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B A Cooke

Publications and source records attributed to B A Cooke.

At least 19 recordsLinked to original sources
PubMed2003

Ovarian modulators of type 1 11beta-hydroxysteroid dehydrogenase (11betaHSD) activity and intra-follicular cortisol:cortisone ratios correlate with the clinical outcome of IVF.

BACKGROUND: Follicular fluid (FF) contains compounds that can modulate NADP(+)-dependent oxidation of cortisol by type 1 11beta-hydroxysteroid dehydrogenase (11betaHSD). The objective of this study was to investigate the relationships between levels of the ovarian modulators of type 1 11betaHSD, intra-follicular cortisol:cortisone ratios and the clinical outcome of IVF cycles. METHODS: A single random sample of FF was aspirated from each of 132 patients undergoing gonadotrophin-stimulated IVF. Components of FF, resolved using C18 column chromatography, were evaluated for effects on NADP(+)-dependent cortisol oxidation in rat kidney homogenates. Intra- follicular steroid concentrations were measured by radioimmunoassays. Clinical pregnancies were confirmed by ultrasonography at 6 weeks post-embryo transfer. RESULTS: Levels of the hydrophilic ovarian 11betaHSD stimuli were significantly lower (P<0.0001) and levels of the hydrophobic ovarian 11betaHSD inhibitors were significantly higher (P<0.002) in conception versus non-conception cycles. Intra-follicular cortisol:cortisone ratios increased with the degree of inhibition of 11betaHSD by the hydrophobic FF fractions. FF obtained from conception cycles had significantly higher cortisol:cortisone ratios than samples from non-conception cycles (12.9+/-0.3 versus 8.5+/-0.2, respectively; P<0.0001). CONCLUSIONS: Conception by IVF is associated with elevated intra-follicular cortisol:cortisone ratios, which reflect low levels of ovarian stimuli and/or high levels of ovarian inhibitors of type 1 11betaHSD.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed2002

Ovarian modulators of 11beta-hydroxysteroid dehydrogenase (11betaHSD) activity in follicular fluid from gonadotrophin-stimulated assisted conception cycles.

In the ovary, cortisol-cortisone interconversion is catalysed by isoforms of 11beta-hydroxysteroid dehydrogenase (11betaHSD). The objective of this study was to establish whether human follicular fluid (hFF), obtained after controlled ovarian hyperstimulation, contains paracrine modulators of 11betaHSD activity. Of 274 hFF samples tested for effects in rat kidney homogenates, 206 hFF samples significantly inhibited NADP(+)-dependent oxidation of cortisol within 1 h (by 11-67% of control 11betaHSD activity), whereas 42 hFF samples significantly stimulated 11betaHSD activity (16-210% increase relative to control). Although charcoal-stripping of hFF prevented the inhibition and potentiated the stimulation of NADP(+)-dependent cortisol oxidation in a renal homogenate, effects of individual hFF samples on NADP(+)-dependent cortisol oxidation were independent of intrafollicular progesterone concentrations. Hydrophilic fractions of hFF samples, isolated by C18 column chromatography, stimulated both the NADP(+)-dependent oxidation of cortisol (by 55+/-5%, n=98) and the NADPH-dependent reduction of cortisone (by 86+/-22%, n= 5). In contrast, the hydrophobic fractions of hFF (eluted at 65-85% methanol) inhibited both NADP(+)-dependent 11beta-dehydrogenase and NADPH-dependent 11-ketosteroid reductase activities (by 63+/-2% and 74+/-4%, respectively). None of the C18 column fractions of 50 hFF samples had any significant effect on NAD(+)-dependent 11beta-dehydrogenase activities. The hydrophobic inhibitors of NADP(H)-dependent cortisol-cortisone metabolism did not co-elute with several candidate compounds (prostaglandins E(2) and F(2alpha), cortisol, cortisone, oestradiol, testosterone, progesterone, pregnenolone or cholesterol). Hence, hFF aspirated from women undergoing controlled ovarian hyperstimulation for assisted conception contains both hydrophilic stimuli and hydrophobic inhibitors of glucocorticoid metabolism which appear to be selective for the NADP(H)-dependent, type 1 isoform of 11betaHSD.

11-beta-Hydroxysteroid Dehydrogenase Type 2↗
PubMed1999

The effects of different culture media, glucose, pyridine nucleotides and adenosine on the activity of 11beta-hydroxysteroid dehydrogenase in rat Leydig cells.

11Beta-hydroxysteroid dehydrogenase (11betaHSD) reversibly converts glucocorticoids into inert 11-ketosteroids. The direction of the reaction has been found to vary with the cell type and sub-cellular preparation used. We have investigated if the directionality of 11betaHSD can be influenced by the nature of the culture medium and compounds added during incubation of rat testis Leydig cells. We found that when the cells were cultured in Dulbecco's Modified Eagle Medium (DMEM) that the dehydrogenase (11betaDH) activity was higher than the reductase (11KSR) activity (11betaDH:11KSR ratio approximately 2:1). When glucose was omitted from the DMEM a higher 11betaDH:11KSR ratio (approximately 33:1) was obtained. However, when the cells were cultured in a combination of DMEM/Ham's F12 (1:1, v/v), a ninefold increase in 11KSR activity was obtained whereas 11betaDH activity was inhibited by 64% compared with cells incubated in DMEM alone. Consequently, the predominant activity changed from a dehydrogenase to a reductase (11betaDH:11KSR ratio 1:15). Addition of the individual components of the Ham's F12 medium to DMEM showed that only pyruvate and/or the amino acids were able to mimic the effects of DMEM/Ham's F12. Similar differential effects were found when NAD+, NADH or adenosine were added to the Leydig cells incubated in DMEM (three to fivefold increases and 20-50% decreases in 11KSR and 11betaDH activities, respectively). In contrast, NADP+ was found to increase 11betaDH activity (up to threefold) but NADPH had no effect on 11KSR activity. Cells incubated with DMEM/Ham's F12, NAD+, NADP+ and adenosine were found to have higher ATP levels (four to sixfold) than those incubated in DMEM alone. These results illustrate that the relative 11betaDH and 11KSR activities of 11betaHSD in Leydig cells are markedly and differentially altered by the nature of the incubation medium and compounds added.

11-beta-Hydroxysteroid Dehydrogenase Type 1↗
PubMed1999

Signal transduction involving cyclic AMP-dependent and cyclic AMP-independent mechanisms in the control of steroidogenesis.

The control of steroidogenesis via signal transduction mechanisms involving cAMP-dependent and cAMP-independent mechanisms is reviewed. Several structurally unrelated factors that are potent stimulators of steroidogenesis whose actions do not require cAMP and/or synthesis of proteins have been identified. These include various interleukins, a lipophilic factor from macrophages, a steroidogenic inducing protein from follicular fluid and an imidazole compound, calmidazolium. All of these factors are capable of inducing maximum steroidogenesis. Calcium is required for steroidogenesis in all steroidogenic cells. With the exception of the effects of angiotensin II, there is little evidence for a role of IP3 in the stimulation of the release of calcium from intracellular stores in steroidogenic cells under physiological conditions. There may however, be a cAMP-mediated activation of a plasma membrane calcium channel. Chloride channels that can be regulated by cAMP-dependent and -independent mechanisms, are present in steroidogenic cells. Chloride ions exert a negative effect on steroidogenesis because exclusion of chloride from the extracellular medium markedly enhances cAMP-stimulated steroidogenesis. Arachidonic acid and its lipoxygenase products are involved in the control of steroidogenesis via cAMP mediated processes. An arachidonic acid related thioesterase has been isolated that is activated by ACTH and which may be involved in the release of arachidonic acid. It is concluded that while cAMP is a second messenger for LH/ACTH in the control of steroidogenesis, other signalling systems exist which are potentially equally effective in controlling steroidogenesis. In addition, the action of cAMP requires other signalling pathways involving calcium and chloride ions, as well as arachidonic acid and its lipoxygenase products.

Animals↗
PubMed1999

Relationship between ovarian cortisol:cortisone ratios and the clinical outcome of in vitro fertilization and embryo transfer (IVF-ET).

OBJECTIVE: Previously, we have reported an association between low levels of intraovarian cortisol metabolism, mediated by 11beta-hydroxysteroid dehydrogenase (11betaHSD), and the establishment of pregnancies by in vitro fertilization and embryo transfer (IVF-ET). The objective of the present study was to investigate the relationship between the clinical outcome of IVF-ET and the intraovarian concentrations of cortisol and cortisone and the cortisol:cortisone ratios in random samples of ovarian follicular fluid (FF). DESIGN: Retrospective, double-blind correlation analyses. PATIENTS: FF samples (n = 41) were obtained from 23 women undergoing gonadotrophin-stimulated IVF-ET cycles at the Cardiff Assisted Reproduction Unit. MEASUREMENTS: Clinical pregnancy was confirmed by ultrasonography. Intrafollicular steroid concentrations were measured by radioimmunoassays. RESULTS: Concentrations of both cortisol and cortisone were significantly lower in FF samples obtained from 6 patients that conceived than in samples obtained from 17 patients that did not achieve pregnancy (cortisol (mean +/- SEM) = 304 +/- 29 vs. 407 +/- 26 nmol/l, P = 0. 0411; cortisone = 32 +/- 3 vs. 65 +/- 7 nmol/l, P = 0.0002). Intrafollicular cortisol:cortisone ratios were significantly higher in samples from conception cycles than in those samples obtained from nonconception cycles (9.7 +/- 0.7 vs. 6.9 +/- 0.5, respectively, P = 0.0060). Whereas 5 of 10 women with intrafollicular cortisol:cortisone ratios greater than the outcome-independent mean of 7.7 became pregnant, only 1 of the 13 patients with intrafollicular cortisol:cortisone ratios < 7.7 conceived (chi2 = 5. 247, P = 0.0220). CONCLUSIONS: Concentrations of both cortisol and cortisone were significantly lower in FF samples obtained from patients that conceived by IVF-ET than in those obtained from nonconception cycles. Conception by gonadotrophin-stimulated IVF-ET was associated with an elevated intrafollicular ratio of cortisol:cortisone, consistent with a low level of intraovarian cortisol oxidation by 11betaHSD.

Buserelin↗
PubMed1998

Effect of a dopamine agonist on luteinizing hormone receptors, cyclic AMP production and steroidogenesis in rat Leydig cells.

Dopamine agonists are known to increase the incidence of Leydig cell hyperplasia/adenomas when administered to rats over periods of 1-2 years. We have examined the early changes in factors affecting luteinizing hormone (LH)-controlled signal transduction pathways and steroidogenesis in Leydig cells in vitro after chronic oral administration of one of these dopamine agonists, Mesulergine (CU327-085) (N-(1-6,dimethylergolin-8a-yl)-N',N'-dimethylsulphamide hydrochloride) to Sprague-Dawley (SD) rats. Eight-week-old rats were given this dopamine agonist (2 mg/kg body wt/day) in food for 1, 5, or 12 weeks. The Leydig cells from control and treated rats were purified by elutriation and density gradient centrifugation. The dopamine agonist treatment was found to decrease the specific binding of 125I-human chorionic gonadotrophin (hCG) binding to the Leydig cells: a decrease was detected as early as 1 week after treatment and was more pronounced after 5 and 12 weeks. This was found to be due to a decrease in the LH/hCG receptor numbers and not to a decrease in LH/hCG-receptor binding affinity. Both basal and LH-stimulated cAMP and testosterone production were also decreased; cAMP production was decreased by approximately 50% by all concentrations of LH added whereas testosterone production was only decreased with submaximum stimulating concentrations of LH. The formation of testosterone in response to dibutyryl cAMP was also decreased by approximately 50%, indicating additional lesions in the signal transduction pathway. The addition of the cell permeant 22R-hydroxycholesterol (22R) demonstrated that testosterone but not pregnenolone production was decreased by treatment with the dopamine agonist, thus indicating that the 17 alpha-hydroxylase/C17-20 lyase may have been inhibited. Supporting evidence for this was found because the dopamine agonist also increased aromatase activity in the Leydig cells and thus the potential to produce estrogens; previous studies have shown that estradiol is an inhibitor of the 17-20 lyase enzyme. The addition of the dopamine agonist directly to the Leydig cells did not inhibit cAMP production or testosterone production except at high concentrations. It is concluded that treatment of rats with the dopamine agonist indirectly (i.e., via the pituitary) affects Leydig cell function resulting in a rapid decrease in LH receptors and cAMP and testosterone production. Aromatase activity is increased and thus the capacity to produce estrogens. These early changes in the signal transduction pathways and steroidogenesis may be involved in the Leydig cell hyperplasia/adenoma formation that subsequently occurs.

Administration, Oral↗
PubMed1997

Leukocytes modulate 11beta-hydroxysteroid dehydrogenase (11beta-HSD) activity in human granulosa-lutein cell cultures.

It is well established that there are interactions between the immune and reproductive systems. The ovary contains indigenous macrophages, as well as other classes of leukocytes in smaller numbers. Cytokines secreted by these cells have been shown to have the ability to regulate ovarian steroidogenesis. In the present study, the effect of leukocytes on 11beta-hydroxysteroid dehydrogenase (11beta-HSD) in human granulosa-lutein cells was examined. In addition, individual cytokines were also tested for their ability to regulate this enzyme. The follicular aspirates of patients undergoing IVF treatment were used as a source of granulosa cells. Cells isolated from these aspirates were found to contain between 15 and 60% leukocytes as assessed by flow cytometry (FACS). Leukocytes were removed from the sample preparations by the use of immunomagnetic beads coated with CD45 antibody, which recognises a surface antigen on all classes of leukocyte. Removal of leukocytes significantly decreased the 11beta-HSD activity in the granulosa cells, assayed after 3 days of culture, from 7.3 (2-20) to 3.5 (1-10) pmol cortisone formed/50000 cells/4 h (medians and ranges, n = 15). Addition of IL-5 and IL-6 significantly increased the 11beta-HSD activity in granulosa cell cultures both in the presence and absence of leukocytes. Addition of IL-4 and IFN-gamma increased 11beta-HSD activity only in the leukocyte-depleted granulosa cell cultures, whereas IL-2 had no effect on either of the cultures. The data suggests that leukocytes interact with the ovarian cells through cytokine secretion and/or cell-cell contact to increase the 11beta-HSD activity in human granulosa cells.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1997

Isoforms of 11beta-hydroxysteroid dehydrogenase in human granulosa-lutein cells.

To date, two isoforms of 11beta-hydroxysteroid dehydrogenase (11betaHSD) have been characterized: a low affinity, NADP+-dependent isoform (11betaHSD1) and a high affinity, NAD+-dependent isoform which metabolizes dexamethasone and is inhibited by cortisone (11betaHSD2). Having previously reported a relationship between ovarian 11betaHSD activities and conception in women undergoing in vitro fertilization (IVF-ET), the objective of the present study was to identify which isoforms of 11betaHSD metabolize glucocorticoids in cultures of human granulosa-lutein cells. In both intact cells and cell homogenates, two distinct 11betaHSD activities were identified with differing affinities for cortisol (Km = 490 nM and 2.6 microM). Even at low concentrations, cortisol oxidation was preferentially supported by NADP+ and was independent of NAD+. Although inhibited by the hemisuccinate ester of glycyrrhetinic acid, carbenoxolone, the predominant 11betaHSD activity in intact cells was resistant to end-product inhibition. Intact cells were also able to reduce [3H]cortisone (Km = 190 nM) but did not metabolize [3H]dexamethasone. 11BetaHSD1 mRNA was expressed in 23 of 28 cell cultures whereas 11betaHSD2 mRNA was not expressed in any of the 22 independent cultures studied by reverse transcriptase-polymerase chain reaction (RT-PCR). We conclude that human granulosa-lutein cells express both type 11betaHSD and a novel isoform of this enzyme. While the low affinity 11beta-dehydrogenase and 11-ketosteroid reductase activities exhibit properties consistent with 11betaHSD1, the high affinity 11beta-dehydrogenase differs from 11betaHSD2 in that it is NADP+-dependent, does not metabolize dexamethasone and is resistant to end-product inhibition.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1997

Modulation of steroidogenesis by chloride ions in MA-10 mouse tumor Leydig cells: roles of calcium, protein synthesis, and the steroidogenic acute regulatory protein.

It has previously been shown that omission of extracellular chloride ions during culture of rat Leydig cells markedly enhances LH-stimulated steroidogenesis. In the present study, the mechanisms of the effect of chloride omission on (Bu)2cAMP-stimulated steroidogenesis in MA-10 mouse Leydig tumor cells have been investigated. It was found that chloride omission enhanced progesterone production 2- and 4-fold in the absence and presence, respectively, of submaximally stimulating levels of (Bu)2cAMP (0.1 mM) during incubation for 2 h. This enhancement of stimulation increased continuously with time, because after 6 h, (Bu)2cAMP-stimulated progesterone production was 15-fold higher in the absence of chloride. These effects were not found in the presence of maximum stimulating levels of (Bu)2cAMP (1 mM). Omission of calcium from the incubation medium decreased (Bu)2cAMP-stimulated progesterone production by over 70% in the presence and absence of chloride. Progesterone production was still enhanced by the omission of chloride in the absence of calcium, but the effects were less marked than those in the presence of calcium. Addition of the protein synthesis inhibitor, cycloheximide, completely inhibited (Bu)2cAMP-stimulated, but not basal, steroidogenesis in the absence and presence of chloride ions during 2- and 6-h incubation. Total protein synthesis (measured by the incorporation of [3H]methionine) was 4-fold higher in cells incubated in chloride-free medium compared with that in cells incubated in chloride-replete medium in the presence of 0.1 mM (Bu)2cAMP. No effects were found on basal levels. Several proteins specific to the steroidogenic machinery were quantified in mitochondria isolated from cells incubated with and without chloride by Western blot analysis after separation by PAGE. Omission of chloride increased (4-fold) the level of the steroidogenic acute regulatory (StAR) protein in the cells incubated with (Bu)2cAMP (0.1 mM). There was no increase in either the levels or activities of cytochrome P450 cholesterol side-chain cleavage enzyme (cytP450scc) or 3beta-hydroxysteroid dehydrogenase. No effects were found on the basal level of any of the proteins measured. These results are consistent with a cAMP-dependent regulatory role of chloride ion efflux in the control of steroidogenesis, which requires protein synthesis. It is proposed that this occurs by increases in StAR protein synthesis via a general increase in cAMP-dependent protein synthesis and/or by enhancement of the steroidogenic effects of StAR.

3-Hydroxysteroid Dehydrogenases↗
PubMed1996

Effect of a dopamine agonist on the development of Leydig cell hyperplasia in Sprague-Dawley rats.

Dopamine agonists are known to increase the incidence of Leydig cell hyperplasia/adenomas when administered to rats over periods of 1-2 years. We have examined the early changes in factors affecting Leydig cell growth/hyperplasia after chronic oral administration of one of these dopamine agonists, Mesulergine (CU32-085) [N-(1-6-dimethylergolin-8 alpha-yl)-N,N-dimethylsulphamide hydrochloride), to Sprague-Dawley (SD) rats. Eight-week-old rats were given the dopamine agonist (2 mg/kg body weight/day) in food for 5 or 57 weeks. The dopamine agonist treatment had no significant effect on food intake, body weight, and testis and seminal vesicle size, but significantly decreased testicular interstitial fluid volume at 5 weeks (by 51%). Leydig cells isolated from rats treated with the dopamine agonist for 5 weeks exhibited an increase in the rate of protein synthesis compared with the controls (by 28%). This treatment, however, had no significant effect on the number of Leydig cells or macrophages as assessed by histological examination of testicular sections. Treatment with the dopamine agonist for 57 weeks caused a 36 and 28% increase in the number of Leydig cells and macrophages, respectively. Nodules of Leydig cells, indicating the first signs of tumor development, were present in testes from the 57- but not the 5-week-treated animals or the controls of both groups, although an increase in the number of Leydig cells occurred with aging. Thick-walled arterioles were found in the intertubular spaces of the testis sections from rats treated for 57 weeks. These findings suggest that chronic treatment of male SD rats with the dopamine agonist causes hypertrophy of Leydig cells within 5 weeks (as assessed by [3H]methionine incorporation), followed by hyperplasia within 2 years, prior to the development of Leydig cell adenomas, which occur within 1-2 years after the initiation of treatment.

Adenoma↗
PubMed1996

Follicular variation in ovarian 11 beta-hydroxysteroid dehydrogenase (11 beta HSD) activities: evidence for the paracrine inhibition of 11 beta HSD in human granulosa-lutein cells.

We have previously shown that detectable metabolism of cortisol to cortisone by 11 beta-hydroxysteroid dehydrogenase (11 beta HSD) in human granulosa-lutein cells, pooled for each patient from all aspirated ovarian follicles, is associated with failure to conceive by in vitro fertilization and embryo transfer. The aims of the present study were to assess: (1) the variation in the 11 beta HSD activities of granulosa-lutein cells obtained from individual follicles in relation to oocyte maturity and (2) whether the 11 beta HSD activity of pooled granulosa-lutein cells reflects the 11 beta HSD activities of the individual follicles for a given patient. 11 beta HSD activities were measured in intact cells in serum-free medium by a radiometric conversion assay (100 nmol/l [3H]cortisol to [3H]cortisone). Follicular 11 beta HSD activities ranged from < 10 (undetectable) to 514 pmol/mg protein per 4 h (n = 105 follicles from 12 patients) and did not correlate with oocyte maturity. In three separate patients, the follicular 11 beta HSD activities ranged from < 10 to 117 pmol/mg protein per 4 h (n = 8 follicles), 19 to 514 pmol/mg per 4 h (n = 9) and 60 to 390 pmol/mg per 4 h (n = 8). The 11 beta HSD activities of the corresponding multi-follicular pools of cells were < 10, < 10 and 44 pmol/mg per 4 h respectively, all of which were significantly lower (P < 0.05) than the arithmetic means for the activities in the individual follicles (52, 132 and 215 pmol/mg per 4 h respectively). Likewise, the 11 beta HSD activities of two independent multi-patient pools of cells were significantly lower than the mean values of the 11 beta HSD activities of the appropriate individual patients. We conclude that ovarian 11 beta HSD activity varies between follicles and that co-culture of granulosa-lutein cells with low enzyme activity can suppress the ovarian 11 beta HSD activity in cells from different follicles (or patients) with high rates of cortisol metabolism. Hence, these data indicate the potential for paracrine inhibition of ovarian 11 beta HSD activity in human granulosa-lutein cells.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1995

Characterization of site-directed antibodies to the LH receptor in functionally active gonadal cells and their differential effects on LH-stimulated signal transduction in Leydig tumour (MA10) cells.

In this study site-directed antibodies have been used to investigate the structure/activity relationships of the LH receptor in functionally active gonadal cells. Polyclonal antibodies were raised in rabbits against synthetic peptides corresponding to regions within both the extracellular N-terminal domain (antibodies 1 and 2 against residues 48-65 and 187-206, respectively) and the cytoplasmic C-terminal domain (antibody 3 against residues 622-636) of the LH receptor. Following affinity purification by chromatography on columns of immobilised peptides the antibodies were demonstrated to be peptide specific both by ELISA and by dot-blotting assays. On Western blots of membranes proteins prepared from superovulated rat ovaries, mouse Leydig tumour (MA10) cells, and rat testes, all three antibodies recognised a single broad band of apparent M(r) 95,000-100,000 corresponding to the putative LH receptor. The protein of apparent M(r) 95,000-100,000 also bound 125I-hCG on ligand blots, and binding was displaced by excess unlabelled hCG. The binding of 125I-hCG in the ligand blots was completely inhibited by excess unlabelled hCG. The two N-terminal antibodies (antibodies 1 and 2 (10 micrograms/ml)) also inhibited 125I-hCG binding to a greater extent than the C-terminal antibody (antibody 3 (10 micrograms/ml)). Antibody 1 (1 and 10 micrograms/ml) also potently inhibited the binding of 125I-hCG to MA10 cells. A lesser but still significant inhibition of binding was produced by antibody 2 (with 10 micrograms/ml), whereas at the concentrations tested antibody 3 exerted no greater inhibition than that yielded by pre-immune IgG. At 0.1 micrograms/ml antibody 1 significantly inhibited and at 10 micrograms/ml completely inhibited LH-stimulated cAMP and progesterone production by MA10 cells. With antibody 2, 10 micrograms/ml was required to give a significant inhibition, whereas neither antibody 3 nor pre-immune IgG had a significant effect. The antibodies had no effect on cAMP or progesterone production when added to the MA10 cells in the absence of LH. These results indicate that binding of antibody 1 and, to a lesser extent, antibody 2 interferes with ligand binding which consequently affects signal transduction. In view of the ability of the antibodies to recognise the LH receptors both in the ovary and the testis and in more than one rodent species, and their greater apparent potency than previously available antisera, the anti-peptide antibodies raised in the present study will therefore be useful to study LH receptors in normal, functionally active gonadal cells.

Animals↗
PubMed1995

Ovarian 11 beta-hydroxysteroid dehydrogenase activity is inversely related to the outcome of in vitro fertilization-embryo transfer treatment cycles.

OBJECTIVES: To determine the relationship of ovarian 11 beta-hydroxysteroid dehydrogenase (11 beta-HSD) activity to the outcome of IVF-ET and to establish whether 11 beta-HSD activities vary for a given patient between consecutive treatment cycles. DESIGN: Retrospective correlation analysis. SETTING: Patients were treated and ovarian 11 beta-HSD activities were measured at independent fertility and biochemistry departments, respectively. PATIENTS: Random series of 137 women undergoing a total of 172 treatment cycles for IVF-ET. INTERVENTIONS: Subcutaneous buserelin acetate plus IM hMG and hCG for controlled ovarian hyperstimulation; blood samples were collected for plasma LH and steroid determinations. MAIN OUTCOME MEASURES: Presence or absence of detectable ovarian 11 beta-HSD activity, oocyte fertilization rates, and the identification of clinical pregnancies. RESULTS: None of the 101 cycles associated with 11 beta-HSD positive granulosa-lutein cells resulted in clinical pregnancies, whereas the pregnancy rate for the 71 patients with undetectable ovarian 11 beta-HSD activity was 63.4% per cycle. The incidence of total fertilization failure was lower and the median oocyte fertilization rate was higher in cycles characterized by 11 beta-HSD-negative cells. Plasma concentrations of LH and E2 were higher in cycles yielding 11 beta-HSD-negative cells, whereas plasma P, patient age, and the number of oocytes retrieved did not relate to ovarian 11 beta-HSD activity. For the 35 patients studied in repeat cycles, ovarian 11 beta-HSD activities did not relate to those in the initial cycles. CONCLUSIONS: Ovarian 11 beta-HSD activity is associated with failure to conceive by IVF-ET. For a given patient, ovarian 11 beta-HSD activity varies between consecutive treatment cycles. Hence, ovarian 11 beta-HSD activities may predict the outcome of this assisted reproduction protocol independently in each treatment cycle.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1994

A working hypothesis for the regulation of steroidogenesis and germ cell development in the gonads by glucocorticoids and 11 beta-hydroxysteroid dehydrogenase (11 beta HSD).

The relationship between glucocorticoid secretion from the adrenal gland and gonadal function has previously been attributed to central inhibition by the adrenal steroids of pituitary gonadotropin output. This review focuses on the direct actions of glucocorticoids within the gonads, including positive effects on germ cell maturation and both positive and negative effects on the stimulation of gonadal steroidogenesis by LH and FSH. In addition, we address the role in the gonads of 11 beta-hydroxysteroid dehydrogenase (11 beta HSD), which interconverts the glucocorticoids with their inactive 11-ketosteroid derivatives. To date, two isoforms of 11 beta HSD have been described. 11 beta HSD1, purified and cloned from the liver, has a relatively low affinity for glucocorticoids and acts instead as an 11-oxoreductase, whereas the high affinity 11 beta HSD2, first identified in the kidney, acts as an efficient 11 beta-dehydrogenase to inactivate physiological concentrations of glucocorticoid. We propose that in the gonads, 11 beta HSD1 promotes the positive effects of glucocorticoids on germ cell maturation (by increasing the local concentration of active glucocorticoids), whereas a high affinity 11 beta-dehydrogenase activity, consistent with that of 11 beta HSD2, inactivates glucocorticoids and so protects luteal cells from the inhibitory effects of these steroids during the luteal phase of the ovarian cycle.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1993

Ovarian 11 beta-hydroxysteroid dehydrogenase: potential predictor of conception by in-vitro fertilisation and embryo transfer.

Cortisol is converted to the inactive glucocorticoid, cortisone, in several tissues by 11 beta-hydroxysteroid dehydrogenase (11 beta HSD). We have recently measured 11 beta HSD activity in cultured human granulosa-lutein cells recovered from patients undergoing in-vitro fertilisation and embryo transfer (IVF-ET). We now report an association between the outcome of IVF-ET and 11 beta HSD activity in these cells. Of the 64 patients studied, 32 had detectable 11 beta HSD activity and none became pregnant; whereas 76% of the remaining "11 beta HSD-negative" patients achieved pregnancies. Hence 11 beta HSD activity may predict the outcome of IVF-ET.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1993

Direct inhibition of ovarian steroidogenesis by cortisol and the modulatory role of 11 beta-hydroxysteroid dehydrogenase.

OBJECTIVE: The association of adrenal hyperactivity with ovarian dysfunction may involve direct inhibition of ovarian steroidogenesis by glucocorticoids. Therefore, the objectives of this study were to investigate the direct effects of cortisol on luteinizing hormone (LH) action in human granulosa-lutein cells and the modulation of this interaction by ovarian 11 beta-hydroxysteroid dehydrogenase (11 beta HSD). DESIGN AND PATIENTS: Effects were investigated in cultured human granulosa-lutein cells isolated from the follicular aspirates of 14 patients undergoing oocyte collection for in-vitro fertilization and embryo transfer. MEASUREMENTS: Pregnenolone production and 3H-cortisol oxidation to 3H-cortisone (11 beta HSD activity) by cultured cells were measured. RESULTS: In cells from nine (of 14) patients, cortisol inhibited LH-stimulated steroidogenesis in a concentration dependent manner with an ID50 of 1250 +/- SEM 377 nmol/l. In these cultures, the 11 beta HSD activities were high (133 +/- SEM 23 pmol/mg protein/4h) and inhibition of the enzyme with carbenoxolone potentiated the action of cortisol. Conversely, cells from the remaining five patients lacked detectable 11 beta HSD activity and exhibited an increased sensitivity to the inhibitory action of cortisol (ID50 = 158 +/- SEM 41 nmol/l in the absence of carbenoxolone). CONCLUSIONS: Cortisol acts directly in human granulosa-lutein cells to inhibit the support of steroidogenesis by LH and this interaction is modulated by ovarian 11 beta HSD in the majority of patients.

11-beta-Hydroxysteroid Dehydrogenases↗