Molecular cloning of estrogen receptor and its function on vitellogenesis in pompano (Trachinotus ovatus).

Estrogen receptors (ERs) play a critical role in vitellogenesis (Vtgs). However, the contribution of each ER for the regulation of vtgs expression was not analyzed clearly in teleosts. In the present study, three ers isoforms (erα, erß1, and erß2) were cloned in pompano (Trachinotus ovatus). Real-time PCR and enzyme-linked immunosorbent assay (ELISA) was used to detect the effects of 17ß-estradiol (E2) on ERs and Vtgs in the liver of pompano. In vivo injection experiments showed that E2 significantly increased the expressions of ers and vtgs. ER broad spectrum antagonist Fulvestrant significantly attenuated the E2- induced up-regulation of ers and vtgs in a dose-dependent manner. ERα antagonist Methyl-piperidino pyrazole (MPP) significantly attenuated the up-regulation of erα, erß2, vtg-B and vtg-C, and promoted the expressions of erß1 and vtg-A. ERß antagonist Cyclofenil significantly inhibited the expressions of erß1, erß2, vtg-A and vtg-C, and promoted the expressions of erα and vtg-B. In addition, E2 significantly increased the protein level of Vtg, while Fulvestrant, MPP and Cyclofenil significantly inhibited the protein level of Vtg in a dose-dependent manner. Our results indicate that E2 may regulate the expression of each vtg with different subtypes of ERs, and shows a distinct compensatory expression effect on the regulation for ers and vtgs, which provides a theoretical basis for reproductive endocrinology study in pompano.

Role of miR-21-5p/FilGAP axis in estradiol alleviating the progression of monocrotaline-induced pulmonary hypertension.

BACKGROUND: Aberrant expression of microRNAs (miRNAs) has been associated with the pathogenesis of pulmonary hypertension (PH). It is, however, not clear whether miRNAs are involved in estrogen rescue of PH. METHODS: Fresh plasma samples were prepared from 12 idiopathic pulmonary arterial hypertension (IPAH) patients and 12 healthy controls undergoing right heart catheterization in Shanghai Pulmonary Hospital. From each sample, 5 µg of total RNA was tagged and hybridized on microRNA microarray chips. Monocrotaline-induced PH (MCT-PH) male rats were treated with 17ß-estradiol (E2 ) or vehicle. Subgroups were cotreated with estrogen receptor (ER) antagonist or with antagonist of miRNA. RESULTS: Many circulating miRNAs, including miR-21-5p and miR-574-5p, were markedly expressed in patients and of interest in predicting mean pulmonary arterial pressure elevation in patients. The expression of miR-21-5p in the lungs was significantly upregulated in MCT-PH rats compared with the controls. However, miR-574-5p showed no difference in the lungs of MCT-PH rats and controls. miR-21-5p was selected for further analysis in rats as E2 strongly regulated it. E2 decreased miR-21-5p expression in the lungs of MCT-PH rats by ERß. E2 reversed miR-21-5p target gene FilGAP downregulation in the lungs of MCT-PH rats. The abnormal expression of RhoA, ROCK2, Rac1 and c-Jun in the lungs of MCT-PH rats was inhibited by E2 and miR-21-5p antagonist. CONCLUSIONS: miR-21-5p level was remarkably associated with PH severity in patients. Moreover, the miR-21-5p/FilGAP signaling pathway modulated the protective effect of E2 on MCT-PH through ERß.

Establishment of an analytical method for simultaneous quantitation of CDK4/6 inhibitors, aromatase inhibitors, and an estrogen receptor antagonist in human plasma using LC-ESI-MS/MS.

Cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors (palbociclib, abemaciclib, and ribociclib) are used to treat human epithelial growth factor receptor (HER)-2 negative and hormone receptor (HR) positive advanced breast cancer in combination with aromatase inhibitors (letrozole, anastrozole) or an estrogen receptor antagonist (fulvestrant). Administration of these drugs frequently causes severe side effects, such as neutropenia and diarrhea. Therefore, therapeutic drug monitoring (TDM) of CDK4/6 inhibitors, aromatase inhibitors, and the estrogen receptor antagonist is considered important for ensuring the efficacy and safety of these drugs. In this study, we describe a simple, highly sensitive, and specific liquid chromatography/electrospray ionization tandem mass spectrometry method for simultaneous quantitation of the concentrations of palbociclib, abemaciclib, ribociclib, letrozole, anastrozole, and fulvestrant. In addition, we analyzed plasma samples from patients with HER2-negative and HR-positive advanced breast cancer treated with these compounds using this novel method. In our method, the intra-assay relative error (RE) values ranged from -12.8% to 12.9%, the inter-assay RE values ranged from -4.8% to 6.2%, and the coefficient of variation (CV) values for intra- and inter-assay were ≤8.6% and ≤13.3%, respectively. The analytes showed good stability with RE values ranging from -13.5% to 13.6% and CV values <10.4%. Moreover, all the samples from patients were successfully quantified, and were within the range of measurement. This method can be used for TDM of routine anticancer drugs in clinical practice and for pharmacokinetics/pharmacodynamics research in future studies.

Pemafibrate, a novel selective PPARα modulator, attenuates tamoxifen-induced fatty liver disease.

Estrogen receptor (ER) antagonists, such as tamoxifen and toremifene, are widely used as adjuvant therapies for ER-positive breast cancer. These agents sometimes cause hepatosteatosis and steatohepatitis and it is problematic whether these agents should be withdrawn due to fatty liver disease and liver dysfunction. We herein describe a patient with fatty liver disease and hypertriglyceridemia during tamoxifen treatment, which significantly improved by adding pemafibrate, a novel PPARα activator designated as a selective PPARα modulator. Serial analysis during pemafibrate treatment revealed significant increases in circulating ketone bodies, which are indicators of hepatic fatty acid (FA) ß-oxidation. As far as we know, this is the first report demonstrating the beneficial effect of pemafibrate on tamoxifen-induced fatty liver disease, which is likely due to enhanced hepatic FA ß-oxidation by PPARα stimulation. Future large-scale studies will be needed to verify the current observation.

Altering rat sexual behavior to teach hormonal regulation of brain imprinting.

In this teaching laboratory, students design and perform an experiment to determine estrogen's role in imprinting the brain of neonatal rats to express either male or female sexual behavior. A discussion question is provided before the laboratory exercise in which each student is asked to search the literature and provide written answers to questions and to formulate an experiment to test the role of estrogen in imprinting the mating behavior of male and female rats. Students discuss their answers to the questions in laboratory with the instructor and design an experiment to test their hypothesis. In male rats, testosterone is converted by aromatase expressed by neurons in the brain to estrogen. Production of estrogen in the brain of neonatal rats imprints mating behavior in males, where a lack of estrogen action in the brain imprints female sexual behavior. The model involves administering exogenous testosterone to imprint male behavior in female pups or administration of an aromatase inhibitor (letrozole) or an estrogen receptor antagonist (ICI 182,780) to imprint female sexual behavior in male pups. In the model, litters of neonatal pups are treated with either carrier (control), testosterone propionate, aromatase inhibitor (letrozole), or an estrogen receptor antagonist (ICI 182,780) postnatally on days 1 and 3. Alteration of mating behavior is evaluated through the numbers of males and females that breed and establish pregnancy. This is a very simple protocol that provides an excellent experiment for student discussion on the effects of hormone action on imprinting brain sexual behavior.

Long-time qingyan formula extract treatment exerts estrogenic activities on reproductive tissues without side effects in ovariectomized rats and via active ER to ERE-independent gene regulation.

The reproductive tissues are negatively influenced by estrogens in hormone therapy. Qingyan formula ethanol extract (QYFE)'s estrogenic effects and safety on reproductive tissues after long-term administration and its mechanism via estrogen receptor (ER) pathway haven't been studied. Here, we characterized its estrogenic effects using ovariectomized rats together with in vitro studies for further molecular characterization. Ovariectomized rats were treated with QYFE at doses of 0.7, 1.4, and 2.8g/kg for 12 weeks. The results showed QYFE has a potent estrogenic activity, as indicated by restoring the disappeared estrous cycle, antagonizing the atrophy of uterus, vagina and mammary gland, and the estrogen decline in circulation caused by ovariectomy. In addition, QYFE upregulated ERα and ERß expressions and had a less stimulatory effect on PCNA and ki-67 antigen in reproductive tissues compared with estradiol valerate. QYFE components can bind to ERα and ERß, significantly increased ERα/ß-ERE luciferase reporter gene expression, upregulated the expressions of ERs, PR and pS2 in MCF-7 cells at protein and gene level. All these activities were significantly inhibited by the ER antagonist ICI182,780. QYFE's estrogenic activity maybe mediated by stimulating biosynthesis of estrogen and increasing the quantity of ERs in target tissue and via active ER to ERE-independent gene regulation.

Mechanism and therapeutic window of a genistein nanosuspension to protect against hematopoietic-acute radiation syndrome.

There are no FDA-approved drugs that can be administered prior to ionizing radiation exposure to prevent hematopoietic-acute radiation syndrome (H-ARS). A suspension of synthetic genistein nanoparticles was previously shown to be an effective radioprotectant against H-ARS when administered prior to exposure to a lethal dose of total body radiation. Here we aimed to determine the time to protection and the duration of protection when the genistein nanosuspension was administered by intramuscular injection, and we also investigated the drug's mechanism of action. A single intramuscular injection of the genistein nanosuspension was an effective radioprotectant when given prophylactically 48 h to 12 h before irradiation, with maximum effectiveness occurring when administered 24 h before. No survival advantage was observed in animals administered only a single dose of drug after irradiation. The dose reduction factor of the genistein nanosuspension was determined by comparing the survival of treated and untreated animals following different doses of total body irradiation. As genistein is a selective estrogen receptor beta agonist, we also explored whether this was a central component of its radioprotective mechanism of action. Mice that received an intramuscular injection of an estrogen receptor antagonist (ICI 182,780) prior to administration of the genistein nanosuspension had significantly lower survival following total body irradiation compared with animals only receiving the nanosuspension (P < 0.01). These data define the time to and duration of radioprotection following a single intramuscular injection of the genistein nanosuspension and identify its likely mechanism of action.

Treatment satisfaction in women receiving palbociclib combination therapies for advanced/metastatic breast cancer.

AIM: To understand treatment satisfaction in patients with advanced or metastatic breast cancer receiving palbociclib plus an aromatase inhibitor or palbociclib plus fulvestrant in a real-world setting. PATIENTS & METHODS: We performed an observational, cross-sectional, web-based survey of 604 patients with self-reported hormone receptor-positive (HR+)/HER2-negative (HER2-) ABC/mBC in six countries. RESULTS: Overall, more than 96% of patients reported the benefits of their palbociclib combination therapy met or exceeded their expectations. Patient expectations and satisfaction with therapy did not differ between patients on palbociclib plus letrozole and palbociclib plus fulvestrant, or between patients with visceral and nonvisceral metastases. CONCLUSION: The patients on palbociclib combination therapy reported high satisfaction scores across multiple countries.

Radix Paeoniae Alba increases serum estrogen level and up-regulates estrogen receptor expression in uterus and vagina of immature/ovariectomized mice.

Radix Paeoniae Alba (RPA) is widely used in clinical treatment for gynecological diseases, particularly abnormal menstruation, menstrual pain, and breast tenderness; however, no scientific evidence base links RPA to estrogen replacement therapy. In this study, we characterize estrogenic activity of RPA using immature and ovariectomized (OVX) mice together with in vitro studies focus on estrogen receptor (ER) pathway for molecular mechanism. RPA treatments demonstrated significant estrogenic activity, as indicated by promoting the development of uterus and vagina in immature mice, reversing the atrophy of uterus and vagina in OVX mice, up-regulating the expressions of ERα and ERß at protein and mRNA level in reproductive tissues. Meanwhile, RPA significantly increased serum estradiol and clearly decreased serum luteinizing hormone and follicle-stimulating hormone of immature/OVX mice. Moreover, RPA could induce ER positive MCF-7 cell from S-phase to G2 stage and induce proliferation and no influence on ER negative MDA-MB-231 cell. RPA could bind with ERα and ERß and significantly stimulate ERα/ß-estrogen response element (ERE) luciferase reporter gene expression. All activities were inhibited by the ER antagonist ICI 182,780. This study illustrates RPA exerts estrogenic effects by stimulating biosynthesis of estrogen in circulation, up-regulating ERs in target tissues, and mimicking the estrogen through ER-ERE-dependent pathway.

[Gender-dependent Expression of ERß in AEC â ¡ of Fetal Mice Exposed to Arsenic and Estrogen Receptor Antagonist].

OBJECTIVE: To determine the effects of arsenic and estrogen receptor antagonist (ICI182, 780) on the expression of estrogen receptor beta (ERß) in alveolar Ⅱ epithelial cells (AECⅡ) of female and male mice. METHODS: Nineteen or twenty day fetus mice were obtained through caesarean section of ICR mice. Purified AECⅡ cells were separated from the female and male fetus,respectively,and confirmed using immunofluorescence staining. The cells were exposed to sodium arsenite (NaAsO2) at a low,medium,or high dosage determined by MTT and cultured for 24 h. The NaAsO2 (5 µmol/L) exposed cells were compared with those treated (for 24 h) with dimethyl sulfoxide (DMSO) or ICI182, 780 (1×10-4 mol/L). Apoptosis rates of the cells were measured by flow cytometry. Real-time fluorescence quantitative PCR method and Western blot technique were used to detect the expression ofERßmRNA and protein in AECⅡ. RESULTS: Purity of AECⅡ cells reached (87.0±2.5)%. NaAsO2 exposure was set at a concentration of 0.5 (low),1.25 (medium),and 5 (high) µmol/L. The cells exposed to medium and high dosage of NaAsO2 had higher apoptosis rates than the blank controls (P<0.05),without sex differences. Female cells exposed to medium and high dosage of NaAsO2 had higher levels of expressions ofERßmRNA and protein than the blank controls (P<0.05) and male cells exposed to the same dosage of NaAsO2 (P<0.05). No significant differences were found in the expressions ofERßmRNA and protein between the exposed male cells and the blank controls. ICI182, 780 lowered the expression levels ofERßmRNA and protein in the female exposed cells (P<0.01). CONCLUSION: Arsenic exposure increases expressions of AECⅡ's ERß,more so in female cells than in male cells. This can be blocked by estrogen receptor antagonists.

Veratrum nigrum inhibits the estrogenic activity of salvia miltiorrhiza bunge in vivo and in vitro.

BACKGROUND: As recorded in the 18 incompatible medicaments of Traditional Chinese Medicine theory, the combined use of Salvia miltiorrhiza bunge (SM) and Veratrum nigrum (VN) could induce toxicity and has been prohibited for thousands of years in China. However, the theory has been validated due to lack of evidence. Previous studies have focused on the chemical constituents that are responsible for the toxicity of the two agents. PURPOSE: This study offers preliminary insight into the pharmacodynamics and mechanism of estrogenic activity responsible for their incompatibility. STUDY DESIGN: We undertook a characterization of the interaction between estrogenic activities of SM and VN using in vivo models of immature and ovariectomized (OVX) mice, and in vitro studies focused on the estrogen receptor (ER) pathway for further mechanism. METHODS: Immature and OVX mice were treated intragastrically with SM at doses of 1.6, 3.2 g/kg, or combine with 0.045 g/kg VN and 0.005 g/kg the ER antagonist ICI182, 780 for elucidating the effects on estrogenic activity in reproductive tissues, E2 secretion, and the ER mechanism. ERα/ß binding experiments and ERα/ß transcriptional activity were performed in order to evaluate the biological action exerted through ERs. RESULTS: VN decreased the estrogenic efficacy of SM in promoting the development of the uterus and vagina in immature mice, and reversing the atrophy of reproductive tissues in OVX mice. VN interfered with the estrogenic efficacy of SM by decreasing the serum estradiol and the upregulation of ERα and ERß expressions in reproductive tissues by treatment with SM. VN antagonized the estrogenic efficacy of SM in promoting the viability of MCF-7 cells and stimulating the binding ability with ERα and ERß, and increasing ERα/ß-estrogen response element (ERE) luciferase activity. CONCLUSIONS: This study provided evidence that the combined use of SM and VN could induce unfavorable effects. VN decreased the estrogenic activity of SM, which might be related to the regulation of estrogen secretion and ERs through the ER-ERE pathway.

Utilizing a rat delayed implantation model to teach integrative endocrinology and reproductive biology.

In this teaching laboratory, the students are directed in an exercise that involves designing and performing an experiment to determine estrogen's role in regulating delayed implantation (diapause) in female rats. To encourage active participation by the students, a discussion question is provided before the laboratory exercise in which each student is asked to search the literature and provide written answers to questions and to formulate an experiment to test the role of ovarian estrogen in inducing implantation in female rats. One week before the laboratory exercise, students discuss their answers to the questions with the instructor to develop an experiment to test their hypothesis that estrogen is involved with inducing implantation in the rat. A rat delayed implantation model was established that utilizes an estrogen receptor antagonist (ICI 182,780), which inhibits the action of ovarian estrogens. Groups of mated females are treated with either carrier (control) or ICI 182,780 (ICI) every other day, starting on day 2 postcoitus (pc) until day 8 pc. One-half of the females receiving ICI are injected with estradiol-17ß on day 8 pc to induce implantation 4 days after the controls. If the ICI-treated females are not administered estradiol, embryo implantation occurs spontaneously ~4 days after the last ICI injection on day 8. This is a very simple protocol that is very effective and provides an excellent experiment for student discussion on hormone action and the use of agonists and antagonists.

Effects of estrogen receptor antagonist on the expressions of Erα, Erβ and p57kip2 in JEC cells of human endometrial carcinoma / 临床与实验病理学杂志

Purpose To explore the effects of estrogen receptor antagonist on the expression of estrogen receptor subtype (ERα, ERβ), and p57kip2 protein in human endometrioid carcinoma cells named JEC. Methods The JEC cells (moderately differentiated EC cells) cultured in vitro were treated with β-Estradiol (E2) (10～6 mol/L) and two types of estrogen receptor antagonists, tamoxifen (TAM) and fulvestrant (ICI182780) (10-6 mol/L). After 24, 48, 72 h, MTT was used to detect the growth condition of JEC cells, and the light microscopy and electron microscopy were used to observe the growth condition and morphological changes of cells, Western blot was used to detect the expression of ERα, ERβ, PR-A, PR-B and P57kip2 protein in JEC cells. Results MTT results: Compared with the control group, E2 could promote the proliferation of JEC cells significantly (P＜0.05), and ICI182780 could inhibit the proliferation of JEC cells obviously (P＜0.05). Compared with the E2 group, the proliferation ability of JEC cells in E2 + ICI182780 group were lower(P＜0.05). Morphological change: Compared with the control group, the cells density of E2 group increased obviously, and the pathologic mitosis was easy to seen in some cells. The cells density decreased obviously in ICI182780 group. Compared with E2 group, the cells density of E2 + TAM group and E2 + ICI182780 group were decreased, and pathological mitotic figures were difficult to seen. Western blot results: Compared with the control group, the expression of ERβ protein increased, and the expression of p57kip2 protein decreased in E2 group (P＜0.05). The expression of ERβ protein decreased, and the expression of p57kip2 protein increased in ICI182780 group and TAM group, and the difference was statistically significant between ICI182780 group and control group (P＜0.05). Compared with the E2 group, the expression of ERβ protein decreased, and the expression of p57kip2 protein increased in E2 + ICI182780 group and E2 + TAM group, and the difference was statistically significant between E2 + ICI182780 group and E2 group (P＜0.05). ERa protein of JEC cells did not expressed in experimental group or control group. Conclusion ERa protein are not expressed in JEC cells. ICI182780 have a stronger role in antagonizing estrogen, and may induce the expression of p57kip2 protein by down-regulating the expression of ERβ protein in JEC cells, block the cell cycle progression and inhibit the growth of tumor cells. TAM has a weaker estrogen like effect on the growth of JEC cells. It is possible that combined detection of the expression of ERa and p57kip2 protein in EC has an important reference value for individualized selection of endocrine therapy for EC patients.

Immunotherapeutic target expression on breast tumors can be amplified by hormone receptor antagonism: a novel strategy for enhancing efficacy of targeted immunotherapy.

Immunotherapy has historically been successful in highly antigenic tumors but has shown limited therapeutic efficacy in non-antigenic tumors such as breast cancers. Our previous studies in autoimmunity have demonstrated that increased antigen load within a tissue enhances immune reactivity against it. We therefore hypothesized that enhancing expression of target proteins on breast tumors can increase efficacy of targeted immunotherapy. We hypothesized that antagonism of the estrogen receptor (ER) can increase expression of targets that are hormonally regulated and facilitate enhanced tumor recognition by targeted immunotherapy. We used a lactation protein α-Lactalbumin, a known immunotherapeutic target on breast tumors, as our model target antigen. Enhancement of target protein expression in human and murine breast tumors was tested in vitro and in vivo by ER antagonism using clinically established ER modulators, Tamoxifen and Fulvestrant. We show that antagonism of the ER can induce a 2-3 fold increase in expression of target proteins on tumors leaving the normal breast tissue unaffected. Tumor progression studies in 4T1 tumor-bearing mice show that efficacy of adoptively transferred cell based targeted immunotherapy was enhanced by target antigen amplification resulting in significantly higher tumor inhibition. However, in spite of increased target expression, anti-tumor efficacy of direct immunization was not enhanced probably due to other limiting factors involved in the immune priming process. Our study provides a novel combinatorial clinical strategy for enhancing efficacy of immunotherapy not only on breast tumors but potentially also for other hormonally driven tumors such as those of the prostate, testis and ovary.

Cloning, expression and functional characterization on vitellogenesis of estrogen receptors in Scatophagus argus.

Estrogen receptors (Er) play a critical role in vitellogenesis. Three ers (erα, erß1 and erß2) and vitellogenins (vtg-A, vtg-B and vtg-C) subtypes were isolated in various fish species, while the contribution of each Er to the regulation of vtgs expression was not analyzed in detail. Here, erα, erß1 and erß2 were cloned and all were found to be expressed in female liver in Scatophagus argus. During proteic vitellogenesis stage, erα was simultaneously up-regulated, while erß1 and erß2 were not, with three vtgs in female liver. The effects of 17ß-estradiol (E2) alone or combined with Er antagonists on ers, vtgs mRNA expressions and Vtg protein content in incubated male liver were examined by real-time PCR and enzyme-linked immunosorbent assay (ELISA), respectively. The expressions of erα, erß1, vtgs mRNA and Vtg protein increased significantly after 24h incubation with E2 (0.1, 1 and 10µM), while Er nonselective antagonist ICI 182 780 (0.01, 0.1 and 1µM) significantly attenuated the up-regulation effects of E2 on ers, vtgs mRNA and Vtg protein in a dose-dependent manner. Erα selective antagonist Methyl-piperidinopyrazole (MPP) (0.01, 0.1 and 1µM) significantly attenuated the up-regulation effects of E2 on erα, vtg-B, vtg-C mRNA and Vtg protein, while promoted the expression of erß1 and vtg-A. Erß selective antagonist Cyclofenil (0.01, 0.1 and 1µM) attenuated the up-regulation effects of E2 on erß1, erß2, vtg-A, vtg-C mRNA and Vtg protein while promoted the expression of erα and vtg-B. Our results suggest that the regulation of Ers on different vtgs was divergent in S. argus.

Salvia miltiorrhiza bunge increases estrogen level without side effects on reproductive tissues in immature/ovariectomized mice.

Salvia miltiorrhiza bunge(SM) is a popular herb for alleviating menopausal symptoms, although the scientific evidence of applying SM to estrogen replacement therapy is limited. In this study, we characterized the estrogenic activity of SM using in vivo models of immature and ovariectomized (OVX) mice and performed in vitro studies focusing on the estrogen receptor (ER) pathway for further molecular characterizations. SM treatments demonstrated significant estrogenic activity by promoting the development of uterus and vagina in immature mice, restoring the estrus cycle and reversing the atrophy of reproductive tissues in OVX mice, as well as increasing the expressions of ERα and ERß at protein and mRNA level in the reproductive tissues. Meanwhile, SM significantly increased estradiol in serum, and decreased follicle-stimulating hormone (FSH) and luteinizing hormone (LH) in the circulation of immature and OVX mice. SM could stimulate the binding effect of ERα and ERß, and significantly induce ERα/ß-estrogen response element (ERE) luciferase reporter gene expression. All these activities were inhibited by the ER antagonist ICI182, 780. This study demonstrates SM exerts estrogenic effects by stimulating biosynthesis of estrogen and increasing ERs in target tissues without side effects on reproductive tissues and through ER-ERE-dependent pathway.

17ß-Estradiol augments antidepressant efficacy of escitalopram in ovariectomized rats: Neuroprotective and serotonin reuptake transporter modulatory effects.

The prevalence or recurrence of depression is seriously increased in women during the transition to and after menopause. The chronic hypo-estrogenic state of menopause may reduce the response to antidepressants; however the influence of estrogen therapy on their efficacy is still controversial. This study aimed at investigating the effects of combining escitalopram with 17ß-estradiol on depression and cognitive impairment induced by ovariectomy, an experimental model of human menopause. Young adult female Wistar rats were subjected to either sham operation or ovariectomy. Ovariectomized animals were treated chronically with escitalopram (10mg/kg/day, i.p) alone or with four doses of 17ß-estradiol (40µg/kg, s.c) given prior to the behavioral tests. Co-administration of 17ß-estradiol improved escitalopram-induced antidepressant effect in forced swimming test verified as more prominent decrease in the immobility time without opposing its memory enhancing effect in Morris water maze. 17ß-estradiol augmented the modulatory effects of escitalopram on the hippocampal levels of brain-derived neurotrophic factor and serotonin reuptake transporter as well as tumor necrosis factor-alpha without altering its effects on the gene expressions of serotonin receptor 1A, estrogen receptors alpha and beta, or acetylcholinestearase content. This combined therapy afforded synergistic protective effects on the brain histopathological architecture, particularly, the hippocampus. The antidepressant effect of 17ß-estradiol was abolished by pretreatment with estrogen receptor antagonist, tamoxifen (10mg/kg, p.o). In conclusion, 17ß-estradiol-induced antidepressant effect was confined to intracellular estrogen receptors activation. Moreover, 17ß-estradiol enhanced escitalopram's efficiency in ameliorating menopausal-like depression, via exerting synergistic neuroprotective and serotonin reuptake transporter modulatory effects, without impeding escitalopram-mediated cognitive improvement.

Fulvestrant for the treatment of endometrial cancer.

INTRODUCTION: About one third of patients with endometrial cancer (EC) relapse and face a limited prognosis, if surgery or radiotherapy are not feasible. The remaining therapeutic options are chemotherapy and endocrine therapy. AREAS COVERED: This review summarizes the development of the first selective estrogen receptor (ER) down-regulator fulvestrant. This article provides its mechanism of action, pharmacokinetics and the available preclinical and clinical data. Furthermore, this review provides an overview of the market of treatments for recurrent or metastatic EC (RMEC) while also taking into account studies of fulvestrant in metastatic breast cancer. EXPERT OPINION: Even if fulvestrant showed only marginal activity in two phase II trials, it shouldn't be abandoned but instead further developed in EC. Firstly, the dose of fulvestrant used in these trials was too low from today's point of view. Secondly, the available literature on other endocrine agents is full of limitations and does not provide a gold standard. Furthermore, given the activity of mTOR inhibitors in EC, there may also be synergistic effects, given the cross-regulation of ER and the PI3K/AKT/mTOR pathway. The authors suggest that a prospective, phase II trial in ER positive RMEC would help to further explore the efficacy and tolerability of fulvestrant together with a mTOR inhibitor.

Role of p38 MAPK pathway in 17ß-estradiol-mediated attenuation of hemorrhagic shock-induced hepatic injury.

Although 17ß-estradiol (E2) treatment following hemorrhagic shock or ischemic reperfusion prevents organs from dysfunction and injury, the precise mechanism remains unknown. We hypothesize that the E2-mediated attenuation of liver injury following hemorrhagic shock and fluid resuscitation occurs via the p38 mitogen-activated protein kinase (MAPK)-dependent heme oxygenase (HO)-1 pathway. After a 5-cm midline laparotomy, male rats underwent hemorrhagic shock (mean blood pressure ∼40 mmHg for 90 min) followed by fluid resuscitation. At the onset of resuscitation, rats were treated with vehicle, E2 (1 mg/kg) alone, or E2 plus p38 MAPK inhibitor SB-203580 (2 mg/kg), HO-1 inhibitor chromium mesoporphyrin-IX chloride (2.5 mg/kg) or estrogen receptor antagonist ICI 182,780 (3 mg/kg). At 2 h after hemorrhagic shock and fluid resuscitation, the liver injury markers were significantly increased compared with sham-operated control. Hemorrhagic shock resulted in a significant decrease in p38 MAPK phosphorylation compared with the shams. Administration of E2 following hemorrhagic shock normalized liver p38 MAPK phosphorylation, further increased HO-1 expression, and reduced cleaved caspase-3 levels. Coadministration of SB-203580 abolished the E2-mediated attenuation of the shock-induced liver injury markers. In addition, administration of chromium mesoporphyrin-IX chloride or ICI 182,780 abolished E2-mediated increases in liver HO-1 expression or p38 MAPK activation following hemorrhagic shock. Our results collectively suggest that the salutary effects of E2 on hepatic injury following hemorrhagic shock and resuscitation are in part mediated through an estrogen-receptor-related p38 MAPK-dependent HO-1 upregulation.

The role of TGF-ß/Smad signaling in dopamine agonist-resistant prolactinomas.

BACKGROUND: Prolactinomas are the most common secretory pituitary adenomas. The first line of treatment involves dopamine agonists (DAs); however, a subset of patients is resistant to such therapy. Recent studies suggest that dopamine can up-regulate TGF-ß1 synthesis in rat pituitary lactotrophs whereas estradiol down-regulates TGF-ß1. To date, the role of TGF-ß/Smad signaling in DAs-resistant prolactinomas has not been explored. METHODS: High-content screening (HCS) techniques, qRT-PCR, Western blot, immunofluorescence and ELISA, were performed to determine the role of TGF-ß/Smad signaling in DAs-resistant prolactinomas. RESULTS: We reported a significant down-regulation of TGF-ß/Smad signaling cascade in DAs-resistant prolactinomas compared to normal human anterior pituitaries. Following treatment with TGF-ß1, the dopamine agonist, bromocriptine, and the estrogen antagonist (ER), fulvestrant in GH3 cells, we found that TGF-ß1 and fulvestrant caused significant cytotoxicity in a dose- and time-dependent manner and activated Smad3 was detected following exposure to TGF-ß1 and fulvestrant. In addition, treating GH3 cells with fulvestrant increased active TGF-ß1 levels and decreased PRL levels in a dose-dependent manner. CONCLUSION: TGF-ß/Smad signaling pathway may play an important role in DA-resistant prolactinomas and has the potential to be a viable target for the diagnosis and treatment of prolactinomas, particularly in patients who are resistant to DAs.