Current strategies for quantification of estrogens in clinical research

Denver, Nina; Khan, Shazia; Homer, Natalie; MacLean, Margaret R.; Andrew, Ruth

doi:10.1016/j.jsbmb.2019.04.022

Cited by 59 publications

(37 citation statements)

References 94 publications

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“…After the cells were 40–50% confluent, the medium was replaced with phenol red-free DMEM (Gibco-BRL) supplemented with 1% charcoal-stripped FBS (Gibco-BRL). 17β-estradiol (E 2 ; 7.14 nmol/l; Sigma-Aldrich), which is the most active form of estrogen [20], was added to the culture medium at gradient concentrations (0, 10 − 9 , 10 − 7 and 10 − 5 mol/l) for 24 h.…”

Section: Methodsmentioning

confidence: 99%

Estrogen stimulates SREBP2 expression in hepatic cell lines via an estrogen response element in the SREBP2 promoter

Meng

Zong

2019

Cell Mol Biol Lett

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ObjectiveHypoestrogenism in women is strongly associated with menopause and it can lead to lipid disorder, which predisposes people to premature cardiovascular disease. However, the mechanism of lipid disorder remains unclear. Sterol regulatory element-binding protein 2 (SREBP2) is the key transcription factor regulating cholesterol metabolism. We hypothesize that estrogen regulates SREBP2 transcription through an estrogen response element (ERE) in the SREBP2 promoter region.MethodsHuman hepatoblastoma cells (HepG2) were treated with dose-dependent concentrations of estradiol (E2) for 24 h. Then, SREBP2 expression was determined via real-time PCR and immunofluorescence. The expressions of the SREBP2 downstream target genes HMGCR and LDLR were determined via real-time PCR. Lipid secretion in the culture media of HepG2 cells was measured using ELISA. Through bioinformatics analysis, we identified high-scoring ERE-like sequences in the SREBP2 gene promoter. Chromatin immunoprecipitation analysis was used to confirm the ERE. DNA fragments of the putative or mutated ERE-like sequence were synthesized and ligated into pGL3-basic plasmid to construct the SREBP2 promoter luciferase reporter systems. SREBP2-Luciferase (SREBP2-Luc), SREBP2-Mutation (SREBP2-Mut) and the blank control were transfected into hepatic cell lines. Luciferase activities were measured using the dual-luciferase reporter assay system. Chromatin immunoprecipitation analysis and the luciferase reporter assay were repeated in human hepatoma cells (HuH-7).ResultsWe found that E2 dose-dependently increased the expression of SREBP2 in HepG2 cells and that the increased levels were blocked when treated with an estrogen receptor-alpha antagonist. Additionally, E2 increased both HMGCR and LDLR expression and lipid secretion in HepG2 cells. Notably, we identified a functional ERE in the SREBP2 gene promoter, to which E2 could specifically bind and induce transcription.ConclusionsAn ERE was identified in the SREBP2 gene promoter. It mediates the regulation of SREBP2 expression by estrogen in hepatocytes. This study provides a mechanism to link cardiovascular disease with estrogen.

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Section: Methodsmentioning

confidence: 99%

Estrogen stimulates SREBP2 expression in hepatic cell lines via an estrogen response element in the SREBP2 promoter

Meng

Zong

2019

Cell Mol Biol Lett

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“…As a mobile phase, an inert gas (usually helium) is used in gas chromatography, while the mixture of polar solvents (most commonly methanol, water, and acetonitrile) which can be enriched with the addition of formic acid, ammonium formate, ammonium fluoride, or some other additive enhancing the ionization of analytes, is used in liquid chromatography [ 69 ]. The correct choice of a mobile phase gradient is often crucial in the separation of the analytes.…”

Section: Determination Of Steroid Hormonesmentioning

confidence: 99%

Determination of Intraprostatic and Intratesticular Androgens

Šimková

Heráček

Drašar

et al. 2021

IJMS

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Androgens represent the main hormones responsible for maintaining hormonal balance and function in the prostate and testis. As they are involved in prostate and testicular carcinogenesis, more detailed information of their active concentration at the site of action is required. Since the introduction of the term intracrinology as the local formation of active steroid hormones from inactive precursors of the adrenal gland, mainly dehydroepiandrosterone (DHEA) and DHEA-S, it is evident that blood circulating levels of sex steroid hormones need not reflect their actual concentrations in the tissue. Here, we review and critically evaluate available methods for the analysis of human intraprostatic and intratesticular steroid concentrations. Since analytical approaches have much in common in both tissues, we discuss them together. Preanalytical steps, including various techniques for separation of the analytes, are compared, followed by the end-point measurement. Advantages and disadvantages of chromatography-mass spectrometry (LC-MS, GC-MS), immunoanalytical methods (IA), and hybrid (LC-IA) are discussed. Finally, the clinical information value of the determined steroid hormones is evaluated concerning differentiating between patients with cancer or benign hyperplasia and between patients with different degrees of infertility. Adrenal-derived 11-oxygenated androgens are mentioned as perspective prognostic markers for these purposes.

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“…Estradiol concentrations are best measured via liquid chromatography-mass spectrometry (LC-MS) given that immunoassays lack selectivity and precision, particularly at low estradiol concentrations. 13 The clinician must also consider the estradiol concentration in relation to the timing of the estradiol dose, given that peak estradiol concentration is generally 4-5 hours post-dose with an elimination half-life of 14-22 h. 14 In one study in which transfeminine individuals were treated with 2 mg oral estradiol valerate, there was a peak median estradiol concentration of 189 (99) pmol/l with 24-h post-dose concentration of 56 (154) pmol/l. 15 No large study has evaluated the pharmacokinetic parameters at doses used in feminising hormone therapy regimens.…”

Section: Estradiol Concentrations and Consensus Guidelinesmentioning

confidence: 99%

Relationships between body mass index with oral estradiol dose and serum estradiol concentration in transgender adults undergoing feminising hormone therapy

Nolan

Brownhill²,

Bretherton

et al. 2020

Therapeutic Advances in Endocrinology

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Aim: Feminising hormone therapy with estradiol is used to align an individual’s physical characteristics with their gender identity. Given considerable variations in doses of estradiol therapy administered as gender-affirming hormone therapy (GAHT), we aimed to assess if body mass index (BMI) correlated with estradiol dose/concentration and assess the correlation between estradiol dose and estradiol concentrations. Methods: In a retrospective cross-sectional study, we analysed transgender individuals attending a primary or secondary care clinic in Melbourne, Australia who were prescribed oral estradiol valerate for at least 6 months and had estradiol dose and concentration available. Estradiol concentration was measured by immunoassay. Outcomes were the correlation between estradiol dose and BMI, and estradiol dose and estradiol concentration. Results: Data were available for 259 individuals {median age 25.8 [interquartile range (IQR) 21.9, 33.5] years}. Median duration of estradiol therapy was 24 (15, 33) months. Median estradiol concentration was 328 (238, 434) pmol/l [89 (65, 118) pg/ml] on 6 (4, 8) mg estradiol valerate. Median BMI was 24.7 (21.8, 28.6) kg/m2. There was a weak positive correlation between estradiol dose and estradiol concentration ( r = 0.156, p = 0.012). There was no correlation between BMI and estradiol concentration achieved ( r = −0.063, p = 0.413) or BMI and estradiol dose ( r = 0.048, p = 0.536). Estradiol concentrations were within the target range recommended in consensus guidelines in 172 (66%) individuals. Conclusion: Estradiol dose was only weakly correlated with estradiol concentration, suggesting significant interindividual variability. Prescription of estradiol dose should not be based upon an individual’s BMI, which did not correlate with estradiol concentration achieved. In all, 66% of individuals achieved estradiol concentrations recommended in Australian consensus guidelines with a relatively high oral estradiol dose.

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Current strategies for quantification of estrogens in clinical research

Cited by 59 publications

References 94 publications

Estrogen stimulates SREBP2 expression in hepatic cell lines via an estrogen response element in the SREBP2 promoter

Estrogen stimulates SREBP2 expression in hepatic cell lines via an estrogen response element in the SREBP2 promoter

Determination of Intraprostatic and Intratesticular Androgens

Relationships between body mass index with oral estradiol dose and serum estradiol concentration in transgender adults undergoing feminising hormone therapy

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