G Protein-coupled Estrogen Receptor Activation Research Articles

Estradiol contributes to sex differences in resilience to sepsis-induced metabolic dysregulation and dysfunction in the heart via GPER-1-mediated PPARδ/NLRP3 signaling

Background and aimClinically, septic males tend to have higher mortality rates, but it is unclear if this is due to sex differences in cardiac dysfunction, possibly influenced by hormonal variations. Cardiac dysfunction significantly contributes to sepsis-related mortality, primarily influenced by metabolic imbalances. Peroxisome proliferator-activated receptor delta (PPARδ) is a key player in cardiac metabolism and its activation has been demonstrated to favor sepsis outcomes. While estradiol (E2) is abundant and beneficial in females, its impact on PPARδ-mediated metabolism in the heart with regards to sex during sepsis remains unknown. Methods and resultsHere, we unveil that while sepsis diminishes PPARδ nuclear translocation and induces metabolic dysregulation, oxidative stress, apoptosis and dysfunction in the heart thereby enhancing mortality, these effects are notably more pronounced in males than females. Mechanistic experiments employing ovariectomized(OVX) mice, E2 administration, and G protein-coupled estrogen receptor 1(GPER-1) knockout (KO) mice revealed that under lipopolysaccharide (LPS)-induced sepsis, E2 acting via GPER-1 enhances cardiac electrical activity and function, promotes PPARδ nuclear translocation, and subsequently ameliorates cardiac metabolism while mitigating oxidative stress and apoptosis in females. Furthermore, PPARδ specific activation using GW501516 in female GPER-1−/− mice reduced oxidative stress, ultimately decreasing NLRP3 expression in the heart. Remarkably, targeted GPER-1 activation using G1 in males mirrors these benefits, improving cardiac electrical activity and function, and ultimately enhancing survival rates during LPS challenge. By employing NLRP3 KO mice, we demonstrated that the targeted GPER-1 activation mitigated injury, enhanced metabolism, and reduced apoptosis in the heart of male mice via the downregulation of NLRP3. ConclusionOur findings collectively illuminate the sex-specific cardiac mechanisms influencing sepsis mortality, offering insights into physiological and pathological dimensions. From a pharmacological standpoint, this study introduces specific GPER-1 activation as a promising therapeutic intervention for males under septic conditions. These discoveries advance our understanding of the sex differences in sepsis-induced cardiac dysfunction and also present a novel avenue for targeted interventions with potential translational impact.

GPER1 Activation Upregulates Renal Purinergic P2Y2 Receptor Expression and Lowers Blood Pressure in Ovariectomized Rats Fed High Salt Diet

Renal purinergic P2Y2 receptor promotes urinary Na+ excretion via inhibiting epithelial Na+ channel (ENaC) activity. Recent evidence implicates that ovarian hormones regulate P2Y2/ENaC natriuretic pathway, however the underlying mechanism remains unclear. Estradiol activates the G protein-coupled estrogen receptor 1 (GPER1), which promotes natriuresis in females. Taken together, we hypothesized that GPER1 activation regulates renal P2Y2 receptor expression and maintains blood pressure in ovariectomized (OVX) rats. To test our hypothesis, we determined the effect of pharmacologic activation GPER1 on (i) blood pressure and (ii) mRNA expression of renal P2Y2 receptor. In particular, female (16–20 weeks of age) Sprague Dawley rats were implanted with telemeters. After recording baseline blood pressure, rats were ovariectomized and simultaneously implanted with an osmotic minipump to deliver either G1, selective GPER1 agonist (OVX+G1), or Vehicle (OVX+Veh) for 3 weeks. Rats were fed a normal salt (NS, 0.49% NaCl) throughout the study and challenged with a high salt (HS, 4% NaCl) during the last week. Finally, animals were euthanized, renal cortex and medulla were assessed for P2Y2 and αENaC mRNA expression. We found that OVX+Veh rats fed a HS diet had greater mean arterial pressure (MAP) when compared to their baseline values prior to ovariectomy (113 ± 1 vs. 105 ± 1 mmHg; p < 0.01; n = 6). Interestingly, OVX+G1 rats fed a HS diet did not elicit significant changes in MAP when compared to their baseline values (106 ± 3 vs. 104 ± 2 mmHg; p = 0.25; n = 6). Moreover, renal cortical P2Y2 mRNA expression in OVX+G1 rats was higher compared to corresponding vehicle-treated rats (1.95 ± 0.21 vs. 1.00 ± 0.17 fold change from OVX+Veh; p < 0.01; n = 6-7/group). Renal outer and inner medullary P2Y2 mRNA expression elicited similar trends; however, it did not reach statistical significance. No differences were evident in the renal αENaC mRNA expression in response to G1 treatment. We also extended the study in female mice to determine the effect of genetic deletion of GPER1 on renal P2Y2 receptor mRNA expression. We found that P2Y2 mRNA expression in kidneys obtained from GPER1 KO mice was markedly lower when compared to wildtype litter mates. However, no differences were detected in the mRNA expression of αENaC in response to GPER1 deletion. The current results indicate that GPER1 activity upregulates the renal P2Y2 expression which may contribute to preventing the increase in blood pressure in HS-fed OVX animals, possibly by promoting natriuresis. Additional studies are required to improve our understanding for GPER1/P2Y2/ENaC signaling pathway in the postmenopausal female population. NIH/NIDDK R00DK119413 to EYG. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Effect of astrocyte GPER on the optic nerve inflammatory response following optic nerve injury in mice

Activated astrocytes are a primary source of inflammatory factors following traumatic optic neuropathy (TON). Accumulation of inflammatory factors in this context leads to increased axonal damage and loss of retinal ganglion cells (RGCs). Therefore, in the present study, we explored the role of the astrocyte G protein-coupled estrogen receptor (GPER) in regulating inflammatory factors following optic nerve crush (ONC), and analyzed its potential regulatory mechanisms. Overall, our results showed that GPER was abundantly expressed in the optic nerve, and co-localized with glial fibrillary acidic proteins (GFAP). Exogenous administration of G-1 led to a significant reduction in astrocyte activation and expression of inflammation-related factors (including IL-1β, TNF-α, NFκB, and p-NFκB). Additionally, it dramatically increased the survival of RGCs. In contrast, astrocytes were activated to a greater extent by exogenous G15 administration; however, RGCs survival was significantly reduced. In vitro, GPER activation significantly reduced astrocyte activation and the release of inflammation-related factors. In conclusion, activation of astrocyte GPER significantly reduced ONC inflammation levels, and should be explored as a potential target pathway for protecting the optic nerve and RGCs after TON.

Modulation of GPER1 alleviates early brain injury via inhibition of A1 reactive astrocytes activation after intracerebral hemorrhage in mice

BackgroundEarly brain injury (EBI) caused by inflammatory responses in acute phase of Intracerebral hemorrhage (ICH) plays a vital role in the pathological progression of ICH. Increasing evidences demonstrate A1 reactive astrocytes are associated with the severity of EBI. G-protein coupled estrogen receptor 1 (GPER1) has been proved mediating the neuroprotective effects of estrogen in central nervous system (CNS) disease. However, whether GPER1 plays a protective effect on ICH and A1 reactive astrocytes activation is not well studied. MethodsICH model was established by infused the autologous whole blood into the right basal ganglia in wild type and GPER1 knockout mice. GPER1 specific agonist G1 and antagonist G15 were administered by intraperitoneal injection at 1 h or 0.5 h after ICH. Neurological function was detected on day 1 and day 3 by open field test and corner turn test following ICH. Besides, A1 reactive astrocytes were determined by immunofluorescence staining after ICH on day 3. To further identify the possible mechanism of GPER1 mediated neuroprotective effect, Western blot assays was performed after ICH on day 3. ResultsAfter ICH, G1 treatment alleviated mice neurobehavior deficits on day 1 and day 3. Meanwhile, G1 treatment also significantly reduced the GFAP positive astrocytes and the C3 positive cells after ICH. Interestingly, G15 reversed the protective effect of G1 on the neurobehavior of ICH mice. Meanwhile, the expression of GFAP+C3+ A1 reactive astrocytes were also reduced by activation of GPER1. Mechanistic studies indicated TLR4 and NF-κB mediated the neuroprotective effect of GPER1. ConclusionGenerally, activation of GPER1 alleviated the EBI through inhibiting A1 reactive astrocytes activation via TLR4/NF-κB pathway after ICH in mice. Additionally, GPER1may be a promising target for ICH treatment.

G protein–coupled estrogen receptor 1 ameliorates nonalcoholic steatohepatitis through targeting AMPK-dependent signaling

Nonalcoholic fatty liver disease (NAFLD), especially nonalcoholic steatohepatitis (NASH), has emerged as a prevalent cause of liver cirrhosis and hepatocellular carcinoma, posing severe public health challenges worldwide. The incidence of NASH is highly correlated with an increased prevalence of obesity, insulin resistance, diabetes, and other metabolic diseases. Currently, no approved drugs specifically targeted for the therapies of NASH partially due to the unclear pathophysiological mechanisms. G protein-coupled estrogen receptor 1 (GPER1) is a membrane estrogen receptor involved in the development of metabolic diseases such as obesity and diabetes. However, the function of GPER1 in NAFLD/NASH progression remains unknown. Here, we show that GPER1 exerts a beneficial role in insulin resistance, hepatic lipid accumulation, oxidative stress, or inflammation in vivo and in vitro. In particular, we observed that the lipid accumulation, inflammatory response, fibrosis, or insulin resistance in mouse NAFLD/NASH models were exacerbated by hepatocyte-specific GPER1 knockout but obviously mitigated by hepatic GPER1 activation in female and male mice. Mechanistically, hepatic GPER1 activates AMP-activated protein kinase signaling by inducing cyclic AMP release, thereby exerting its protective effect. These data suggest that GPER1 may be a promising therapeutic target for NASH.

GPER activation attenuates cardiac dysfunction by upregulating the SIRT1/3-AMPK-UCP2 pathway in postmenopausal diabetic rats.

Postmenopausal diabetic women are at higher risk to develop cardiovascular diseases (CVD) compared with nondiabetic women. Alterations in cardiac cellular metabolism caused by changes in sirtuins are one of the main causes of CVD in postmenopausal diabetic women. Several studies have demonstrated the beneficial actions of the G protein-coupled estrogen receptor (GPER) in postmenopausal diabetic CVD. However, the molecular mechanisms by which GPER has a cardioprotective effect are still not well understood. In this study, we used an ovariectomized (OVX) type-two diabetic (T2D) rat model induced by high-fat diet/streptozotocin to investigate the effect of G-1 (GPER-agonist) on sirtuins, and their downstream pathways involved in regulation of cardiac metabolism and function. Animals were divided into five groups: Sham-Control, T2D, OVX+T2D, OVX+T2D+Vehicle, and OVX+T2D+G-1. G-1 was administrated for six weeks. At the end, hemodynamic factors were measured, and protein levels of sirtuins, AMP-activated protein kinase (AMPK), and uncoupling protein 2 (UCP2) were determined by Western blot analysis. In addition, cardiac levels of oxidative stress biomarkers were measured. The findings showed that T2D led to left ventricular dysfunction and signs of oxidative stress in the myocardium, which were accompanied by decreased protein levels of Sirt1/2/3/6, p-AMPK, and UCP2 in the heart. Moreover, the induction of the menopausal state exacerbated these changes. In contrast, treatment with G-1 ameliorated the hemodynamic changes associated with ovariectomy by increasing Sirt1/3, p-AMPK, UCP2, and improving oxidative status. The results provide evidence of the cardioprotective effects of GPER operating through Sirt1/3, p-AMPK, and UCP2, thereby improving cardiac function. Our results suggest that increasing Sirt1/3 levels may offer new therapeutic approaches for postmenopausal diabetic CVD.

Activation of GPER1 in macrophages ameliorates UUO-induced renal fibrosis

Numerous studies have proven the critical role of macrophages in the renal fibrosis process. Notably, G Protein-coupled Estrogen Receptor 1 (GPER1), a novel estrogen receptor, has been shown to play a ubiquitous role in regulating macrophage activities and proinflammatory pathways. However, the precise role of GPER1 in macrophage-mediated renal fibrosis is unknown. In this study, we aimed to investigate the function of macrophage GPER1 in the UUO-induced renal fibrosis model. Compared to vehicle-treated ovariectomized (OVX) female and male unilateral ureteral obstruction (UUO) models, we observed that G-1 (GPER1 agonist)-treated OVX female and male UUO mice had fewer renal fibrotic lesions and less M1 and M2 macrophage infiltration in the kidney tissues. Conversely, Gper1 deletion in male UUO mice accelerated renal fibrosis and increased inflammation. In vitro studies also revealed that GPER1 activation reduced M0 macrophage polarization towards M1 or M2 phenotypes. The RNA-sequencing analysis and immunoblotting indicated that GPER1 activation was primarily involved in downregulating immune pathways activation and inactivating MAPK pathways. Tubular epithelial cells co-cultured with G-1-pretreated M1 macrophages exhibited fewer injuries and immune activation. In addition, fibroblasts co-cultured with G-1-pretreated M2 macrophages showed downregulated extracellular matrix expression. Overall, this is the first study to demonstrate the effect of GPER1 on macrophage-mediated renal fibrosis via inhibition of M1 and M2 macrophage activation. These findings indicate that GPER1 may be a promising therapeutic target for treating renal fibrosis.

GPER1 Activation Exerts Anti-Tumor Activity in Multiple Myeloma.

G protein-coupled estrogen receptor 1 (GPER1) activation is emerging as a promising therapeutic strategy against several cancer types. While GPER targeting has been widely studied in the context of solid tumors, its effect on hematological malignancies remains to be fully understood. Here, we show that GPER1 mRNA is down-regulated in plasma cells from overt multiple myeloma (MM) and plasma cell leukemia patients as compared to normal donors or pre-malignant conditions (monoclonal gammopathy of undetermined significance and smoldering MM); moreover, lower GPER1 expression associates with worse overall survival of MM patients. Using the clinically applicable GPER1-selective agonist G-1, we demonstrate that the pharmacological activation of GPER1 triggered in vitro anti-MM activity through apoptosis induction, also overcoming the protective effects exerted by bone marrow stromal cells. Noteworthy, G-1 treatment reduced in vivo MM growth in two distinct xenograft models, even bearing bortezomib-resistant MM cells. Mechanistically, G-1 upregulated the miR-29b oncosuppressive network, blunting an established miR-29b-Sp1 feedback loop operative in MM cells. Overall, this study highlights the druggability of GPER1 in MM, providing the first preclinical framework for further development of GPER1 agonists to treat this malignancy.

Activation of G-Protein-Coupled Estrogen Receptor 1 (GPER1) Reduces Progression of Vulvar Carcinoma Cells.

Whether G protein-coupled estrogen receptor 1 (GPER1) is tumor-promoting or tumor-suppressive depends in part on tumor entity. Little is known about the function of GPER1 in vulvar carcinoma. In this work, we aim to clarify what role GPER1 plays in vulvar cancer, tumor-promoting or tumor-suppressive. Localization of GPER1 in A431 and CAL-39 vulvar carcinoma cells was examined by immunofluorescence. Using a tissue microarray of vulvar neoplasias, the correlation between GPER1 expression and grade of malignancy was investigated. A431 and CAL-39 cells were treated either with GPER1 agonist G1 or antagonist G36. Proliferation was quantified by BrdU assay and viability examined using Resazurin assay. Morphological changes were analyzed by microscopy and measured using ImageJ. Cell migration was analyzed by gap closure assay. Clonogenic potential was tested by colony and sphere formation. Expression of estrogen receptors was examined by Western blot. GPER1 was found consistently expressed in vulvar neoplasia tissues. The immune-reactive score was found to be significantly higher in tissue samples of lymph node metastases and neoplasias with grade 3. In A431 and CAL-39 vulvar carcinoma cells, GPER1 expression was mainly found in the cytoplasm and nuclei. Treatment of A431 and CAL-39 cells with GPER1 agonist G1 resulted in a decrease in proliferation and migration. In addition, colony formation and tumor sphere formation were reduced. Furthermore, morphological signs of necrosis and reduction in cell viability after G1 treatment were observed. The GPER1 antagonist G36 did not have significant effects on vulvar carcinoma cells. Neither agonist G1 nor antagonist G36 treatment resulted in altered expression of estrogen receptors. Activation of GPER1 with GPER1 agonist G1 reduces the tumorigenic potential of the vulvar carcinoma cells. It can be deduced from this that GPER1 appears to have a tumor-suppressive effect in vulvar carcinoma.

The G protein-coupled estrogen receptor of the trigeminal ganglion regulates acute and chronic itch in mice.

Itch is an unpleasant sensation that severely impacts the patient's quality of life. Recent studies revealed that the G protein-coupled estrogen receptor (GPER) may play a crucial role in the regulation of pain and itch perception. However, the contribution of the GPER in primary sensory neurons to the regulation of itch perception remains elusive. This study aimed to investigate whether and how the GPER participates in the regulation of itch perception in the trigeminal ganglion (TG). Immunofluorescence staining results showed that GPER-positive (GPER+ ) neurons of the TG were activated in both acute and chronic itch. Behavioral data indicated that the chemogenetic activation of GPER+ neurons of the TG of Gper-Cre mice abrogated scratching behaviors evoked by acute and chronic itch. Conversely, the chemogenetic inhibition of GPER+ neurons resulted in increased itch responses. Furthermore, the GPER expression and function were both upregulated in the TG of the dry skin-induced chronic itch mouse model. Pharmacological inhibition of GPER (or Gper deficiency) markedly increased acute and chronic itch-related scratching behaviors in mouse. Calcium imaging assays further revealed that Gper deficiency in TG neurons led to a marked increase in the calcium responses evoked by agonists of the transient receptor potential ankyrin A1 (TRPA1) and transient receptor potential vanilloid V1 (TRPV1). Our findings demonstrated that the GPER of TG neurons is involved in the regulation of acute and chronic itch perception, by modulating the function of TRPA1 and TRPV1. This study provides new insights into peripheral itch sensory signal processing mechanisms and offers new targets for future clinical antipruritic therapy.

G protein-coupled estrogen receptor (GPER) selective agonist G1 attenuates the neurobehavioral, molecular and biochemical alterations induced in a valproic acid rat model of autism

AimsAutism Spectrum Disorder (ASD) is a neurodevelopmental disorder with a rising prevalence in boys rather than girls. G protein-coupled estrogen receptor (GPER) activation by its agonist G1 showed a neuroprotective effect, similar to estradiol. The present study aimed to examine the potential of the selective GPER agonist G1 therapy on the behavioral, histopathological, biochemical, and molecular alterations induced in a valproic acid (VPA)-rat model of autism. Main methodsVPA (500 mg/kg) was intraperitoneally administered to female Wistar rats (on gestational day 12.5) to induce the VPA-rat model of autism. The male offspring were intraperitoneally administered with G1 (10 and 20 μg/kg) for 21 days. After the treatment process, rats performed behavioral assessments. Then, sera and hippocampi were collected for biochemical and histopathological examinations and gene expression analysis. Key findingsGPER agonist G1 attenuated behavioral deficits, including hyperactivity, declined spatial memory and social preferences, anxiety, and repetitive behavior in VPA rats. G1 improved neurotransmission and reduced oxidative stress and histological alteration in the hippocampus. G1 reduced serum free T levels and interleukin-1β and up-regulated GPER, RORα, and aromatase gene expression levels in the hippocampus. SignificanceThe present study suggests that activation of GPER by its selective agonist G1 altered the derangements induced in a VPA-rat model of autism. G1 normalized free T levels via up-regulation of hippocampal RORα and aromatase gene expression. G1 provoked estradiol neuroprotective functions via up-regulation of hippocampal GPER expression. The G1 treatment and GPER activation provide a promising therapeutic approach to counteract the autistic-like symptoms.

Chronic GPER activation prompted the proliferation of ileal stem cell in ovariectomized mice depending on Paneth cell-derived Wnt3.

Menopausal women often face long-term estrogen treatment. G protein-coupled estrogen receptor (GPER) expressed in intestinal crypt was activated by estrogen therapy, but it was unclear whether chronic GPER activation during menopause had an effect on intestinal stem cells (ISCs). We tested the effect of chronic GPER activation on ISCs of ovariectomized (OVX) mice by injection of the selective GPER agonist G-1 for 28 days, or G-1 stimulation of organoids derived from crypts of OVX mice. G-1 up-regulated crypt depth, the number of Ki67+, bromodeoxyuridine+ cells and Olfm4+ ISCs, and the expression of ISCs marker genes (Lgr5, Olfm4 and Axin2). G-1 administration promoted organoid growth, increased the number of EdU+ cells per organoid and protein expression of Cyclin D1 and cyclin B1 in organoids. After G-1 treatment in vivo or in vitro, Paneth cell-derived Wnt3, Wnt3 effector β-catenin and Wnt target genes c-Myc and Cyclin D1 increased in ileum or organoids. Once blocking the secretion of Wnt3 from Paneth cells, the effects of G-1 on organoids growth, ISCs marker genes and Wnt/β-catenin signaling were abolished. G-1 did not affect the number of Paneth cells in ex vivo organoids, while activated Mmp7/cryptdin program in Paneth cells, promoted their maturation, and increased the expression of lysozyme protein. G-1 pretreatment in OVX mice inhibited radiation-induced ISCs proliferation injury and enhanced the resistance of mice to intestinal injury. In conclusion, chronic GPER activation prompted the Wnt3 synthesis in Paneth cells, thus increased the proliferation of ISCs via activation of Wnt3/β-catenin signaling in OVX mice.

Molecular identification of bulbospinal ON neurons by GPER, which drives pain and morphine tolerance.

The rostral ventromedial medulla (RVM) exerts bidirectional descending modulation of pain attributable to the activity of electrophysiologically identified pronociceptive ON and antinociceptive OFF neurons. Here, we report that GABAergic ON neurons specifically express G protein-coupled estrogen receptor (GPER). GPER+ neurons exhibited characteristic ON-like responses upon peripheral nociceptive stimulation. Optogenetic activation of GPER+ neurons facilitated, but their ablation abrogated, pain. Furthermore, activation of GPER caused depolarization of ON cells, potentiated pain, and ameliorated morphine analgesia through desensitizing μ-type opioid receptor-mediated (MOR-mediated) activation of potassium currents. In contrast, genetic ablation or pharmacological blockade of GPER attenuated pain, enhanced morphine analgesia, and delayed the development of morphine tolerance in diverse preclinical pain models. Our data strongly indicate that GPER is a marker for GABAergic ON cells and illuminate the mechanisms underlying hormonal regulation of pain and analgesia, thus highlighting GPER as a promising target for the treatment of pain and opioid tolerance.

In vitro and in silico assessment of GPER-dependent neurocytotoxicity of emerging bisphenols

To rapidly assess the toxicity of bisphenols (BPs) via the activation of G protein-coupled estrogen receptor (GPER), eight BPs action on GPER were evaluated by molecular docking and molecular dynamics (MD) simulation and then confirmed with IMR-32 cells. The target BPs significantly promoted the production of reactive oxygen species (ROS), reduced cell viability, activated the expression of apoptosis-related proteins and increased the apoptosis rate of IMR-32 cells. Intracellular Ca2+ level increased significantly after the treatments with bisphenol A (BPA), bisphenol E (BPE), bisphenol C (BPC) and bisphenol AP (BPAP), suggesting the activation of GPER. Moreover, the stable binding conformations between GPER and BPA, BPE, BPC and BPAP and their dynamic changes of GPER-BPs via MD simulation also suggest that these BPs may activate GPER. The interaction between bisphenol G/bisphenol P/bisphenol PH and GPER are weak, which is consistent with their low GPER activity in vitro. Notably, after the pretreatment of GPER antagonist, Ca2+ accumulation and ROS production induced by BPA, BPE, BPC and BPAP in IMR-32 cells were attenuated. Overall, MD simulation and in vitro results mutually verified the activation of GPER by BPs, and MD simulation can rapidly evaluate the neurocytotoxicity of BPs.

LBODP005 Gper Modulates Multiple Functions In Adipose Tissue To Promote An Anti-obese Phenotype

Abstract Obesity and metabolic syndrome are major health issues of modern society and their incidence is increasing at alarming rates worldwide. Obesity and associated chronic diseases exhibit sexual disparity with pre-menopausal females displaying better protection compared to age-matched males. These beneficial effects on metabolism in females are predominantly attributed to the female hormone estrogen, specifically 17β-estradiol (E2). E2 exerts its actions via multiple receptors, nuclear and extra-nuclear estrogen receptor (ER) α and ERβ, and the G protein-coupled estrogen receptor (GPER). GPER is present in diverse cells types and tissues and has been implicated in numerous physiological processes via multiple cellular signaling pathways. We have combined in vivo studies in an ovariectomized (OVX) mouse model along with in vitro assays in 3T3-L1 cells to investigate the role of GPER in modulating adipocyte function. Our studies reveal that loss of GPER increased adiposity in female mice on high fat diet and activation of GPER by its selective agonist G-1 attenuates adiposity in OVX mice. G-1 activated AMPK, increased cellular respiration and expression of mitochondrial and antioxidant genes and reduced lipid deposition in adipocytes. Furthermore, G-1 treatment promoted an anti-inflammatory phenotype in mice. Thus, our findings establish a role of GPER agonist G-1 in exerting anti-lipogenic and anti-inflammatory effects in adipocytes, which may have potential therapeutic implications in treatment of obesity. Presentation: No date and time listed

Targeted activation of GPER enhances the efficacy of venetoclax by boosting leukemic pyroptosis and CD8+ T cell immune function in acute myeloid leukemia

Acute myeloid leukemia (AML) is a rapidly progressing and often fatal hematopoietic malignancy. Venetoclax (VEN), a recent FDA-approved BCL-2 selective inhibitor, has high initial response rates in elderly AML patients, but the majority of patients eventually acquire resistance. Multiple studies have demonstrated that the female sex is associated with better outcomes in patients with AML, which are predominantly attributed to estrogen signaling. As a novel membrane estrogen receptor, G protein-coupled estrogen receptor (GPER)-mediated-rapid estrogen effects have attracted considerable attention. However, whether targeting GPER enhances the antileukemic activity of VEN is unknown. In this study, we first demonstrated that GPER expression was dramatically reduced in AML cells owing to promoter hypermethylation. Furthermore, pharmacological activation of GPER by G-1 combined with VEN resulted in synergistic antileukemic activity in vitro and in vivo. Mechanistically, G-1/VEN combination synergistically triggered concurrent mitochondria-related apoptosis and gasdermin E (GSDME)-dependent pyroptosis by activating p38-MAPK/myeloid cell leukemia 1 (MCL-1) axis. Importantly, leukemic pyroptosis heightened CD8+ T cell immune function by releasing interleukin (IL)-1β/18 into the tumor microenvironment. Our study corroborates that GPER activation shows a synergistic antileukemic effect with VEN, making it a promising therapeutic regimen for AML.

Involvement of the G-Protein-Coupled Estrogen Receptor-1 (GPER) Signaling Pathway in Neurodegenerative Disorders: A Review.

The G-protein-coupled estrogen receptor-1 (GPER) is an extranuclear estrogen receptor that regulates the expression of several downstream signaling pathways with a variety of biological actions including cell migration, proliferation, and apoptosis in different parts of the brain area. It is endogenously activated by estrogen, a steroidal hormone that binds to GPER receptors which help in maintaining cellular homeostasis and neuronal integrity as well as influences neurogenesis. In contrast, neurodegenerative disorders are a big problem for society, and still many people suffer from motor and cognitive impairments. Research to date reported that GPER has the potential to whittle down motor abnormalities and cognitive dysfunction by limiting the progression of neurodegenerative disorders. Although several findings suggest that GPER activation accelerated transcription of the PI3K/Akt/Gsk-3β and ERK1/2 signaling pathway that halt disease progression by decreasing oxidative stress, neuroinflammation, and apoptosis. Accordingly, the goal of this review is to highlight the basic mechanism of GPER signaling pathway-mediated neuroprotection in various neurodegenerative disorders including Parkinson's disease (PD), Huntington's disease (HD), Tardive dyskinesia (TD), and Epilepsy. This review also discusses the role of the GPER activators which might be a promising therapeutic target option to treat neurodegenerative disorders. All the data were obtained from published articles in PubMed (353), Web of Science (788), and Scopus (770) databases using the search terms: GPER, PD, HD, TD, epilepsy, and neurodegenerative disorders.

Neonicotinoid insecticides promote breast cancer progression via G protein-coupled estrogen receptor: In vivo, in vitro and in silico studies

Neonicotinoid insecticides (NIs) have been widely detected in environmental media and human body with concentrations reaching hundreds of nanomolar to micromolar levels. However, the information about their human health toxicology and mechanism is deficient. Previous studies have implied that NIs might exert estrogenic disruption and promote breast cancer progression, but the molecular mechanism is unclear, especially the molecular initiating event. G protein-coupled estrogen receptor (GPER), as a candidate therapeutic target, plays vital roles in the development of breast cancer. This work aimed to reveal the potential mechanism through GPER pathway. Firstly, we screened the activities of seven most common NIs on GPER signal pathway by calcium mobilization assay. Clothianidin, acetamiprid (ACE), and dinotefuran activated GPER most potently and ACE displayed the highest agonistic activity with the lowest observed effective concentration (LOEC) of 1 μM. The molecular docking and dynamics simulation showed favored interaction trend between the NIs and GPER. The three NIs with GPER activity induced 4T1 breast cancer cells migration and ACE showed the highest potency with LOEC of 100 nM. ACE also induced 4T1 cells proliferation at high concentration of 50 μM and up-regulated GPER expression in a dose-dependent manner. We speculated that both the induction effects of ACE on 4T1 cells proliferation and migration might be owing to the activation and up-regulation of GPER. By using 4T1-Luc cells injected orthotopic tumor model, we found that ACE also promoted in-situ breast cancer growth and lung metastasis in normal mouse dependent on GPER. However, ACE only promoted in-situ breast cancer growth through GPER but not lung metastasis in ovariectomized mice, implying that the ACE-induced lung metastasis should be related to endogenous estrogen from ovary. Overall, we demonstrated that NIs promoted breast cancer progression via GPER pathway at human related exposure levels and their female health risks need urgent concerns.

Expression and estrogen regulation of G protein-coupled estrogen receptor in human glioblastoma cells.

Glioblastoma (GB) is the most frequent primary brain tumor with a very poor prognosis. Sex hormones are crucial players in the development of GBs. 17 β-estradiol (E2) signaling is involved through its corresponding intracellular receptors [estrogen receptor α (ERα) and β (ERβ)] in GB cell proliferation and progression. E2 activates G-protein coupled estrogen receptor (GPER), leading to rapidly occurring effects, independently of gene transcription. GPER activation is involved in tumor progression in various cancer types. Currently, available data concerning the occurrence and role of GPER in GB are very limited. In the present study, it was observed that GPER was expressed in human brain tumor cell lines [U251 (astrocytoma-derived cell line), U87, LN229 and T98 (glioblastoma-derived cell line)]. Immunofluorescence assays revealed that GPER localizes in the plasma membrane, cytoplasm and nucleus. An in silico analysis identified two potential E2 response elements in the promoter region of the GPER gene. E2 increased GPER expression in the U251, U87 and LN229 cell lines. Molecular modeling data derived from in silico analysis predicted the three-dimensional conformation of GPER, and docking analysis identified potential binding sites of E2 and its specific agonist, G1. Taken together, these results indicate that GPER may be differentially expressed in human GB cell lines with E2 possibly upregulating GPER expression. The present study raises further questions about the implications of GPER-mediated E2 signaling in the biology of GBs.

Sex differences in the participation of endothelial mediators and signaling pathways involved in the vasodilator effect of a selective GPER agonist in resistance arteries of gonadectomized Wistar rats

AimEndothelial mechanisms underlying the vascular effects of estrogen modulated by the G protein-coupled estrogen receptor (GPER) are not well understood, especially in gonadal sex hormone deprivation. Thus, we investigated vascular function and endothelial signaling pathways involved in the selective activation of GPER in resistance arteries of gonadectomized rats. MethodsGonadectomy was performed in Wistar rats of both sexes. After 21 days, the animals were euthanized. Concentration-response curves were obtained by cumulative additions of G-1 in third-order mesenteric arteries. The vasodilatory effects of G-1 were evaluated before and after endothelium removal or incubation with pharmacological inhibitors. Tissue protein expression was measured by western blotting. Assays with 4-amino-5-methylamino-2′,7′-difluorofluorescein diacetate (DAF-FM) and 2′,7′ dichlorodihydrofluorescein-diacetate (H2DCF-DA) were performed in the arteries investigated. Immunolocalization was assessed by immunofluorescence. ResultsG-1 induced partially endothelium-dependent relaxation in both sexes. The three isoforms of the enzyme nitric oxide synthase contributed to the production and release of nitric oxide in both gonadectomized groups, but the role of inducible nitric oxide synthase is more expressive in males. The mechanistic pathway by which endothelial nitric oxide synthase is phosphorylated appears to differ between sexes, with the rapid signaling pathway phosphatidylinositol-3-kinase/protein kinase B/endothelial nitric oxide synthase (PI3k-Akt-eNOS) being identified for males and mitogen-activated protein kinase/extracellular signal-regulated kinase/endothelial nitric oxide synthase (MEK-ERK-eNOS) for females. The contribution of hydrogen peroxide as an endothelial relaxation mediator seems to be greater in females. ConclusionThese results provide new insights into the effects of estrogen-induced responses via GPER on vascular function in gonadal sex hormone deprivation.