Selective Estrogen Receptor Agonists Research Articles

MiR-206 inhibits estrogen signaling and ovarian cancer cell migration without affecting GPER

AimsEstrogen-regulated pathways are involved in the etiology and progression of epithelial ovarian cancer (EOC), but the relative contribution of estrogen receptor isoforms is unclear. Only a subset of patients responds to antiestrogens including tamoxifen. Based on our previous evidence that miR-206 behaves as an oncosuppressor in EOC, we hypothesized that miR-206 would interfere with G protein-coupled estrogen receptor (GPER)-mediated signaling and cell motility. Main methodsPFKFB3 and FAK proteins from OC cells challenged with selective estrogen receptor agonist and antagonist were measured by Western blotting. Cell proliferation and motility were analyzed by MTT and Boyden chamber, respectively. Estrogen-dependent cells were transfected with miR-206 mimic or control using Lipofectamine. Key findingsThe migration of SKOV3 and OVCAR5 cells significantly increased following treatment with 17β-estradiol (E2) and the selective GPER agonist G1. However, tamoxifen failed to inhibit E2 effect and even promoted SKOV3 cell migration. Estrogen receptor ligands did not affect SKOV3 proliferation. The GPER antagonist G15 significantly prevented E2-mediated upregulation of PFKFB3 expression, while G1 concentration-dependently upregulated PFKFB3 levels. Consistent with the functional link between PFKFB3 and FAK activation, E2 and G1 increased FAK phosphorylation at Tyr397. Transfection with miR-206 abolished estrogen-induced EOC migration and down-regulated PFKFB3 protein levels. Notably, miR-206 transfection reduced ERα protein abundance, whereas GPER amount was unchanged. SignificanceBy blocking estrogen signaling and G1-induced EOC cell invasiveness with no direct interference with GPER levels, miR-206 mimics have the potential to act as pathway-selective antagonists and deserve further testing as RNA therapeutics in estrogen-dependent EOC.

Endocrinology for the hepatologist.

Endocrinology for the hepatologist.

Cyclopia extracts act as selective estrogen receptor subtype downregulators in estrogen receptor positive breast cancer cell lines: Comparison to standard of care breast cancer endocrine therapies and a selective estrogen receptor agonist and antagonist.

Breast cancer is the most diagnosed type of cancer amongst women in economically developing countries and globally. Most breast cancers express estrogen receptor alpha (ERα) and are categorized as positive (ER+) breast cancer. Endocrine therapies such as, selective estrogen receptor modulators (SERMs), aromatase inhibitors (AIs), and selective estrogen receptor downregulators (SERDs) are used to treat ER+ breast cancer. However, despite their effectiveness, severe side-effects and resistance are associated with these endocrine therapies. Thus, it would be highly beneficial to develop breast cancer drugs that are as effective as current therapies, but less toxic with fewer side effects, and less likely to induce resistance. Extracts of Cyclopia species, an indigenous South African fynbos plant, have been shown to possess phenolic compounds that exhibit phytoestrogenic and chemopreventive activities against breast cancer development and progression. In the current study, three well characterized Cyclopia extracts, SM6Met, cup of tea (CoT) and P104, were examined for their abilities to modulate the levels of the estrogen receptor subtypes, estrogen receptor alpha and estrogen receptor beta (ERβ), which have been recognized as crucial to breast cancer prognosis and treatment. We showed that the Cyclopia subternata Vogel (C. subternata Vogel) extracts, SM6Met and cup of tea, but not the C. genistoides extract, P104, reduced estrogen receptor alpha protein levels while elevating estrogen receptor beta protein levels, thereby reducing the ERα:ERβ ratio in a similar manner as standard of care breast cancer endocrine therapies such as fulvestrant (selective estrogen receptor downregulator) and 4-hydroxytamoxifen (elective estrogen receptor modulator). Estrogen receptor alpha expression enhances the proliferation of breast cancer cells while estrogen receptor beta inhibits the proliferative activities of estrogen receptor alpha. We also showed that in terms of the molecular mechanisms involved all the Cyclopia extracts regulated estrogen receptor alpha and estrogen receptor beta protein levels through both transcriptional and translational, and proteasomal degradation mechanisms. Therefore, from our findings, we proffer that the C. subternata Vogel extracts, SM6Met and cup of tea, but not the C. genistoides extract, P104, selectively modulate estrogen receptor subtypes levels in a manner that generally supports inhibition of breast cancer proliferation, thereby demonstrating attributes that could be explored as potential therapeutic agents for breast cancer.

Anticancer Drug Conjugates Incorporating Estrogen Receptor Ligands.

Hormone-dependent cancers, such as certain types of breast cancer are characterized by over-expression of estrogen receptors (ERs). Anticancer drug conjugates combining ER ligands with other classes of anticancer agents may not only benefit from dual action at both anti-cancer targets but also from selective delivery of cytotoxic agents to ER-positive tumor cells resulting in less toxicity and adverse effects. Moreover, they could also take advantage of overcoming resistance typical for anti-hormonal monotherapy such as tamoxifen. In this review, we discuss the design, structures and pharmacological effects of numerous series of drug conjugates containing ER ligands such as selective ER modulators (tamoxifen, 4-hydroxytamoxifen, endoxifen), selective ER degraders (ICI-164384) and ER agonists (estradiol) linked to diverse anti-cancer agents including histone-deacetylase inhibitors, DNA-alkylating agents, antimitotic agents and epidermal growth factor receptor inhibitors.

Selective ER-β agonists alleviate neuronal deficits in insulin-resistant estrogen-deficient rats

Objective The present study aimed to determine the effect of estrogen receptor (ER) agonists on depression and memory impairment in insulin-resistant ovariectomized (OVX) rats. Methods Rats underwent bilateral ovariectomy, and low-dose streptozotocin (STZ) and a high-fat diet (58% fat, 25% protein, and 17% carbohydrates as a percentage of kilocalories) were administered to induce an estrogen-deficient insulin-resistant state. After 1 week of STZ administration, rats were treated with 17β-estradiol (17βE2) and selective ER-α (propylpyrazoletriol) and ER-β (diarylpropionitrile) agonists (10 μg/kg subcutaneously). Memory was evaluated using the Morris water maze and depression using the forced swim test. Results Treatment with selective ER-β agonist and 17βE2 but not with selective ER-α agonist significantly modulated the neurobehavioral deficits in insulin-resistant OVX rats. These neurobehavioral parameters were further correlated with brain-derived neurotrophic factor (BDNF) levels and acetylcholinesterase (AChE) activity. Selective ER-β agonist and 17βE2 significantly modulated BDNF levels and AChE activity in insulin-resistant OVX rats. Significant increases in estradiol and uterine weight were observed in 17βE2-treated rats, but selective ER agonists did not show any effect. Conclusion ER-β agonist can be an effective strategy for the mitigation of memory loss and depression in an estrogen-deficient insulin-resistant state without all of the deleterious feminizing effects that occur with the use of 17βE2.

The genitourinary syndrome of menopause.

Genitourinary syndrome of menopause (GSM) refers to a collection of symptoms resulting from diminished hormonal, primarily estrogenic stimulation to the vulvovaginal or lower urinary tract and may affect up to 50% of postmenopausal women. Symptoms, which are typically progressive and unlikely to resolve spontaneously, may include, but are not limited to, vulvovaginal dryness, burning or irritation, dyspareunia, or urinary symptoms of urgency, dysuria or recurrent urinary tract infection. These symptoms are typically progressive and unlikely to resolve spontaneously. Diagnosis is clinical. Telemedicine may play a role in diagnosis, initiation of treatment, and follow-up of women with GSM. Effective treatments include moisturizers and lubricants, local hormonal therapy with estrogen or dehydroepiandrosterone, and oral selective estrogen receptor agonists. Laser or radiofrequency procedures, although currently utilized, are being studied to comprehensively understand their overall effectiveness and safety. Additionally, the influence and effect of the vaginal microbiome, as well as potential of treatment via its manipulation, is being studied. We performed a literature search of PubMed, Google Scholar, and Ovid with search terms of vulvovaginal atrophy and GSM and reviewed major US Society Guidelines to create this narrative review of this topic. The literature suggests that healthcare providers can make a significant impact of the health and quality of life of women by being proactive about discussing and providing interventions for GSM. A systematic approach with consideration of current guidelines and attention to developing protocols for interventions should be employed.

Role of sex hormones and their receptors on gastric Nrf2 and neuronal nitric oxide synthase function in an experimental hyperglycemia model

BackgroundGastroparesis, a condition of abnormal gastric emptying, is most commonly observed in diabetic women. To date, the role of ovarian hormones and/or gastric hormone receptors on regulating nitrergic-mediated gastric motility remains inconclusive.AimThe purpose of this study is to investigate whether sex hormones/their receptors can attenuate altered Nuclear factor (erythroid-derived 2)-like 2 (Nrf2), neuronal Nitric Oxide Synthase (nNOS) expression and nitrergic relaxation in gastric neuromuscular tissues exposed to in-vitro hyperglycemia (HG).MethodsGastric neuromuscular sections from adult female C57BL/6 J mice were incubated in normoglycemic (NG, 5 mM) or hyperglycemic (30 mM or 50 mM) conditions in the presence or absence of selective estrogen receptor (ER) agonists (ERα /PPT or ERβ: DPN); or non-selective sex hormone receptor antagonists (ER/ICI 182,780, or progesterone receptor (PR)/ RU486) for 48 h. mRNA, protein expression and nitrergic relaxation of circular gastric neuromuscular strips were assessed.ResultsOur findings in HG, compared to NG, demonstrate a significant reduction in ER, Nrf2, and nNOS expression in gastric specimens. In addition, in-vitro treatment with sex hormones and/or their agonists significantly (*p < 0.05) restored Nrf2/nNOSα expression and total nitrite production. Conversely, ER, but not PR, antagonist significantly reduced Nrf2/nNOSα expression and nitrergic relaxation.ConclusionsOur data suggest that ER’s can regulate nitrergic function by improving Nrf2/nNOS expression in experimental hyperglycemia.

Impact of estrogen receptor agonists and model of menopause on enzymes involved in brain metabolism, acetyl-CoA production and cholinergic function

Our goal is to understand how loss of circulating estrogens and estrogen replacement affect brain physiology and function, particularly in brain regions involved in cognitive processes. We recently conducted a large metabolomics study characterizing the effects of rodent models of menopause and treatment with estrogen receptor (ER) agonists on neurochemical targets in hippocampus, frontal cortex, and striatum. Here we characterize effects on levels of several key enzymes involved in glucose utilization and energy production, specifically phosphofructokinase, glyceraldehyde 3-phosphate dehydrogenase, and pyruvate dehydrogenase. We also evaluated effects on levels of β-actin and α-tubulin, choline acetyltransferase (ChAT) activity, and levels of ATP citrate lyase. All experiments were conducted in young adult rats. Experiment 1 compared the effects of ovariectomy (OVX), a model of surgical menopause, and 4-vinylcyclohexene diepoxide (VCD)-treatments, a model of transitional menopause, with tissues collected at proestrus and at diestrus. Experiment 2 used a separate cohort of rats to evaluate the same targets in OVX and VCD-treated rats treated with estradiol or with selective ER agonists. Differences in the expression of metabolic enzymes between cycling animals and models of surgical and transitional menopause were detected. These differences were model-, region- and time- dependent, and were modulated by selective ER agonists. Collectively, the findings demonstrate that loss of ovarian function and ER agonist treatments have differing effects in OVX vs. VCD-treated rats. Differences may help to explain differences in the effects of estrogen treatments on brain function and cognition in women who have experienced surgical vs. transitional menopause.

Sex‐Specific Estrogen Regulation of Hypothalamic Astrocyte Estrogen Receptor and Glycogen Metabolic Enzyme Protein Expression, and Glycogen Content in Rats

Brain astrocytes are implicated in estrogenic neuroprotection against bio‐energetic insults, which may involve their glycogen energy reserve. Forebrain estrogen receptor (ER)‐alpha (ERα) and ‐beta (ERβ) exert differential control of glycogen metabolic enzyme [glycogen synthase (GS); phosphorylase (GP)] expression in hypoglycemic male versus female rats. Studies were conducted using a rat hypothalamic astrocyte primary culture model along with selective ER agonists to investigate the premise that estradiol (E2) exerts sex‐dimorphic control over astrocyte glycogen mass and metabolism. Female astrocyte GS and GP profiles are more sensitive to E2 stimulation than the male. E2 did not regulate expression of phospho‐GS (inactive enzyme form) in either sex. Data also show that transmembrane G protein‐coupled ER‐1 (GPER) signaling is implicated in E2 control of GS profiles in each sex, and regulates GP expression in females. E2 increases total 5′‐AMK‐activated protein kinase (AMPK) protein in female astrocytes, but stimulated pAMPK (active form) expression with equivalent potency via GPER in females and ERα in males. In female astrocytes, ERα protein was up‐regulated at a lower E2 concentration and over a broader dosage range compared to males, whereas ERβ was increased after exposure to 1–10 nM versus 100 pM E2 levels in females and males, respectively. GPER profiles were stimulated by E2 in female, but not male astrocytes. E2 increased astrocyte glycogen content in female, but not male astrocytes; selective ERβ or ERα stimulation elevated glycogen levels in the female and male, respectively. Outcomes imply that dimorphic astrocyte protein and glycogen responses to E2 may reflect, in part, differential steroid induction of ER variant expression and/or regulation of post‐receptor signaling in each sex.Support or Funding InformationThis research was supported by NIH DK 109382.

Rapid effects of estradiol and its receptor agonists on object recognition and object placement in adult male zebrafish

In recent years, there has been a growing appreciation that 17β-estradiol (E2) can rapidly modulate learning and memory processes by binding to membrane estrogen receptors and cause the activation of a number of signaling cascades within the central nervous system. In this study, we sought to investigate the effects of post-training administration of E2 (100 ng/g, 1 μg/g, 10 μg/g) and involvement of the estrogen receptors (ERs) using selective ER agonists on the consolidation of object recognition (OR) and object placement memory (OP) in adult male zebrafish. The general activation of ERs with the highest E2 dose improved consolidation of memory in both learning tasks within 1.45 h of administration. Activation of classical ERs (ERα and ERβ) improved consolidation of OR memory, but had no effect on fish performance in OP task. On the other hand, activation of G protein-coupled ER1 impaired and enhanced consolidation of OR and OP memories, respectively. Memory improvement in both tasks was accompanied by a marked up-regulation in the expression of genes encoding ionotropic and metabotropic glutamate receptors in a task-dependent manner. In contrast, the down-regulation in the expression of certain ionotropic glutamate receptors was observed in fish with impaired OR memory. Moreover, our study also revealed an increase in the transcript abundance of genes associated with synaptic protein synthesis (brain-derived neurotrophic factor, synaptophysin, and the mechanistic target of rapamycin). These results suggest that E2 may affect consolidation of memory in zebrafish likely through rapid changes in synaptic morphology and function.

Sex-specific estrogen regulation of hypothalamic astrocyte estrogen receptor expression and glycogen metabolism in rats

Brain astrocytes are implicated in estrogenic neuroprotection against bio-energetic insults, which may involve their glycogen energy reserve. Forebrain estrogen receptors (ER)-alpha (ERα) and -beta (ERβ) exert differential control of glycogen metabolic enzyme [glycogen synthase (GS); phosphorylase (GP)] expression in hypoglycemic male versus female rats. Studies were conducted using a rat hypothalamic astrocyte primary culture model along with selective ER agonists to investigate the premise that estradiol (E2) exerts sex-dimorphic control over astrocyte glycogen mass and metabolism. Female astrocyte GS and GP profiles are more sensitive to E2 stimulation than the male. E2 did not regulate expression of phospho-GS (inactive enzyme form) in either sex. Data also show that transmembrane G protein-coupled ER-1 (GPER) signaling is implicated in E2 control of GS profiles in each sex and alongside ERα, GP expression in females. E2 increases total 5′-AMP-activated protein kinase (AMPK) protein in female astrocytes, but stimulated pAMPK (active form) expression with equivalent potency via GPER in females and ERα in males. In female astrocytes, ERα protein was up-regulated at a lower E2 concentration and over a broader dosage range compared to males, whereas ERβ was increased after exposure to 1–10 nM versus 100 pM E2 levels in females and males, respectively. GPER profiles were stimulated by E2 in female, but not male astrocytes. E2 increased astrocyte glycogen content in female, but not male astrocytes; selective ERβ or ERα stimulation elevated glycogen levels in the female and male, respectively. Outcomes imply that dimorphic astrocyte ER and glycogen metabolic responses to E2 may reflect, in part, differential steroid induction of ER variant expression and/or regulation of post-receptor signaling in each sex.

Estradiol and selective estrogen receptor agonists differentially affect brain monoamines and amino acids levels in transitional and surgical menopausal rat models

Estrogens have many beneficial effects in the brain. Previously, we evaluated the effects of two models of menopause (surgical vs. transitional) on multiple monoaminergic endpoints in different regions of the adult rat brain in comparison with levels in gonadally intact rats. Here we evaluated the effects of estrogen receptor (ER) agonist treatments in these same two models of menopause. Neurochemical endpoints were evaluated in the hippocampus (HPC), frontal cortex (FCX), and striatum (STR) of adult ovariectomized (OVX) rats and in rats that underwent selective and gradual ovarian follicle depletion by daily injection of 4-vinylcyclohexene-diepoxide (VCD), after 1- and 6-weeks treatment with 17β-estradiol (E2), or with selective ERα (PPT), ERβ (DPN), or GPR30 (G-1) agonists. Endpoints included serotonin (5-HT) and 5-Hydroxyindoleacetic acid, dopamine (DA), 3,4-Dihydroxyphenylacetic acid and homovanillic acid, norepinephrine (NE) and epinephrine, as well as the amino acids tryptophan (TRP) and tyrosine (TYR). Significant differences between the models were detected. OVX rats were much more sensitive to ER agonist treatments than VCD-treated rats. Significant differences between brain regions also were detected. Within OVX rats, more agonist effects were detected in the HPC than in any other region. One interesting finding was the substantial decrease in TRP and TYR detected in the HPC and FCX in response to agonist treatments, particularly in OVX rats. This is on top of the substantial decreases in TRP and TYR previously reported one week after OVX or VCD-treatments in comparison with gonadally intact controls. Other interesting findings included increases in the levels of 5-HT, DA, and NE in the HPC of OVX rats treated with DPN, increases in DA detected in the FCX of OVX rats treated with any of the ER agonists, and increases in 5-HT and DA detected in the STR of OVX rats treated with E2. Many effects that were observed after 1-week of treatment were no longer observed after 6-weeks of treatment, demonstrating that effects were temporary despite continued agonist treatment. Collectively, the results demonstrate significant differences in the effects of ER agonists on monoaminergic endpoints in OVX vs. VCD-treated rats that also were brain region-specific and time dependent. The fact that agonist treatments had lesser effects in VCD treated rats than in OVX rats may help to explain reports of lesser effects of estrogen replacement on cognitive performance in women that have undergone transitional vs. surgical menopause.

203 - Estradiol and estrogen receptor-α and β agonist’s effects on ROS and RNS depend on cell culture oxygen levels

Oxygen affects various aspects of mammalian cell biology including reactive oxygen species (ROS) and reactive nitrogen species (RNS) metabolism, which are both important in normal cell signaling and in mediating pathological damage to macromolecules. Under standard culture conditions, cells are grown in near-atmospheric oxygen levels (~18% O2; hyperoxia), and thus exposed to significantly higher levels of oxygen than most cell types experience in vivo (1-5% O2; physioxia) [1]. The consequences of growing cells in non-physiological hyperoxia have been under-appreciated by researchers. The hormone estradiol (E2) and the selective estrogen receptor-β agonist diarylpropiontrile (DPN) have been shown to alter cellular ROS and RNS metabolism in vitro in mammalian cells [2]. However, much of our understanding of this is based on data obtained from cell culture using standard, hyperoxic, conditions. Here we measured cellular H2O2 production in C2C12 mouse myoblasts treated for 48h with either E2, DPN, or the estrogen receptor-α-specific agonist PPT, at either 18% or 5% O2. Our results were remarkably dependent on the prevailing O2 levels during the experiment. For example, while DPN and E2 both decreased cellular H2O2 production in experiments performed at 18% they increased it at 5% O2. Concomitantly, the effects of E2, DPN and PPT on the expression of cellular NADPH oxidases (NOXs) at 18% O2 were opposite to the effects at 5%. This result, and others like it, indicate the importance of physiological O2 levels for studying the effects of E2 and selective estrogen receptor agonists in cell culture.

Relaxant Effects of the Selective Estrogen Receptor Modulator, Bazedoxifene, and Estrogen Receptor Agonists in Isolated Rabbit Basilar Artery.

We have previously shown that the selective estrogen receptor modulator, bazedoxifene, improves the consequences of ischemic stroke. Now we aimed to characterize the effects and mechanisms of action of bazedoxifene in cerebral arteries. Male rabbit isolated basilar arteries were used for isometric tension recording and quantitative polymerase chain reaction. Bazedoxifene relaxed cerebral arteries, as 17-β-estradiol, 4,4',4″-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol [estrogen receptor (ER) α agonist], and G1 [G protein-coupled ER (GPER) agonist] did it (4,4',4″-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol > bazedoxifene = G1 > 17-β-estradiol). 2,3-Bis(4-hydroxyphenyl)-propionitrile (ERβ agonist) had no effect. Expression profile of genes encoding for ERα (ESR1), ERβ (ESR2), and GPER was GPER > ESR1 > ESR2. As to the endothelial mechanisms, endothelium removal, N-nitro-L-arginine methyl ester, and indomethacin, did not modify the relaxant responses to bazedoxifene. As to the K channels, both a high-K medium and the Kv blocker, 4-aminopyridine, inhibited the bazedoxifene-induced relaxations, whereas tetraethylammonium (nonselective K channel blocker), glibenclamide (selective KATP blocker) or iberiotoxin (selective KCa blocker) were without effect. Bazedoxifene also inhibited both Ca- and Bay K8644-elicited contractions. Therefore, bazedoxifene induces endothelium-independent relaxations of cerebral arteries through (1) activation of GPER and ERα receptors; (2) increase of K conductance through Kv channels; and (3) inhibition of Ca entry through L-type Ca channels. Such a profile is compatible with the beneficial effects of estrogenic compounds (eg, SERMs) on vascular function and, specifically, that concerning the brain. Therefore, bazedoxifene could be useful in the treatment of cerebral disorders in which the cerebrovascular function is compromised (eg, stroke).

Dietary Genistein Prevents Denervation-Induced Muscle Atrophy in Male Rodents via Effects on Estrogen Receptor-α

Background: Genistein has high estrogenic activity. Previous studies have shown beneficial effects of estrogen or hormone replacement therapy on muscle mass and muscle atrophy.Objective: We investigated the preventive effects and underlying mechanisms of genistein on muscle atrophy.Methods: In Expt. 1, male Wistar rats were fed a diet containing no genistein [control (CON)] or 0.05% genistein (GEN; wt:wt diet) for 24 d. On day 14, the sciatic nerve in the left hind leg was severed, and the right hind leg was sham-treated. In Expt. 2, male C57BL6J mice were subcutaneously administered a vehicle (Veh group) or the estrogen receptor (ER) antagonist ICI 182,780 (ICI group) via an osmotic pump for 27 d, and each group was subsequently fed CON or GEN diets from day 3 to day 27. Muscle atrophy was induced on day 17 as in Expt. 1. In Expt. 3, male C57BL6J mice were subcutaneously administered vehicle or a selective ER agonist—ER-α [4,4′,4′-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol (PPT)] or ER-β [2,3-bis(4-hydroxyphenyl)-propionitrile (DPN)]—or genistein (GEN-sc-i) via an osmotic pump for 13 d, and muscle atrophy was induced on day 3 as in Expt. 1. The ratio of denervated soleus muscle weight to sham-operated soleus muscle weight (d/s ratio) was used as the index of muscle atrophy.Results: Expt. 1: The d/s ratio in the GEN group was 20% higher than that in the CON group (P < 0.05). Expt. 2: The d/s ratio in the Veh-GEN group was 14% higher than that in the Veh-CON group (P < 0.05), although there was no significant difference between ICI-CON and ICI-GEN groups (P = 0.69). Expt. 3: The d/s ratio in the PPT-treated group was 20% greater than that in the Veh group (P < 0.05), but DPN and GEN-sc-i had no effect on the d/s ratio (P ≥ 0.05 compared with vehicle).Conclusion: Genistein intake mitigated denervation-induced soleus muscle atrophy. ER-α was related to the preventive effect of genistein on muscle atrophy.

Estrogen replacement therapy-induced neuroprotection against brain ischemia-reperfusion injury involves the activation of astrocytes via estrogen receptor β.

The incidence of ischemic stroke is significantly increased in postmenopausal women. However, the neuroprotective effects of estrogen replacement therapy (ERT) against stroke remain controversial, and the role of astrocytes in ERT has rarely been explored. In this study, we investigated the effects of estrogen and selective estrogen receptor (ER) agonists on astrocytes activation and neuronal apoptosis in mice under conditions of cell culture oxygen and glucose deprivation and reperfusion (OGD-R), and global cerebral ischemia (GCI). We demonstrated that hippocampal astrocytes primarily express ERβ. In astrocytes, 2.5–20 nM 17β-estradiol (E2) or 10 nM DPN (ERβ agonist) not 10 nM PPT (ERα agonist), significantly increased GFAP expression. And 10 nM E2, DPN or E2+MPP (ERα antagonist), but not PPT or E2+PHTPP (ERβ antagonist), significantly reduced neuronal apoptosis following the subjection of astrocyte and neuronal cocultures to OGD-R. We also found that either 50 μg/kg E2 or 8 mg/kg DPN replacement (3 weeks) significantly increased GFAP expression and reduced GCI-induced neuronal apoptosis in hippocampal CA1 region of ovariectomized mice. These results indicate that estrogen-induced neuroprotection against ischemia-reperfusion injury involves activation of astrocytes via ERβ. Thus, the discovery and design of astrocyte-selective ERβ modulators may offer a new strategy for ERT of ischemic stroke.

Differential role of estrogen receptor modulators in depression-like behavior and memory impairment in rats with postmenopausal diabetes.

Diabetes and menopause are frequent comorbidities. The objective of the present study was to delineate the effects of nonselective and selective estrogen receptor (ER) agonists (α and β) on cognitive function and depressive behavior in ovariectomized diabetic (Ovx-Dia) rats. Bilateral ovariectomy was performed in female Sprague-Dawley rats (200-250 g), and streptozotocin was used to induce experimental diabetes. Rats were administered (10 μg/kg SC) a nonselective ER agonist, 17β-estradiol (E2); a selective ER-α agonist, propionitrile (PPT; 4,4',4″-(4-propyl-[1H] pyrazole-1,3,5-triyl) tris phenol); and a selective ER-β agonist, 2,3-bis(4-hydroxyphenyl)-propionitrile (DPN) for 4 weeks after streptozotocin injection. Marked impairment in memory indicated by increased transfer latency on the fifth day (363%) and increased immobility time (90.5%), coupled with a marked decrease in brain-derived neurotrophic factors (22.5%) and increase in acetylcholinesterase activity (58.1%), were observed in Ovx-Dia rats compared with sham rats. However, a partial change in all these parameters was observed in ovariectomized or diabetic rats compared with sham rats. Treatment with DPN and E2 markedly prevented--whereas treatment with PPT partially prevented--changes induced in Ovx-Dia rats. To assess feminizing action, we measured serum estradiol levels and uterine weights. E2 reversed the ovariectomy-induced decrease in serum estradiol levels and uterine weights, but PPT and DPN treatment did not show any effect. Results reveal that specific ER-β agonists can ameliorate memory dysfunction and depressive behavior associated with postmenopausal diabetes and are devoid of the feminizing adverse effects of nonselective ER agonists.

Pharmacological activation of estrogen receptors-α and-β differentially modulates keratinocyte differentiation with functional impact on wound healing.

Estrogen deprivation is considered responsible for many age-related processes, including poor wound healing. Guided by previous observations that estradiol accelerates re-epithelialization through estrogen receptor (ER)-β, in the present study, we examined whether selective ER agonists [4,4′,4″-(4-propyl [1H] pyrazole-1,3,5-triyl)-trisphenol (PPT), ER-α agonist; 2,3-bis(4-hydroxyphenyl)-propionitrile (DPN), ER-β agonist] affect the expression of basic proliferation and differentiation markers (Ki-67, keratin-10, -14 and -19, galectin-1 and Sox-2) of keratinocytes using HaCaT cells. In parallel, ovariectomized rats were treated daily with an ER modulator, and wound tissue was removed 21 days after wounding and routinely processed for basic histological analysis. Our results revealed that the HaCaT keratinocytes expressed both ER-α and -β, and thus are well-suited for studying the effects of ER agonists on epidermal regeneration. The activation of ER-α produced a protein expression pattern similar to that observed in the control culture, with a moderate expression of Ki-67 being observed. However, the activation of ER-β led to an increase in cell proliferation and keratin-19 expression, as well as a decrease in galectin-1 expression. Fittingly, in rat wounds treated with the ER-β agonist (DPN), epidermal regeneration was accelerated. In the present study, we provide information on the mechanisms through which estrogens affect the expression patterns of selected markers, thus modulating keratinocyte proliferation and differentiation; in addition, we demonstrate that the pharmacological activation of ER-α and -β has a direct impact on wound healing.

Abstract 5047: Role of estrogen receptors alpha and beta in the balance between proliferation and surveillance

Abstract Estrogens have paradoxical effects in both promoting and preventing breast cancer. The proliferative actions of estrogens promote the expansion of both normal mammary epithelium and breast cancer. But estrogen also activates p53-mediated surveillance and apoptotic pathways to eliminate cells with potential oncogenic mutations. This is exemplified by tumor regression in response to high dose estrogen-treatment in postmenopausal breast cancer. Estrogens mediate their effects through the activation of two estrogen receptor subtypes: Estrogen Receptor alpha (ERα)and Estrogen Receptor beta (ERβ). Our objective is to examine the contribution of these receptors in mediating surveillance as opposed to proliferation by comparing selective estrogen receptor agonists with 17β-estradiol (E2) for induction of proliferation and surveillance in three model systems: (1) in vivo mouse model, (2) in vitro MCF7-tet-off-ERβ model, and (3) an ex-vivo human mammary explant model. Selective activation of ERβ with the agonist diarylpropionitrile (DPN) in vivo enhances p53-mediated apoptosis in the mouse mammary epithelium without stimulating proliferation. In addition, radiation-induced apoptosis is significantly reduced in mice lacking ERβ (βERKO). E2 or the ERα selective agonist pyrazole triol (PPT) induced the expression of estrogen response genes including progesterone receptor and amphiregulin in all three models. DPN-mediated activation of ERβ failed to induce the expression of these genes. The ERβ agonist DPN selectively induced the expression of genes that repress proliferation including TGFβ and inhibitors of canonical WNT signaling, WNT5a and AXIN2. DPN was also more potent in stimulating EGR1, a modulator of p53 activity, and CEBPd, a pro-apoptotic gene. Conclusions: The ability of DPN and ERβ to potentiate surveillance pathways while limiting proliferation suggests that ERβ agonists may provide a new approach for chemoprevention of breast cancer. Citation Format: D. Joseph Jerry, Karen A. Dunphy, Amy L. Roberts, Margarita Brown. Role of estrogen receptors alpha and beta in the balance between proliferation and surveillance. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5047. doi:10.1158/1538-7445.AM2015-5047

Activation of G-protein-coupled receptor 30 is sufficient to enhance spatial recognition memory in ovariectomized rats

In ovariectomized rats, administration of estradiol, or selective estrogen receptor agonists that activate either the α or β isoforms, have been shown to enhance spatial cognition on a variety of learning and memory tasks, including those that capitalize on the preference of rats to seek out novelty. Although the effects of the putative estrogen G-protein-coupled receptor 30 (GPR30) on hippocampus-based tasks have been reported using food-motivated tasks, the effects of activation of GPR30 receptors on tasks that depend on the preference of rats to seek out spatial novelty remain to be determined. Therefore, the aim of the current study was to determine if short-term treatment of ovariectomized rats with G-1, an agonist for GPR30, would mimic the effects on spatial recognition memory observed following short-term estradiol treatment. In Experiment 1, ovariectomized rats treated with a low dose (1μg) of estradiol 48h and 24h prior to the information trial of a Y-maze task exhibited a preference for the arm associated with the novel environment on the retention trial conducted 48h later. In Experiment 2, treatment of ovariectomized rats with G-1 (25μg) 48h and 24h prior to the information trial of a Y-maze task resulted in a greater preference for the arm associated with the novel environment on the retention trial. Collectively, the results indicated that short-term treatment of ovariectomized rats with a GPR30 agonist was sufficient to enhance spatial recognition memory, an effect that also occurred following short-term treatment with a low dose of estradiol.