Neuroprotective Effect Of 17β-estradiol Research Articles

Therapeutic Assessment and Neuroprotective Effects of a Novel G Protein‐Coupled Estrogen Receptor (GPR30/GPER) Agonist

The G protein‐coupled estrogen receptor (GPER) has been implicated in the neuroprotective effects of 17β‐estradiol and selective GPER agonist, G‐1. Currently, G‐1 remains the standard for GPER agonists, necessitating the design and identification of novel candidates. Previously, our group synthesized and biologically evaluated a series of GPER antagonists. Modification of an aromatic moiety on the scaffold altered the pharmacological profile from antagonists to agonists. Using a fluorescence‐based binding assay for the classical estrogen receptors, ERa and ERb, we discovered off‐target binding. Due to the similarity in functionalization to estradiol, we hypothesized that the hydrogen bonding abilities of the initial candidate were responsible for the observed off‐target binding. To mitigate off‐target binding, modifications were made to reduce the hydrogen bonding ability. For each resulting candidate, the EC50 value was determined through in vitro biochemical screening. From these studies, we identified a lead compound that exhibited an EC50 value below that of our experimentally determined EC50 value of G‐1. Using E18 rat embryonic hippocampal neurons, the lead compound was shown to increase synaptogenesis. These results suggest that the identified lead compound for GPER may be a potential candidate for neurodegenerative diseases.Support or Funding InformationSaint Louis UniversityThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Neuroprotective Effects of 17β-Estradiol against Thrombin-Induced Apoptosis in Primary Cultured Cortical Neurons

Aims: 17β-estradiol (E2) is a powerful neuroprotective agent in the central nervous system; however, little is known about its effects on intracerebral hemorrhage. This study examined the effects of E2 on thrombin-induced apoptosis in vitro and investigated the potential mechanisms. Methods: Primary cultured cortical neurons were treated with E2 or vehicle and then the cells were exposed to thrombin. Neuronal apoptosis was assessed by flow cytometry. The phosphorylated c-Jun-N-terminal kinase (p-JNK), phosphorylated extracellular signal-regulated kinases 1/2 (p-ERK1/2), B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax) and caspase-3 were assayed by western blot. Results: Consequently, we found that E2 has significantly reduced the apoptosis in thrombin-treated neurons. E2 also exhibited a downregulation in the ratio of Bax/Bcl-2, caspase-3 and p-JNK. However, E2 had little effect on p-ERK1/2 proteins activation. Conclusion: Taken together, E2 has shown neuroprotective effects on thrombin-induced neuronal apoptosis, and the molecular mechanisms may correlate with the inhibition of the JNK signaling pathway.

The Effects of Estrogen Receptors' Antagonist on Brain Edema, Intracranial Pressure and Neurological Outcomes after Traumatic Brain Injury in Rat.

Background:In previous studies, the neuroprotective effect of 17β-estradiol in diffuse traumatic brain injury has been shown. This study used ICI 182,780, a non-selective estrogen receptor antagonist, to test the hypothesis that the neuroprotective effect of 17β-estradiol in traumatic brain injury is mediated by the estrogen receptors. Methods: The ovariectomized rats were divided into eight groups. Brain injury was induced by Marmarou’s method. Estrogen was injected 30 minutes after traumatic brain injury, and ICI 182,780 was injected before traumatic brain injury and also before estrogen treatment. In one group only ICI 182,780 was injected. The brain water content and Evans blue dye content were measured 24 and 5 hours after traumatic brain injury, respectively. The neurologic outcomes and intracranial pressure were assessed before, 4, and 24 hours after traumatic brain injury. Results: Brain water content and Evans blue content were less in estrogen-treated group comparison to vehicle group. ICI 182,780 eliminated the effects of estrogen on brain edema and brain blood barrier permeability. Intracranial pressure was increased significantly after trauma, and estrogen decreased intracranial pressure at 4 and 24 hours after traumatic brain injury in comparison to vehicle. This inhibitory effect was also eliminated by treatment with ICI182,780. ICI 182,780 also inhibited the estrogen induced increase in neurologic outcomes following traumatic brain injury. However, the use of ICI 182,780 alone had no neuroprotective effect after traumatic brain injury. Conclusion: The results suggest that classical estrogen receptors have probably a role in the neuroprotective function of estrogen following traumatic brain injury.

A Combination Therapy of 17β-Estradiol and Memantine Is More Neuroprotective Than Monotherapies in an Organotypic Brain Slice Culture Model of Traumatic Brain Injury.

Combination therapies are a promising therapeutic option for traumatic brain injury (TBI) owing to the clinical failure of monotherapy treatments, such as progesterone. Organotypic hippocampal slice cultures (OHSCs) from Sprague-Dawley rats were subjected to an in vitro TBI, and the neuroprotective effects of 17β-estradiol (E2) or memantine (MEM) monotherapies were quantified. Several combination treatments at different concentrations of both drugs were tested, with 100 pM of E2 and 10 μM of MEM statistically and significantly reducing cell death over either monotherapy when administered immediately after injury. This combination was also significantly neuroprotective when administered 1 h postinjury, possibly supporting future in vivo studies. Further, we hypothesized that this synergy could be the result of MEM blocking a potentially deleterious effect of E2, specifically E2 enhancement of N-methyl-D-aspartate (NMDA) currents. Evoked electrophysiological responses in OHSCs were potentiated by E2 treatment, whereas this potentiation was significantly reduced by MEM. In conclusion, a combination therapy of E2 and memantine was significantly more neuroprotective than both monotherapy treatments, and this synergy may be the result of MEM blocking a deleterious E2-mediated enhancement of NMDA receptors.

The neuroprotective effect of 17β-estradiol is independent of its antioxidative properties

Introduction17β-Estradiol (E2) is neuroprotective in experimental models of stroke. While some postulate a mainly antioxidative action due to E2׳s free C3 hydroxyl group at its A-ring, others suggest that neuroprotection is mediated by a hormonal, receptor mediated effect. The aim of the current study was to clarify this issue by testing whether E2 analogues lacking hormonal activity are also neuroprotective following cerebral ischemia. Material & MethodsFocal cerebral ischemia was induced in male C57/BL6 mice by laser-Doppler-controlled endovascular occlusion of the middle cerebral artery for 40min. Mice received either 1) memantine, a NMDA-receptor antagonist, as a positive control, 2) E2 (1400µg/kg b.w.), or 3) 2,4,6-trimethylphenol (TMP), an E2 analogue without hormonal activity (1400, 140, or 14µg/kg b.w.). Motor function was tested 3h and 24h after ischemia. Thereafter mice were sacrificed and brain damage was quantified by histomorphometry. ResultsTreatment with memantine or E2 significantly reduced infarct volume by >40% and significantly improved neurological function while treatment with TMP had no effect. ConclusionE2 is equally neuroprotective as antagonization of NMDA receptors while E2 analogues without hormonal activity are not neuroprotective. Therefore the current data suggest that the neuroprotection activity of E2 is independent of its free-radical scavenging properties.

The selective estrogen receptor modulator, bazedoxifene, reduces ischemic brain damage in male rat

While the estrogen treatment of stroke is under debate, selective estrogen receptor modulators (SERMs) arise as a promising alternative. We hypothesize that bazedoxifene (acetate, BZA), a third generation SERM approved for the treatment of postmenopausal osteoporosis, reduces ischemic brain damage in a rat model of transient focal cerebral ischemia. For comparative purposes, the neuroprotective effect of 17β-estradiol (E2) has also been assessed. Male Wistar rats underwent 60min middle cerebral artery occlusion (intraluminal thread technique), and grouped according to treatment: vehicle-, E2- and BZA-treated rats. Optimal plasma concentrations of E2 (45.6±7.8pg/ml) and BZA (20.7±2.1ng/ml) were achieved 4h after onset of ischemia, and maintained until the end of the procedure (24h). Neurofunctional score and volume of the damaged brain regions were the main end points. At 24h after ischemia–reperfusion, neurofunctional examination of the animals did not show significant differences among the three experimental groups. By contrast, both E2- and BZA-treated groups showed significantly lower total infarct volumes, BZA acting mainly in the cortical region and E2 acting mainly at the subcortical level. Our results demonstrate that: (1) E2 at physiological plasma levels in female rats is neuroprotective in male rats when given at the acute stage of the ischemic challenge and (2) BZA at clinically relevant plasma levels mimics the neuroprotective action of E2 and could be, therefore, a candidate in stroke treatment.

Neuroprotective Effects of Nonfeminizing Estrogens in Retinal Photoreceptor Neurons

Retinal diseases such as macular degeneration and glaucoma are disorders that target specific retinal neurons that can ultimately lead to vision loss. Under these conditions and pathologies, retinal neurons can die via apoptosis that may be due to increased oxidative stress. The neuroprotective effects of 17β-estradiol (E2) and three synthetic nonfeminizing estrogen analogs (ZYC-26, ZYC-23, and ZYC-3) were investigated to examine their abilities to protect retinal neurons against glutamate toxicity. Using an in vitro model of glutamate-induced cell death in 661W cells, a mouse cone photoreceptor cell line, shown to express both estrogen receptors (ERs) via immunoblotting, was pretreated with E2 and its analogs and cell viability were assessed. It was observed that E2 and estrogen analogs, ZYC-26 and ZYC-3, were protective against a 5 mM glutamate insult in 661W cells. The neuroprotective abilities of ZYC-26 and ZYC-3 were autonomous of estrogen receptor-α (ERα) and ERβ demonstrated by their ability to protect in the presence of ICI 182780, a pan-ER antagonist with a high affinity for the estrogen receptor. Treatment with PPT and DPN, ERα- and ERβ-specific agonists, respectively, did not protect the 661W cells from the glutamate insult. Studying the membrane ER (mER) or GPR30 did show that activation of the receptor by G1 protected the retinal neuron from insult, whereas G15, an antagonist of the mER was not able to antagonize the protection previously seen. These data demonstrate that nonfeminizing estrogens may emerge as useful compounds for neuroprotection of retinal cells.

Estrogen protection against mitochondrial toxin-induced cell death in hippocampal neurons: Antagonism by progesterone

Previously we demonstrated that mitochondrial dysfunction plays a critical role in the pathogenesis of Alzheimer's disease. Further, we have shown that the neuroprotective effects of 17β-estradiol (E2) are dependent upon mitochondrial function. In the current study, we sought to identify mitochondrial sites of E2 action that mediate neuroprotection by assessing the efficacy of E2 to protect neurons against inhibitors of mitochondrial respiration which target specific complexes within the respiratory chain. Subsequently, the impact of progesterone (P4) on E2-induced prevention against mitochondrial toxins was investigated. Mitochondrial inhibitors, rotenone, 3-NPA, antimycin, KCN, and oligomycin, exhibited concentration dependent toxicity in primary hippocampal neurons. The concentration inducing 30% cell death (LD30) was selected for analyses assessing the neuroprotective efficacy of ovarian hormones (E2 and P4). Pretreatment of hippocampal neurons with E2 significantly protected against 3-NPA (7.5 mM) and antimycin (125 μM) induced cell death and was moderately neuroprotective against rotenone (3 μM). E2 was ineffective against KCN and oligomycin-induced cell death. Pretreatment with P4 was without effect against these mitochondrial inhibitors. Co-administration of P4 with E2 abolished E2 induced neuroprotection against 3-NPA and antimycin. Additional metabolic analyses indicated that E2 and P4 separately increased mitochondrial respiratory capacity whereas the co-administration of E2 and P4 resulted in diminished mitochondrial respiration. These findings indicate that E2 protects against mitochondrial toxins that target complexes I, II and III whereas P4 was without effect. The data also predict that continuous combined co-administration of estrogen and progesterone common to many hormone therapy regimens is unlikely to prevent the deficits in mitochondrial function.

Mechanisms of Estrogen Receptor Neuroprotection in Dopamine Neurons

Estrogen involvement in neuroprotection is now a widely accepted research area, although the mechanisms of estrogen action in neuroprotection remain unclear. To investigate the neuroprotective effects of 17‐β estradiol on dopaminergic neurons we utilized primary mesencephalic neurons cultured from embryos of time‐mated C57 Bl/6 mice. The primary mesencephalic cultures allow for analysis of both direct estrogen‐mediated neuorprotection on the dopamine neuron and/or indirect actions through surrounding glial cells. We demonstrate the existence of both ERα and ERβ with a predominance of ERα on both dopamine neurons and astrocytes. In primary cultures of mouse mesencephalic neurons estrogen protects dopamine neurons from injury induced by 1‐methyl‐4‐phenyl pyridium (MPP+). ERα mediated the protection afforded by estrogen since only the ERα agonist HPTE protected against dopamine cell loss. The ERβ agonist DPN, alone did not protect against MPP+ induced neuronal death. Since glial cells were shown to express ERα we investigated an indirect mechanism of estrogen action on dopamine neuron survival. Treating the cultures with the mitotic inhibitor, 5‐Fluoro‐2’‐deoxyuridine (dFUR) abolished glial cells and significantly reduced the amount of neuroprotection seen with estrogen. These data indicate that neuroprotection provided by estrogen is mediated by ERα and involves a interplay among several cell types.

Dose dependence and therapeutic window for the neuroprotective effects of 17β-estradiol when administered after cerebral ischemia

The present study was undertaken to determine if the neuroprotective effect of 17β-estradiol (E 2) when administrated after ischemia is dose-dependent and if the therapeutic window for estrogen can be prolonged. Ischemic injury was induced by permanent middle cerebral artery occlusion (p-MCAO). Administration of E 2 at 30 min after ischemia resulted in a reduction in lesion volume. A higher dose of E 2 extended the therapeutic window to 6 h after cerebral ischemia in 33% of the rats. These findings suggest that postischemic treatment with estrogen affords protection against ischemic damage and that it acts within a clinically useful therapeutic window.

Estrogens influence behavioral responses in a kainic acid model of neurotoxicity

The behavioral and neuroprotective effects of 17β-estradiol (E2), on ovariectomized rats treated with a subconvulsive dose (7 mg/kg bw, ip) of kainic acid (KA), were examined. Estradiol was administered either acutely (150 μg/rat, ip) along with KA, 14 days post-ovariectomy, or chronically (sc capsules providing proestrus estrogen levels in serum) starting at ovariectomy. Exploratory behavior, as deduced by sniffing in the open field test, was reduced in KA-treated rats. Both hormonal schemes partially restored sniffing behavior in KA-lesioned subjects. Moreover, acute and chronic E2 administration in KA-treated rats resulted in increased vertical and horizontal activity of these animals in the open field test. Memory for object recognition was reduced following KA and was not restored by hormonal treatments. Acute, but not chronic, E2 coadministration with KA significantly impaired spatial performance in the water maze task, while KA alone had no effect. Both acute and chronic estradiol administration rescued hilar and CA1 neurons from KA-induced cell death. Chronic, but not acute, E2 increased neurofilament immunoreactivity in the mossy fibers of the dentate gyrus neurons, similarly to KA. Our results show that although estradiol administration in KA-treated rats has beneficial effects on cell survival, it has diverse effects on exploratory behavior, object, and spatial memory. Estradiol effects on KA-lesioned animals depended on the duration and timing of exposure to the hormone, implying different mechanisms of hormone actions.

Neuroprotective and neurotoxic effects of estrogens

The ovarian hormone 17β-estradiol (E2) is neuroprotective in animal models of neurodegenerative diseases. Some studies suggest that the neuroprotective effects of 17β-estradiol are a consequence of its antioxidant activity that depend on the hydroxyl group in the C3 position of the A ring. As in other tissues, 17β-estradiol is metabolized in the brain to 2-hydroxyestradiol (2OHE2) and 2-methoxyestradiol (2MEOHE2). These two molecules present the hydroxyl group in the A ring and have a higher antioxidant activity than 17β-estradiol. To test the hypothesis that conversion to 2-hydroxyestradiol and 2-methoxyestradiol may mediate neuroprotective actions of 17β-estradiol in vivo, we have assessed whether these molecules protect hilar hippocampal neurons from kainic acid toxicity. Ovariectomized Wistar rats received an i.p. injection of 1, 10 or 100 μg 17β-estradiol, 2-hydroxyestradiol or 2-methoxyestradiol followed by an i.p. injection of kainic acid (7 mg/kg) or vehicle. Treatment with kainic acid resulted in a significant loss of hilar neurons. Only the highest dose tested of 17β-estradiol (100 μg/rat) prevented kainic acid-induced neuronal loss. 2-Hydroxyestradiol and 2-methoxyestradiol did not protect hilar neurons from kainic acid, suggesting that the mechanism of neuroprotection by 17β-estradiol in vivo is not mediated by its metabolism to catecholestrogens or methoxycatecholestrogens. Furthermore, 2-methoxyestradiol (100 μg/rat), by itself, resulted in a significant neuronal loss in the hilus that was detected 96 h after the treatment with the steroid. This finding suggests that endogenous metabolism of 17β-estradiol to 2-methoxyestradiol may counterbalance the neuroprotective effects of the hormone.