Hydroxysteroid (17ß)-dehydrogenase 1-deficient female mice present with normal puberty onset but are severely subfertile due to a defect in luteinization and progesterone production.

Hydroxysteroid (17ß)-dehydrogenase type 1 (HSD17B1) catalyzes the conversion of low active 17-ketosteroids, androstenedione (A-dione) and estrone (E1) to highly active 17-hydroxysteroids, testosterone (T) and E2, respectively. In this study, the importance of HSD17B1 in ovarian estrogen production was determined using Hsd17b1 knockout (HSD17B1KO) mice. In these mice, the ovarian HSD17B enzyme activity was markedly reduced, indicating a central role of HSD17B1 in ovarian physiology. The lack of Hsd17b activity resulted in increased ovarian E1:E2 and A-dione:T ratios, but we also observed reduced progesterone concentration in HSD17B1KO ovaries. Accordingly with the altered steroid production, altered expression of Star, Cyp11a1, Lhcgr, Hsd17b7, and especially Cyp17a1 was observed. The ovaries of HSD17B1KO mice presented with all stages of folliculogenesis, while the corpus luteum structure was less defined and number reduced. Surprisingly, bundles of large granular cells of unknown origin appeared in the stroma of the KO ovaries. The HSD17B1KO mice presented with severe subfertility and failed to initiate pseudopregnancy. However, the HSD17B1KO females presented with normal estrous cycle defined by vaginal smears and normal puberty appearance. This study indicates that HSD17B1 is a key enzyme in ovarian steroidogenesis and has a novel function in initiation and stabilization of pregnancy.

Human ovarian cancer stroma contains luteinized theca cells harboring tumor suppressor gene GT198 mutations.

Ovarian cancer is a highly lethal gynecological cancer, and its causes remain to be understood. Using a recently identified tumor suppressor gene, GT198 (PSMC3IP), as a unique marker, we searched for the identity of GT198 mutant cells in ovarian cancer. GT198 has germ line mutations in familial and early onset breast and ovarian cancers and recurrent somatic mutations in sporadic fallopian tube cancers. GT198 protein has been shown as a steroid hormone receptor coregulator and also as a crucial factor in DNA repair. In this study, using GT198 as a marker for microdissection, we find that ovarian tumor stromal cells harboring GT198 mutations are present in various types of ovarian cancer including high and low grade serous, endometrioid, mucinous, clear cell, and granulosa cell carcinomas and in precursor lesions such as inclusion cysts. The mutant stromal cells consist of a luteinized theca cell lineage at various differentiation stages including CD133(+), CD44(+), and CD34(+) cells, although the vast majority of them are differentiated overexpressing steroidogenic enzyme CYP17, a theca cell-specific marker. In addition, wild type GT198 suppresses whereas mutant GT198 protein stimulates CYP17 expression. The chromatin-bound GT198 on the human CYP17 promoter is decreased by overexpressing mutant GT198 protein, implicating the loss of wild type suppression in mutant cells. Together, our results suggest that GT198 mutant luteinized theca cells overexpressing CYP17 are common in ovarian cancer stroma. Because first hit cancer gene mutations would specifically mark cancer-inducing cells, the identification of mutant luteinized theca cells may add crucial evidence in understanding the cause of human ovarian cancer.

Pregnane glycosides interfere with steroidogenic enzymes to down-regulate corticosteroid production in human adrenocortical H295R cells.

A group of bioactive steroidal glycosides (pregnanes) with anorectic activity in animals was isolated from several genera of milkweeds including Hoodia and Asclepias. In this study, we investigated the effects, structure-activity relationships, and mechanism of action of pregnane glycosides on steroidogenesis in human adrenocortical H295R cells. Administration of pregnane glycosides for 24 h suppressed the basal and forskolin-stimulated release of androstenedione, corticosterone, and cortisone from H295R cells. The conversion of progesterone to 11-deoxycorticosterone and 17-hydroxyprogesterone to either androstenedione or 11-deoxycortisol was most strongly affected, with 12-cinnamoyl-, benzoyl-, and tigloyl-containing pregnanes showing the highest activity. Incubation of pregnane glycosides for 24 h had no effect on mRNA transcripts of CYP11A1, CYP21A1, CYP11B1 cytochrome enzymes and steroidogenic acute regulatory protein (StaR) protein, yet resulted in twofold decrease in HSD3B1 mRNA levels. At the same time, pregnane glycosides had no effect on the CYP1, 2, or 3 drug and steroid metabolism enzymes and showed weak Na(+) /K(+) ATPase and glucocorticoid receptor binding. Taken together, these data suggest that pregnane glycosides specifically suppress steroidogenesis through strong inhibition of 11ß-hydroxylase and steroid 17-alpha-monooxygenase, and weak inhibition of cytochrome P450 side chain cleavage enzyme and 21ß-hydroxylase, but not 3ß-hydroxysteroid dehydrogenase/isomerase.

The AP-1 family member FOS blocks transcriptional activity of the nuclear receptor steroidogenic factor 1.

Steroid production in the adrenal zona glomerulosa is under the control of angiotensin II (Ang II), which, upon binding to its receptor, activates protein kinase C (PKC) within these cells. PKC is a potent inhibitor of the steroidogenic enzyme CYP17. We have demonstrated that, in the ovary, PKC activates expression of FOS, a member of the AP-1 family, and increased expression of this gene is linked to CYP17 downregulation. However, the pathway and the molecular mechanism responsible for the inhibitory effect of PKC on CYP17 expression are not defined. Herein, we demonstrated that Ang II inhibited CYP17 through PKC and ERK1/2-activated FOS and that blocking FOS expression decreased PKC-mediated inhibition. Although CYP17 transcription was activated by the nuclear receptor SF-1, expression of FOS resulted in a decrease in SF-1-mediated gene transcription. FOS physically interacted with the hinge region of SF-1 and modulated its transactivity, thus preventing binding of cofactors such as SRC1 and CBP, which were necessary to fully activate CYP17 transcription. Collectively, these results indicate a new regulatory mechanism for SF-1 transcriptional activity that might influence adrenal zone-specific expression of CYP17, a mechanism that can potentially be applied to other steroidogenic tissues.

Restart of steroidogenesis in dogs during recrudescence of testicular function following downregulation with a GnRH-agonist implant.

To date, no details are available concerning the restart of steroidogenesis following the downregulation of testicular endocrine and germinative function by gonadotrophin-releasing hormone (GnRH)-agonist implants. This restart was assessed by determining the expression of steroidogenic acute regulatory (StAR) protein, cytochrome P450 side-chain cleavage enzyme (P450scc) and cytochrome P450 17α-hydroxylase,17,20-lyase (P450c17). The re-establishment of steroidogenesis was initiated by the removal of the GnRH-agonist implant (18.5 mg azagly nafarelin, Gonazon) at 5 months after treatment. Testes were removed at 3-week intervals (weeks 0-24) and four groups were formed according to the stage of spermatogenesis as revealed by the most developed germ cells observed (developmental group [DG] spermatocytes to DG elongated spermatids). Five dogs served as untreated controls. Positive immunostaining for StAR, P450scc and P450c17 was restricted to Leydig cells. Western blot indicated the specifity of the respective antibodies with hints of a expression of canine-specific P450scc and P450c17 proteins. A significant effect of group was observed for a percentage of the immunopositive area (PIA) as an indicator of active Leydig cells for StAR (P<0.05), P450scc (P<0.001) and P450c17 (P<0.001), with PIA being lowest for the DG spermatocytes. With regard to the strength of the immunopositive signal, a significant effect of group was found for P450scc (P<0.01) and P450c17 (P<0.05), with the lowest intensity being observed in DG spermatocytes. At the mRNA level, the upregulation from DG spermatocytes to DG round spermatids was clearly evident but was only significant for P450scc (P<0.05). Thus, downregulation affects the whole cascade of steroidogenesis, whereas withdrawal of inhibition results in a rapid restart, in part indicating a rebound phenomenon.

Expression and phylogeny of candidate genes for sex differentiation in a primitive fish species, the Siberian sturgeon, Acipenser baerii.

The molecular mechanisms underlying testis differentiation in basal actinopterygian fish remains poorly understood. The sex differentiation period was investigated in the Siberian sturgeon, Acipenser baerii, by expression profiling of Sertoli cell transcription factors (dmrt1, sox9) that control testis differentiation in vertebrates; Leydig cell factors (cyp17a1, star) affecting androgen production; the androgen receptor (ar); a growth factor controlling testis development (igf1); and a gene coding for a gonadotropin hormone (lh). Two genes were characterised for the first time in the Siberian sturgeon (dmrt1, cyp17a1), while the others came from public databases. Sturgeon gonad development is very slow, with a late sexual differentiation time during their juvenile stage, and are still immature at 3 years of age. Immature fish showed a sex-dimorphic pattern; all the genes studied displayed a higher expression level in male gonads. We took advantage of the presence of juvenile fish with pre- and post-differentiated gonads (16 and 18 months old) to characterise them at the molecular level. The post-differentiated fish displayed a sex dimorphism of gene expression in their gonads for all genes studied, with the exception of sox9. The trends in undifferentiated fish lead us to propose that sturgeons undergoing male differentiation express high levels of Sertoli cell factors (dmrt1, sox9) and of genes involved in the production and receptivity of androgens (cyp17a1, star and ar) together with lh. Expression profiles and phylogenetic studies suggest that these genes are potential regulators of testis development in the Siberian sturgeon.

Prenatal and concurrent exposure to halogenated organic compounds and gene expression of CYP17A1, CYP19A1, and oestrogen receptor alpha and beta genes.

OBJECTIVE: To determine whether prenatal exposure to dichlorodiphenyl ethylene (DDE) and polychlorinated biphenyls (PCBs) and concurrent exposure to DDE, PCBs and polybrominated diphenylethers (PBDEs) affect gene expression of aromatase (CYP19A1), 17-α-hydroxylase (CYP17A1), and oestrogen receptors α and ß (ESR 1 and ESR2). METHODS: Based on maternal PCB and DDE levels in the parent generation of the Michigan Fisheater Cohort determined between 1973 and 1991, individual prenatal exposures were estimated and have been published. In 2007, female adult offspring of this cohort were examined. Gene expression and concurrent lipid-adjusted exposures to DDE, PCBs and PBDEs were measured in blood and serum, respectively. Using mixed models and path analyses, gene-expression data were regressed on prenatal and concurrent exposures controlling for confounders. RESULTS: 139 daughters of Michigan fisheaters (65.3%) participated in the investigation. While prenatal PCB levels were statistically significantly associated with decreased expression of the aromatase and 17-α-hydroxylase genes, prenatal DDE levels were significantly related to increased gene expression of aromatase but not of 17-α-hydroxylase. The DDE association seems to be mediated by concurrent lipid-adjusted p,p'-DDE serum levels. Prenatal and concurrent exposure of both PCBs and DDE had comparable effects. No association was found for PBDEs or for the gene expression of ESR 1 and ESR2. CONCLUSIONS: A 40-year antecedent prenatal exposure and concurrent levels of PCBs and DDE are associated with the expression of aromatase and 17-α-hydroxylase genes. Prenatal exposures to organochlorines may instigate long-term alterations of gene expression. Mechanisms of prenatal induction of persistent gene-expression alterations are speculated to be epigenetic in nature.

Testicular steroidogenesis is suppressed during experimental autoimmune encephalomyelitis in rats.

Multiple sclerosis (MS) is an autoimmune disease that usually occurs during the reproductive years in both sexes. Many male patients with MS show lower blood testosterone levels, which was also observed in male rats during experimental autoimmune encephalomyelitis (EAE), an animal model of MS. To better understand the causes of decreased testosterone production during EAE, we investigated the expression status of genes and proteins associated with steroidogenesis in the testes. No changes in the number of interstitial cells were observed in EAE animals, but the expression of the insulin-like 3 gene was reduced at the peak of the disease, implying that the Leydig cell functional capacity was affected. Consistent with this finding, the expression of most steroidogenic enzyme genes and proteins was reduced during EAE, including StAR, CYP11A1, CYP17A1 and HSD3B. No signs of testicular inflammation were observed. Recovery of steroidogenesis was observed after injection of hCG, the placental gonadotropin, or buserelin acetate, a gonadotropin-releasing hormone analogue, at the peak of EAE. Together, our results are consistent with the hypothesis that impaired testicular steroidogenesis originates upstream of the testes and that low serum LH is the main cause of decreased testosterone levels during EAE.

Aldosterone-Regulating Receptors and Aldosterone-Driver Somatic Mutations.

Background: Somatic gene mutations that facilitate inappropriate intracellular calcium entrance have been identified in most aldosterone-producing adenomas (APAs). Studies suggest that angiotensin II and adrenocorticotropic hormone (ACTH) augment aldosterone production from APAs. Little is known, however, regarding possible variations in response to hormonal stimuli between APAs with different aldosterone-driver mutations. Objective: To analyze the transcript expression of type 1 angiotensin II receptors (AGTR1), ACTH receptors (MC2R), and melanocortin 2 receptor accessory protein (MRAP) in APAs with known aldosterone-driver somatic mutations. Methods: RNA was isolated from APAs with mutations in: KCNJ5 (n = 14), ATP1A1 (n = 14), CACNA1D (n = 14), and ATP2B3 (n = 5), and from normal adjacent adrenal tissue (n = 45). Transcript expression of MC2R, MRAP, AGTR1, aldosterone synthase (CYP11B2), 17α-hydroxylase/17,20-lyase (CYP17A1), and 11ß-hydroxylase (CYP11B1) were quantified using quantitative RT-PCR and normalized to ß-actin. Results: Compared to adjacent normal adrenal tissue, APAs had higher transcript levels of CYP11B2 (2,216.4 [1,112.0, 2,813.5]-fold, p < 0.001), MC2R (2.88 [2.00, 4.52]-fold, p < 0.001), and AGTR1 (1.80 [1.02, 2.80]-fold, p < 0.001]), and lower transcript levels of MRAP, CYP17A1, and CYP11B1 (0.28-0.36, p < 0.001 for all). MC2R and CYP11B2 transcripts were lower in APAs with KCNJ5 vs. other mutations (p < 0.01 for both). MC2R expression correlated positively with that of AGTR1 in APAs harboring KCNJ5 and CACNA1D mutations, and with MRAP expression in APAs harboring ATPase mutations. Conclusions: While MC2R and AGTR1 are expressed in all APAs, differences were observed based on the underlying aldosterone-driver somatic mutations. In tandem, our findings suggest that APAs with ATPase-mutations are more responsive to ACTH than KCNJ5-mutated APAs.

Polychlorinated biphenyls have inhibitory effect on testicular steroidogenesis by downregulation of P450(17alpha) and P450(scc).

Polychlorinated biphenyls (PCBs) are environmental pollutants that are quite toxic to biological systems. This study examined the inhibitory effect of PCB126 and PCB114 on testicular steroidogenesis in male rats. Male Sprague Dawley rats received weekly intraperitoneal injections of PCB126 (0.2 mg/kg) or PCB114 (20 mg/kg) or vehicle (corn oil). Animals from each group were sacrificed at 2, 5 and 8 weeks after the injections. Blood and testis tissue samples were collected for the hormone assay, Western blotting and reverse transcriptase polymerase chain reaction (RT-PCR). The testosterone, luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels were assayed, and the expression levels of the mRNA and proteins associated with the testosterone biosynthesis pathway were measured to determine the effect of PCB126 and PCB114 on testicular steroidogenesis. The results showed that the testis weight was significantly higher in the PCB126-treated rats given eight shots. Moreover, the serum testosterone levels were significantly lower in the PCB126 and PCB114-treated groups than the control. The transcription and translation levels of P450(17alpha) and P450(scc) were significantly lower in the PCB126-treated groups than the control. These results suggest that PCB126 may affect testicular steroidogenesis by downregulating P450(17alpha), P450(scc) and have inhibitory effect on the testicular functions.

Targeting CYP17: established and novel approaches in prostate cancer.

There is a growing body of evidence that although medical or surgical castration blocks the generation of gonadal testosterone in prostate cancer, androgens originating from other sources may continue to drive androgen receptor (AR) signaling. Recent studies have demonstrated high intratumoral levels of androgens and continued AR signaling in castration-resistant prostate cancer (CRPC), suggesting that androgens may also be synthesized de novo. Inhibiting the systemic biosynthesis of androgens in CRPC by targeting CYP17 may thus represent a rational therapeutic approach since this enzyme catalyses two key steroid reactions involving 17alpha-hydroxylase and C(17,20)-lyase in the androgen biosynthesis pathway. This review will discuss the rationale for and implications of targeting CYP17 in CRPC and focus on established and novel CYP17 inhibitors, including ketoconazole, abiraterone acetate, and VN/124-1, which are agents currently at different stages of development.

Effects of lentivirus-mediated CYP17A1 gene silencing on the biological activity of glioma.

Gliomas are the most common malignant primary brain tumors with poor prognosis. We attempted to explore the role of CYP17A1 in glioma progression. We demonstrated that the expression of CYP17A1 was significantly higher in the glioma tissues than the normal brain tissues, especially in malignant glioma. Moreover, the expression of CYP17A1 gene was positively correlative with glioma pathological grades. In vitro, CYP17A1 gene silence inhibited the proliferation and invasion of glioma cells and promoted the apoptosis in glioma cells. Also, the subcutaneously transplanted tumour in BALB/C-nu showed that CYP17A1 gene silence inhibited glioma growth. These results reveal that CYP17A1 plays a major role in the progress of glioma.

SIRT2 plays a novel role on progesterone, estradiol and testosterone synthesis via PPARs/LXRα pathways in bovine ovarian granular cells.

SIRT2 has been shown to possess NAD+-dependent deacetylase and desuccinylase enzymatic activities, it also regulates metabolism homeostasis in mammals. Previous data has suggested that resveratrol, a potential activator of Sirtuins, played a stimulation role in steroidogenesis. Unfortunately, to date, the physiological roles of SIRT2 in ovarian granular cells (GCs) are largely unknown. Here, we studied the function and molecular mechanisms of SIRT2 on steroid hormone synthesis in GCs from Qinchuan cattle. Immunohistochemistry and western blotting showed that SIRT2 was expressed not only in GCs and cumulus cells, but also in oocytes and theca cells. We found that the secretion of progesterone was induced, whereas that of estrogen and testosterone secretion was suppressed by treatment with the SIRT2 inhibitor (Thiomyristoyl or SirReal2) or siRNA. Additionally, the PPARs/LXRα signaling pathways were suppressed by SIRT2 siRNA or inhibitors. The mRNA expression of CYP17, aromatase and StAR was suppressed, but the abundance of CYP11A1 mRNA was induced by SIRT2 inhibition. Furthermore, the PPARα agonist or PPARγ antagonist could mimic the effects of SIRT2 inhibition on hormones levels and gene expression associated with steroid hormone biosynthesis. In turn, those effects were abolished by the LXRα agonist (LXR-623). Together, these data support the hypothesis that SIRT2 regulates steroid hormone synthesis via the PPARs/LXRα pathways in GCs.

Development of an early biomarker for the ovarian liability of selective estrogen receptor modulators in rats.

Selective estrogen receptor modulators (SERMs) have the potential to treat estrogen sensitive diseases such as uterine leiomyoma and endometriosis, which are prevalent in reproductive age women. However, SERMs also increase the risk of developing ovarian cysts in this population, a phenomenon that is not seen in postmenopausal women. It is believed that current SERMs partially block estradiol's ability to downregulate gonadotropin-releasing hormone (GnRH) secretion from the hypothalamus thereby interfering with estradiol's negative feedback, leading to increased ovarian stimulation by gonadotropins, and cyst formation. It has been postulated that a SERM with poor brain exposure would have less negative effect on the HPO axis, therefore reducing the risk of developing ovarian cysts. In order to test this hypothesis, we identified an early marker of SERM-dependent ovarian effects: upregulation of Cyp17a1 mRNA. SERMs known to cause ovarian cysts upregulate Cyp17a1 after only 4 days of dosing and suppression of the HPO axis prevented this regulation, indicating that ovarian expression of Cyp17a1 was secondary to SERM's effect on the brain. We then characterized three SERMs with similar binding affinity and antagonist effects on the uterus for their relative brain/plasma exposure and ovarian effects. We found that the degree of brain exposure correlated very well with Cyp17a1 expression.

Real-time PCR array to study effects of chemicals on the Hypothalamic-Pituitary-Gonadal axis of the Japanese medaka.

This paper describes the development and validation of a PCR array for studying chemical-induced effects on gene expression of selected endocrine pathways along the hypothalamic-pituitary-gonadal (HPG) axis of the small, oviparous fish, the Japanese medaka (Oryzias latipes). The Japanese medaka HPG-PCR array combines the quantitative performance of SYBR Green-based real-time PCR with the multiple gene profiling capabilities of a microarray to examine expression profiles of 36 genes associated with endocrine pathways in brain, liver and gonad. The performance of the Japanese medaka HPG-PCR array was evaluated by examining effects of two model compounds, the synthetic estrogen, 17alpha-ethinylestradiol (EE2) and the anabolic androgen, 17beta-trenbolone (TRB) on the HPG axis of the Japanese medaka. Four-month-old medaka was exposed to three concentrations of EE2 (5, 50, 500 ng/L) or TRB (50, 500, 5000 ng/L) for 7d in a static renewal exposure system. A pathway-based approach was implemented to analyze and visualize concentration-dependent mRNA expression in the HPG axis of Japanese medaka. The compensatory response to EE2 exposure included the down-regulation of male brain GnRH RI and testicular CYP17. The down-regulation of AR-alpha expression in brain of EE2-exposed males was associated with suppression of male sexual behavior. Compensatory responses to TRB in the female HPG axis included up-regulation of brain GnRH RII and ovary steroidogenic CYP19A. Overall, the results suggested that the Japanese medaka HPG-PCR array has potential not only as a screening tool of potential endocrine-disrupting chemicals but also in elucidating mechanisms of action.

CYP17 inhibitors. Annulations of additional rings in methylene imidazole substituted biphenyls: synthesis, biological evaluation and molecular modelling.

Twenty-one novel compounds originating from two classes of annulated biphenyls were synthesized as mimetics of the steroidal A- and C-rings and examined for their potency as inhibitors of human CYP17. Selected compounds were tested for inhibition of the hepatic CYP enzyme 3A4. Potent CYP17 inhibitors were found for each class, compound 9 (17 and 71% at 0.2 and 2 microM, respectively) and 21 (591 nM). Compound 21 showed only weak inhibition of CYP3A4 (32 and 64% at 2 and 10 microM, respectively). Both compounds, however, exhibited moderate to strong inhibition of the glucocorticoid-forming enzyme CYP11B1. The most interesting compounds were docked into our protein model. They bound into one of the modes which we have previously published. New interaction regions were identified.

Expression and regulation of CYP17A1 and 3ß-hydroxysteroid dehydrogenase in cells of the nervous system: Potential effects of vitamin D on brain steroidogenesis.

Steroids are reported to have diverse functions in the nervous system. Enzymatic production of steroid hormones has been reported in different cell types, including astrocytes and neurons. However, the information on some of the steroidogenic enzymes involved is insufficient in many respects. Contradictory results have been reported concerning the relative importance of different cell types in the nervous system for expression of CYP17A1 and 3ß-hydroxysteroid dehydrogenase (3ß-HSD). 3ß-HSD is important in all basic steroidogenic pathways and CYP17A1 is required to form sex hormones. In the current investigation we studied the expression of these enzymes in cultured primary rat astrocytes, in neuron-enriched cells from rat cerebral cortex and in human neuroblastoma SH-SY5Y cells, a cell line often used as an in vitro model of neuronal function and differentiation. As part of this study we also examined potential effects on CYP17A1 and 3ß-HSD by vitamin D, a compound previously shown to have regulatory effects in steroid hormone-producing cells outside the brain. The results of our study indicate that astrocytes are a major site for expression of 3ß-HSD whereas expression of CYP17A1 is found in both astrocytes and neurons. The current data suggest that neurons, contrary to some previous reports, are not involved in 3ß-HSD reactions. Previous studies have shown that vitamin D can influence gene expression and hormone production by steroidogenic enzymes in some cells. We found that vitamin D suppressed CYP17A1-mediated activity by 20% in SH-SY5Ycells and astrocytes. Suppression of CYP17A1 mRNA levels was considerably stronger, about 50% in SH-SY5Y cells and 75% in astrocytes. In astrocytes 3ß-HSD was also suppressed by vitamin D, about 20% at the enzyme activity level and 60% at the mRNA level. These data suggest that vitamin D-mediated regulation of CYP17A1 and 3ß-HSD, particularly on the transcriptional level, may play a role in the nervous system.

Alterations in the steroid biosynthetic pathways in the human prefrontal cortex in mood disorders: A post-mortem study.

Altered levels of steroids have been reported in the brain, cerebral spinal fluid and plasma of patients with mood disorders. Neuroimaging studies have reported both functional and structural alterations in mood disorders, for instance in the anterior cingulate cortex (ACC) and dorsolateral prefrontal cortex (DLPFC). In order to determine whether the endogenous production of steroids is altered in the ACC and DLPFC of patients with major depressive disorder (MDD) or bipolar disorder (BPD), quantitative real-time PCR was performed to detect mRNA expression level of key enzymes in the steroid biosynthetic pathways. In MDD, a significant decrease in mRNA level of cytochrome P450 17A1 (CYP17A1, synthesizing C19 ketosteroids) in the ACC and a significant increase in mRNA levels of hydroxysteroid sulfotransferase 2A1 [SULT2A1, catalyzing the sulfate conjugation of dehydroepiandrosterone (DHEA)] were observed in the DLPFC, suggesting alterations in DHEA and its sulfate metabolite DHEAS levels. Decreased intensity and distribution of CYP17A1 immunohistochemical staining was found in the ACC of MDD patients. Interestingly, there was a significant positive correlation between the mRNA levels of CYP17A1 and tyrosine-related kinase B (TrkB) full length isoform. In a unique post-mortem human brain slice culture paradigm, BDNF mRNA expression was found to be significantly increased following incubation with DHEA. Together, these data indicate a close relationship between DHEA and BDNF-TrkB pathways in depression. Furthermore, in the DLPFC, higher mRNA levels of 11ß-hydroxysteroid dehydrogenase-1 (HSD11B1, reducing cortisone to the active hormone cortisol) and steroidogenic acute regulatory protein (STAR, facilitating the shuttle of cholesterol through the intermembrane space) were found in the MDD patients and BPD patients, respectively. In conclusion, this study suggests the presence of a disturbance in the endogenous synthesis of DHEA and DHEAS in mood disorders, which has a close relationship with BDNF-TrkB signaling.

Maternal fructose intake disturbs ovarian estradiol synthesis in rats.

AIMS: Recent increases in fructose consumption have raised concerns regarding the potential adverse intergenerational effects, as maternal fructose intake may induce physiological dysfunction in offspring. However, no reports are available regarding the effect of excess maternal fructose on reproductive tissues such as the ovary. Notably, the maternal intrauterine environment has been demonstrated to affect ovarian development in the subsequent generation. Given the fructose is transferred to the fetus, excess fructose consumption may affect offspring ovarian development. As ovarian development and its function is maintained by 17ß-estradiol, we therefore investigated whether excess maternal fructose intake influences offspring ovarian estradiol synthesis. Rats received a 20% fructose solution during gestation and lactation. After weaning, offspring ovaries were isolated. KEY FINDINGS: Offspring from fructose-fed dams showed reduced StAR and P450(17α) mRNA levels, along with decreased protein expression levels. Conversely, attenuated P450arom protein level was found in the absence of mRNA expression alteration. Consistent with these phenomena, decreased circulating levels of estradiol were observed. Furthermore, estrogen receptor α (ERα) protein levels were also down-regulated. In accordance, the mRNA for progesterone receptor, a transcriptional target of ERα, was decreased. These results suggest that maternal fructose might alter ovarian physiology in the subsequent generation.

The mechanism for protein kinase C inhibition of androgen production and 17alpha-hydroxylase expression in a theca cell tumor model.

INTRODUCTION: In polycystic ovary syndrome (PCOS), there is increased formation of androgens by thecal cells. Moreover, PCOS ovaries have been shown to have decreased levels of c-fos transcription factor. We hypothesize that c-fos expression inhibits 17alpha-hydroxylase 17,20 lyase (CYP17) activity in the human ovary, and its decreased expression seen in PCOS may lead to elevated CYP17 transcription, resulting in increased androgen production. OBJECTIVE: Our objective was to define the role of the activator protein-1 transcription factors, namely c-fos, in the regulation of CYP17 expression in theca cells. METHODS: Human ovarian thecal-like tumor cells were used for all experiments. The following techniques were used: steroid quantification, mRNA extraction, microarray analysis, transfection, small interfering RNA, and immunohistochemistry. RESULTS: Stimulation of human ovarian thecal-like tumor cells with the protein kinase A pathway activator forskolin resulted in stimulation of C19 androgen production. In contrast, treatment with the protein kinase C pathway activator tetradecanoylphorbol acetate (TPA) resulted in decreased androgen production with a shift toward C21 progesterone production. TPA also led to complete inhibition of CYP17. Microarray data showed a 37-fold increase in c-fos after treatment with TPA. Transfection with steroidogenic factor 1 resulted in an increase in CYP17 promoter activity, which was significantly inhibited in the presence of c-fos. c-fos gene silencing led to an increase in CYP17 mRNA levels. Immunohistochemical staining for c-fos in ovaries demonstrated strong staining in granulosa cells, but not theca. CONCLUSIONS: The activator protein-1 transcription factor c-fos plays a role in the inhibition of CYP17 expression. The decreased levels of c-fos expression in polycystic ovaries may be responsible for increased CYP17 levels in PCOS.