Endogenous Metabolite; steroid hormone

Estradiol (10 nM, 7 d) stimulates neuronal development and enhances axonal branching in human endometrial stem cells (hEnSCs) [1]. Estradiol (17β-estradiol, 10 nM, 7 days) boosts the expression of neuron-like cell markers (Tuj-1, nestin and NF-H) in neuronal-like cells generated from hEnSCs [1].

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Human endometrial stem cells (hEnSCs) that can be differentiated into various neural cell types have been regarded as a suitable cell population for neural tissue engineering and regenerative medicine. Considering different interactions between hormones, growth factors, and other factors in the neural system, several differentiation protocols have been proposed to direct hEnSCs towards specific neural cells. The 17β-estradiol (E2) plays important roles in the processes of development, maturation, and function of nervous system. In the present research, the impact of 17β-estradiol (estrogen, E2) on the neural differentiation of hEnSCs was examined for the first time, based on the expression levels of neural genes and proteins. In this regard, hEnSCs were differentiated into neuron-like cells after exposure to retinoic acid (RA), epidermal growth factor (EGF), and also fibroblast growth factor-2 (FGF2) in the absence or presence of 17β-estradiol. The majority of cells showed a multipolar morphology. In all groups, the expression levels of nestin, Tuj-1 and NF-H (neurofilament heavy polypeptide) (as neural-specific markers) increased during 14 days. According to the outcomes of immunofluorescence (IF) and real-time PCR analyses, the neuron-specific markers were more expressed in the estrogen-treated groups, in comparison with the estrogen-free ones. These findings suggest that 17β-estradiol along with other growth factors can stimulate and upregulate the expression of neural markers during the neuronal differentiation of hEnSCs. Moreover, our findings confirm that hEnSCs can be an appropriate cell source for cell therapy of neurodegenerative diseases and neural tissue engineering [1].

Estradiol (1 nM, hippocampus slices from FBN-ARO-KO mice) restores LTP amplitude [1]. Estradiol (0.0167 mg, subcutaneously implanted) corrects molecular and functional abnormalities in FBN-ARO-KO mice [1].

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Depletion of neuron-derived 17β-estradiol (E2) leads to a significant decrease in dendritic spine density. Loss of neuron-derived E2 leads to a significant decrease in synapse number. Functional synaptic plasticity is significantly impaired in FBN-ARO-KO mice and rescued by acute E2 treatment. In vivo exogenous E2 replacement rescues the molecular and functional deficits in FBN-ARO-KO mice. [2]

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17β-estradiol (E2) is produced from androgens via the action of the enzyme aromatase. E2 is known to be made in neurons in the brain, but its precise functions in the brain are unclear. Here, we used a forebrain-neuron-specific aromatase knock-out (FBN-ARO-KO) mouse model to deplete neuron-derived E2 in the forebrain of mice and thereby elucidate its functions. FBN-ARO-KO mice showed a 70–80% decrease in aromatase and forebrain E2 levels compared with FLOX controls. Male and female FBN-ARO-KO mice exhibited significant deficits in forebrain spine and synaptic density, as well as hippocampal-dependent spatial reference memory, recognition memory, and contextual fear memory, but had normal locomotor function and anxiety levels. Reinstating forebrain E2 levels via exogenous in vivo E2 administration was able to rescue both the molecular and behavioral defects in FBN-ARO-KO mice. Furthermore, in vitro studies using FBN-ARO-KO hippocampal slices revealed that, whereas induction of long-term potentiation (LTP) was normal, the amplitude was significantly decreased. Intriguingly, the LTP defect could be fully rescued by acute E2 treatment in vitro. Mechanistic studies revealed that FBN-ARO-KO mice had compromised rapid kinase (AKT, ERK) and CREB-BDNF signaling in the hippocampus and cerebral cortex. In addition, acute E2 rescue of LTP in hippocampal FBN-ARO-KO slices could be blocked by administration of a MEK/ERK inhibitor, further suggesting a key role for rapid ERK signaling in neuronal E2 effects. In conclusion, the findings provide evidence of a critical role for neuron-derived E2 in regulating synaptic plasticity and cognitive function in the male and female brain.

For the in vitro 17β-estradiol (E2) rescue experiment, E2 was dissolved in DMSO, diluted to working concentration (1 nm) in oxygenated ACSF (Di Mauro et al., 2015). A DMSO (0.001%) vehicle control was also included in the experiment. In addition, the MEK inhibitor, U0126 (Cell Signaling Technology, catalog #9903S, 10 μm) was also dissolved in DMSO and diluted to working concentration in oxygenated ACSF. U0126 was coadministered with E2. The drugs were applied for all the recording period 20 min before the application of the stimulation protocol.[2]

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Measurement of 17β-estradiol (E2) levels. [2]
17β-estradiol (E2) levels in hippocampal CA1, cortex, and serum were measured using a high-sensitivity ELISA kit, as we described previously (Zhang et al., 2014). Briefly, 100 μl of sample was added into the bottom of an appropriate well coated with a donkey anti-sheep polyclonal antibody. Subsequently, 50 μl of E2 conjugate was added, followed by 50 μl of sheep polyclonal antibody, to E2. The plate was then sealed and incubated at room temperature with shaking speed of ∼500 rpm for 2 h. After washing 3 times with 400 μl of wash buffer each time, 200 μl of pNpp substrate was added into each well and incubated for 1 h at room temperature without shaking. Afterward, 50 μl of stop solution was added into each well and the optical density was read at 405 nm.

Cell Differentiation Assay[1]
Cell Types: Isolated human endometrial stem cells (hEnSCs) from human endometrial tissue
Tested Concentrations: 10 nM
Incubation Duration: 7 days
Experimental Results: Increased the number of neurite processes including neural differentiation and neurite branching.

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Immunofluorescence[1]
Cell Types: Isolated human endometrial stem cells (hEnSCs) from human endometrial tissue
Tested Concentrations: 10 nM
Incubation Duration: 7 days
Experimental Results: Increased the percentage of neural marker (Tuj-1, nestin and NF-H)-positive cells of 62.2±1.3%, 71.5±4% and 51.2±1.5% respectively.

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Neuronal differentiation of hEnSCs [1]
To induce neuronal differentiation, about 3 × 104 hEnSCs were seeded on each well of 24-well culture plates and incubated with two different differentiation media during 14 days. Cells were initially incubated with DMEM/F12 complete medium containing 1% Pen-Strep and also 10% FBS for 24 h at 37°C. In the first group, to induce neuronal differentiation of hEnSCs, we replaced the culture medium with the first step induction medium (DMEM/F12 containing EGF and FGF2 [each at 20 ng/ml concentration] and B27 [1%]) for 7 days. To continue neuronal differentiation, cells were subsequently exposed to the second step induction medium (DMEM/F12 supplemented with 1% ITS, 0.5 µM RA, and 20 ng/ml FGF2) for the next 7 days. In the second group, after 24 h of initial incubation, the expansion medium was replaced with the first step induction medium (DMEM/F12 containing EGF and FGF2 [each at 20 ng/ml concentration] and B27 [1%]) for 7 days and after that, cells were treated with the second step induction medium (DMEM/F12 supplemented with 1% ITS, 0.5 µM RA, 20 ng/ml FGF2, and 10 nM 17β-estradiol (E2) [Kang et al., 2007]) until day 14. Cells in the control groups were cultured on TCP in the presence of DMEM/F12 supplemented with 1% Pen-Strep and 10% FBS for 14 days. All media were changed every 2 days (Figure 1).

Animal/Disease Models: FBN-ARO-KO Mice[2]
Doses: 1 nM
Route of Administration: Treated for the hippocampal slices
Experimental Results: Rescued long-term potentiation (LTP) amplitude of both male and female mice.

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Animal/Disease Models: FBN-ARO-KO Mice [2]
Doses: 0.0167 mg
Route of Administration: Alzet minipumps with Estradiol (implanted sc), examined 7 days after minipump implantation.
Experimental Results: Restored hippocampal and cortical E2 levels to 93%, phosphorylation of AKT, ERK and CREB in the hippocampus and cortex to 90-95%, BDNF level to 80-90%, restored both synaptophysin and PSD95 in the forebrain. Rescued the spatial learning and memory defects.

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n vivo 17β-estradiol (E2) rescue experiment. [2]
Three-month-old ovx female mice were used in this experiment, which were divided into four groups: FLOX + placebo, FLOX + E2, FBN-ARO-KO + placebo, and FBN-ARO-KO + E2. Alzet minipumps osmotic minipumps; model 1007D, 7 d release; Durect) with placebo or E2 (0.0167 mg) were implanted subcutaneously in the upper midback region at the time of ovariectomy. The dose of E2 used here yields stable serum levels of 26.52 ± 0.89 pg/ml, which represents a physiological diestrus II–proestrus level of E2 (Nelson et al., 1981). Molecular and functional endpoints were examined 7 d after minipump implantation.

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Experimental design and statistical analyses. [2]
All quantitative analyses were performed on age-matched FLOX control and FBN-ARO-KO mice. Except for the in vivo 17β-estradiol (E2) rescue experiment, both male and female mice were used. Only female mice were used for in vivo E2 replacement. For behavioral tests, 8–10 mice were used for each group; otherwise, 4–6 samples from each group were analyzed. SigmaStat 3.5 software was used to analyze all data. Data represented in bar graphs were expressed as mean ± SE. A Student's t test was performed when only comparing two groups. Statistical data from the Barnes maze training trial, fear acquisition test, electrophysiological measurements, and part of the in vivo E2 replacement experiments requiring multiple groups comparisons were analyzed with two-way ANOVA followed by Tukey's all pairwise comparisons test to determine group differences. A value of p < 0.05 was considered statistically significant.

Absorption, Distribution and Excretion
The absorption of several formulations of estradiol is described below: Oral tablets and injections First-pass metabolism in the gastrointestinal tract rapidly breaks down estradiol tablets before entering the systemic circulation. The bioavailability of oral estrogens is said to be 2-10% due to significant first-pass effects. The esterification of estradiol improves the administration (such as with estradiol valerate) or to sustain release from intramuscular depot injections (including estradiol cypionate) via higher lipophilicity. After absorption, the esters are cleaved, which leads to the release of endogenous estradiol, or 17β-estradiol. Transdermal preparations The transdermal preparations slowly release estradiol through intact skin, which sustains circulating levels of estradiol during a 1 week period of time. Notably, the bioavailability of estradiol after transdermal administration is about 20 times higher than after oral administration. Transdermal estradiol avoids first pass metabolism effects that reduce bioavailability. Administration via the buttock leads to a Cmax of about 174 pg/mL compared to 147 pg/mL via the abdomen. Spray preparations After daily administration, the spray formulations of estradiol reach steady state within 7-8 days. After 3 sprays daily, Cmax is about 54 pg/mL with a Tmax of 20 hours. AUC is about 471 pg•hr/mL. Vaginal ring and cream preparations Estradiol is efficiently absorbed through the mucous membranes of the vagina. The vaginal administration of estrogens evades first-pass metabolism. Tmax after vaginal ring delivery ranges from 0.5 to 1 hour. Cmax is about 63 pg/mL. The vaginal cream preparation has a Cmax of estradiol (a component of Premarin vaginal estrogen conjugate cream) was a Cmax of 12.8 ± 16.6 pg/mL, Tmax of 8.5 ± 6.2 hours, with an AUC of 231 ± 285 pg•hr/mL.
Estradiol is excreted in the urine with both glucuronide and sulfate conjugates.
Estrogens administered exogenously distribute in a similar fashion to endogenous estrogens. They can be found throughout the body, especially in the sex hormone target organs, such as the breast, ovaries and uterus.
In one pharmacokinetic study, the clearance of orally administered micronized estradiol in postmenopausal women was 29.9±15.5 mL/min/kg. Another study revealed a clearance of intravenously administered estradiol was 1.3 mL/min/kg.
Estrogens used in therapeutics are well absorbed through the skin, mucous membranes, and the gastrointestinal (GI) tract. The vaginal delivery of estrogens circumvents first-pass metabolism.
The Estradiol Transdermal System Continuous Delivery (Once-Weekly) continuously releases estradiol which is transported across intact skin leading to sustained circulating levels of estradiol during a 7 day treatment period. The systemic availability of estradiol after transdermal administration is about 20 times higher than that after oral administration. This difference is due to the absence of first-pass metabolism when estradiol is given by the transdermal route.
In a Phase I study of 14 postmenopausal women, the insertion of ESTRING (estradiol vaginal ring) rapidly increased serum estradiol (E2) levels. The time to attain peak serum estradiol levels (Tmax) was 0.5 to 1 hour. Peak serum estradiol concentrations post-initial burst declined rapidly over the next 24 hours and were virtually indistinguishable from the baseline mean (range: 5 to 22 pg/mL). Serum levels of estradiol and estrone (E1) over the following 12 weeks during which the ring was maintained in the vaginal vault remained relatively unchanged
Table: PHARMACOKINETIC MEAN ESTIMATES FOLLOWING SINGLE ESTRING APPLICATION [Table#4649]

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Metabolism / Metabolites
Exogenously administered estrogens are metabolized in the same fashion as endogenous estrogens. Metabolic transformation occurs primarily in the liver and intestine. Estradiol is metabolized to estrone, and both are converted to estriol, which is later excreted in the urine. Sulfate and glucuronide conjugation estrogens also take place in the liver. Biliary secretion of metabolic conjugates are released into the intestine, and estrogen hydrolysis in the gut occurs, followed by reabsorption. The CYP3A4 hepatic cytochrome enzyme is heavily involved in the metabolism of estradiol. CYP1A2 also plays a role.
Exogenous estrogens are metabolized in the same manner as endogenous estrogens. Circulating estrogens exist in a dynamic equilibrium of metabolic interconversions. These transformations take place mainly in the liver. Estradiol is converted reversibly to estrone, and both can be converted to estriol, which is the major urinary metabolite. Estrogens also undergo enterohepatic recirculation via sulfate and glucuronide conjugation in the liver, biliary secretion of conjugates into the intestine, and hydrolysis in the gut followed by reabsorption. In postmenopausal women, a significant proportion of the circulating estrogens exist as sulfate conjugates, especially estrone sulfate, which serves as a circulating reservoir for the formation of more active estrogens.
Variations in estradiol metabolism ... depend upon the stage of the menstrual cycle ... In general, the hormone undergoes rapid hepatic biotransformation with a plasma half-life measured in minutes.
Estradiol is primarily converted ... to estriol, which is the major urinary metabolite. A variety of sulfate and glucuronide conjugates also are excreted in the urine.
The metabolism of estradiol-17beta and estrone is similar in rats and in humans, in that both species transform these steroids mainly by (aromatic) 2-hydroxylation, and also by 16alpha-hydroxylation. Glucuronides of the various metabolites are excreted in the bile. Differences in the metabolism of estrogens by humans and rats lie mostly in the type of conjugation. A relatively large proportion of administered estrone, estradiol-17beta and estriol is transformed in rats to metabolites oxygenated both at C-2 and C-16. When estriol is administered to rats, glucuronides and, to a lesser extent, sulfates of 16-ketooestradiol and of 2- and 3-methyl ethers of 2-hydroxyoestriol and 2-hydroxy-16-ketooestradiol are excreted in the bile. In contrast, hydroxylations at C-6 or C-7 of ring B of estradiol-17beta and estrone are a minor pathway in rats. 2-Hydroxyoestrogens ('catechol estrogens') are further transformed by various routes, including covalent binding to proteins.
For more Metabolism/Metabolites (Complete) data for ESTRADIOL (8 total), please visit the HSDB record page.
17-beta-estradiol has known human metabolites that include 2-hydroxyestradiol, 4-Hydroxyestradiol, 17-beta-Estradiol-3-glucuronide, and 17-beta-Estradiol glucuronide.
Exogenous estrogens are metabolized using the same mechanism as endogenous estrogens. Estrogens are partially metabolized by cytochrome P450.
Route of Elimination: Estradiol, estrone and estriol are excreted in the urine along with glucuronide and sulfate conjugates.
Half Life: 36 hours

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Biological Half-Life
The terminal half-lives for various estrogen products post oral or intravenous administration has been reported to range from 1-12 hours. One pharmacokinetic study of oral estradiol valerate administration in postmenopausal women revealed a terminal elimination half-life of 16.9 ± 6.0 h. A pharmacokinetic study of intravenous estradiol administration in postmenopausal women showed an elimination half-life of 27.45 ± 5.65 minutes. The half-life of estradiol appears to vary by route of administration.
... After oral administration ... the terminal half life was 20.1 hr ...

Toxicity Summary
Estradiol enters target cells freely (e.g., female organs, breasts, hypothalamus, pituitary) and interacts with a target cell receptor. When the estrogen receptor has bound its ligand it can enter the nucleus of the target cell, and regulate gene transcription which leads to formation of messenger RNA. The mRNA interacts with ribosomes to produce specific proteins that express the effect of estradiol upon the target cell. Estrogens increase the hepatic synthesis of sex hormone binding globulin (SHBG), thyroid-binding globulin (TBG), and other serum proteins and suppress follicle-stimulating hormone (FSH) from the anterior pituitary.

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Interactions
Estrogens may interfere with the effects of bromocriptine; dosage adjustment may be necessary. /Estrogens/
A combination of testosterone and estradiol-B17 after treatment with methylnitrosurea also resulted in the development of adenocarciomas of the prostate.
Concurrent use with estrogens may increase calcium absorption and exacerbate nephrolithiasis in susceptible individuals; this can be used to therapeutic advantage to increase bone mass. /Estrogens/
Concurrent use /of glucocorticoid corticosteroids/ with estrogens may alter the metabolism and protein binding of the glucocorticoids, leading to decreased clearance, increased elimination half-life, and increased therapeutic and toxic effects of the glucocorticoids; glucocorticoid dosage adjustment may be required during and following concurrent use. /Estrogens/
For more Interactions (Complete) data for ESTRADIOL (11 total), please visit the HSDB record page.

[1]. Elham Hasanzadeh. Defining the role of 17β-estradiol in human endometrial stem cells differentiation into neuron-like cells. Cell Biol Int. 2021 Jan;45(1):140-153.

[2]. Neuron-Derived Estrogen Regulates Synaptic Plasticity and Memory. J Neurosci. 2019 Apr 10;39(15):2792-2809.

Therapeutic Uses
Estradiol tablets are indicated in the treatment of moderate to severe vasomotor symptoms associated with the menopause. /Included in US product label/
Estradiol tablets are indicated in the treatment of moderate to severe symptoms of vulvar and vaginal atrophy associated with the menopause. When prescribing solely for the treatment of symptoms of vulvar and vaginal atrophy, topical vaginal products should be considered. /Included in US product label/
Estradiol tablets are indicated in the treatment of hypoestrogenism due to hypogonadism, castration or primary ovarian failure. /Included in US product label/
Estradiol tablets are indicated in the treatment of breast cancer (for palliation only) in appropriately selected women and men with metastatic disease. /Included in US product label/
For more Therapeutic Uses (Complete) data for ESTRADIOL (7 total), please visit the HSDB record page.

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Drug Warnings
ESTROGENS INCREASE THE RISK OF ENDOMETRIAL CANCER- Close clinical surveillance of all women taking estrogens is important. Adequate diagnostic measures, including endometrial sampling when indicated, should be undertaken to rule out malignancy in all cases of undiagnosed persistent or recurring abnormal vaginal bleeding. There is no evidence that the use of "natural" estrogens results in a different endometrial risk profile than "synthetic" estrogens at equivalent estrogen doses.
CARDIOVASCULAR AND OTHER RISKS- Estrogens with or without progestins should not be used for the prevention of cardiovascular disease.
The Women's Health Initiative (WHI) study reported increased risks of myocardial infarction, stroke, invasive breast cancer, pulmonary emboli, and deep vein thrombosis in postmenopausal women (50 to 79 years of age) during 5 years of treatment with oral conjugated estrogens (CE 0.625 mg) combined with medroxyprogesterone acetate (MPA 2.5 mg) relative to placebo.
The Women's Health Initiative Memory Study (WHIMS), a substudy of WHI, reported increased risk of developing probable dementia in postmenopausal women 65 years of age or older during 4 years of treatment with oral conjugated estrogens plus medroxyprogesterone acetate relative to placebo. It is unknown whether this finding applies to younger postmenopausal women or to women taking estrogen alone therapy.
For more Drug Warnings (Complete) data for ESTRADIOL (48 total), please visit the HSDB record page.

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Pharmacodynamics
Estradiol acts on the on the estrogen receptors to relieve vasomotor systems (such as hot flashes) and urogenital symptoms (such as vaginal dryness and dyspareunia). Estradiol has also been shown to exert favorable effects on bone density by inhibiting bone resorption. Estrogen appears to inhibit bone resorption and may have beneficial effects on the plasma lipid profile. Estrogens cause an increase in hepatic synthesis of various proteins, which include sex hormone binding globulin (SHBG), and thyroid-binding globulin (TBG). Estrogens are known to suppress the formation of follicle-stimulating hormone (FSH) in the anterior pituitary gland. **A note on hyper-coagulable state, cardiovascular health, and blood pressure** Estradiol may cause an increased risk of cardiovascular disease, DVT, and stroke, and its use should be avoided in patients at high risk of these conditions. Estrogen induces a hyper-coagulable state, which is also associated with both estrogen-containing oral contraceptive (OC) use and pregnancy. Although estrogen causes an increase in levels of plasma renin and angiotensin. Estrogen-induced increases in angiotensin, causing sodium retention, which is likely to be the mechanism causing hypertension after oral contraceptive treatment.

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17β-estradiol (E2) is produced from androgens via the action of the enzyme aromatase. E2 is known to be made in neurons in the brain, but its precise functions in the brain are unclear. Here, we used a forebrain-neuron-specific aromatase knock-out (FBN-ARO-KO) mouse model to deplete neuron-derived E2 in the forebrain of mice and thereby elucidate its functions. FBN-ARO-KO mice showed a 70-80% decrease in aromatase and forebrain E2 levels compared with FLOX controls. Male and female FBN-ARO-KO mice exhibited significant deficits in forebrain spine and synaptic density, as well as hippocampal-dependent spatial reference memory, recognition memory, and contextual fear memory, but had normal locomotor function and anxiety levels. Reinstating forebrain E2 levels via exogenous in vivo E2 administration was able to rescue both the molecular and behavioral defects in FBN-ARO-KO mice. Furthermore, in vitro studies using FBN-ARO-KO hippocampal slices revealed that, whereas induction of long-term potentiation (LTP) was normal, the amplitude was significantly decreased. Intriguingly, the LTP defect could be fully rescued by acute E2 treatment in vitro Mechanistic studies revealed that FBN-ARO-KO mice had compromised rapid kinase (AKT, ERK) and CREB-BDNF signaling in the hippocampus and cerebral cortex. In addition, acute E2 rescue of LTP in hippocampal FBN-ARO-KO slices could be blocked by administration of a MEK/ERK inhibitor, further suggesting a key role for rapid ERK signaling in neuronal E2 effects. In conclusion, the findings provide evidence of a critical role for neuron-derived E2 in regulating synaptic plasticity and cognitive function in the male and female brain.SIGNIFICANCE STATEMENT The steroid hormone 17β-estradiol (E2) is well known to be produced in the ovaries in females. Intriguingly, forebrain neurons also express aromatase, the E2 biosynthetic enzyme, but the precise functions of neuron-derived E2 is unclear. Using a novel forebrain-neuron-specific aromatase knock-out mouse model to deplete neuron-derived E2, the current study provides direct genetic evidence of a critical role for neuron-derived E2 in the regulation of rapid AKT-ERK and CREB-BDNF signaling in the mouse forebrain and demonstrates that neuron-derived E2 is essential for normal expression of LTP, synaptic plasticity, and cognitive function in both the male and female brain. These findings suggest that neuron-derived E2 functions as a novel neuromodulator in the forebrain to control synaptic plasticity and cognitive function. [2]

C18H24O2

272.38

272.177

C, 79.37; H, 8.88; O, 11.75

50-28-2

Alpha-Estradiol;57-91-0;Estradiol (Standard);50-28-2;Estradiol-d3;79037-37-9;Estradiol-d4;66789-03-5;Estradiol-d5;221093-45-4;Estradiol-13C2;82938-05-4;Estradiol (cypionate);313-06-4;Estradiol benzoate;50-50-0;Estradiol enanthate;4956-37-0;Estradiol hemihydrate;35380-71-3;Estradiol-d2;53866-33-4;Estradiol-13C6;Estradiol-d2-1;3188-46-3;rel-Estradiol-13C6; 979-32-8 (valerate); 113-38-2 (dipropionate); 57-63-6 (ethinyl); 172377-52-5 (sulfamate); 3571-53-7 (undecylate)

5757

White to off-white solid powder

1.2±0.1 g/cm3

445.9±45.0 °C at 760 mmHg

173ºC

209.6±23.3 °C

0.0±1.1 mmHg at 25°C

1.599

4.13

2

2

0

20

382

5

O([H])[C@@]1([H])C([H])([H])C([H])([H])[C@@]2([H])[C@]3([H])C([H])([H])C([H])([H])C4C([H])=C(C([H])=C([H])C=4[C@@]3([H])C([H])([H])C([H])([H])[C@@]21C([H])([H])[H])O[H]

VOXZDWNPVJITMN-ZBRFXRBCSA-N

InChI=1S/C18H24O2/c1-18-9-8-14-13-5-3-12(19)10-11(13)2-4-15(14)16(18)6-7-17(18)20/h3,5,10,14-17,19-20H,2,4,6-9H2,1H3/t14-,15-,16+,17+,18+/m1/s1

(8R,9S,13S,14S,17S)-13-methyl-6,7,8,9,11,12,14,15,16,17-decahydrocyclopenta[a]phenanthrene-3,17-diol

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Solubility in Formulation 1: ≥ 2.5 mg/mL (9.18 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: 2.5 mg/mL (9.18 mM) in 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (9.18 mM) in 5% DMSO + 95% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 4: ≥ 2.08 mg/mL (7.64 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 5: ≥ 2.08 mg/mL (7.64 mM)(saturation unknown) in ≥ 2.5 mg/mL (5.35 mM) (add these co-solvents sequentially from left to right, and one by one),clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 6: 12.5 mg/mL (45.89 mM) in Corn Oil (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.

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 (Please use freshly prepared in vivo formulations for optimal results.)