Ginkgo biloba extract protects against tartrazine-induced testicular toxicity in rats: involvement of antioxidant, anti-inflammatory, and anti-apoptotic mechanisms.

The use of additives, especially colorants, in food and pharmaceutical industry is increasing dramatically. Currently, additives are classified as contaminants of emerging concern (CECs). Concerns have been raised about the potential hazards of food additives to reproductive organs and fertility. The present study investigates the reproductive toxicity of tartrazine (TRZ), a synthetic colorant, in male rats and aims to explore the curative effect of Ginkgo biloba extract (EGb) against TRZ-induced testicular toxicity. Twenty-four rats were divided into four groups: the control (0.5 ml distilled water), the EGb group (100 mg/kg EGb alone), the TRZ group (7.5 mg/kg TRZ alone), and the TRZ-EGb group (7.5 mg/kg TRZ plus 100 mg/kg EGb). The doses were administered orally in distilled water once daily for 28 days. Toxicity studies of TRZ investigated testicular redox state, serum gonadotropins, and testosterone levels, testicular 17 ß-hydroxysteroid dehydrogenase activity, sperm count and quality, levels of inflammatory cytokines, and caspase-3 expression as an apoptotic marker. Also, histopathological alterations of the testes were examined. TRZ significantly affected the testicular redox status as indicated by the increase in malondialdehyde and the decrease in reduced glutathione, superoxide dismutase, and catalase. It also disrupted serum gonadotropins (follicle stimulating hormone and luteinizing hormone) and testosterone levels and the activity of testicular 17ß-hydroxysteroid dehydrogenase. Additionally, TRZ adversely affected sperm count, motility, viability, and abnormality. Levels of tumor necrosis factor-α, interleukin-1ß, interleukin-6, and expression of caspase-3 were increased in the testes. Histopathological examination of the testes supported the alterations mentioned above. Administration of EGb significantly ameliorated TRZ-induced testicular toxicity in rats. In conclusion, EGb protected against TRZ-induced testicular toxicity through antioxidant, anti-inflammatory, and anti-apoptotic mechanisms.

Carbon Chain Length Determines Inhibitory Potency of Perfluoroalkyl Sulfonic Acids on Human Placental 3ß-Hydroxysteroid Dehydrogenase 1: Screening, Structure-Activity Relationship, and In Silico Analysis.

Objective: This study aimed to compare 9 perfluoroalkyl sulfonic acids (PFSA) with carbon chain lengths (C4-C12) to inhibit human placental 3ß-hydroxysteroid dehydrogenase 1 (3ß-HSD1), aromatase, and rat 3ß-HSD4 activities. Methods: Human and rat placental 3ß-HSDs activities were determined by converting pregnenolone to progesterone and progesterone secretion in JEG-3 cells was determined using HPLC/MS-MS, and human aromatase activity was determined by radioimmunoassay. Results: PFSA inhibited human 3ß-HSD1 structure-dependently in the order: perfluorooctanesulfonic acid (PFOS, half-maximum inhibitory concentration, IC 50: 9.03 ± 4.83 µmol/L) > perfluorodecanesulfonic acid (PFDS, 42.52 ± 8.99 µmol/L) > perfluoroheptanesulfonic acid (PFHpS, 112.6 ± 29.39 µmol/L) > perfluorobutanesulfonic acid (PFBS) = perfluoropentanesulfonic acid (PFPS) = perfluorohexanesulfonic acid (PFHxS) = perfluorododecanesulfonic acid (PFDoS) (ineffective at 100 µmol/L). 6:2FTS (1H, 1H, 2H, 2H-perfluorooctanesulfonic acid) and 8:2FTS (1H, 1H, 2H, 2H-perfluorodecanesulfonic acid) did not inhibit human 3ß-HSD1. PFOS and PFHpS are mixed inhibitors, whereas PFDS is a competitive inhibitor. Moreover, 1-10 µmol/L PFOS and PFDS significantly reduced progesterone biosynthesis in JEG-3 cells. Docking analysis revealed that PFSA binds to the steroid-binding site of human 3ß-HSD1 in a carbon chain length-dependent manner. All 100 µmol/L PFSA solutions did not affect rat 3ß-HSD4 and human placental aromatase activity. Conclusion: Carbon chain length determines inhibitory potency of PFSA on human placental 3ß-HSD1 in a V-shaped transition at PFOS (C8), with inhibitory potency of PFOS > PFDS > PFHpS > PFBS = PFPS = PFHxS = PFDoS = 6:2FTS = 8:2FTS.

Effect of peripheral neural stimulation with allopregnanolone on ovarian physiology.

Neuroactive steroids can rapidly regulate multiple physiological functions in the central and peripheral nervous systems. The aims of the present study were to determine whether allopregnanolone (ALLO), administered in low nanomolar and high micromolar concentrations, can: (i) induce changes in the ovarian progesterone (P4) and estradiol (E2) release; (ii) modify the ovarian mRNA expression of Hsd3b1 (3ß-hydroxysteroid dehydrogenase, 3ß-HSD)3ß-, Akr1c3 (20α-hydroxysteroid dehydrogenase, 20α-HSD), and Akr1c14 (3α-hydroxy steroid oxidoreductase, 3α-HSOR)); and (iii) modulate the ovarian expression of progesterone receptors A and B, α and ß estrogenic receptors, luteinizing hormone receptor (LHR) and follicle-stimulating hormone receptor (FSHR). To further characterize ALLO peripheral actions, the effects were evaluated using a superior mesenteric ganglion-ovarian nervous plexus-ovary (SMG-ONP-O) and a denervated ovary (DO) systems. ALLO SMG administration increased P4 concentration in the incubation liquid by decreasing ovarian 20α-HSD mRNA, and it also increased ovarian 3α-HSOR mRNA expression. In addition, ALLO neural peripheral modulation induced an increase in the expression of ovarian LHR, PRA, PRB, and ERα. Direct ALLO administration to the DO decreased E2 and increased P4 concentration in the incubation liquid. The mRNA expression of 3ß-HSD decreased and 20α-HSD increased. Further, ALLO in the OD significantly changed ovarian FSHR and PRA expression. This is the first evidence of ALLO's direct effect on ovarian steroidogenesis. Our results provide important insights about how this neuroactive steroid interacts both with the PNS and the ovary, and these findings might help devise some of the pleiotropic effects of neuroactive steroids on female reproduction. Moreover, ALLO modulation of ovarian physiology might help uncover novel treatment approaches for reproductive diseases.

Carbon Chain Length Determines Inhibitory Potency of Perfluoroalkyl Sulfonic Acids on Human Placental 3β-Hydroxysteroid Dehydrogenase 1: Screening, Structure-Activity Relationship, and In Silico Analysis / 生物医学与环境科学（英文）

OBJECTIVE@#This study aimed to compare 9 perfluoroalkyl sulfonic acids (PFSA) with carbon chain lengths (C4-C12) to inhibit human placental 3β-hydroxysteroid dehydrogenase 1 (3β-HSD1), aromatase, and rat 3β-HSD4 activities.@*METHODS@#Human and rat placental 3β-HSDs activities were determined by converting pregnenolone to progesterone and progesterone secretion in JEG-3 cells was determined using HPLC/MS-MS, and human aromatase activity was determined by radioimmunoassay.@*RESULTS@#PFSA inhibited human 3β-HSD1 structure-dependently in the order: perfluorooctanesulfonic acid (PFOS, half-maximum inhibitory concentration, IC 50: 9.03 ± 4.83 μmol/L) > perfluorodecanesulfonic acid (PFDS, 42.52 ± 8.99 μmol/L) > perfluoroheptanesulfonic acid (PFHpS, 112.6 ± 29.39 μmol/L) > perfluorobutanesulfonic acid (PFBS) = perfluoropentanesulfonic acid (PFPS) = perfluorohexanesulfonic acid (PFHxS) = perfluorododecanesulfonic acid (PFDoS) (ineffective at 100 μmol/L). 6:2FTS (1H, 1H, 2H, 2H-perfluorooctanesulfonic acid) and 8:2FTS (1H, 1H, 2H, 2H-perfluorodecanesulfonic acid) did not inhibit human 3β-HSD1. PFOS and PFHpS are mixed inhibitors, whereas PFDS is a competitive inhibitor. Moreover, 1-10 μmol/L PFOS and PFDS significantly reduced progesterone biosynthesis in JEG-3 cells. Docking analysis revealed that PFSA binds to the steroid-binding site of human 3β-HSD1 in a carbon chain length-dependent manner. All 100 μmol/L PFSA solutions did not affect rat 3β-HSD4 and human placental aromatase activity.@*CONCLUSION@#Carbon chain length determines inhibitory potency of PFSA on human placental 3β-HSD1 in a V-shaped transition at PFOS (C8), with inhibitory potency of PFOS > PFDS > PFHpS > PFBS = PFPS = PFHxS = PFDoS = 6:2FTS = 8:2FTS.

HB-EGF upregulates StAR expression and stimulates progesterone production through ERK1/2 signaling in human granulosa-lutein cells.

BACKGROUND: Heparin-binding epidermal growth factor-like growth factor (HB-EGF) belongs to the epidermal growth factor (EGF) family of growth factors. HB-EGF and its receptors, epidermal growth factor receptor (EGFR) and HER4, are expressed in the human corpus luteum. HB-EGF has been shown to regulate luteal function by preventing cell apoptosis. Steroidogenesis is the primary function of the human corpus luteum. Steroidogenic acute regulatory protein (StAR) plays a critical role in steroidogenesis. StAR expression and progesterone (P4) production in human granulosa-lutein (hGL) cells have been shown to be upregulated by a ligand of EGFR, amphiregulin. However, whether HB-EGF can achieve the same effects remains unknown. METHODS: A steroidogenic human ovarian granulosa-like tumor cell line, KGN, and primary culture of hGL cells obtained from patients undergoing in vitro fertilization treatment were used as experimental models. The underlying molecular mechanisms mediating the effects of HB-EGF on StAR expression and P4 production were explored by a series of in vitro experiments. RESULTS: Western blot showed that EGFR, HER2, and HER4 were expressed in both KGN and hGL cells. Treatment with HB-EGF for 24 h induced StAR expression but did not affect the expression of steroidogenesis-related enzymes, P450 side chain cleavage enzyme, 3ß-hydroxysteroid dehydrogenase, and aromatase. Using pharmacological inhibitors and a siRNA-mediated knockdown approach, we showed that EGFR, HER4, but not HER2, were required for HB-EGF-stimulated StAR expression and P4 production. In addition, HB-EGF-induced upregulations of StAR expression and P4 production were mediated by the activation of the ERK1/2 signaling pathway. CONCLUSION: This study increases the understanding of the physiological role of HB-EGF in human luteal functions. Video Abstract.

Apoptosis and glucocorticoid-related genes mRNA expression is modulated by coenzyme Q10 supplementation during in vitro maturation and vitrification of bovine oocytes and cumulus cells.

Oocyte in vitro maturation (IVM) and vitrification procedures lead to detrimental effects on the overall oocyte quality. The addition of antioxidants during IVM, such as the coenzyme Q10 (Q10), has been demonstrated to positively impact on the cumulus-oocyte complexes due to its role in protection from oxidative damage and modulating gene transcription. Furthermore, glucocorticoids (GC) regulate gene transcription, energy metabolism and apoptosis during the early steps of reproduction. In this sense, most GC actions are mediated by the glucocorticoid receptor (NR3C1), a transcription factor. However, the specific roles of GC in ovarian physiology and oocyte maturation are still unknown. In this regard, a better knowledge on the expression of GC-related and apoptosis-related genes during IVM and cryopreservation procedures could potentially benefit the refinement of assisted reproductive techniques in the bovine species. The present study aims to explore the expression of NR3C1 mRNA in fresh and vitrified bovine oocytes and cumulus cells in response to Q10 (50 µM), and the effect of cortisol addition (0.25 µM, 0.5 µM) on the expression of NR3C1. We also studied the mRNA expression of NR3C1-related genes belonging to the GC regulation pathway, such as hydroxysteroid dehydrogenases (HSD11B1; HSD11B2), immunophilins (FKBP4; FKBP5), signal transducers and activators of transcription (STAT3; STAT5A), the mineralocorticoid receptor (NR3C2), and to the apoptosis pathway, such as the anti- (BCL2) and pro-apoptotic (BAX) mRNA transcripts in oocytes and cumulus cells 1) after IVM, and 2) after vitrification, both in presence or absence of Q10 supplementation during IVM. Our results show that there is an increase in the NR3C1 receptor expression after vitrification of oocytes, but not after exogenous cortisol supplementation during IVM. In addition, Q10 reduces the mRNA expression of HSD11B1 and FKBP5 in oocytes at levels of immature oocytes (HSD11B1 mRNA expression also in cumulus cells), and the BAX:BCL2 ratio mRNA expression. After vitrification in the presence of Q10, HSD11B2 mRNA expression increases in cumulus cells, while HSD11B1 and BAX:BCL2 mRNA expression decreases significantly both in oocytes and cumulus cells. In conclusion, our results show for the first time the effect of IVM, vitrification and Q10 supplementation on the mRNA relative expression of GC-related and apoptosis genes, and the effect of vitrification in the protein expression of NR3C1.

Umbelliferone ameliorates oxidative stress and testicular injury, improves steroidogenesis and upregulates peroxisome proliferator-activated receptor gamma in type 2 diabetic rats.

OBJECTIVES: Diabetes mellitus (DM) is a chronic disease associated with serious complications, including male infertility. Umbelliferone (UMB) is a coumarin with promising antioxidant, anti-inflammatory and other beneficial effects. This study investigated the ameliorative effect of UMB against testicular injury, oxidative stress and altered steroidogenesis in rats with type 2 DM. METHODS: Rats received a high fat diet for 4 weeks followed by a single injection of streptozotocin. Diabetic rats were treated with UMB or pioglitazone (PIO) for 6 weeks and samples were collected for analysis. KEY FINDINGS: Diabetic rats exhibited hyperglycemia, insulin resistance and dyslipidemia associated with increased serum pro-inflammatory cytokines, and decreased gonadotropins and testosterone. UMB significantly ameliorated metabolic alterations, decreased pro-inflammatory cytokines, and increased gonadotropins and testosterone levels. UMB prevented testicular injury, suppressed lipid peroxidation and nitric oxide and increased antioxidants in diabetic rats. In addition, UMB upregulated testicular gonadotropins receptors, steroidogenesis markers (steroidogenic acute regulatory protein, cytochrome P450 family 17 subfamily A member 1 [CYP17A1], 3ß-hydroxysteroid dehydrogenase [3ß-HSD] and 17ß-hydroxysteroid dehydrogenase [17ß-HSD]), and peroxisome proliferator-activated receptor gamma (PPARγ) expression. CONCLUSIONS: UMB prevents testicular injury by preventing metabolic alterations, suppressing oxidative damage and inflammation, and boosting antioxidant defenses in diabetic rats. UMB enhanced pituitary-gonadal axis and steroidogenesis and upregulated testicular PPARγ in diabetic rats. Thus, UMB may represent a protective agent against testicular injury and sexual dysfunction associated with chronic hyperglycemia.

HSD17B1 gene polymorphisms and risk of endometrial and breast cancer.

Estrogen exposure influences breast and endometrial cancer risk. The HSD17B1 gene produces an enzyme that catalyzes the conversion of estrone to estradiol. We hypothesized that genetic variations in HSD17B1 gene may alter endogenous estrogen levels and, thus, influence endometrial and breast cancer risk. We validated and genotyped polymorphisms in the HSD17B1 gene and assessed whether these single nucleotide polymorphisms (SNPs), or the imputed haplotypes, were associated with endometrial and breast cancer risk. We also assessed whether a priori risk factors modified the associations between HSD17B1 genotype and cancer risk, and whether HSD17B1 genotypes were associated with plasma estrogen levels among postmenopausal women not using hormone replacement therapy. Ten SNPs of HSD17B1 gene were validated in 30 women from the Nurses' Health Study. Using the expectation maximization algorithm, three common (>5% frequency) haplotypes accounted for 97% of the chromosomes at this locus, and seven SNPs were in complete linkage disequilibrium. We identified and genotyped two haplotype-tagging SNPs (+1004C/T and +1322C/A), and genotyped an additional SNP [+1954A/G (Ser312Gly)] in nested case-control studies of endometrial cancer (cases = 222, controls = 666) and breast cancer (cases = 1007, controls = 1441) in the prospective Nurses' Health Study. Although no overall association by SNP or haplotype analysis was observed with endometrial or breast cancer risk, the +1954A/A genotype was associated with higher estradiol levels in lean women (P = 0.01) and interaction between the +1954 genotype with body mass index in postmenopausal breast cancer (P = 0.05) was suggested. These findings suggest that the HSD17B1 may be associated with circulating estradiol levels and interact with body mass index in postmenopausal breast cancer.

Estren (4-estren-3alpha,17beta-diol) is a prohormone that regulates both androgenic and estrogenic transcriptional effects through the androgen receptor.

Alternative mechanisms of steroid action, through both traditional nuclear receptors and indirect pathways of gene activation, are emerging. Recent studies suggest that the synthetic steroid, 4-estrene-3alpha,17beta-diol (estren), has nongenotropic as well as sex-nonspecific osteogenic effects in ovariectomized and orchidectomized mice. We found limited estrogen receptor-dependent effects by estren on gene expression in primary osteoblast cultures and showed that it binds poorly to estrogen and androgen receptors in vitro. However, estren potently regulated direct and indirect androgen receptor-dependent effects on gene expression by osteoblasts. Consistent with this, osteoblasts produced the potent androgen 19-nortestosterone from estren by way of a 3alpha-hydroxysteroid dehydrogenase-like activity. Moreover, recombinant 3alpha-hydroxysteroid dehydrogenase (AKR1C9) and osteoblast-derived cell lysate each effectively converted estren to 19-nortestosterone in vitro, and mRNA encoding this enzyme occurs in osteoblasts. In addition to its androgenic activity, estren potently stimulated androgen receptor-dependent effects on gene expression through conventional estrogen-sensitive transcriptional elements in osteoblasts. Therefore, through local metabolism, estren indirectly activates the androgen receptor to regulate both androgen- and estrogen-like transcriptional responses by bone-forming cells.

Blood steroid profile and in vitro steroidogenesis by ovarian follicles and testis fragments of adult sea lamprey, Petromyzon marinus.

The main purpose of the study was to identify the principal gonadal steroids synthesized by male and female sea lampreys, Petromyzon marinus. To achieve this, we used high performance liquid chromatography to separate the steroids in the serum of sexually mature animals, and to separate the steroids produced by gonadal tissue incubated in the presence of radiolabelled precursor steroids, as a means of identifying the major steroidogenic pathways. We were unable to detect evidence of the 'classical' steroids, such as 17beta-estradiol (E(2)) or testosterone (T) in the serum of either male or female lampreys. Instead, the principal chromatographic peaks contained very polar compounds that had elution times consistent with 15alpha-hydroxylated estrogens and androgens, and there were sex-specific differences in the chemical nature and the quantity of these compounds. Testis fragments or ovarian follicles co-incubated with tritium-labelled pregnenolone ([3H]P(5)), 17-hydroxyprogesterone ([3H]17OHP(4)), or androstenedione ([3H]A(4)), provided additional confirmation that the gonads synthesize a range of very polar steroids, and the metabolites found were consistent with the presence of a 15alpha-hydroxylated (15alphaOH) metabolic pathway common to testis and ovary. For ovarian tissue, the major 'end product' metabolites from all three precursors were 15alphaOH-estrogens, and for testis tissue 15alpha-hydroxyprogesterone (15alphaOHP(4)) and 15alpha-hydroxytestosterone (15alphaOHT) and small amounts of 15alphaOH estrogen. Small amounts of E(2) were also produced by both ovarian (all substrates) and testicular tissue (some substrates). Although it was assumed that the E(2) was synthesized via the aromatization of T, [3H]T was not found as an intermediate metabolite. The study suggests that the principal gonadal steroids in sea lamprey are 15alpha-OH compounds, and that only small amounts of E(2) or T are synthesized by the gonads at this stage of reproductive development. There was no direct evidence of progesterone (P(4)) synthesis from [3H]P(5), although the metabolites synthesized by both testis and ovary were indicative of a metabolic pathway that involved P(4) as an intermediate.

11Beta-hydroxysteroid dehydrogenase 1 transforms 11-dehydrocorticosterone into transcriptionally active glucocorticoid in neonatal rat heart.

The ability of cells to directly respond to glucocorticoids and aldosterone is a function of GR and MR expression, and coexpression of 11beta-hydroxysteroid dehydrogenases (11betaHSDs), which convert glucocorticoids and their 11-ketometabolites into either receptor inactive or active derivatives. The aim of the present study was to determine the cellular expression of GR, MR, 11betaHSD1, and 11betaHSD2 in neonatal rat heart and determine the role these enzymes play in modulating glucocorticoid and aldosterone action. Ribonuclease protection analysis and steroid binding assays showed that GR is expressed in both cardiac myocytes and fibroblasts, whereas MR is expressed only in myocytes. 11betaHSD2 was not detected in cardiac cells, but 11betaHSD1 was expressed at high levels in both cardiac myocytes and fibroblasts. Enzyme activity studies demonstrated that 11betaHSD1 acted as a reductase only, converting biologically inactive 11-dehydrocorticosterone to corticosterone, which then stimulated serum and glucocorticoid-induced kinase gene transcription via GR. In both cardiac myocytes and fibroblasts, aldosterone stimulated serum and glucocorticoid-induced kinase gene expression exclusively via GR, but not MR, indicating that aldosterone can have glucocorticoid-like actions in heart. The ability of cardiac cells to use both circulating corticosterone and 11-dehydrocorticosterone as a source of glucocorticoid suggests that the heart is under tonic glucocorticoid control, implying that glucocorticoids play important homeostatic roles in the heart.

Functional adrenocorticotropic hormone receptor in cultured human vascular endothelial cells : possible role in control of blood pressure.

Hypertension is a prominent feature of patients with Cushing's disease and ectopic adrenocorticotropic hormone (ACTH) syndrome, who have elevated ACTH levels. Chronic administration of ACTH (1-24) also raises blood pressure in humans. This effect has been postulated to be due to ACTH-induced increases in cortisol secretion in the adrenal gland. It is well known that cortisol increases vascular tone by potentiating the vasoconstrictor action of a number of pressor hormones. In the present study, we show direct evidence that human aortic endothelial cells possess the ACTH receptor. 11beta-Dehydrogenation, converting cortisol to its inactive metabolite, cortisone, mediated by vascular 11beta-hydroxysteroid dehydrogenase type 2 is essential for the control of vascular tone, and the reduced activity may be relevant to the pathogenesis of hypertension. We found that ACTH (1-24) dose-dependently decreased the gene expression and enzyme activity of 11beta-hydroxysteroid dehydrogenase type 2 in these cells, and the decrease was partially abolished by a selective ACTH receptor antagonist. This may indicate that ACTH potentiates the action of cortisol through its direct effect on the vasculature. Therefore, the present study provides important information for understanding the mechanism of ACTH-induced hypertension.

Regulation of the action of hydrocortisone in airway epithelial cells by 11beta-hydroxysteroid dehydrogenase.

11beta-hydroxysteroid dehydrogenase (11betaHSD) reversibly converts hydrocortisone, the predominant active endogenous glucocorticoid in humans, to its inactive metabolite cortisone by oxidizing the 11-hydroxy group to an 11-keto group. Because this enzyme is highly expressed in human bronchial epithelial cells, we hypothesized that it regulates epithelial responses to glucocorticoids by reducing levels of hydrocortisone available to bind to the glucocorticoid receptor. Primary human bronchial epithelial cells (PBECs) were isolated from seven autopsy specimens and cultured in F12/Dulbecco's modified Eagle's medium with 5% fetal bovine serum until approximately 80% confluent. Cells were preincubated with 10(-9) M to 10(-5) M hydrocortisone for 24 h in the presence or absence of 10(-6) M of the 11betaHSD inhibitor glycyrrhetinic acid, after which the cells were stimulated with 5 ng/ml interleukin-1beta for 24 h. Granulocyte macrophage colony-stimulating factor (GM-CSF) levels were quantitated in the resulting supernatants by enzyme-linked immunosorbent assay. Hydrocortisone inhibited GM-CSF release in stimulated PBEC with a concentration that produces 50% inhibition of maximum effect (IC(1/2)max) of 5.0 x 10(-8) M. In the presence of glycyrrhetinic acid, the potency of hydrocortisone was increased approximately 33-fold (IC(1/2)max with glycyrrhetinic acid, 1.5 x 10(-9) M). Hydrocortisone activity was maximally enhanced at concentrations between 10(-9) M and 10(-8) M, levels that are comparable to plasma levels of hydrocortisone not bound to plasma proteins. Glycyrrhetinic acid had no effect on the suppression of GM-CSF release by hydrocortisone in the transformed cell line BEAS-2B, which does not express the 11betaHSD enzyme. Glycyrrhetinic acid also had no effect on the inhibition of GM-CSF release in PBECs by the synthetic glucocorticoids budesonide, beclomethasone dipropionate, fluticasone propionate, mometasone furoate, and triamcinolone acetonide, steroids not metabolized by 11betaHSD. Together, these findings suggest that metabolism of hydrocortisone by 11betaHSD may regulate glucocorticoid activity in human airway epithelial cells.

Effect of growth hormone, insulin and dexamethasone on 11 beta-hydroxysteroid dehydrogenase activity on a primary culture of rat hepatocytes.

11 beta-hydroxysteroid dehydrogenase (11 beta-HSD) is a microsomal enzyme present in the peripheral tissues of the rat, including the liver, and is mediated by a number of factors in animal in vivo studies. However, the effect of peptide hormones and glucocorticoids on the activity of 11 beta-HSD in isolated rat hepatocytes is not clear. To investigate these effects, we determined 11 beta-HSD activity in a primary culture of rat hepatocytes by adding various concentrations of growth hormone, insulin and dexamethasone (Dex). 11 beta-HSD activity increased significantly after treatment with Dex (10(-9)M-10(-6)M) for 48h. Dex (100nM) treated hepatocytes, incubated for 12h to 48h, resulted in a significant two-to four-fold rise in 11 beta-HSD activity compared to control (p < 0.01), which was in contrast to GH (10(-9)M-10(-6)M) and insulin (10(-8)M-10(-5)M), which inhibited 11 beta-HSD activity (p < 0.05). These results suggest that the 11 beta-HSD of rat hepatocytes is under multifactorial regulation; Dex stimulates and GH and insulin inhibit 11 beta-HSD activity in primary cultures of rat hepatocytes.

11 beta-Hydroxysteroid dehydrogenase activity and corticosteroid hormone action.

In normal physiology 11 beta-hydroxysteroid dehydrogenase (11 beta-OHSD) protects the mineralocorticoid receptor (MR) from glucocorticoid excess. In the rat, however, 11 beta-OHSD mRNA and activity is widespread, suggesting that it may also play a role in regulating ligand access to the glucocorticoid receptor (GR). We have studied the role of the 11 beta-OHSD in modulating corticosteroid hormone action in rat pituitary GH3 cells (glucocorticoids inhibit prolactin gene transcription) and renal epithelial NRK-52E cells (mineralocorticoids increase Na-K ATPase subunit gene expression) in culture. Both cell lines express high levels of 11 beta-OHSD activity, and Northern/Western blot analyses using a rat cDNA probe and antisera raised against rat liver 11 beta-OHSD reveal a single 1.4 Kb mRNA encoding an enzyme of molecular size 34 kDa. In GH3 cells, prolactin gene transcription was unaffected by corticosterone (B) in doses of 10(-8) to 10(-6) M. When 11 beta-OHSD activity was inhibited with the licorice derivative, glycyrrhetinic acid (GE); however, 10(-6) M B inhibited prolactin (PRL) mRNA levels to the same degree as an equimolar concentration of the GR agonist RU 28362. This effect was blocked by co-incubation with the GR antagonist RU 38486. In NRK-52E cells, co-incubation with B and GE resulted in a marked increase in alpha 1/beta 1 Na-K ATPase subunit mRNA levels when compared with GE and/or B alone and this effect could be blocked by administration of the MR antagonist RU 26752.(ABSTRACT TRUNCATED AT 250 WORDS)

Inactivation of corticosteroids in intestinal mucosa by 11 beta-hydroxysteroid: NADP oxidoreductase (EC 1.1.1.146).

Activity of the enzyme 11 beta-hydroxysteroid:NADP oxidoreductase (EC 1.1.1.146) in human intestinal mucosa was determined by incubating scraped mucosa with 3H-cortisone and 14C-cortisol; these steroids were then extracted, separated chromatographically, and the radioactivity assayed to determine simultaneously both reductase and dehydrogenase activities. This was the only significant metabolic alteration which the substrate underwent. Only two cases had slight (5 and 13%) reductase activity. In 35 patients, 16 male and 19 female, including seven cases of Crohn's disease, three ulcerative colitis, five diverticulitis, two undergoing surgery for repair of injuries and 18 for carcinoma of colon or rectum, cortisol was converted to cortisone in 15 min with a wide range of values distributed uniformly up to 85% dehydrogenation, with a mean of 42%. When tissue homogenates were fortified with coenzymes, excess NADPH lowered dehydrogenase activity 81%; excess NADP increased dehydrogenase activity 2-fold in three cases. It is possible that a value is characteristic of an individual but perhaps more likely enzyme activity varies with metabolic events involving changes in the coenzyme levels in mucosa, and a random sampling might be expected to yield such a distribution of values. In any event, where activity is high most of the cortisol is inactivated within minutes. It is suggested that synthetic corticoids which escape such metabolic alteration might, except during pregnancy, prove superior in the treatment of conditions such as inflammatory bowel disease.

Effect of cholesterol oxidation on (Na+, K+) ATPase activity of erythrocyte membranes.

By incubation of human erythrocyte ghosts with cholesterol oxidase (EC 1.1.3.6) part of the cholesterol of the membrane is replaced by 4-cholesten-3-one. This alteration in the sterol composition is accompanied by an inhibition of the (Na+, K+) ATPase of the erythrocyte membrane.