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PubMed1992

Immunohistochemical localization of 3 beta-hydroxy-5-ene-steroid dehydrogenase/delta 5----delta 4 isomerase in human placenta and fetal membranes throughout gestation.

The regulation of steroid production by the placenta and fetal membranes is important for both the maintenance of pregnancy and the timing of parturition. 3 beta-Hydroxy-5-ene-steroid dehydrogenase/delta 5----delta 4-isomerase (3 beta HSD) catalyzes an obligatory step in the biosynthesis of steroid hormones. We have determined the localization of 3 beta HSD in the human placenta, fetal membranes, and umbilical cord throughout gestation by immunohistochemical analysis, using a polyclonal antibody raised in rabbits against a purified preparation of human placental 3 beta HSD. In placenta, immunoreactive (IR-) 3 beta HSD was localized in the syncytiotrophoblast and intermediate trophoblast cells at both villous and extravillous sites, but not in cytotrophoblast cells from 6 weeks gestation to term. At 6-7 weeks gestation, IR-3 beta HSD was distributed in the cytoplasm of syncytiotrophoblast in about half of placental villi. By 12-14 weeks, the syncytiotrophoblast of all placental villi stained positively for 3 beta HSD. In the fetal membranes, strong IR-3 beta HSD staining was found in the trophoblast and reticular layers of chorion and in invasive trophoblast cells in decidua, and weakly in decidual stromal cells and amniotic epithelium. No IR-3 beta HSD was found in amnion on the placental plate, but in the umbilical cord, IR-3 beta HSD was present in the amniotic epithelium and also in fibroblast cells in Warton's jelly. These observations demonstrate that the localization of 3 beta HSD immunoreactivity and, therefore, the presumed sites of delta 5- to delta 4-steroid interconversion throughout gestation are principally the syncytiotrophoblast and intermediate trophoblast cells in placenta and the trophoblast cells in chorion and decidua in fetal membranes.

Extraembryonic Membranes↗
PubMed1993

Localization of cytochrome P450 cholesterol side-chain cleavage, cytochrome P450 17 alpha-hydroxylase/17, 20-lyase, and 3 beta-hydroxysteroid dehydrogenase isomerase steroidogenic enzymes in human and rhesus monkey fetal adrenal glands: reappraisal of functional zonation.

We examined the in situ localization of key steroidogenic enzymes in adrenal gland sections from midgestation (17-24 weeks) human fetuses and late gestation (130-142 days; term = 165 days) rhesus monkey fetuses. The rhesus monkey fetal adrenals were used as a model for the late gestation human fetal adrenal. The enzymes examined were cytochrome P450 cholesterol side-chain cleavage (P450scc), cytochrome P450 17 alpha-hydroxylase/17,20-lyase (P450c17), and 3 beta-hydroxysteroid dehydrogenase/isomerase (3 beta HSD). In human fetal adrenals, P450scc and P450c17 proteins and mRNAs were detected only in fetal zone (innermost cortical zone) and transitional zone (between the fetal and definitive zone) cells, not in definitive zone cells. Expression of 3 beta HSD was not detected in any cortical zone cells in midgestation human fetal adrenals. In rhesus monkey fetal adrenals, a similar pattern of P450scc and P450c17 expression was observed in the fetal and transitional zones. In the definitive zone cells of rhesus monkey fetal adrenals, expression of both P450scc and 3 beta HSD was detected. In addition, low levels of 3 beta HSD expression could be detected in some transitional zone cells. P450c17 expression was lacking in definitive zone cells from rhesus monkey fetal adrenals. These data suggest that early in gestation, cortisol is not produced by the human fetal adrenal cortex in vivo (because it does not express 3 beta HSD), whereas androgen production occurs in the transitional and fetal zones (which express P450scc and P450c17). Later in gestation, the definitive zone may produce minearlocorticoids (because it expresses P450scc and 3 beta HSD, but lacks P450c17), and the transitional zone may produce glucocorticoids (it expresses P450scc, P450c17, and 3 beta HSD), whereas the fetal zone continues to produce androgens. Thus, late in gestation the functional zonation of the human fetal adrenal cortex may be similar to that of the adult, with the definitive zone being analogous to the nascent zona glomerulosa, the transitional zone analogous to the zona fasciculata, and the fetal zone analogous to the zona reticularis.

Adrenal Glands↗
PubMed1994

Plasma 3 beta-hydroxy-delta 5-steroids in patients with congenital adrenal hyperplasia due to 21-hydroxylase deficiency.

There is little information about the plasma concentrations of 3 beta-hydroxy-delta 5-steroids (delta 5-steroids) in untreated patients with congenital adrenal hyperplasia due to 21-hydroxylase deficiency. To further study the delta 5 pathway, we measured plasma levels of delta 5- and delta 4-steroids in 21 adult patients with different degrees of 21-hydroxylase deficiency (11 salt-wasters, 5 simple virilizers, and 5 patients with the nonclassical form of the disease). In all patients, investigations were performed after withdrawal of steroid treatment for at least 10 days. In addition, catheterization of gonadal and adrenal veins was performed in two salt-wasting male patients displaying bilateral testicular tumors to study adrenal secretion of delta 5- and delta 4-steroids. In one of them, surgical resection of the intratesticular adrenal rests gave the opportunity to measure 3 beta-hydroxysteroid dehydrogenase (3 beta HSD) activity. In all untreated patients, an increase in plasma delta 4-steroids was observed. In contrast, although plasma levels of dehydroepiandrosterone (DHEA) and dehydroepiandrosterone sulfate (DHEAS) were not significantly modified in simple virilizers, a paradoxical decrease in all delta 5-steroids was observed in salt-wasters. Catheterization of the adrenal veins confirmed the decrease in delta 5-steroids, particularly DHEA and DHEAS. The androstenedione/DHEA ratio was increased in all patients proportionally to the severity of the disease, suggesting an increase in adrenal 3 beta HSD. In vitro analysis of 3 beta HSD activity showed a 4-fold increase in intratesticular adrenal tissue compared to that in normal adrenals. A positive correlation between the androstenedione/DHEA ratio and plasma ACTH levels was observed, suggesting a long term stimulatory effect of ACTH on 3 beta HSD. Angiotensin-II could have an additive effect on ACTH-induced 3 beta HSD activity.

Adrenal Hyperplasia, Congenital↗
PubMed1994

No evidence of mutations in the genes for type I and type II 3 beta-hydroxysteroid dehydrogenase (3 beta HSD) in nonclassical 3 beta HSD deficiency.

Nonclassical 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4-isomerase deficiency (NC3 beta HSDD) has been diagnosed in hyperandrogenic women with an increasing frequency during the last 14 yr. Fifteen menarcheal women with androgen excess syndrome, diagnosed with NC3 beta HSDD previously were restudied, in 12 after discontinuation of glucocorticoid treatment, in 2 patients never treated with glucocorticoids, and in 1 both before and after glucocorticoid therapy. Each of the 15 patients underwent ACTH stimulation testing, in some cases on multiple occasions. Although some (very few) patients seem to have improved with time, others remained the same or got worse. Molecular DNA analysis was also performed in 6 of the patients, using the strategy successfully used to detect point mutations in the type II 3 beta-hydroxysteroid dehydrogenase (3 beta HSD) gene, which are responsible for classical 3 beta HSD deficiency. This strategy consists of the direct sequencing of polymerase chain reaction-amplified DNA fragments corresponding to the complete coding sequence and all intron-exon junctions and to the 5'- and 3'-noncoding region of this gene. We were unable to demonstrate any mutation of the type II 3 beta HSD gene in these 6 patients. To gain additional information about potential mutations, direct sequencing of the type I 3 beta HSD gene was also performed using this same strategy, and no mutations were found. The present study strongly suggests that unlike the salt-losing and nonsalt-losing forms of classical 3 beta HSD deficiency, NC3 beta HSDD is not due to a mutant type II 3 beta HSD enzyme. However, the possibility remains of a mutation(s) in the unsequenced regions of the type II 3 beta HSD gene or elsewhere, such as in a gene for modulatory protein, playing a specific role in the expression of the type II 3 beta HSD gene. On the other hand, knowing the multiple hormonal controls to which 3 beta HSD activity is subject, it cannot be excluded that at least in some cases, NC3 beta HSDD may be an acquired defect, the result of endogenous or environmental factors.

3-Hydroxysteroid Dehydrogenases↗
PubMed1999

Medroxyprogesterone acetate and dexamethasone are competitive inhibitors of different human steroidogenic enzymes.

Medroxyprogesterone acetate (MPA), a widely used progestin, can suppress the hypothalamic-pituitary-gonadal axis but can also directly inhibit gonadal steroidogenesis; the success of MPA as a treatment for gonadotropin-independent sexual precocity derives from its direct action on steroidogenic tissues. Dexamethasone, a widely used glucocorticoid, can suppress the hypothalamic-pituitary-adrenal axis, but its potential effect directly on the adrenal is unclear. Previous reports suggested that these two drugs may act on the initial steps in the rodent steroidogenic pathway; therefore, we investigated their abilities to inhibit the first three human enzymes in steroidogenesis: the cholesterol side-chain cleavage enzyme (P450scc), the 17alpha-hydroxylase/17,20-lyase (P450c17), and type II 3beta-hydroxysteroid dehydrogenase/isomerase (3betaHSDII). We found no effect of either drug on P450scc in intact human choriocarcinoma JEG-3 cells. Using microsomes from yeast expressing human P450c17 or microsomes from human adrenals, we found that dexamethasone inhibited P450c17 with a Ki of 87 micromol/L, which is about 1000 times higher than typical therapeutic concentrations, but that MPA has no detectable action on P450c17. Using microsomes from yeast expressing human 3betaHSDII, we found that this enzyme has indistinguishable apparent Km values of 5.2-5.5 micromol/L and similar maximum velocities of 0.34-0.56 pmol steroid/min x microg microsomal protein for the three principal endogenous substrates, pregnenolone, 17-hydroxypregnenolone, and dehydroepiandrosterone. In this system, MPA inhibited 3betaHSDII with a Ki of 3.0 micromol/L, which is near concentrations achieved by high therapeutic doses of 5-20 mg MPA/kg x day. These data establish the mechanism of action of MPA as an inhibitor of human steroidogenesis, and are in contrast with the results of earlier studies indicating that MPA inhibited both P450c17 and 3betaHSD in rat Leydig cells. These studies establish the "humanized yeast" system as a model for studying the actions of drugs on human steroidogenic enzymes and suggest that 3betaHSDII may be an appropriate target for pharmacological interventions in human disorders characterized by androgen excess or sex steroid dependency.

Animals↗
PubMed2005

GATA factors and the nuclear receptors, steroidogenic factor 1/liver receptor homolog 1, are key mutual partners in the regulation of the human 3beta-hydroxysteroid dehydrogenase type 2 promoter.

The human HSD3B2 gene encodes the 3beta-hydroxysteroid dehydrogenase/delta5-delta4 isomerase type 2 (3beta-HSD2) enzyme that is required for steroid hormone biosynthesis. Mutations in the hHSD3B2 gene are responsible for a form of congenital adrenal hyperplasia and male pseudohermaphroditism whereas overexpression of hHSD3B2 has been recently associated with polycystic ovarian syndrome. Despite the importance of the hHSD3B2 gene, the molecular mechanisms that regulate its expression remain poorly understood. Transcription factors belonging to the GATA family are emerging as novel regulators of steroidogenesis. Indeed, GATA-4 and GATA-6 are abundantly expressed in steroidogenic cells of the gonads and adrenals. We now report that the human HSD3B2 promoter (hHSD3B2), which contains four consensus GATA elements, constitutes an important target for GATA factors. GATA-4 and GATA-6 by themselves are sufficient to activate transcription (up to 15-fold) from a -1073 bp hHSD3B2 promoter fragment and blockade of endogenous GATA expression and/or activity blunts hHSD3B2 promoter activity in steroidogenic cells. Deletion studies showed that the proximal GATA element located at -196 bp is sufficient to confer GATA responsiveness of the hHSD3B2 promoter and is required for full hHSD3B2 promoter activity in steroidogenic cells. Moreover, we report that GATA-4 and GATA-6 can physically interact with the nuclear receptors, steroidogenic factor 1 and liver receptor homolog 1, to synergistically activate hHSD3B2 promoter activity in both homologous and heterologous cells. Aberrant expression of transcription factors essential for hHSD3B2 expression might also be involved in some pathologies/syndromes associated with deregulated hHSD3B2 expression.

Animals↗
PubMed1990

Structural analysis of the gene encoding human 3 beta-hydroxysteroid dehydrogenase/delta 5----4-isomerase.

The structural gene encoding human 3 beta-hydroxysteroid dehydrogenase/delta 5----4-isomerase (3 beta HSD) was isolated from a human EMBL3 genomic library. The gene encompasses approximately 8 kilobases of DNA and is comprised of two large introns and three exons encoding amino acid residues 1-48, 49-103, and 104-373, respectively. The exonic sequence is identical to that of the cDNA that we previously isolated and expressed in COS 1 cells. DNA sequence analysis reveals a putative TATA (TATATAA) motif 26 basepairs up-stream of the beginning of exon I, as determined by S1 nuclease protection analysis. However, primer extension analysis using poly(A)+ RNA isolated from both placenta and corpora lutea indicates that the RNA initiates up-stream of the putative TATA motif, and that an additional 53-basepair exon, which is untranslated, is present 5' to the first coding exon. Southern hybridization analysis of genomic DNA using a single exon probe suggests that there may be more than one copy of the gene in the human genome. In addition, we confirm from Southern analysis of genomic DNA isolated from human x hamster somatic cell hybrids that the gene is located on human chromosome 1. These findings will provide a foundation for the characterization of apparent 3 beta HSD clinical deficiencies when these are due to a mutation in the structural gene.

Amino Acid Sequence↗
PubMed1991

Structure and expression of a new complementary DNA encoding the almost exclusive 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4-isomerase in human adrenals and gonads.

The 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4 isomerase (3 beta HSD) enzyme catalyzes the oxidation and isomerization of delta 5-3 beta-hydroxysteroid precursors into delta 4-ketosteroids, thus leading to the formation of all classes of steroid hormones. In addition, 3 beta HSD catalyzes the interconversion of 3 beta-hydroxy- and 3-keto-5 alpha-androstane steroids. Clinical observations in patients with 3 beta HSD deficiency as well as our recent data obtained by Southern blot analysis using a human placental 3 beta HSD cDNA (type I) as probe suggested the existence of multiple related 3 beta HSD isoenzymes. We now report the isolation and characterization of a second type of cDNA clone (arbitrarily designated type II) encoding 3 beta HSD after screening of a human adrenal lambda gt22A library. The nucleotide sequence of 1676 basepairs of human 3 beta HSD type II cDNA predicts a protein of 371 amino acids with a calculated molecular mass of 41,921 daltons, which displays 93.5% and 96.2% homology with human placental type I and rhesus macaque ovary 3 beta HSD deduced proteins, respectively. To characterize and compare the kinetic properties of the two isoenzymes, plasmids derived from pCMV and containing type I or type II 3 beta HSD full-length cDNA inserts were transiently expressed in HeLa human cervical carcinoma cells. In vitro incubation with NAD+ and 3H-labeled pregnenolone or dehydroepiandrosterone shows that the type I protein possesses a 3 beta HSD/delta 5-delta 4 isomerase activity higher than type II, with respective Km values of 0.24 vs. 1.2 microM for pregnenolone and 0.18 vs. 1.6 microM for dihydroepiandrosterone, while the specific activity of both types is equivalent. Moreover, incubation in the presence of NADH of homogenates from cells transfected with type I or type II 3 beta HSD indicates that dihydrotestosterone is converted into 5 alpha-androstane-3 beta, 17 beta-diol, with Km values of 0.26 and 2.7 microM, respectively. Ribonuclease protection assay using type I- and type II-specific cRNA probes revealed that type II transcripts are the almost exclusive 3 beta HSD mRNA species in the human adrenal gland, ovary, and testis, while type I transcripts correspond to the almost exclusive 3 beta HSD mRNA species in the placenta and skin and represent the predominantly expressed species in mammary gland tissue. The present data show for the first time that adrenals and gonads express a type of 3 beta HSD isoenzyme that is distinct from the type expressed in the placenta.(ABSTRACT TRUNCATED AT 400 WORDS)

Adrenal Glands↗
PubMed1999

Stat5-mediated regulation of the human type II 3beta-hydroxysteroid dehydrogenase/delta5-delta4 isomerase gene: activation by prolactin.

Altered PRL levels are associated with infertility in women. Molecular targets at which PRL elicits these effects have yet to be determined. These studies demonstrate transcriptional regulation by PRL of the gene encoding the final enzymatic step in progesterone biosynthesis: 3beta-hydroxysteroid dehydrogenase/delta5-delta4 isomerase (3beta-HSD). A 9/9 match with the consensus Stat5 response element was identified at -110 to -118 in the human Type II 3beta-HSD promoter. 3beta-HSD chloramphenicol acetyltransferase (CAT) reporter constructs containing either an intact or mutated Stat5 element were tested for PRL activation. Expression vectors for Stat5 and the PRL receptor were cotransfected with a -300 --> +45 3beta-HSD CAT reporter construct into HeLa cells, which resulted in a 21-fold increase in reporter activity in the presence of PRL. Promoter activity showed an increased response with a stepwise elevation of transfected Stat5 expression or by treatment with increasing concentrations of PRL (max, 250 ng/ml). This effect was dramatically reduced when the putative Stat5 response element was removed by 5'-deletion of the promoter or by the introduction of a 3-bp mutation into critical nucleotides in the element. Furthermore, 32P-labeled promoter fragments containing the Stat5 element were shifted in electrophoretic mobility shift assay experiments using nuclear extracts from cells treated with PRL, and this complex was supershifted with antibodies to Stat5. These results demonstrate that PRL has the ability to regulate expression of a key human enzyme gene (type II 3beta-HSD) in the progesterone biosynthetic pathway, which is essential for maintaining pregnancy.

DNA-Binding Proteins↗
PubMed2000

Multiple signaling pathways mediate interleukin-4-induced 3beta-hydroxysteroid dehydrogenase/delta5-delta4 isomerase type 1 gene expression in human breast cancer cells.

The 3beta-hydroxysteroid dehydrogenase/delta5-delta4 isomerase (3beta-HSD) isoenzymes catalyze an essential step in the formation of all classes of active steroid hormones. We have recently shown that 3beta-HSD type 1 gene expression is specifically induced by interleukin (IL)-4 and IL-13 in breast human cancer cell lines and in normal human mammary epithelial cells in primary culture. There is evidence that IL-4 stimulates bifurcating signaling pathways in which the signal transducer and activator of transcription-6 (Stat6)-signal pathway is involved in differentiation and gene regulation, whereas insulin receptor substrate (IRS) proteins mediate the mitogenic action of IL-4. In fact, we have shown that Stat6 was activated by IL-4 in all cell lines studied where IL-4 induced 3beta-HSD expression, but not in those that failed to respond to IL-4. The present study was designed to investigate the potential contribution of IRS proteins and their downstream targets to IL-4-induced 3beta-HSD type 1 gene expression. IL-4 rapidly induced IRS-1 and IRS-2 phosphorylation in ZR-75-1 human breast cancer cell lines. Moreover, insulin-like growth factor (IGF)-I and insulin, which are well known to cause IRS-1 and IRS-2 phosphorylation, increased the stimulatory effect of IL-4 on 3beta-HSD activity. IRS-1 and IRS-2 are adapter molecules that provide docking sites for different SH2-domain-containing proteins such as the phosphatidylinositol (PI) 3-kinase. In this light, the inhibition of IL-4-induced 3beta-HSD expression by wortmannin and LY294002, two potent PI 3-kinase inhibitors, indicates the probable involvement of the PI 3-kinase signaling molecules in this response to IL-4. Furthermore, it has been suggested that the IRS proteins are part of the signaling complexes that lead to activation of the mitogen-activated protein (MAP) kinase by insulin; thus we investigated the potential role of the MAP kinase (MAPK) cascade in the IL-4 action. In ZR-75-1 cells, both the activation of MAPK by IL-4 and the IL-4-induced 3beta-HSD activity were completely blocked by PD98059, an inhibitor of MAPK activation. Wortmannin also blocked MAPK activation by IL-4, IGF-I, and insulin, suggesting that the MAPK cascade acts as a downstream effector of PI 3-kinases. To further understand the cross-talk between signaling pathways involved in IL-4 action, we investigated the possible involvement of protein kinase C (PKC). The potential role of PKC was suggested by the observation that the well known PKC activator phorbol-12-myristate-13-acetate (PMA) potentiated the IL-4-induced 3beta-HSD activity. Taken together, these findings suggest the existence of a novel mechanism of gene regulation by IL-4. This mechanism would involved the phosphorylation of IRS-1 and IRS-2, which transduce the IL-4 signal through a PI 3-kinase- and MAPK-dependent signaling pathway. The inability of IGF-I, insulin, and PMA to stimulate 3beta-HSD expression by themselves in the absence of IL-4 makes obvious the absolute requirement of an IL-4-specific signaling molecule. Our findings thus suggest that the multiple pathways downstream of IRS-1 and IRS-2 must act in cooperation with the IL-4-specific transcription factor Stat6 to mediate the induction of 31beta-HSD type 1 gene expression in ZR-75-1 human breast cancer cells.

Androstadienes↗
PubMed2002

Gonadal differentiation, sex determination and normal Sry expression in mice require direct interaction between transcription partners GATA4 and FOG2.

In mammals, Sry expression in the bipotential, undifferentiated gonad directs the support cell precursors to differentiate as Sertoli cells, thus initiating the testis differentiation pathway. In the absence of Sry, or if Sry is expressed at insufficient levels, the support cell precursors differentiate as granulosa cells, thus initiating the ovarian pathway. The molecular mechanisms upstream and downstream of Sry are not well understood. We demonstrate that the transcription factor GATA4 and its co-factor FOG2 are required for gonadal differentiation. Mouse fetuses homozygous for a null allele of Fog2 or homozygous for a targeted mutation in Gata4 (Gata4(ki)) that abrogates the interaction of GATA4 with FOG co-factors exhibit abnormalities in gonadogenesis. We found that Sry transcript levels were significantly reduced in XY Fog2(-/-) gonads at E11.5, which is the time when Sry expression normally reaches its peak. In addition, three genes crucial for normal Sertoli cell function (Sox9, Mis and Dhh) and three Leydig cell steroid biosynthetic enzymes (p450scc, 3betaHSD and p450c17) were not expressed in XY Fog2(-/-) and Gata(ki/ki) gonads, whereas Wnt4, a gene required for normal ovarian development, was expressed ectopically. By contrast, Wt1 and Sf1, which are expressed prior to Sry and necessary for gonad development in both sexes, were expressed normally in both types of mutant XY gonads. These results indicate that GATA4 and FOG2 and their physical interaction are required for normal gonadal development.

Animals↗
PubMed1998

Concentration of mRNA encoding 3 beta-hydroxysteroid dehydrogenase/delta 5,delta 4 isomerase (3 beta-HSD) and 3 beta-HSD enzyme activity following treatment of ewes with prostaglandin F2 alpha.

The objectives of these experiments were (1) to determine if prostaglandin F2 alpha (PGF2 alpha) decreased mRNA encoding 3 beta-hydroxysteroid dehydrogenase/d5,delta 4 isomerase (3 beta-HSD) specifically in large steroidogenic luteal cells, which contain the high affinity receptors for PGF2 alpha; and (2) to determine if the decreased concentration of mRNA encoding 3 beta-HSD following administration of PGF2 alpha was associated with a decrease in 3 beta-HSD enzyme activity. Ewes on days 11 or 12 of the estrous cycle were administered PGF2 alpha (25 mg i.v. followed by 10 mg i.m. 2 h later) and corpora lutea collected 4, 12, 24, or 48 h later (n = 4-5/time). Corpora lutea were also collected from non-injected (n = 4) or saline-injected (n = 4) control ewes. Administration of PGF2 decreased (P < 0.05) steady-state concentrations of mRNA encoding 3 beta-HSD to 35, 15, 9, and 5 percent of the concentrations in the control group at 4, 12, 24, and 48 h, respectively. Concentrations of mRNA encoding 3 beta-HSD in large luteal cells were decreased to 43% of controls 4 h following injection, which was similar to the decrease seen in steady-state concentrations of this mRNA in total luteal mRNA (35%). However, 3 beta-HSD enzyme activity was not significantly decreased by 48 h after PGF2 alpha injection. Thus, the dramatic decreased in mRNA encoding 3 beta-HSD was not associated with an immediate decrease in 3 beta-HSD enzyme activity and, therefore, does not appear to be responsible for the acute decrease in secretion of progesterone from ovine luteal tissue during PGF2 alpha-induced luteolysis.

Animals↗
PubMed1994

[Endocrine pathological analysis of primary pigmented nodular adrenocortical disease].

We have studied eleven cases of the adrenal with primary pigmented nodular adrenocortical disease (PPNAD) by immunohistochemistry of all steroidogenic enzymes involved in cortisol biosynthesis, by in situ hybridization of P450C17 in seven cases in order to localize the sites of steroidogenesis and by chromosome suppression in situ hybridization of genomic DNA in five cases in order to determine possible genetic abnormalities of the disorder. Immunoreactivity of all the enzymes examined was intense in almost all the cells in adrenocortical nodules while internodular adrenal cortex, including the cases without cortical atrophy, was negative for the enzymes with the exception of 3 beta HSD. In situ hybridization studies of P450C17 yielded results consistent with those of immunohistochemistry. These results may be consistent with autonomous cortisol production by the nodular cells and indicate that almost all of the cells in the cortical nodules produce cortisol, which can explain the presence of hypercortisolism despite small sizes of adrenals in PPNAD. Chromosome suppression in situ hybridization analysis demonstrated possible genetic defects in chromosome 16 in this disorder.

Adolescent↗
PubMed1994

[The effects of GnRH agonist on steroidogenesis in the rat ovary].

The effects of GnRH agonist (buserelin) on in vitro ovarian steroidogenesis were studied using DES-treated immature rats and PMS-treated immature rats. The estradiol and progesterone secreted by the cultured ovarian cells and the activities of various enzymes of steroid-metabolism were examined with or without gonadotropins (FSH or hCG), and the effects of 10(-6)-10(-12) M of buserelin on those indices were observed for 3-72 hours. In addition, the kinetic study of ovarian GnRH receptor was performed using 125I-labelled buserelin and crude ovarian cell membrane fraction of PMS-treated rats. The Scatchard analysis revealed the specific high affinity and low capacity ovarian GnRH receptor (Kd = 0.92 nM and Bmax = 0.57 fmol/mg tissue). The FSH-stimulated cholesterol side chain cleavage enzyme (CSCC) activity of the DES-treated rats was suppressed in a dose-dependent manner by buserelin. Estradiol release and aromatase activity were increased by 10(-8) M buserelin within 48 hours from the beginning of the incubation of the PMS-treated rat ovarian cells, but were suppressed after 48 hours. Buserelin increased basal progesterone secretion and both activities of CSCC and of 3 beta-hydroxysteroid dehydrogenase of PMS-treated rat ovarian cells incubated without hCG, which were suppressed by buserelin co-incubated with 100 IU/ml of hCG. These results suggested that GnRH plays a physiological role in ovarian steroidogenesis binding the specific receptor and that GnRH promotes the development of the follicle through increased estrogen synthesis in the early stage of the folliculogenesis and the luteinization in the late stage of the follicular development through increased progesterone and decreased estradiol production and the luteolysis in the luteinized cells by hCG through decreased progesterone secretion.

Animals↗
PubMed1991

Inhibitory effects of gossypol on adrenal function.

Gossypol, an antifertility agent, has inhibitory actions on many membrane-associated enzymes, suggesting that this agent might have a generalized effect on cell membranes. This hypothesis was examined in the present study using membranes and dispersed cells prepared from human and rat adrenal glands. Four parameters were determined: microviscosity as measured by fluorescence polarization of human adrenal microsomal- and mitochondrial-enriched membranes, adrenal steroidogenic enzymes; and cAMP and cortisol responses to ACTH. It was found that gossypol increased the polarization constants of microsomes and mitochondria in a dose-dependent manner. Of the three adrenal enzymes tested, both 3 beta-hydroxysteroid dehydrogenase delta 5-delta 4 isomerase and 11-hydroxylase were inhibited by gossypol, but not 21-hydroxylase. Using intact human adrenocortical cells, high doses of gossypol also inhibited the ACTH-stimulated cAMP and cortisol levels. The in vivo corticosterone response to ACTH in rats subjected to chronic gossypol treatment was also found to be reduced. These findings suggest that gossypol has multiple effects on adrenal function. Its effects on membrane microviscosity, adrenal steroidogenesis, cAMP and corticosterone responses to ACTH stimulation probably occur through a generalized membrane effect.

Adrenal Glands↗
PubMed1976

The influence of prostaglandin F-2alpha on pregnenolone metabolism by the autotransplanted ovary of the ewe.

Eight ewes each with an autotransplanted ovary received infusions of tritium-labelled pregnenolone (41 muCi/hr) for 8 hr into the artery supplying the ovary, together with prostaglandin (PG) F-2alpha (30 mug/hr) for 3 hr beginning 2 hr after the start of the pregnenolone infusion. All animals exhibited oestrus 2-3 days after the start of the experiment. During the PGF-2alpha infusion blood flow through the ovaries was increased by 13%, but subsequently returned to pre-infusion levels. Secretion rates of endogenous progesterone and 20alpha-hydroxypregn-4-en-3-one dropped rapidly 5 hr after the PGF-2alpha infusion had started from values of 250 mug/hr and 25 mug/hr to values below 60 mug/hr and 8 mug/hr, respectively. At this time the conversion of radioactive pregnenolone to progesterone was reduced by 50% of its initial value, but the secretion of endogenous pregnenolone and the formation of radioactive metabolites other than progesterone were not diminished. In 4 control animals, which received pregnenolone only, no changes in ovarian blood flow, steroid secretion rates, or in the conversion of labelled pregnenolone were observed. These results suggest a possible involvement of PGF-2alpha in the regulation of progesterone biosynthesis by an action on the 3beta-hydroxysteroid oxidoreductase-delta5(-4) isomerase enzyme system.

Animals↗