Dehydroepiandrosterone Sulfate Secretion Research Articles

Aging induces region-specific dysregulation of hormone synthesis in the primate adrenal gland.

Adrenal glands, vital for steroid secretion and the regulation of metabolism, stress responses and immune activation, experience age-related decline, impacting systemic health. However, the regulatory mechanisms underlying adrenal aging remain largely uninvestigated. Here we established a single-nucleus transcriptomic atlas of both young and aged primate suprarenal glands, identifying lipid metabolism and steroidogenic pathways as core processes impacted by aging. We found dysregulation in centripetal adrenocortical differentiation in aged adrenal tissues and cells in the zona reticularis region, responsible for producing dehydroepiandrosterone sulfate (DHEA-S), were highly susceptible to aging, reflected by senescence, exhaustion and disturbed hormone production. Remarkably, LDLR was downregulated in all cell types of the outer cortex, and its targeted inactivation in human adrenal cells compromised cholesterol uptake and secretion of dehydroepiandrosterone sulfate, as observed in aged primate adrenal glands. Our study provides crucial insights into endocrine physiology, holding therapeutic promise for addressing aging-related adrenal insufficiency and delaying systemic aging.

Job demands and DHEA-S levels: a study on healthcare workers.

The intricate interplay between work-related stress and its physiological impact has drawn extensive research attention. Dehydroepiandrosterone sulphate (DHEA-S) emerges as a potential biomarker reflecting stress-related endocrine changes. This cross-sectional study aimed to examine the association between job demands and DHEA-S levels among healthcare workers. The study also explored potential correlations between DHEA-S levels and psychophysical symptoms commonly linked to work-related stress. A sample of 488 healthcare workers from a local health authority participated. Job demands were measured using the Demands scale of the Health and Safety Management Standards Indicator Tool. DHEA-S levels and symptom prevalence were assessed through serum analysis and questionnaires, respectively. Workers exposed to high job demands exhibited significantly lower DHEA-S levels compared to those with low job demands. Psychophysical symptoms, including sleep disorders, depression, and headache, were more prevalent in the high-demands group. DHEA-S levels showed significant negative correlations with the prevalence of all considered symptoms. The study shows the inverse relationship between job demands and DHEA-S levels among healthcare workers, indicating that high job demands correlate with reduced DHEA-S secretion and increased symptom prevalence. The findings suggest DHEA-S as a potential biomarker for assessing the physiological consequences of work-related stress. Proactive interventions in managing job demands are crucial for promoting employee well-being and productivity in demanding work environments. By recognizing DHEA-S as a stress biomarker, organizations can effectively address stress-related health risks and implement targeted interventions for enhancing employees' overall health and work performance.

Treatment with Intravenous Methylprednisolone in Patients with Graves' Orbitopathy Significantly Affects Adrenal Function: Assessment of Serum, Salivary Cortisol and Serum Dehydroepiandrosterone Sulfate.

Treatment of active, moderate-to-severe Graves’ orbitopathy (GO) is the administration of intravenous methylprednisolone (IVMP). IVMP may be followed by additional therapy with oral prednisone. The aim of this study was to analyze the impact of IVMP on adrenal function by evaluation of serum, salivary cortisol and serum dehydroepiandrosterone sulfate (DHEA-S). Fourteen patients received IVMP treatment (cumulative dose of 4.5 g in 12 weekly infusions) followed by oral prednisone (for three months). All patients showed normal adrenal function before the 12th IVMP pulse and one patient was diagnosed with secondary adrenal insufficiency (AI) after prednisone treatment. DHEA-S was significantly lower before the 12th IVMP pulse and after oral prednisone (p = 0.015 and p = 0.00002, respectively) in comparison to evaluation before therapy. DHEA-S levels were below the reference range in one and three patients before the 12th IVMP pulse and after prednisone therapy, respectively. We observed decreased serum (p = 0.05) and salivary (p = 0.011) cortisol levels after oral prednisone therapy in comparison to evaluation before therapy. Treatment with IVMP in a cumulative dose of 4.5 g affects adrenal function, causing more severe impairment of DHEA-S secretion than that of cortisol but does not cause secondary AI. Additional therapy with oral glucocorticoids after IVMP can cause secondary AI.

Interleukin-6 increases adrenal androgen release by regulating the expression of steroidogenic proteins in NCI-H295R cells.

The purpose of this study was to investigate the potential mechanism of interleukin-6 (IL-6) on the stimulation of excessive androgen secretion in human NCI-H295R adrenocortical cells. We performed transcriptome sequencing of cancer and paracancerous tissues obtained from functional adrenal cortical adenomas. The secretion of dehydroepiandrosterone sulfate (DHEAS) in NCI-H295R cells was detected by a chemiluminescence assay. The expression of messenger RNA (mRNA) was detected by real-time polymerase chain reaction and that of protein was detected by western blotting. The expression of secretogranin II (SCG2) and IL-6 were significantly increased in cancer tissues. Upregulation of mRNA and protein levels of AKR1C3, CYP11A, CYP17A1, 3βHSD, and SULT2A1 was observed after stimulation with IL-6. IL-6 could also increase the expression of StAR mRNA and proteins. Our results suggest that IL-6 can promote androgen secretion by regulating the expression of genes related to androgen pathways.

Особенности секреции андрогенов у женщин с угрозой самопроизвольного выкидыша

Objective. To assess androgen secretion and its possible effect on pregnancy in women with threatened miscarriage in the first trimester. Patients and methods. This prospective observational study included 120 pregnant women divided into four groups. Group I comprised 32 patients with threatened miscarriage and hyperandrogenism who received corticosteroids; Group II was composed of 28 patients with threatened miscarriage and hyperandrogenism who did not receive corticosteroids; Group III included 30 patients with threatened miscarriage and no hyperandrogenism; and Group IV comprised 30 women with normal pregnancy. Serum levels of dehydroepiandrosterone sulfate (DHEA-S), 17-hydroxyprogesterone (17-OHP), and total testosterone were measured on the following weeks of gestation: 5–8, 9–12, 13–18, 19–24, and 25–32. We also evaluated clinical outcomes of pregnancy. Results. We observed no significant differences in 17-ОНР and DHEA-S secretion between women from Group III and controls. Patients from Group II demonstrated higher hormone levels than controls; however, their dynamics of 17-ОНР and testosterone secretion was similar to that in women without hyperandrogenism, so their DHEA-S levels decreased and reached control values by the third trimester. Corticosteroids reduced 17-ONR secretion in the second and third trimesters and DHEA-S secretion in the third trimester. Women receiving corticosteroids demonstrated the poorest clinical pregnancy outcomes. Conclusion. Hyperandrogenism should be considered as one of the risk factors for poor pregnancy outcomes. Administration of corticosteroids to reduce androgen levels impairs normal dynamics of their secretion, does not improve pregnancy outcomes, and is potentially harmful; therefore, these drugs should not be used for such purposes. Key words: pregnancy, hyperandrogenism, corticosteroid therapy, pregnancy outcomes, pregnancy loss, androgen secretion, threatened miscarriage

Adrenal Function in Adolescence is Related to Intrauterine and Postnatal Growth.

Background and objectives: Intrauterine growth restriction is thought to be implicated in long-term programming of hypothalamic–pituitary–adrenal axis activity. We investigated adrenal function in adolescents born small for gestational age (SGA) in relation to their postnatal growth and cardiovascular parameters. Materials and Methods: Anthropometric parameters, blood pressure, heart rate, dehydroepiandrosterone sulfate (DHEAS), and cortisol levels were assessed in 102 adolescents aged 11–14 years followed from birth (47 SGA and 55 born appropriate for gestational age (AGA)). Results: Mean DHEAS levels were higher in SGA adolescents with catch-up growth (SGACU+), compared with AGA. Second-year height velocity and body mass index (BMI) gain during preschool years were positively related to DHEAS levels. Morning cortisol levels and systolic and diastolic blood pressure were higher in SGA adolescents without catch-up growth (SGACU−) compared with AGA. Second-year BMI gain was inversely, and 2–12 years increase in subscapular skinfold thickness was directly associated with cortisol levels. Size at birth and postnatal growth explained 47.8% and 38.2% of variation in DHEAS and cortisol levels, respectively. Conclusion: Adrenal function in adolescence is affected by prenatal and postnatal growth: small size at birth with postnatal catch-up growth is related to higher DHEAS secretion, whereas increased cortisol levels and blood pressure are higher in short SGA adolescents.

Involvement of serum dehydroepiandrosterone sulfate in erythropoietic activity

Purpose The aim of the present study was to determine how age-related decreases in serum dehydroepiandrosterone sulfate (DHEAS) levels affect various physiological processes. Methods We retrospectively reviewed the medical records of patients in whom serum DHEAS levels were measured in our department and assessed the relationships between serum DHEAS levels and various patient characteristics. Results Among the 149 patients included in our analysis (mean age: 52.7 ± 17.6 years, range: 15–84 years), 54 (36.2%) were men. Serum DHEAS levels inversely correlated with age in men (R = −0.810, p < .01) and to a lesser extent in women (R = −0.391, p < .01). Of note, there were significant positive associations between DHEAS levels and hemoglobin (R = 0.317, p ≤ .01) and hematocrit (R = 0.324, p ≤ .01) levels, which were observed in men, but not in women. Conclusions Our study suggests that reduced DHEAS secretion inhibits erythropoietic activity in aging men, perhaps owing to the erythropoietic androgenic actions of DHEAS. Importantly, it suggests that the age-associated decline in DHEAS secretion might decrease erythropoietic activity in aging men. It is also possible that the adrenal cortex, the source of DHEAS, is dysfunctional in anemic men.

Estrogen Regulation of Fetal Adrenal Cortical Zone-Specific Development in the Nonhuman Primate Impacts Adrenal Production of Androgen and Cortisol and Response to ACTH in Females in Adulthood.

We showed that the volume of the fetal zone of the fetal adrenal gland and serum dehydroepiandrosterone sulfate (DHAS) levels at term were increased in baboons in which estradiol levels were suppressed by treatment with aromatase inhibitor 4,4-[1,2,3-triazol-1yl-methylene] bis-benzonitrite (letrozole). The fetal zone remodels postnatally into the reticular zone and DHAS production, and serum levels decline with age. Therefore, we determined whether the trajectory of reticular zone DHAS secretion and response to ACTH were altered in offspring deprived of estrogen in utero. Female offspring were delivered to baboons untreated or treated daily throughout the second half of gestation with letrozole (estradiol reduced >95%) or letrozole plus estradiol and cortisol and DHAS determined in blood samples obtained bimonthly between 4 and 125 months and after iv bolus of ACTH. The slope/rate of decline in serum DHAS with advancing age was greater (P < .01) in letrozole-treated (-0.54 ± 0.005) than untreated (-0.32 ± 0.003) baboons and partially restored by letrozole-estradiol (-0.43 ± 0.004). Serum cortisol was similar and relatively constant in all offspring. Moreover, in letrozole-treated offspring, serum DHAS at 61-66, 67-95, and 96-125 months were lower (P < .05), and cortisol to DHAS ratio was greater (P < .05) than in untreated offspring. ACTH at high level increased cortisol and DHAS in untreated baboons and cortisol but not DHAS in letrozole-treated offspring. We propose that postnatal development of the primate adrenal cortex, including the decline in reticular zone DHAS production, response to ACTH and maintenance of cortisol to DHAS ratio with advancing age is modulated by exposure of the fetal adrenal to estradiol.

The effect of mitotane on viability, steroidogenesis and gene expression in NCI-H295R adrenocortical cells

Mitotane, also known as o,p'‑DDD or (RS)‑1‑chl-oro‑2‑[2,2‑dichloro‑1‑(4‑chlorophenyl)‑ethyl]‑benzene, is an adrenal cortex-specific cytotoxic drug used in the therapy of adrenocortical carcinoma (ACC). The drug also inhibits steroidogenesis, however, the mechanisms of its anticancer and antisteroidogenic effects remain unknown. At present, data on the impact of mitotane on cell viability and the regulation of genes encoding proteins associated with steroids synthesis in the adrenal cortex, including cortisol and dehydroepiandrosterone sulfate (DHEAS), are limited and contradictory. In the present study, the effect of 24‑h mitotane treatment on viability of the ACC cell line, NCI‑H295R, was analyzed, identifying a decrease in cell viability and an increase in caspase‑3 and ‑7 activities. Mitotane treatment also led to decreased cortisol and DHEAS concentration in the culture media. Concomitantly, mitotane resulted in decreased mRNA levels of two cytochromes P450(CYP11A1 and CYP17A1), mRNAs encoding proteins involved in the synthesis of cortisol and DHEAS. Mitotane did not affect mRNA levels of cyclin dependent kinase inhibitor 1A (encoding p21) and MYC (encoding cMyc). cMyc and p21 are key transcription factors associated with cell cycle regulation. However, mitotane inhibited expression of transforming growth factor β1 gene, encoding a potent inhibitor of cell proliferation and steroidogenesis. PRKAR1A, a protein kinase A regulatory subunit, is involved in the activation of steroidogenesis. PRKAR1A mRNA levels were reduced following 24‑h treatment with mitotane. Results indicate that mitotane markedly inhibited expression of genes involved in steroidogenesis, secretion of cortisol and DHEAS. Reduced expression of TGFB1 cannot account fully for the effect of mitotane on CYP11A1 and CYP17A1. We hypothesized that reduced viability of NCI‑H295R cells in the presence of mitotane may be a result of apoptosis triggered by increased caspase‑3 and ‑7 activities. Since p21 and cMyc mRNA levels were stable in the presence of mitotane, the mechanism by which caspase‑3 and ‑7 are induced remains unknown.

Acute and chronic stress increase DHEAS concentrations in rhesus monkeys

Most studies on the stress-responsiveness of the hypothalamic-pituitary-adrenal (HPA) axis have focused on glucocorticoids, while few studies have investigated the adrenal secretion of dehydroepiandrosterone sulfate (DHEAS), which is unique to primates. Monkeys were chair-restrained for 2h per day for seven consecutive days, and blood samples were collected upon placement in the chair, and at 15, 30, 60 and 120 min later. Like cortisol, DHEAS concentrations increased throughout the initial session of chair restraint (acute stress). Unlike the cortisol response, which decreased after repeated exposure to the stressor, the DHEAS response was sustained throughout the seventh session of restraint (chronic stress) and response to the seventh session of restraint did not differ from the DHEAS response to the initial session. Like cortisol, DHEAS concentrations showed a diurnal rhythm with higher concentrations in the morning compared to the evening and a decrease in response to dexamethasone (DEX) administration. After repeated exposure to the stressor, the suppression of DHEAS in response to dexamethasone was more complete, suggesting an increase in negative feedback sensitivity. These data show that DHEAS concentrations increase in response to both acute and chronic (repeated) stress and provide another measure of HPA activity that parallels cortisol during acute responses to stress but diverges in chronic or repeated stress.

Effects of Chromium(III) Picolinate on Cortisol and DHEAs Secretion in H295R Human Adrenocortical Cells

Dietary chromium(III) picolinate (CrPic) effects on circulating steroid hormones have been reported in various experimental animals. However, direct effects of CrPic on adrenocortical steroidogenesis are uncertain. Therefore, the objective was to determine the effects of CrPic on cortisol and dehydroepiandrosterone sulfate (DHEAs) secretion from H295R cells. In experiment 1, a 24-h exposure to CrPic (0 to 200 microM) had both linear (p < 0.001) and quadratic (p < 0.001) effects on cortisol secretion from forskolin-stimulated cells with the highest cortisol secretion at 0.1 microM of CrPic and the lowest at 200 microM of CrPic. In experiment 2, a 48-h exposure to CrPic (200 microM) decreased cortisol (p < 0.07) release from forskolin-stimulated cells during a 24-h collection period. In experiment 3, a 48-h exposure to CrPic (100 microM) decreased cortisol (p < 0.05) and DHEAs (p < 0.01) from forskolin-stimulated cells during a 24-h sampling period. In experiment 4, a 24-h exposure to forskolin followed by a 24-h exposure to both forskolin and CrPic (100 and 200 microM) decreased both cortisol and DHEAs secretion (p < 0.01). This study suggests that at high concentrations, CrPic inhibits aspects of steroidogenesis in agonist-stimulated adrenocortical cells.

Effects of mirtazapine on dehydroepiandrosterone-sulfate and cortisol plasma concentrations in depressed patients

Background Among the neuroactive steroids, dehydroepiandrosterone sulfate (DHEA-S) is at least in part produced in the adrenal gland and is therefore under the control of the hypothalamic–pituitary–adrenocortical (HPA)-system. In the present study, the impact of mirtazapine on DHEA-S and cortisol (COR) levels was investigated in relation to clinical response in depressed patients. Methods A total of 23 inpatients suffering from a major depressive episode (DSM-IV criteria) underwent 5-week treatment with mirtazapine (45 mg/day). Plasma samples were taken weekly at 0800 h and quantified for COR and DHEA-S levels. Results Mirtazapine significantly reduced both COR and DHEA-S concentrations, but had no impact on the COR/DHEA-S ratio. The percentage decrease of DHEA-S, but not that of COR was significantly and positively correlated with the percentage reduction in the sum score of the Hamilton Depression Rating Scale at week 5, suggesting a relationship between DHEA-S reduction and clinical efficacy of mirtazapine. There was a significant positive correlation between the decline in COR and DHEA-S levels. Conclusions Apparently, the decrease in COR and DHEA-S concentrations conjointly reflects an attenuating impact of mirtazapine on HPA axis activity, thereby decreasing the adrenal secretion of COR and DHEA-S.

Dehydroepiandrosterone levels vary according as heart failure condition in patients with idiopathic dilated cardiomyopathy

The secretion of dehydroepiandrosterone sulfate (DHEAS) decreases with age, and the incidence of heart failure rises in the elderly population. We measured plasma DHEAS levels in 50 male patients (mean 66.7 ± 9.1 years old) with congestive heart failure due to idiopathic dilated cardiomyopathy before and after treatment. The study included 50 age-matched control subjects with coronary spastic angina (mean 65.5 ± 8.8 years old). DHEAS levels were significantly lower in patients with congestive heart failure than in controls (82.2 ± 9.9 vs. 122.7 ± 18.6 µg/dL, respectively, p < 0.01), whereas there was no difference in cortisol levels between the 2 groups. After 3 months of treatment, NYHA functional class improved in all patients, and DHEAS levels increased (from 82.2 ± 9.9 to 106.2 ± 21.1 µg/dL, p < 0.01). DHEAS levels vary according as heart failure condition in patients with idiopathic dilated cardiomyopathy.

Relationship Between Low Levels of Anabolic Hormones and 6-Year Mortality in Older Men&lt;subtitle&gt;The Aging in the Chianti Area (InCHIANTI) Study&lt;/subtitle&gt;

Aging in men is characterized by a progressive decline in levels of anabolic hormones, such as testosterone, insulinlike growth factor 1 (IGF-1), and dehydroepiandrosterone sulfate (DHEA-S). We hypothesized that in older men a parallel age-associated decline in bioavailable testosterone, IGF-1, and DHEA-S secretion is associated with higher mortality independent of potential confounders. Testosterone, IGF-1, DHEA-S, and demographic features were evaluated in a representative sample of 410 men 65 years and older enrolled in the Aging in the Chianti Area (InCHIANTI) study. A total of 126 men died during the 6-year follow-up. Thresholds for lowest-quartile definitions were 70 ng/dL (to convert to nanomoles per liter, multiply by 0.0347) for bioavailable testosterone, 63.9 ng/mL (to convert to nanomoles per liter, multiply by 0.131) for total IGF-1, and 50 microg/dL (to convert to micromoles per liter, multiply by 0.027) for DHEA-S. Men were divided into 4 groups: no hormone in the lowest quartile (reference) and 1, 2, and 3 hormones in the lowest quartiles. Kaplan-Meier survival and Cox proportional hazards models adjusted for confounders were used in the analysis. Compared with men with levels of all 3 hormones above the lowest quartiles, having 1, 2, and 3 dysregulated hormones was associated with hazard ratios for mortality of 1.47 (95% confidence interval [CI], 0.88-2.44), 1.85 (95% CI, 1.04-3.30), and 2.29 (95% CI, 1.12-4.68), respectively (test for trend, P <.001). In the fully adjusted analysis, only men with 3 anabolic hormone deficiencies had a significant increase in mortality (hazard ratio, 2.44; 95% CI, 1.09-5.46 (test for trend, P <.001). Age-associated decline in anabolic hormone levels is a strong independent predictor of mortality in older men. Having multiple hormonal deficiencies rather than a deficiency in a single anabolic hormone is a robust biomarker of health status in older persons.

Increased Steroidogenic Factor-1 Dosage Triggers Adrenocortical Cell Proliferation and Cancer

Steroidogenic factor-1 (SF-1/Ad4BP; NR5A1), a nuclear receptor transcription factor, has a pivotal role in adrenal and gonadal development in humans and mice. A frequent feature of childhood adrenocortical tumors is SF-1 amplification and overexpression. Here we show that an increased SF-1 dosage can by itself augment human adrenocortical cell proliferation through concerted actions on the cell cycle and apoptosis. This effect is dependent on an intact SF-1 transcriptional activity. Gene expression profiling showed that an increased SF-1 dosage regulates transcripts involved in steroid metabolism, the cell cycle, apoptosis, and cell adhesion to the extracellular matrix. Consistent with these results, increased SF-1 levels selectively modulate the steroid secretion profile of adrenocortical cells, reducing cortisol and aldosterone production and maintaining dehydroepiandrosterone sulfate secretion. As a model to understand the mechanisms of transcriptional regulation by increased SF-1 dosage, we studied FATE1, coding for a cancer-testis antigen implicated in the control of cell proliferation. Increased SF-1 levels increase its binding to a consensus site in FATE1 promoter and stimulate its activity through modulation of the recruitment of specific cofactors. On the other hand, sphingosine, which can compete with phospholipids for binding to SF-1, had no effect on the SF-1 dosage-dependent increase of adrenocortical cell proliferation and expression of the FATE1 promoter. In mice, increased Sf-1 dosage produces adrenocortical hyperplasia and formation of tumors expressing gonadal markers (Amh, Gata-4), which originate from the subcapsular region of the adrenal cortex. Gene expression profiling revealed that genes involved in cell adhesion and the immune response and transcription factor signal transducer and activator of transcription-3 (Stat3) are differentially expressed in Sf-1 transgenic mouse adrenals compared with wild-type adrenals. Our studies reveal a critical role for SF-1 dosage in adrenocortical tumorigenesis and constitute a rationale for the development of drugs targeting SF-1 transcriptional activity for adrenocortical tumor therapy.

Stress reducing regulative effects of integrated mental training with self‐hypnosis on the secretion of dehydroepiandrosterone sulfate (DHEA‐S) and cortisol in plasma: a pilot study

AbstractEarly observations have shown that preventive educational efforts towards experience of stress entail advantages in terms of improved well‐being, health and performance. We examined the influence in healthy subjects of self‐hypnosis and mental training taught in group sessions, with individual training in between according to a standard protocol, and integrated in daily life and work for six months, on the alteration in plasma concentration of the stress hormone cortisol and the ‘anti ageing’ hormone dehydroepiandrosterone sulfate (DHEA‐S). DHEA‐S and cortisol were analysed in twelve healthy men and women, with six individuals randomly divided equally between experimental and control groups. In the experimental group DHEA‐S was significantly increased by 16% (P &lt; 0.05), whereas plasma DHEA‐S in controls followed an expected age‐related decline. Cortisol was reduced by 12.3% (P &lt; 0.05) in the experimental group, but remained unchanged in the control group. The ratio between plasma concentrations of DHEA‐S and cortisol that reflects stress‐related alteration in the adrenal secretion between androgens and glucocorticoids, increased significantly by 27.8% (P &lt; 0.05) in the experimental group with a reduction of 8,2% in controls. The increase in plasma DHEA‐S under experimental conditions was equivalent to a range normally found in individuals 5 to 10 years younger. Copyright © 2006 British Society of Experimental &amp; Clinical Hypnosis. Published by John Wiley &amp; Sons, Ltd.

Effect of chromium (III) picolinate on cortisol and DHEAs secretion in H295R human adrenocortical cells

Dietary chromium (III) picolinate (CP) effects on steroid hormones have been reported in various experimental animals. However, the effects of CP on adrenal steroidogenesis are uncertain. Therefore, the objective of our study was to determine the effects of CP on cortisol and dehydroepiandrosterone sulfate (DHEAs) secretion from H295R cells, a cell line derived from human adrenocortical cells. In experiment 1, 24 hr of exposure to CP dose dependently (0, 0.1, 1, 10, 100, and 200 μM) increased cortisol secretion (p<0.05). When forskolin (10 μM) was included in the incubation media, CP had both a linear (p<0.001) and quadratic (p<0.001) effect on cortisol secretion with the highest cortisol secretion 0.1 μM of CP and the lowest at 200 μM of CP. In experiment 2, 48 hr exposure to CP (200 μM) decreased cortisol (p<0.05) release from forskolin-stimulated cells during a 24 hr of collection period. Additionally, 1 μM of CP increased DHEAs (p<0.05) secretion from these cells. In experiment 3, 48 hr exposure to CP (100 μM) decreased cortisol (p<0.05) and DHEAs (p<0.01) from forskolin-stimulated cells during a 24 hr collection period. In experiment 4, 24 hr exposure to forskolin followed by 24 hr exposure to both froskolin and CP (100 μM) also decreased both cortisol (p<0.001) and DHEAs (p<0.001) secretion. This study suggests that at high concentrations, CP inhibits aspects of steroidogenesis in agonist-stimulated adrenocortical cells.

DNA methylation affects cell proliferation, cortisol secretion and steroidogenic gene expression in human adrenocortical NCI-H295R cells

Aberrant DNA methylation may be involved in human adrenocortical tumorigenesis, which is often accompanied by abnormal hormone production. In this study, we aimed to clarify the effects of DNA methylation on steroidogenesis using the human adrenocortical NCI-H295R cell line as a model. Treatment with the DNA methylation inhibitor 5-aza-2'-deoxycytidine (Azad; 10 microM for 7 days) decreased the proliferation rate to approximately 20% and the cell number to 60% of the control, with a simultaneous increase in the expression of the cyclin-dependent kinase inhibitor p57(KIP2) gene. In addition, Azad treatment increased cortisol secretion dose and time dependently, whereas dehydroepiandrosterone sulfate secretion was not affected. Azad treatment decreased basal and (Bu)2cAMP-induced expression of low- and high-density lipoprotein receptor, steroidogenic acute regulatory protein (StAR), cholesterol side-chain cleavage enzyme, steroid 17alpha-hydroxylase/17,20-lyase and steroid 21-hydroxylase mRNA, as well as the StAR protein level. In contrast, Azad treatment increased the basal expression of steroid 11beta-hydroxylase and 3beta-hydroxysteroid dehydrogenase/Delta5-Delta4-isomerase genes, although it inhibited the (Bu)2cAMP-induced expression of these two genes. The expression of steroidogenic factor-1 (SF-1) and DAX-1 (dosage-sensitive sex reversal-adrenal hypoplasia congenita critical region on the X-chromosome 1) genes (both harboring putative CpG islands in their promoters) and the methylation degree of the HpaII recognition site(s) in the SF-1 gene promoter region were reduced by Azad treatment. The immunostaining pattern of the methyl-CpG-binding protein MeCP2 was also modified by Azad treatment. These results suggest that DNA methylation may be implicated in the regulation of cell proliferation and steroidogenesis in human adrenocortical cells.

Functional luteinizing hormone/chorionic gonadotropin receptors in human adrenal cortical H295R cells.

Previous studies have suggested that activation of normal human adrenal and adrenal tumor luteinizing hormone (LH)/chorionic gonadotropin (hCG) receptors results in an increased secretion of steroid hormones. Since it is not feasible to test this suggestion on normal human adrenal cells, we used human adrenal cortical carcinoma H295R cells, which are similar in some respects to normal adrenal cortical cells. These cells contained LH/hCG receptor transcripts and receptor protein that can bind (125)I-hCG in a hormone-specific manner. Culturing the cells with highly purified hCG resulted in a time- and dose-dependent significant increase in dehydroepiandrosterone sulfate (DHEAS) secretion as compared with the controls. The DHEAS response was hormone as well as steroid specific. Since hCG treatment did not increase DHEA secretion, we suspected that the hCG might increase DHEA sulfotransferase (ST). Consistent with this possibility, hCG treatment increased steady-state DHEA-ST mRNA levels. The hCG effects require its receptors, as inhibition of their synthesis by treatment with antisense phosphorothioate oligodeoxynucleotides (ODN) made from the LH/hCG receptor sequence resulted in loss of DHEA-ST and DHEAS responses. The findings that 1) hCG treatment increased cAMP levels and activated protein kinase A (PKA), 2) 8-bromo cAMP mimicked hCG, and 3) blocking PKA activation prevented hCG as well as 8-bromo cAMP from increasing both DHEA-ST mRNA and DHEAS levels suggested that cAMP/PKA signaling was involved in the hCG actions. In conclusion, H295R cells contain LH/hCG receptors, which are coupled to increasing DHEAS secretion through upregulating the ST enzyme mRNA level. This action is mediated by the cAMP/PKA signaling pathway. These findings support the concept that adrenal function in normal and pathological conditions could be influenced by LH and hCG.

Sex differences due to dehydroepiandrosterone (DHEA) feeding affecting dehydroepiandrosterone sulfate secretion in golden Syrian hamsters.

The metabolism of orally administered dehydroepiandrosterone (DHEA) by male and female golden Syrian hamsters was examined by quantification of DHEA and dehydroepiandrosterone sulfate (DHEAS) in gallbladder bile, urine and feces using high-performance liquid chromatography (HPLC). Plasma levels of DHEA and DHEAS were also determined by radioimmunoassay (RIA). After 5 days of oral DHEA administration (100 mg/kg body weight twice a day), RIA showed that plasma levels of DHEA and DHEAS were increased approximately 3-6 and 4-5 times, respectively, compared to controls. More than 95 % of circulating DHEA (S) in the peripheral blood was DHEAS. There was no significant sex difference in DHEAS plasma levels between male and female animals in the DHEA-supplemented group. However, 0.2 - 0.3 % of ingested DHEA was conjugated to DHEAS and excreted in urine by females, whereas less than 0.002 % was excreted in urine by males (p < 0.005). DHEAS was excreted in bile by males after DHEA supplementation, and the sex differences in DHEAS levels observed in bile were statistically significant (male, 18.7 +/- 7.5 vs. female, 5.6 +/- 3.1 micromol/l) (p < 0.005). Small amounts of ingested DHEA were excreted in an unchanged state in feces, and no sex difference was observed. These results suggest that there is a considerable sex difference in the conjugation and excretion of orally administered DHEA in the hamster.