activation of endothelial NO synthase. The goal of the present work was to determine the respective roles of the 2 ERα activities in endothelial function and cardiac and kidney damage in young and old female mice with hypertension, which is a major risk factor in postmenopausal women. Methods and Results Five- and 18-month-old female mice lacking either ERα (ERα-/-), the nuclear activating function AF2 of ERα (AF2°), or membrane-located ERα (C451A) were treated with angiotensin II (0.5 mg/kg per day) for 1 month. Systolic blood pressure, left ventricle weight, vascular reactivity, and kidney function were then assessed. Angiotensin II increased systolic blood pressure, ventricle weight, and vascular contractility in ERα-/- and AF2° mice more than in wild-type and C451A mice, independent of age. In both the aorta and mesenteric resistance arteries, angiotensin II and aging reduced endothelium-dependent relaxation in all groups, but this effect was more pronounced in ERα-/- and AF2° than in the wild-type and C451A mice. Kidney inflammation and oxidative stress, as well as blood urea and creatinine levels, were also more pronounced in old hypertensive ERα-/- and AF2° than in old hypertensive wild-type and C451A mice. Conclusions The nuclear ERα-AF2 dependent function attenuates angiotensin II-dependent hypertension and protects target organs in aging mice, whereas membrane ERα signaling does not seem to play a role.
Assuntos
Envelhecimento/metabolismo , Receptor alfa de Estrogênio/metabolismo , Hipertensão/prevenção & controle , Hipertrofia Ventricular Esquerda/prevenção & controle , Nefrite/prevenção & controle , Fatores Etários , Envelhecimento/genética , Angiotensina II , Animais , Aorta Torácica/metabolismo , Aorta Torácica/fisiopatologia , Pressão Arterial , Modelos Animais de Doenças , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Feminino , Hipertensão/induzido quimicamente , Hipertensão/metabolismo , Hipertensão/fisiopatologia , Hipertrofia Ventricular Esquerda/etiologia , Hipertrofia Ventricular Esquerda/metabolismo , Hipertrofia Ventricular Esquerda/fisiopatologia , Rim/metabolismo , Rim/fisiopatologia , Artérias Mesentéricas/metabolismo , Artérias Mesentéricas/fisiopatologia , Camundongos Knockout , Nefrite/etiologia , Nefrite/metabolismo , Nefrite/fisiopatologia , Vasodilatação , Função Ventricular Esquerda , Remodelação VentricularRESUMO
OBJECTIVE: Among obesity-associated metabolic diseases, non-alcoholic fatty liver disease (NAFLD) represents an increasing public health issue due to its emerging association with atherogenic dyslipidemia and cardiovascular diseases (CVDs). The lower prevalence of NAFLD in pre-menopausal women compared with men or post-menopausal women led us to hypothesize that the female-inherent ability to counteract this pathology might strongly rely on estrogen signaling. In female mammals, estrogen receptor alpha (ERα) is highly expressed in the liver, where it acts as a sensor of the nutritional status and adapts the metabolism to the reproductive needs. As in the male liver this receptor is little expressed, we here hypothesize that hepatic ERα might account for sex differences in the ability of males and females to cope with an excess of dietary lipids and counteract the accumulation of lipids in the liver. METHODS: Through liver metabolomics and transcriptomics we analyzed the relevance of hepatic ERα in the metabolic response of males and females to a diet highly enriched in fats (HFD) as a model of diet-induced obesity. RESULTS: The study shows that the hepatic ERα strongly contributes to the sex-specific response to an HFD and its action accounts for opposite consequences for hepatic health in males and females. CONCLUSION: This study identified hepatic ERα as a novel target for the design of sex-specific therapies against fatty liver and its cardio-metabolic consequences.
Assuntos
Dieta Hiperlipídica , Receptor alfa de Estrogênio/metabolismo , Lipídeos/administração & dosagem , Fígado/metabolismo , Caracteres Sexuais , Animais , Receptor alfa de Estrogênio/deficiência , Feminino , Masculino , Camundongos , Camundongos Knockout , Hepatopatia Gordurosa não Alcoólica/metabolismoRESUMO
Loss of ovarian hormones leads to increased adiposity and insulin resistance (IR), increasing the risk for cardiovascular and metabolic diseases. The purpose of this study was to investigate whether the molecular mechanism behind the adverse systemic and adipose tissue-specific metabolic effects of ovariectomy requires loss of signaling through estrogen receptor alpha (ERα) or estrogen receptor ß (ERß). We examined ovariectomized (OVX) and ovary-intactwild-type (WT), ERα-null (αKO), and ERß-null (ßKO) female mice (age ~49 weeks; n = 7-12/group). All mice were fed a phytoestrogen-free diet (<15 mg/kg) and either remained ovary-intact (INT) or were OVX and followed for 12 weeks. Body composition, energy expenditure, glucose tolerance, and adipose tissue gene and protein expression were analyzed. INT αKO were ~25% fatter with reduced energy expenditure compared to age-matched INT WT controls and ßKO mice (all P < 0.001). Following OVX, αKO mice did not increase adiposity or experience a further increase in IR, unlike WT and ßKO, suggesting that loss of signaling through ERα mediates OVX-induced metabolic dysfunction. In fact, OVX in αKO mice (i.e., signaling through ERß in the absence of ERα) resulted in reduced adiposity, adipocyte size, and IR (P < 0.05 for all). ßKO mice responded adversely to OVX in terms of increased adiposity and development of IR. Together, these findings challenge the paradigm that ERα mediates metabolic protection over ERß in all settings. These findings lead us to suggest that, following ovarian hormone loss, ERß may mediate protective metabolic benefits.
Assuntos
Adiposidade/genética , Receptor alfa de Estrogênio/genética , Receptor beta de Estrogênio/genética , Resistência à Insulina/genética , Ovariectomia , Adiponectina/genética , Adiponectina/metabolismo , Tecido Adiposo Branco/metabolismo , Animais , Composição Corporal/genética , Metabolismo Energético/genética , Receptor alfa de Estrogênio/deficiência , Receptor beta de Estrogênio/deficiência , Feminino , Expressão Gênica , Humanos , Leptina/genética , Leptina/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais/genéticaRESUMO
Estrogen deficiency reduces estrogen receptor-alpha (ERα) and promotes apoptosis in the hippocampus, inducing learning-memory deficits; however, underlying mechanisms remain less understood. Here, we explored the molecular mechanism in an ovariectomized (OVX) rat model, hypothesizing participation of autophagy and growth factor signaling that relate with apoptosis. We observed enhanced hippocampal autophagy in OVX rats, characterized by increased levels of autophagy proteins, presence of autophagosomes and inhibition of AKT-mTOR signaling. Investigating upstream effectors of reduced AKT-mTOR signaling revealed a decrease in hippocampal heparin-binding epidermal growth factor (HB-EGF) and p-EGFR. Moreover, 17ß-estradiol and HB-EGF treatments restored hippocampal EGFR activation and alleviated downstream autophagy process and neuronal loss in OVX rats. In vitro studies using estrogen receptor (ERα)-silenced primary hippocampal neurons further corroborated the in vivo observations. Additionally, in vivo and in vitro studies suggested the participation of an attenuated hippocampal neuronal HB-EGF and enhanced autophagy in apoptosis of hippocampal neurons in estrogen- and ERα-deficient conditions. Subsequently, we found evidence of mitochondrial loss and mitophagy in hippocampal neurons of OVX rats and ERα-silenced cells. The ERα-silenced cells also showed a reduction in ATP production and an HB-EGF-mediated restoration. Finally in concordance with molecular studies, inhibition of autophagy and treatment with HB-EGF in OVX rats restored cognitive performances, assessed through Y-Maze and passive avoidance tasks. Overall, our study, for the first time, links neuronal HB-EGF/EGFR signaling and autophagy with ERα and memory performance, disrupted in estrogen-deficient condition.
Assuntos
Receptor alfa de Estrogênio/deficiência , Estrogênios/deficiência , Fator de Crescimento Semelhante a EGF de Ligação à Heparina/metabolismo , Transtornos da Memória/metabolismo , Animais , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Estradiol/farmacologia , Feminino , Hipocampo/metabolismo , Mitocôndrias/metabolismo , Neurônios/metabolismo , Ovariectomia , Ratos , Transdução de SinaisRESUMO
MicroRNAs (miRNAs) regulate activities in living organisms through various signaling pathways and play important roles in the development and progression of osteoporosis. The balance between osteogenic and adipogenic differentiation of rBMSCs is closely related to the occurrence of osteoporosis. ERα regulates bone metabolism in various tissues. However, the correlation among ERα, miRNAs, and the differentiation of rBMSCs is still unclear. In this study, we used lentivirus transfection into rBMSCs to construct an ERα-deficient model, analyzed the differences in expressed miRNAs between control and ERα-deficient rBMSCs. The results revealed that the expression of 25 miRNAs were upregulated, 164 miRNAs were downregulated, and some of the regulated miRNAs such as miR-210-3p and miR-214-3p were related to osteogenic or adipogenic differentiation, as well as to particular signaling pathways. Next, we overexpressed miR-210-3p to evaluate its effects on the osteogenic and adipogenic differentiation of rBMSCs, and identified the relationship among miR-210-3p, Wnt signaling pathway, and the differentiation of rBMSCs. The results indicated that ERα-deficient inhibited osteogenic differentiation, promoted adipogenic differentiation, and regulated the expression of some miRNAs. Meanwhile, overexpression of miR-210-3p promoted osteogenic differentiation and inhibited adipogenic differentiation of rBMSCs, processes likely to be related to the Wnt signaling pathway. In conclusion, we identified a group of upregulated and downregulated miRNAs in ERα-deficient rBMSCs that might play a vital role in regulating osteogenic or adipogenic differentiation. One of these, miR-210-3p, inhibited osteogenic differentiation and promoted adipogenic differentiation correlated with the Wnt signaling pathway in ERα-deficient rBMSCs, providing new insight into the regulation of bone metabolism.
Assuntos
Adipócitos/metabolismo , Adipogenia , Receptor alfa de Estrogênio/deficiência , Células-Tronco Mesenquimais/metabolismo , MicroRNAs/metabolismo , Osteoblastos/metabolismo , Osteogênese , Via de Sinalização Wnt , Células Cultivadas , Receptor alfa de Estrogênio/genética , Regulação da Expressão Gênica , Humanos , MicroRNAs/genética , FenótipoRESUMO
The brain regulates fertility through gonadotropin-releasing hormone (GnRH) neurons. Estradiol induces negative feedback on pulsatile GnRH/luteinizing hormone (LH) release and positive feedback generating preovulatory GnRH/LH surges. Negative and positive feedbacks are postulated to be mediated by kisspeptin neurons in arcuate and anteroventral periventricular (AVPV) nuclei, respectively. Kisspeptin-specific ERα knockout mice exhibit disrupted LH pulses and surges. This knockout approach is neither location-specific nor temporally controlled. We utilized CRISPR-Cas9 to disrupt ERα in adulthood. Mice with ERα disruption in AVPV kisspeptin neurons have typical reproductive cycles but blunted LH surges, associated with decreased excitability of these neurons. Mice with ERα knocked down in arcuate kisspeptin neurons showed disrupted cyclicity, associated with increased glutamatergic transmission to these neurons. These observations suggest that activational effects of estradiol regulate surge generation and maintain cyclicity through AVPV and arcuate kisspeptin neurons, respectively, independent from its role in the development of hypothalamic kisspeptin neurons or puberty onset.
Assuntos
Hipotálamo/fisiologia , Neurônios/fisiologia , Reprodução , Comportamento Sexual Animal , Animais , Estradiol/metabolismo , Receptor alfa de Estrogênio/deficiência , Feminino , Técnicas de Inativação de Genes , Kisspeptinas/análise , Camundongos Knockout , Neurônios/químicaRESUMO
Polycystic ovary syndrome (PCOS) is a hypothalamic-pituitary-gonadal (HPG) axis disorder. PCOS symptoms most likely result from a disturbance in the complex feedback regulation system of the HPG axis, which involves gonadotrophic hormones and ovarian steroid hormones. However, the nature of this complex and interconnecting feedback regulation makes it difficult to dissect the molecular mechanisms responsible for PCOS phenotypes. Global estrogen receptor α (ERα) knockout (KO) mice exhibit a disruption of the HPG axis, resulting in hormonal dysregulation in which female ERα KO mice have elevated levels of serum estradiol (E2), testosterone, and LH. The ERα KO females are anovulatory and develop cystic hemorrhagic ovaries that are thought to be due to persistently high circulating levels of LH from the pituitary. However, the role of ERα in the pituitary is still controversial because of the varied phenotypes reported in pituitary-specific ERα KO mouse models. Therefore, we developed a mouse model where ERα is reintroduced to be exclusively expressed in the pituitary on the background of a global ERα-null (PitERtgKO) mouse. Serum E2 and LH levels were normalized in PitERtgKO females and were comparable to wild-type serum levels. However, the ovaries of PitERtgKO adult mice displayed a more overt cystic and hemorrhagic phenotype when compared with ERα KO littermates. We determined that anomalous sporadic LH secretion caused the severe ovarian phenotype of PitERtgKO females. Our observations suggest that pituitary ERα is involved in the estrogen negative feedback regulation, whereas hypothalamic ERα is necessary for the precise control of LH secretion. Uncontrolled, irregular LH secretion may be the root cause of the cystic ovarian phenotype with similarities to PCOS.-Arao, Y., Hamilton, K. J., Wu, S.-P., Tsai, M.-J., DeMayo, F. J., Korach, K. S. Dysregulation of hypothalamic-pituitary estrogen receptor α-mediated signaling causes episodic LH secretion and cystic ovary.
Assuntos
Receptor alfa de Estrogênio/fisiologia , Hipotálamo/fisiopatologia , Hormônio Luteinizante/metabolismo , Ovário/fisiopatologia , Adeno-Hipófise/fisiopatologia , Síndrome do Ovário Policístico/fisiopatologia , Animais , Modelos Animais de Doenças , Estradiol/fisiologia , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Retroalimentação Fisiológica , Feminino , Hemorragia/etiologia , Humanos , Sistema Hipotálamo-Hipofisário/fisiopatologia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Especificidade de Órgãos , Ovário/patologia , Síndrome do Ovário Policístico/sangue , Síndrome do Ovário Policístico/patologia , Proteínas Recombinantes/metabolismoRESUMO
Clinical and experimental studies demonstrate the potential of low-magnitude high-frequency vibration (LMHFV) to enhance bone formation in the intact skeleton and during fracture healing. Moreover, it was shown that the effects of vibration therapy during fracture healing are highly dependent on the estrogen status of the vibrated individual and that estrogen receptor (ER) α signaling plays a major role in mechanotransduction of LMHFV. Because it is known that LMHFV can directly act on osteogenic cells, we hypothesize that the differential effects of LMHFV in the presence and absence of estrogen are mediated by ERα signaling in osteoblasts. To prove this hypothesis, we subjected preosteoblastic MC3T3-E1 cells and primary osteoblasts to LMHFV in vitro. We found increased Cox2 gene expression, cell metabolic activity and cell proliferation after LMHFV in the absence of estrogen, whereas the effects were contrary in the presence of estrogen. Blocking of ERα signaling by Esr1-siRNA knockdown or adding the selective ERα antagonist MPP dihydrochloride abolished the effects of LMHFV on osteoblast proliferation and Cox2 expression. Furthermore, primary osteoblasts isolated from ERα-knockout mice did not show a response towards LMHFV in the presence of estrogen. Additionally, blocking of actin cytoskeletal remodeling by adding the p160ROCK inhibitor Y-27632 abolished the effects of LMHFV. In contrast, expression of primary cilium was not necessary for mechanotransduction of LMHFV. These results suggest that direct effects of LMHFV on osteoblasts are dependent on ERα signaling and cytoskeletal remodeling.
Assuntos
Estradiol/metabolismo , Receptor alfa de Estrogênio/genética , Fraturas Ósseas/terapia , Mecanotransdução Celular , Osteoblastos/metabolismo , Osteogênese/genética , Vibração/uso terapêutico , Citoesqueleto de Actina/efeitos dos fármacos , Citoesqueleto de Actina/metabolismo , Citoesqueleto de Actina/ultraestrutura , Amidas/farmacologia , Animais , Diferenciação Celular , Linhagem Celular , Proliferação de Células , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Estradiol/farmacologia , Receptor alfa de Estrogênio/antagonistas & inibidores , Receptor alfa de Estrogênio/deficiência , Feminino , Consolidação da Fratura/efeitos dos fármacos , Fraturas Ósseas/genética , Fraturas Ósseas/metabolismo , Fraturas Ósseas/patologia , Regulação da Expressão Gênica , Camundongos , Camundongos Endogâmicos C57BL , Osteoblastos/citologia , Osteoblastos/efeitos dos fármacos , Piperidinas/farmacologia , Cultura Primária de Células , Pirazóis/farmacologia , Piridinas/farmacologia , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Quinases Associadas a rho/antagonistas & inibidores , Quinases Associadas a rho/genética , Quinases Associadas a rho/metabolismoRESUMO
Estrogen has been implicated in the regulation of growth and immune function in the kidney, which expresses the full-length estrogen receptor-α (ERα66), its ERα splice variants, and estrogen receptor-ß (ERß). Thus, we hypothesized that these splice variants may inhibit the glomerular enlargement that occurs early in type 1 diabetes (T1D). T1D was induced by streptozotocin (STZ) injection in 8- to 12-wk-old female mice lacking ERα66 (ERα66KO) or all ERα variants (αERKO), and their wild-type (WT) littermates. Basal renal ERα36 protein expression was reduced in the ERα66KO model and was downregulated by T1D in WT mice. T1D did not alter ERα46 or ERß in WT-STZ; however, ERα46 was decreased modestly in ERα66KO mice. Renal hypertrophy was evident in all diabetic mice. F4/80-positive immunostaining was reduced in ERα66KO compared with WT and αERKO mice but was higher in STZ than in Control mice across all genotypes. Glomerular area was greater in WT and αERKO than in ERα66KO mice, with T1D-induced glomerular enlargement apparent in WT-STZ and αERKO-STZ, but not in ERα66KO-STZ mice. Proteinuria and hyperfiltration were evident in ERα66KO-STZ and αERKO-STZ, but not in WT-STZ mice. These data indicate that ERα splice variants may exert an inhibitory influence on glomerular enlargement and macrophage infiltration during T1D; however, effects of splice variants are masked in the presence of the full-length ERα66, suggesting that ERα66 acts in opposition to its splice variants to influence these parameters. In contrast, hyperfiltration and proteinuria in T1D are attenuated via an ERα66-dependent mechanism that is unaffected by splice variant status.
Assuntos
Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Nefropatias Diabéticas/prevenção & controle , Receptor alfa de Estrogênio/metabolismo , Glomérulos Renais/metabolismo , Animais , Glicemia/metabolismo , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Tipo 1/induzido quimicamente , Diabetes Mellitus Tipo 1/genética , Nefropatias Diabéticas/induzido quimicamente , Nefropatias Diabéticas/genética , Nefropatias Diabéticas/metabolismo , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Feminino , Taxa de Filtração Glomerular , Glomérulos Renais/patologia , Glomérulos Renais/fisiopatologia , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Isoformas de Proteínas , Proteinúria/genética , Proteinúria/metabolismo , Proteinúria/prevenção & controle , Estreptozocina , Aumento de PesoRESUMO
Human papillomavirus (HPV) is required but not sufficient for cervical carcinoma (CxCa) development. Oestradiol (E2 ) promotes CxCa development in K14E7 transgenic mice expressing the HPV16 E7 oncoprotein under the control of the keratin (K14) promoter. E2 mainly functions through oestrogen receptor α (ERα). However, the role of ERα in human CxCa has been underappreciated largely because it is not expressed in carcinoma cells. We have shown that deletion of Esr1 (the ERα-coding gene) in the cervical stroma of K14E7 mice promotes regression of cervical intraepithelial neoplasia (CIN), the precursor lesion of CxCa. Here, we deleted Esr1 in the cervical epithelium but not in the stroma. We found that E2 induced cervical epithelial cell proliferation in epithelial ERα-deficient mice. We also found that E2 promoted the development of CIN and CxCa in epithelial ERα-deficient K14E7 mice and that all neoplastic epithelial cells were negative for ERα. In addition, proliferation indices were similar between ERα- and ERα+ CxCa. These results indicate that epithelial ERα is not necessary for E2 -induced CIN and CxCa. Taking these findings together, we conclude that stromal ERα rather than epithelial ERα mediates oncogenic E2 signalling in CxCa. Our results support stromal ERα signalling as a therapeutic target for the disease. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
Assuntos
Células Epiteliais/metabolismo , Estradiol , Receptor alfa de Estrogênio/metabolismo , Displasia do Colo do Útero/metabolismo , Neoplasias do Colo do Útero/metabolismo , Animais , Diferenciação Celular , Proliferação de Células , Transformação Celular Neoplásica/induzido quimicamente , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Transformação Celular Neoplásica/patologia , Modelos Animais de Doenças , Células Epiteliais/patologia , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Feminino , Queratina-14/genética , Camundongos Knockout , Proteínas Oncogênicas Virais/genética , Proteínas Oncogênicas Virais/metabolismo , Ovariectomia , Regiões Promotoras Genéticas , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Transdução de Sinais , Células Estromais/metabolismo , Células Estromais/patologia , Neoplasias do Colo do Útero/induzido quimicamente , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologia , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , Displasia do Colo do Útero/induzido quimicamente , Displasia do Colo do Útero/genética , Displasia do Colo do Útero/patologiaRESUMO
Estrogen has been shown to affect vascular reactivity. Here, we assessed the estrogen receptor-α (ERα) dependency of estrogenic effects on vasorelaxation via a rapid nongenomic pathway in both male and ovary-intact female mice. We compared the effect of a primary estrogen, 17ß-estradiol (E2) or 4,4',4â³-(4-propyl-[1H]pyrazole-1,3,5-triyl)tris-phenol (PPT; selective ERα agonist). We found that E2 and PPT induced greater aortic relaxation in female mice than in male mice, indicating ERα mediation, which was further validated by using ERα antagonism. Treatment with 1,3-bis(4-hydroxyphenyl)-4-methyl-5-[4-(2-piperidinylethoxy)phenol]-1H-pyrazole dihydrochloride (MPP dihydrochloride; ERα antagonist) attenuated PPT-mediated vessel relaxation in both sexes. ERα-mediated vessel relaxation was further validated by the absence of significant PPT-mediated relaxation in aortas isolated from ERα knockout mice. Treatment with a specific ERK inhibitor, PD-98059, reduced E2-induced vessel relaxation in both sexes but to a lesser extent in female mice. Furthermore, PD-98059 prevented PPT-induced vessel relaxation in both sexes. Both E2 and PPT treatment activated ERK as early as 5-10 min, which was attenuated by PD-98059 in aortic tissue, cultured primary vascular smooth muscle cells (VSMCs), and endothelial cells (ECs). Aortic rings denuded of endothelium showed no differences in vessel relaxation after E2 or PPT treatment, implicating a role of ECs in the observed sex differences. Here, our results are unique to show estrogen-stimulated rapid ERα signaling mediated by ERK activation in aortic tissue, as well as VSMCs and ECs in vitro, in regulating vascular function by using side-by-side comparisons in male and ovary-intact female mice in response to E2 or PPT. NEW & NOTEWORTHY Here, we assessed the estrogen receptor-α dependency of estrogenic effects in vasorelaxation of both male and ovary-intact female mice by performing side-by-side comparisons. Also, we describe the connection between estrogen-stimulated rapid estrogen receptor-α signaling and downstream ERK activation in regulating vascular function in male and ovary-intact female mice.
Assuntos
Aorta Torácica/efeitos dos fármacos , Estradiol/farmacologia , Receptor alfa de Estrogênio/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fenóis/farmacologia , Pirazóis/farmacologia , Vasodilatação/efeitos dos fármacos , Vasodilatadores/farmacologia , Animais , Aorta Torácica/enzimologia , Células Cultivadas , Relação Dose-Resposta a Droga , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/enzimologia , Ativação Enzimática , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Feminino , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/enzimologia , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/enzimologia , Fatores Sexuais , Transdução de Sinais/efeitos dos fármacosRESUMO
Estrogen receptor α (ERα) regulates gene transcription through two activation functions (ERα-AF1 and ERα-AF2). We recently found that the protection conferred by 17ß-estradiol against obesity and insulin resistance requires ERα-AF2 but not ERα-AF1. However, the interplay between the two ERα-AFs is poorly understood in vivo and the metabolic influence of a specific ERα-AF1 action remains to be explored. To this end, wild-type, ERα-deficient, or ERα-AF1-deficient ovariectomized female mice were fed a high-fat diet and concomitantly administered with vehicle or tamoxifen, a selective ER modulator that acts as a ERα-AF1 agonist/ERα-AF2 antagonist. In ovariectomized wild-type mice, tamoxifen significantly reduced food intake and totally prevented adiposity, insulin resistance, and steatosis. These effects were abolished in ERα-deficient and ERα-AF1-deficient mice, revealing the specific role of ERα-AF1 activation. Finally, hepatic gene expression changes elicited by tamoxifen in wild-type mice were abrogated in ERα-AF1-deficient mice. The combination of pharmacologic and transgenic approaches thus indicates that selective ERα-AF1 activation by tamoxifen is sufficient to elicit metabolic protection, contrasting with the specific requirement of ERα-AF2 in the metabolic actions of 17ß-estradiol. This redundancy in the ability of the two ERα-AFs to separately mediate metabolic prevention strikingly contrasts with the contribution of both ERα-AFs in breast cancer proliferation, shedding new light on the therapeutic potential of selective ER modulation.