Showing papers on "Hydroxysteroid dehydrogenase published in 1998"
TL;DR: The results suggest that AKR1C3 acts as prostaglandin D2 11-ketoreductase, and that its principal gene in the human has a coding region represented by DBDH cDNA.
Abstract: Human 3alpha-hydroxysteroid dehydrogenase exists in four isoforms, which belong to the aldo-keto reductase (AKR) superfamily and are named AKR1C1-AKR1C4. The properties of AKR1C3 have not been fully characterized compared to the other isoforms. In addition, a cDNA that shows more than 99% homology with AKR1C3 cDNA has been cloned from human myeloblasts. We have here expressed and purified a recombinant enzyme (designated as DBDH) from this cDNA. DBDH oxidized xenobiotic alicyclic alcohols and 3alpha- or 17beta-hydroxy-5beta-androstanes, and catalyzed the reversible conversion between prostaglandin D2 and 9alpha,11beta-prostaglandin F2 more efficiently than that of 3alpha- or 17beta-hydroxysteroids:the respective Km values were 0.6 and 6.8 microM, and kcat/Km values were about 1,000 min-1.mM-1. Anti-inflammatory drugs highly inhibited the enzyme. The recombinant AKR1C3 prepared by site-directed mutagenesis of DBDH also showed the same properties as the wild-type DBDH. Analyses of expression of mRNAs for DBDH and AKR1C3 by reverse transcription-PCR indicated that only one mRNA species for DBDH is expressed in 33 human specimens of liver, kidney, lung, brain, heart, spleen, adrenal gland, small intestine, placenta, prostate, and testis. These results suggest that AKR1C3 acts as prostaglandin D2 11-ketoreductase, and that its principal gene in the human has a coding region represented by DBDH cDNA.
173 citations
TL;DR: The results indicate that 17HSD7 is an enzyme of E2 biosynthesis, which is predominantly expressed in the corpus luteum of the pregnant animal, and it is suggested that rPRAP is rat 17 HSD type 7.
Abstract: 17β-Hydroxysteroid dehydrogenases/17-ketosteroid reductases (17HSDs) modulate the biological activity of certain estrogens and androgens by catalyzing reductase or dehydrogenase reactions between 17-keto- and 17β-hydroxysteroids. In the present study, we demonstrate expression cloning of a novel type of 17HSD, chronologically named 17HSD type 7, from the HC11 cell line derived from mouse mammary gland. The cloned cDNA, 1.7 kb in size, encodes a protein of 334 amino acids with a calculated molecular mass of 37,317 Da. The primary structure contains segments characteristic of enzymes belonging to the short-chain dehydrogenase/reductase superfamily. Strikingly, mouse 17HSD type 7 (m17HSD7) shows 89% identity with a recently cloned rat protein called PRL receptor-associated protein (PRAP). The function of PRAP has not yet been demonstrated. The enzymatic characteristics of m17HSD7 and RT-PCR-cloned rat PRAP (rPRAP) were analyzed in cultured HEK-293 cells, where both of the enzymes efficiently catalyzed conver...
149 citations
TL;DR: It is found that 7-hydroxyflavone and apigenin are the most effective aromatase and 17beta-Hydroxysteroid dehydrogenase inhibitors, respectively.
135 citations
TL;DR: The demonstration of 11beta-HSD-1 expression in adipocytes and its predominant reductase activity in intact 3T3-F442A adipocytes suggests that 11 beta-HSd-1 may play an important role in potentiating glucocorticoid action in these cells.
123 citations
TL;DR: It is shown that Ke 6 is a 17β-hydroxysteroid dehydrogenase and can regulate the concentration of biologically active estrogens and androgens and is found that the Ke 6 gene is expressed within the ovaries and testes.
107 citations
TL;DR: It is hypothesized that the differential expression of the two 17HSD enzymes, with opposite activities in same cell types, could modulate intracellular E2 concentrations during the end of the luteal phase of the menstrual cycle.
Abstract: According to the current hypothesis, 17β-hydroxysteroid dehydrogenases (17HSDs) regulate the extent of estrogen influence in the endometrium by converting estradiol (E2) locally into a biologically less active sex steroid, estrone (E1), and vice versa. Recently, we have shown that both 17HSD type 1 and type 2 are expressed in the human endometrium, and in the present work, using in situ hybridization, we show that 17HSD type 2 is localized in the glandular epithelial cells as previously shown for the type 1 enzyme, but in contrast to type 1, the expression of type 2 is highest at the end of the cycle. Hence, we hypothesize that the differential expression of the two 17HSD enzymes, with opposite activities in same cell types, could modulate intracellular E2 concentrations during the end of the luteal phase of the menstrual cycle. We further analyzed the expression of 17HSD type 1 and type 2 mRNAs in term human placenta. Expression of 17HSD type 1 mRNA was detected in the syncytiotrophoblasts, and signals f...
84 citations
TL;DR: The current findings boost to 16 the number of mutations in the HSD17B3 gene that impair testosterone synthesis and cause male pseudohermaphroditism, and add 1 apparently silent polymorphism to this tally.
Abstract: Isozymes of 17β-hydroxysteroid dehydrogenase (17βHSD) regulate levels of bioactive androgens and estrogens in a variety of tissues. For example, the 17βHSD type 3 isozyme catalyzes the conversion of the inactive C19-steroid androstenedione to the biologically active androgen, testosterone, in the testis. Testosterone is essential for the correct development of male internal and external genitalia; hence, deleterious mutations in the HSD17B3 gene give rise to a rare form of male pseudohermaphroditism termed 17βHSD deficiency. Here, 2 additional missense mutations in the HSD17B3 gene in subjects with 17βHSD deficiency are described. One mutation (A56T) impairs enzyme function by affecting NADPH cofactor binding. A second mutation (N130S) led to complete loss of enzyme activity. Also, a single base pair polymorphism in exon 11 of the HSD17B3 gene is described. The polymorphic A allele encodes a protein with a serine rather than a glycine at position 289 (GGT → AGT). The frequency of the G allele (Gly) was 0....
81 citations
TL;DR: Rat OB cell lines were able to synthesize E1, E2, and testosterone from androstenedione, although activity varied between OB cell types, and data suggest that local synthesis of sex hormones is an important function of OB cells and may play a key role in the modulation of bone turnover independent of circulating hormone concentrations.
Abstract: Postmenopausal loss of 17 beta-estradiol (E2) in women is associated with decreased bone mineral density and increased susceptibility to osteoporotic bone fracture. These changes in bone status are assumed to be due to circulating levels of the hormone; therapeutic replacement of E2 can alleviate the bone disease. However, recent reports have shown that human osteoblastic (OB) cells are able to synthesize estrogens locally, via expression of the enzyme aromatase. In this study, we have characterized the expression and activity of aromatase and 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) in rat OB cell lines. Aromatase activity in ROS 17/2.8, ROS 25/1, and UMR 106 cells was similar to that shown in human OB cells, with the highest levels of activity observed in the more differentiated ROS 17/2.8 cells (Vmax = 45 pmol/h/mg of protein). The rat OB cells also showed 17 beta-HSD activity, with the predominant metabolism in all three cell lines being estrone (E1) to E2. As with aromatase, the highest activity was observed in ROS 17/2.8 cells (Vmax = 800 pmol/h/mg of protein). Northern analyses indicated the variable presence of transcripts corresponding to the type 1, 2, 3, and 4 isoforms of 17 beta-HSD. Further analysis of androstenedione metabolism indicated that the net effect of aromatase and 17 beta-HSD activity varied with cell type and culture treatment. All three OB cell lines were able to synthesize E1, E2, and testosterone from androstenedione, although activity varied between OB cell types. Regulatory effects were observed with 1,25-dihydroxyvitamin D3 (positive) and dexamethasone (negative). These data suggest that local synthesis of sex hormones is an important function of OB cells and may play a key role in the modulation of bone turnover independent of circulating hormone concentrations.
67 citations
TL;DR: The results indicate that in both male and female mice, 17HSD type 2 is expressed mainly in the various epithelial cell types of the gastrointestinal and urinary tracts, and therefore suggest a role for the enzyme in steroid inactivation in a range of tissues and cell types not considered as classical sex steroid target tissues.
Abstract: Hydroxysteroid dehydrogenase (17HSD) type 2e Yciently catalyzes the conversion of the high activity 17‚-hydroxy forms of sex steroids into less potent 17-ketosteroids. In the present study in situ hybridization was utilized to analyze the cellular localization of 17HSD type 2 expression in adult male and female mice. The data indicate that 17HSD type 2 mRNA is expressed in several epithelial cell layers, including both absorptive and secretory epithelia as well as protective epithelium. In both males and females, strong expression of 17HSD type 2 was particularly detected in epithelial cells of the gastrointestinal and urinary tracts. The mRNA was expressed in the stratified squamous epithelium of the esophagus, and surface epithelial cells of the stomach, small intestine and colon. The hepatocytes of the liver and the thick limbs of the loops of Henle in the kidneys, as well as the epithelium of the urinary bladder, also showed strong expression of 17HSD type 2 mRNA in both male and female mice. In the genital tracts, low 17HSD type 2 expression was detected in the seminiferous tubules, the uterine epithelial cells and the surface epithelium of the ovary. Expression of the mRNA was also detected in the sebaceous glands of the skin. The results indicate that in both male and female mice, 17HSD type 2 is expressed mainly in the various epithelial cell types of the gastrointestinal and urinary tracts, and therefore suggest a role for the enzyme in steroid inactivation in a range of tissues and cell types not considered as classical sex steroid target tissues.
60 citations
TL;DR: A new category of 17 β -hydroxysteroid dehydrogenase (17 β -HSD) type 1 inhibitors was developed, found to be a more potent inhibitor than the substrate estrone itself or a panel of three known inhibitors.
44 citations
TL;DR: It is inferred from the phylogenetic analysis that prostaglandin F synthase may represent a recent recruit to the eicosanoid biosynthetic pathway from the hydroxysteroid dehydrogenase pathway and furthermore that, in the context of gene recruitment, Xenopus laevisρ-crystallin may represents a shared gene.
Abstract: The aldo-keto reductase enzymes comprise a functionally diverse gene family which catalyze the NADPH-dependant reduction of a variety of carbonyl compounds. The protein sequences of 45 members of this family were aligned and phylogenetic trees were deduced from this alignment using the neighbor-joining and Fitch algorithms. The branching order of these trees indicates that the vertebrate enzymes cluster in three groups, which have a monophyletic origin distinct from the bacterial, plant, and invertebrate enzymes. A high level of conservation was observed between the vertebrate hydroxysteroid dehydrogenase enzymes, prostaglandin F synthase, and ρ-crystallin of Xenopus laevis. We infer from the phylogenetic analysis that prostaglandin F synthase may represent a recent recruit to the eicosanoid biosynthetic pathway from the hydroxysteroid dehydrogenase pathway and furthermore that, in the context of gene recruitment, Xenopus laevisρ-crystallin may represent a shared gene.
TL;DR: Evidence for an additional steroidogenic lesion induced by gonadotropin is presented and transcripts for type I and type II 3β-HSD were substantially (5- to 8-fold) down-regulated.
Abstract: 3Beta-hydroxysteroid dehydrogenase/delta5-delta4 isomerases (3beta-HSD) are enzymes that catalyze the conversion of delta5 to delta4 steroids in the gonads and adrenal for the biosynthesis of sex steroid and corticoids. In gonadotropin-desensitized Leydig cells, from rats treated with high doses of human CG (hCG), testosterone production is markedly reduced, a finding that was attributed in part to reduction of CYP17 expression. In this study, we present evidence for an additional steroidogenic lesion induced by gonadotropin. Using differential display analysis of messenger RNA (mRNA) from Leydig cells of rats treated with a single desensitizing dose of hCG (2.5 microg), we found that transcripts for type I and type II 3beta-HSD were substantially (5- to 8-fold) down-regulated. This major reduction, confirmed by RNase protection assay, was observed at the high hCG dose (2.5 microg), whereas minor or no change was found at lower doses (0.01 and 0.1 microg). In contrast, 3beta-HSD mRNA transcripts were not changed in luteinized ovaries of pseudopregnant rats treated with 2.5 microg hCG. The down-regulation of 3beta-HSD mRNA in the Leydig cell resulted from changes at the transcriptional level. Western blot analysis showed 3beta-HSD protein was significantly reduced by hCG treatment, with changes that were coincidental with the reduction of enzyme activity and temporally consistent with the reduction of 3beta-HSD mRNA but independent of LH receptor down-regulation. The reduction of 3beta-HSD mRNA resulting from transcriptional inhibition of gene expression, and the consequent reduction of 3beta-HSD activity could contribute to the inhibition of androgen production in gonadotropin-induced steroidogenic desensitization of Leydig cells. The gender-specific regulation of 3beta-HSD by hCG reflects differential transcriptional regulation of the enzymes to accommodate physiological hormonal requirements and reproductive function.
TL;DR: In this paper, the effect of cytokines on 17 β-hydroxysteroid dehydrogenase (17 β -HSD) and 3 β -hydroxsteroid de-hydrogenase/isomerase (3 β − HSD) activities in human breast cancer cells was examined.
TL;DR: Observations provide direct evidence that Tyr253 functions as the general acid (proton donor) in the isomerase reaction mechanism, and the coenzyme-activation profiles support the proposed two-step enzyme mechanism in which NADH produced by the 3beta-HSD activity induces the enzyme to assume theIsomerase conformation.
Abstract: 3β-Hydroxysteroid dehydrogenase/steroid Δ5-isomerase (3β-HSD/isomerase) was expressed by baculovirus in Spodoptera fungiperda (Sf9) insect cells from cDNA sequences encoding the human wild-type I (placental) enzyme and the human type I mutant- Y253F. The wild-type and Y253F enzymes were each purified as a single, homogeneous protein from a suspension of the Sf9 cells. Ultraviolet (UV) spectral analyses showed that the wild-type enzyme induced changes in the UV spectrum of the competitive isomerase inhibitor, 19-nortestosterone, and the Y253F mutant did not. The wild-type isomerase required activaltion by coenzyme to produce the spectral shift. Activation of isomerase by NADH produced a greater change in the 19-nortestosterone spectrum than activation by NAD+. These observations provide direct evidence that Tyr253 functions as the general acid (proton donor) in the isomerase reaction mechanism. Furthermore, the coenzyme-activation profiles support our proposed two-step enzyme mechanism in which NADH produc...
TL;DR: Chronic hypoxemia selectively inhibits renal 11beta-HSD2 mRNA expression and enzyme activity in the ovine fetus, which may contribute, at least in part, to the mechanisms leading to fetal hypertension.
Abstract: The present study was designed to examine the effects of chronic fetal placental embolization on the expression of 11, hydroxysteroid dehydrogenase (11 S-HSD) types 1 and 2, the intracellular enzymes responsible for the metabolism of glucocorticoids. Twelve instrumented fetal sheep were randomly allocated on Day 110 (term = 147 days) to either a control (n = 6) or embolized (n = 6) group. Embolized fetuses received daily injections of nonradioactive microspheres into the abdominal aorta for 21 days to decrease arterial oxygen content by 4050% of the pre-embolization values. At the end of the experiment, fetal liver and kidney tissues as well as placental cotyledons were collected, and tissue levels of 11 -HSD mRNA and activity were determined by standard Northern blot analysis and radiometric conversion assay, respectively. There was a 44% reduction (p < 0.01) in the level of renal 11 -HSD2 mRNA in the embolized group as compared with the control group. Moreover, this reduction in mRNA was carried through to 11 ,-HSD2 protein, since there was a corresponding decrease in the level of 11,P-HSD2 activity (4.5 + 0.2 vs. 2.9 + 0.1 pmol/min per milligram protein, p < 0.01). In contrast, levels of both 11,HSD1 mRNA and activity in the fetal liver remained unchanged. Moreover, both 111-HSD types 1 and 2 mRNA and activity in the placenta were not altered by the fetal placental embolization. In conclusion, chronic hypoxemia selectively inhibits renal 11 P-HSD2 mRNA expression and enzyme activity in the ovine fetus, which may contribute, at least in part, to the mechanisms leading to fetal hypertension.
TL;DR: The aim of this study was to compare the formation of metabolites in freshly isolated epithelial cells and in cells of long‐term cultures and to identify the 5α‐reductase (5α‐R) and 17β‐hydroxysteroid dehydrogenase (17β‐HSD) isoforms responsible for metabolite formation.
Abstract: The aim of this study on testosterone (T) metabolism in benign prostatic hyperplasia (BPH) and prostatic cancer was to compare the formation of metabolites in freshly isolated epithelial cells and in cells of long-term cultures (2 passages) and to identify the 5alpha-reductase (5alpha-R) and 17beta-hydroxysteroid dehydrogenase (17beta-HSD) isoforms responsible for metabolite formation. Androst-4-enedione (A), dihydrotestosterone (DHT) and 5alpha-androstanedione (5alpha-A) formation were measured by high-performance liquid chromatography coupled to a Flo-one HP radioactivity detector. Enzyme isoforms were studied by Northern blot analysis and reverse transcriptase-polymerase chain reaction (RT-PCR). T conversion into A by 17beta-HSD, rather than reduction into DHT by 5alpha-R, was by far the predominant activity in cultured epithelial cells. The metabolic profile did not differ substantially between BPH and cancer cells. Long-term cell culture led to an increase in A formation compared with the level recorded in freshly isolated cells, with no significant incidence on the relative DHT level. According to RT-PCR results, both 5alpha-R isoforms (1 and 2) and 2 17beta-HSD isoforms (2 and 3) are present in epithelial cell cultures and in tissues. According to Northern blot analyses, the mRNAs for 5alpha-R2 and 17beta-HSD4 are expressed in tissue and those for 5alpha-R1 and types 2 and 4 17beta-HSD in isolated cell cultures. Moreover, finasteride, a specific 5alpha-R2 inhibitor, inhibits DHT and 5alpha-A formation in long-term cell culture of adenocarcinoma epithelial cells plated on Matrigel, suggesting a 5alpha-R2 expression. Thus, although 5alpha-R2 is present in freshly isolated epithelial cell cultures and in long-term epithelial cells cultured on Matrigel and predominates in prostate tissue, it is the 5alpha-R1 isoform that is preferentially expressed in epithelial cell cultures.
TL;DR: The effect on the reduction of two ketone and two aldehyde substrates by pig 3alpha/beta,20beta-hydroxysteroid dehydrogenase in which tyrosine-194 has been mutated to phenylalanine and cysteine, and lysine-198 has been mutation to isoleucine and arginine is reported.
Abstract: Pig 3alpha/beta,20beta-hydroxysteroid dehydrogenase is an NADPH-dependent enzyme that catalyses the reduction of ketones on steroids and aldehydes and ketones on various xenobiotics, like its homologue carbonyl reductase. 3alpha/beta,20beta-Hydroxysteroid dehydrogenase and carbonyl reductase are members of the short-chain dehydrogenases/reductase family, in which a tyrosine residue and a lysine residue have been identified as catalytically important. In pig 20beta-hydroxysteroid dehydrogenase these residues are tyrosine-194 and lysine-198. Here we report the effect on the reduction of two ketone and two aldehyde substrates by pig 3alpha/beta,20beta-hydroxysteroid dehydrogenase in which tyrosine-194 has been mutated to phenylalanine and cysteine, and lysine-198 has been mutated to isoleucine and arginine. Mutants with phenylalanine-194 or isoleucine-198 are inactive. Depending on the substrate, the mutant with cysteine-194 has a catalytic efficiency of 0.4-1% and the mutant with arginine-198 has a catalytic efficiency of 4-23% of the wild-type enzyme. We also mutated tyrosine-81 and tyrosine-253 to phenylalanine. Although both tyrosines are conserved in 3alpha/beta,20beta-hydroxysteroid dehydrogenase and carbonyl reductase, depending on the substrate, the mutant enzymes are as active as, or more active than, wild-type enzyme.
TL;DR: The promoter region of the rat ovarian 20 α-HSD gene is identified and partially characterized and it is demonstrated that the regulatory elements for 20α- HSD are present within a 2.5 kb 5′ flanking region of this gene.
TL;DR: It is demonstrated that only 17β‐hydroxysteroid dehydrogenase seems to be involved in the AA action, since nearly 60% inhibition of testosterone production was found when the cells were incubated with androstenedione, and the conversion of AA to its metabolites is not required for its action.
TL;DR: A distinct and previously undescribed syndrome of alopecia totalis, ichthyosis, and male pseudohermaphroditism due to steroid 17b-hydroxysteroid dehydrogenase deficiency was observed in an Israeli-Arab newborn infant.
Abstract: A distinct and previously undescribed syndrome of alopecia totalis, ichthyosis, and male pseudohermaphroditism due to steroid 17b-hydroxysteroid dehydrogenase deficiency was observed in an Israeli-Arab newborn infant.
01 Jan 1998
TL;DR: It is concluded that both the PKA and PKC pathways are involved in the regulation of rat adrenal 11beta-HSD2 gene expression.
Abstract: The regulation of 1 lphydroxysteroid dehydrogenase type I1 (1 1PHSD2) gene expression was studied in primary cultures of rat adrenocortical cells. The protein kinase A (PKA) pathway agonists forskolin, dibutyryl CAMP and ACTH caused a .510 fold increase in l lp-HSD2 mRNA as determined by semiquantitative PCR. The effect of forskolin could be partially inhibited by the addition of the phorbol ester TPA, an activator of the protein kinase C (PKC) pathway. The increase in mRNA encoding 1 lPHSD2 was accompanied by increased synthesis of 1 lp-HSD2 as measured by immunoprecipitation of labeled protein. It is concluded that both the PKA and PKC pathways are involved in the regulation of rat adrenal 1p--HSD2 gene expression.