The Journal of steroid biochemistry and molecular biology.
Publisher:
Pergamon,
Frequency: Six no. a year, 2013-
Country: England
Language: English
Start Year:1990 -
ISSN:
0960-0760 (Print)
1879-1220 (Electronic)
0960-0760 (Linking)
1879-1220 (Electronic)
0960-0760 (Linking)
Impact Factor
4.1
2022
| NLM ID: | 9015483 |
| (DNLM): | SR0069716(s) |
| (OCoLC): | 22512922 |
| LCCN: | 90642733 |
| Classification: | W1 JO904S |
Characterization of equine GST A3-3 as a steroid isomerase. Glutathione transferases (GSTs) comprise a superfamily of enzymes prominently involved in detoxication by making toxic electrophiles more polar and therefore more easily excretable. However some GSTs have developed alternative functions. Thus, a member of the Alpha class GSTs in pig and human tissues is involved in steroid hormone biosynthesis, catalyzing the obligatory double-bond isomerization of Δ-androstene-3,17-dione to Δ-androstene-3,17-dione and of Δ-pregnene-3,20-dione to Δ-pregnene-3,20-dione on the biosynthetic pathways to testosterone and progesterone. The human GST A3-3 is the ...
In vitro simulation of the equine hindgut as a tool to study the influence of phytosterol consumption on the excretion of anabolic-androgenic steroids in horses. Traditionally, steroids other than testosterone are considered to be synthetic, anabolic steroids. Nevertheless, in stallions, it has been shown that β-Bol can originate from naturally present testosterone. Other precursors, including phytosterols from feed, have been put forward to explain the prevalence of low levels of steroids (including β-Bol and ADD) in urine of mares and geldings. However, the possible biotransformation and identification of the precursors has thus far not been investigated in horses. To study the possible endogenous digestive transformation, in vitro simulations of t...
Metabolic study of androsta-1,4,6-triene-3,17-dione in horses using liquid chromatography/high resolution mass spectrometry. Androsta-1,4,6-triene-3,17-dione (ATD) is an irreversible steroidal aromatase inhibitor and is marketed as a supplement. It has been reported to effectively reduce estrogen biosynthesis and significantly increase the levels of endogenous steroids such as dihydrotestosterone and testosterone in human. ATD abuses have been reported in human sports. Its metabolism in human has been studied, and the in vitro metabolic study of ATD in horses has been reported, however, little is known about its biotransformation and elimination in horses. This paper describes the in vitro and in vivo metabolism stu...
Dexamethasone acutely down-regulates genes involved in steroidogenesis in stallion testes. In rodents, livestock and primate species, a single dose of the synthetic glucocorticoid dexamethasone acutely lowers testosterone biosynthesis. To determine the mechanism of decreased testosterone biosynthesis, stallions were treated with 0.1mg/kg dexamethasone 12h prior to castration. Dexamethasone decreased serum concentrations of testosterone by 60% compared to saline-treated control stallions. Transcriptome analyses (microarrays, northern blots and quantitative PCR) of testes discovered that dexamethasone treatment decreased concentrations of glucocorticoid receptor alpha (NR3C1), alpha a...
Ex vivo spontaneous generation of 19-norandrostenedione and nandrolone detected in equine plasma and urine. 19-Norandrostenedione (NAED) and nandrolone are anabolic-androgenic steroids (AASs). Nandrolone was regarded solely as a synthetic AAS until the 1980s when trace concentrations of apparently endogenous nandrolone were detected in urine samples obtained from intact male horses (stallions). Since then, its endogenous origin has been reported in boars and bulls; endogenous NAED and nandrolone have been identified in plasma and urine samples collected from stallions. More recently, however, it was suggested that NAED and nandrolone detected in urine samples from stallions are primarily artifacts d...
The presence of 19-norandrostenedione and its sulphate form in yolk-sac fluid of the early equine conceptus. C(18) neutral steroid formation by cytochrome P450 aromatase has been recorded for several equine and porcine tissues. High activity of P450 aromatase is reflected in the quantities of estrogens in yolk-sac (y-s) fluid of early equine conceptuses. In a previous study of y-s fluid we detected large amounts of androgens by radioimmunoassay (RIA), using an antiserum for androstenedione (A(4)). Here, we report that RIA, following chromatography, gave tentative identification of the major peak as norandrostenedione (19-norA) not as A(4). Furthermore, even greater quantities of 19-norA seemed to be ...
Urinary excretion of 5(10)-estrene-3beta,17alpha-diol and estrone by the female horse: complementary indicators of early pregnancy screened with regard to a putative anabolic doping practice. Rules of horse racing stipulate that pregnant mares may compete under definite conditions of date, because early pregnant status may be misused for the sake of enhancing physical performance by putative anabolic steroid action. Screening for pregnancy is generally performed by plasma equine gonadotrophin (eCG) immunoassay, which covers the period between Days 40 and 120. In common screening for urinary anabolic steroids performed by gas chromatography-mass spectrometry, inclusion of two complementary criteria, i.e. the evaluation of total conjugates of 5(10)-estrene-3beta,17alpha-diol (EED) an...
Analysis of anabolic steroids in the horse: development of a generic ELISA for the screening of 17alpha-alkyl anabolic steroid metabolites. Due to the potential for misuse of a wide range of anabolic steroids in horse racing, a screening test to detect multiple compounds, via a common class of metabolites, would be a valuable forensic tool. An enzyme-linked immunosorbent assay (ELISA) has been developed to detect 17alpha-alkyl anabolic steroid metabolites in equine urine. 16beta-Hydroxymestanolone (16beta,17beta-dihydroxy-17alpha-methyl-5alpha-androstan-3-one) was synthesised in six steps from commercially available epiandrosterone (3beta-hydroxy-5alpha-androstan-17-one). Polyclonal antibodies were raised in sheep, employing mesta...
Effects of pharmacological doses of Vitamin D3 on mineral balance and profiles of plasma Vitamin D3 metabolites in horses. Metabolism and functions of Vitamin D in horses differ from those in humans, pigs and rats. In horses, calcidiol and calcitriol concentrations in blood plasma are remarkably low (<10 nmol L(-1); 20-40 pmol L(-1), respectively). When a toxic amount of Vitamin D(3) is administered, the responsiveness of calcium and calcitriol concentrations in blood plasma is much reduced compared to the other domestic animal species but inorganic phosphate (Pi) response is much more marked, leading to an increase of the Ca x Pi product. Also, soft tissue calcifications have been observed to develop in horses...
Amplified androstenedione enzymeimmunoassay for the diagnosis of cryptorchidism in the male horse: comparison with testosterone and estrone sulphate methods. An amplified enzymeimmunoassay (EIA) was validated for androstenedione in the serum of male horses. We will use the assay as a tool for the diagnosis of equine cryptorchidism. We will compare androstenedione EIA to the currently used methods (testosterone and estrone sulphate determinations). The study was conducted on 115 horses of pure Spanish and Arabian breeds, that included 30 geldings, 60 bilateral cryptorchids and 25 stallions. Androstenedione standard curve covered a range between 0 and 1 ng per well. Low detection limit was 1.54 pg/ml. Intra- and inter-assay coefficients of variation ...
Investigations into the biosynthetic pathways for classical and ring B-unsaturated oestrogens in equine placental preparations and allantochorionic tissues. In on-going studies of 'classical' and ring B-unsaturated oestrogens in equine pregnancy, the products of metabolism of [2,2,4,6,6-2H(5)]-testosterone and [16,16,17-2H(3)]-5,7-androstadiene-3 beta,17 beta-diol with equine placental subcellular preparations and allantochorionic villi have been identified. Using mixtures of unlabelled and [2H]-labelled steroid substrates has allowed the unequivocal identification of metabolites by twin-ion monitoring in gas chromatography-mass spectrometry (GC-MS). Two types of incubation were used: (i) static in vitro and (ii) dynamic in vitro. The latter invol...
Isolation and characterisation of a C(18) neutral steroid, oestra-5(10),7-diene-3,17-diol, from pregnant mare urine and allantoic fluid. Facile oxidation to yield oestra-5(10),6,8-triene-3, 17-diol (diol of Heard’s ketone). Oestradiene-3,17-diol and oestratriene-3,17-diol (or the diol of Heard's ketone (3-hydroxy-5(10),6,8-oestratriene-17-one) have been extracted on a large scale from pooled urines and allantoic fluid obtained from pregnant mares. Initial purification was achieved using column chromatography, and further purification by high performance liquid chromatography or silver nitrate (argentation) thin layer chromatography. The steroids were characterised using gas chromatography-mass spectrometry. Positions of the double bonds in ring B of oestradienediol were deduced on the basis of results of ultravio...
MR 20492 and MR 20494: two indolizinone derivatives that strongly inhibit human aromatase. In this study, we describe the synthesis of a new family of indolizinone derivatives designed to fit an extrahydrophobic pocket within the active site of aromatase and to strongly inhibit human aromatase. This could help improve the specificity of the inhibitors. Equine aromatase, very well characterized biochemically, is used as a comparative model. Indeed, in a previous comparison between both human and equine aromatases, we described the importance of the interaction between the inhibitor and this pocket for the indane derivative MR 20814. MR 20492 and MR 20494 are more potent inhibitors of...
Cannulation in situ of equine umbilicus. Identification by gas chromatography-mass spectrometry (GC-MS) of differences in steroid content between arterial and venous supplies to and from the placental surface. Equine umbilicus was cannulated in utero and a series of cord plasma samples removed for analysis. After steroid extraction and derivatisation, gas chromatographic-mass spectrometric (GC-MS) analysis demonstrated large differences in steroid content between the plasma samples obtained from the umbilical artery and vein, the blood supplies leading to and from the placental surface, respectively. 3Beta-hydroxy-5,7-androstadien-17-one, dehydroepiandrosterone, pregnenolone, 3beta-hydroxy-5alpha-pregnan-20-one, 5-pregnene-3beta,20beta-diol and 5beta-pregnane-3beta,20beta-diol were identified as maj...
Studies into aromatase activity associated with fetal allantochorionic and maternal endometrial tissues of equine placenta. Identification of metabolites by gas chromatography mass spectrometry. Maternal endometrial and fetal allantochorionic tissues were separated manually from the placentae of seven healthy thoroughbred and three pony mares, ranging in gestational age from 100 to 318 days. The homogeneity of subcellular fractions prepared from these tissues was assessed initially using the marker enzymes, succinate dehydrogenase, NADPH cytochrome C reductase and lactate dehydrogenase for the mitochondrial, microsomal and cytosolic fractions, respectively. Light microscopy and histochemical analysis demonstrated that the separated fetal allantochorionic membrane, which is made up of ...
Equine cytochrome P450 aromatase exhibits an estrogen 2-hydroxylase activity in vitro. Aromatase (estrogen synthetase) is a steroidogenic enzyme complex which catalyzes the conversion of androgens to estrogens (termed aromatization). This enzyme was purified from adult equine testis to homogeneity by five chromatographic steps. The ability of purified and reconstituted equine aromatase to exhibit an estrogen 2-hydroxylase activity was tested and compared to testosterone aromatization. Enzymatic activities were assessed by tritiated water release from labelled estradiol and testosterone. Kinetic analysis of estradiol 2-hydroxylation showed an apparent K(m) of 23 microM and a V(ma...
Binding of estrogen-3-sulfates to stallion plasma and equine serum albumin. The binding of estrone-3-sulfate (E1-3-S) and estradiol-3-sulfate (E2-3-S) to adult stallion plasma was determined and compared with the binding to equine serum albumin (ESA). On the ESA molecule, two binding sites for E1-3-S with an association constant of 1.3 x 10(5) M-1 and several sites of weaker affinity were found; the data for E2-3-S showed the existence of four binding sites of moderate affinity (1 x 10(5) M-1) and several sites of weaker affinity. The removal of albumin from the stallion plasma resulted in the absence of binding of E1-3-S or E2-3-S, whereas the removal of glycoprotein...
Steroid 21-hydroxylase activity in equine ovarian follicles evidenced by isotope dilution-mass spectrometry. Steroid 21-hydroxylase activity of the microsome-enriched fraction of follicular linings from equine ovaries has been demonstrated by gas chromatography-mass spectrometry. The 21-hydroxylated metabolites were quantified by isotope dilution with deuterated analogues. The two most abundant potential substrates for follicular steroid 21-hydroxylase, progesterone (P) and 17-hydroxyprogesterone (17OHP), were converted respectively to 11-deoxycorticosterone (DOC) and 11-deoxycortisol with corresponding apparent specific activities of 308 and 24 pmol/mg protein/h and apparent Km values of 1.1 and 6.4...