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1.
Front Endocrinol (Lausanne) ; 12: 644382, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33796077

RESUMO

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.


Assuntos
Hormônio Adrenocorticotrópico/metabolismo , Aldosterona/metabolismo , Proteínas de Membrana/biossíntese , Mutação , Receptor Tipo 1 de Angiotensina/biossíntese , Receptor Tipo 2 de Melanocortina/biossíntese , Adenoma , Glândulas Suprarrenais/metabolismo , Adulto , Idoso , Feminino , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/biossíntese , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Receptor Tipo 1 de Angiotensina/genética , Receptor Tipo 2 de Melanocortina/genética , Receptores da Corticotropina/metabolismo , Esteroide 11-beta-Hidroxilase/biossíntese , Esteroide 11-beta-Hidroxilase/genética , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroide 17-alfa-Hidroxilase/genética , Adulto Jovem
2.
Sci Rep ; 11(1): 8996, 2021 04 26.
Artigo em Inglês | MEDLINE | ID: mdl-33903635

RESUMO

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.


Assuntos
Encefalomielite Autoimune Experimental/metabolismo , Esclerose Múltipla/metabolismo , Testículo/metabolismo , Testosterona/biossíntese , Animais , Enzima de Clivagem da Cadeia Lateral do Colesterol/biossíntese , Encefalomielite Autoimune Experimental/patologia , Regulação Enzimológica da Expressão Gênica , Masculino , Complexos Multienzimáticos/biossíntese , Esclerose Múltipla/patologia , Progesterona Redutase/biossíntese , Ratos , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroide Isomerases/biossíntese , Testículo/patologia
3.
J Steroid Biochem Mol Biol ; 185: 27-38, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30009951

RESUMO

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.


Assuntos
Estradiol/biossíntese , Células da Granulosa/metabolismo , Receptores X do Fígado/metabolismo , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , Progesterona/biossíntese , Sirtuína 2/metabolismo , Testosterona/biossíntese , Acetamidas/farmacologia , Animais , Bovinos , Células Cultivadas , Enzima de Clivagem da Cadeia Lateral do Colesterol/biossíntese , Feminino , Indazóis/farmacologia , Receptores X do Fígado/agonistas , Oócitos/metabolismo , Interferência de RNA , RNA Interferente Pequeno/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Sirtuína 2/antagonistas & inibidores , Sirtuína 2/genética , Esteroide 17-alfa-Hidroxilase/biossíntese , Células Tecais/metabolismo , Tiazóis/farmacologia
4.
Neurosci Lett ; 692: 210-215, 2019 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-30439398

RESUMO

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.


Assuntos
Neoplasias Encefálicas/genética , Glioma/genética , Esteroide 17-alfa-Hidroxilase/genética , Animais , Apoptose , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Proliferação de Células , Progressão da Doença , Feminino , Inativação Gênica , Vetores Genéticos , Glioma/metabolismo , Glioma/patologia , Humanos , Lentivirus , Camundongos Endogâmicos BALB C , Gradação de Tumores , Esteroide 17-alfa-Hidroxilase/biossíntese
5.
Life Sci ; 202: 117-123, 2018 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-29654807

RESUMO

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.


Assuntos
Estradiol/biossíntese , Frutose/farmacologia , Ovário/efeitos dos fármacos , Ovário/metabolismo , Animais , Peso Corporal/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Receptor alfa de Estrogênio/biossíntese , Feminino , Lactação , Fosfoproteínas/biossíntese , Fosfoproteínas/genética , Gravidez , Efeitos Tardios da Exposição Pré-Natal/metabolismo , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Ratos , Ratos Sprague-Dawley , Receptores de Progesterona/biossíntese , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroide 17-alfa-Hidroxilase/genética
6.
Brain Pathol ; 28(4): 536-547, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-28752602

RESUMO

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.


Assuntos
Transtorno Bipolar/metabolismo , Transtorno Depressivo Maior/metabolismo , Transtornos do Humor/metabolismo , Córtex Pré-Frontal/metabolismo , Esteroides/biossíntese , Fator Neurotrófico Derivado do Encéfalo/biossíntese , Feminino , Giro do Cíngulo/metabolismo , Humanos , Masculino , Glicoproteínas de Membrana/biossíntese , RNA Mensageiro/metabolismo , Receptor trkB/biossíntese , Transdução de Sinais , Esteroide 17-alfa-Hidroxilase/biossíntese , Sulfotransferases/biossíntese
7.
Neurochem Int ; 113: 46-55, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29162485

RESUMO

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.


Assuntos
17-Hidroxiesteroide Desidrogenases/biossíntese , Encéfalo/enzimologia , Regulação Enzimológica da Expressão Gênica , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroides/biossíntese , Vitamina D/farmacologia , 17-Hidroxiesteroide Desidrogenases/antagonistas & inibidores , 17-Hidroxiesteroide Desidrogenases/genética , Animais , Encéfalo/efeitos dos fármacos , Linhagem Celular Tumoral , Células Cultivadas , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Humanos , Ratos , Ratos Sprague-Dawley , Esteroide 17-alfa-Hidroxilase/antagonistas & inibidores , Esteroide 17-alfa-Hidroxilase/genética , Esteroides/antagonistas & inibidores
8.
PLoS One ; 12(3): e0174224, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28323907

RESUMO

Resveratrol, a natural compound found in grapes, became very popular for its suggested protective effects against aging. It was reported to have similar positive effects on the human metabolism as caloric restriction. Recently, positive effects of resveratrol on steroid biosynthesis in cell systems and in humans suffering from polycystic ovary syndrome have also been reported, but the exact mechanism of this action remains unknown. Sirtuins seem targeted by resveratrol to mediate its action on energy homeostasis. In this study, we investigated the mechanisms of action of resveratrol on steroidogenesis in human adrenal H295R cells. Resveratrol was found to inhibit protein expression and enzyme activities of CYP17 and CYP21. It did not alter CYP17 and CYP21 mRNA expression, nor protein degradation. Only SIRT3 mRNA expression was found to be altered by resveratrol, but SIRT1, 3 and 5 overexpression did not result in a change in the steroid profile of H295R cells, indicating that resveratrol may not engage sirtuins to modulate steroid production. Previous studies showed that starvation leads to a hyperandrogenic steroid profile in H295R cells through inhibition of PKB/Akt signaling, and that resveratrol inhibits steroidogenesis of rat ovarian theca cells via the PKB/Akt pathway. Therefore, the effect of resveratrol on PKB/Akt signaling was tested in H295R cells and was found to be decreased under starvation growth conditions, but not under normal growth conditions. Overall, these properties of action together with recent clinical findings make resveratrol a candidate for the treatment of hyperandrogenic disorders such as PCOS.


Assuntos
Androgênios/biossíntese , Anti-Inflamatórios não Esteroides/farmacologia , Família 21 do Citocromo P450/antagonistas & inibidores , Biossíntese de Proteínas/efeitos dos fármacos , Esteroide 17-alfa-Hidroxilase/antagonistas & inibidores , Estilbenos/farmacologia , Córtex Suprarrenal/citologia , Córtex Suprarrenal/metabolismo , Linhagem Celular , Família 21 do Citocromo P450/biossíntese , Família 21 do Citocromo P450/metabolismo , Humanos , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Mensageiro/biossíntese , Resveratrol , Sirtuína 3/biossíntese , Sirtuína 3/genética , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroide 17-alfa-Hidroxilase/metabolismo
9.
Gene ; 593(1): 167-171, 2016 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-27511375

RESUMO

It is believed that excess androgen exposure of the fetus, via altered gene expression, causes hyperandrogenism a key feature of polycystic ovary syndrome (PCOS). The aim of this study was to evaluate expression of Cytochrome P450-17 (CYP17), GATA-binding protein (GAGT6) and Steroidogenic acute regulatory protein (StAR), genes of adult female rats prenatally exposed to androgen excess, closely reflect endocrine and ovarian disturbances of PCOS in women, by comparing them during different phases of estrus cycle with those of non-treated rats. Both the adult prenatally testosterone exposed and control rats (n=23, each) were divided into four groups based on their observed vaginal smear (proestrus, estrus, metestrus and diestrus) and the relative expression of CYP17, GATA6 and StAR genes was measured in ovarian theca cells using Cyber-green Real-Time PCR. Serum sex steroid hormones and gonadotropins levels were measured using the ELISA method; a comparison of these two groups showed that there was an overall increase in the studied genes (CYP17; 2.39 fold change, 95% CI: 1.23-3.55; P<0.05, GATA6; 2.08 fold change, 95% CI: 1.62-2.55; P<0.0001, and StAR; 1.4 fold change, 95% CI: 1.02-1.78; P<0.05), despite variations in different phases with maximum elevation for all genes in diestrus. The changes observed may impair the normal development of ovaries that mediate the programming of adult PCOS.


Assuntos
Androgênios/efeitos adversos , Fator de Transcrição GATA6/biossíntese , Regulação da Expressão Gênica/efeitos dos fármacos , Fosfoproteínas/metabolismo , Esteroide 17-alfa-Hidroxilase/biossíntese , Virilismo/metabolismo , Androgênios/farmacologia , Animais , Feminino , Fator de Transcrição GATA6/genética , Masculino , Fosfoproteínas/síntese química , Fosfoproteínas/genética , Síndrome do Ovário Policístico/genética , Síndrome do Ovário Policístico/metabolismo , Gravidez , Efeitos Tardios da Exposição Pré-Natal , Ratos , Esteroide 17-alfa-Hidroxilase/genética , Virilismo/induzido quimicamente , Virilismo/genética
10.
FASEB J ; 29(9): 3806-16, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26018678

RESUMO

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.


Assuntos
17-Hidroxiesteroide Desidrogenases/deficiência , Ciclo Estral , Infertilidade Feminina/enzimologia , Luteinização , Ovário/metabolismo , Progesterona/biossíntese , 17-Hidroxiesteroide Desidrogenases/biossíntese , 17-Hidroxiesteroide Desidrogenases/genética , 17-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Enzima de Clivagem da Cadeia Lateral do Colesterol/biossíntese , Enzima de Clivagem da Cadeia Lateral do Colesterol/genética , Feminino , Infertilidade Feminina/genética , Masculino , Camundongos , Camundongos Knockout , Ovário/patologia , Fosfoproteínas/biossíntese , Fosfoproteínas/genética , Gravidez , Progesterona/genética , Maturidade Sexual/genética , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroide 17-alfa-Hidroxilase/genética
11.
Toxicol In Vitro ; 29(1): 155-61, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25317747

RESUMO

Monocrotophos (MCP) pesticide, listed as a UNEP Prior Informed Consent chemical, has been proved to exert toxic effects on the reproductive system of teleost fishes by changing the balance of sex steroid hormones. To investigate the effects of MCP on steroidogenesis in vitro, the rainbow trout (Oncorhynchus mykiss) gonadal cell line RTG-2 was exposed to different MCP concentrations for 48 h. The levels of 17 ß-estradiol (E(2)) and testosterone in the medium were measured by radioimmunoassay and the expression of steroidogenic acute regulatory protein and cytochrome P450 enzymes CYP11A1, CYP17, and CYP19A was detected by quantitative real-time PCR. The results showed that 1.0 and 10.0 µg/L MCP pesticide induced E(2) levels and promoted steroidogenic enzyme expression. The possible mechanisms of MCP steroidogenic activity were investigated using inhibitors of protein kinase A (PKA) and protein kinase C. The PKA inhibitor H-89 abrogated the 10.0 µg/L MCP-induced transcriptional up-regulation of steroidogenic enzymes, suggesting an involvement of PKA-dependent mechanism in the disruption of steroidogenesis by the MCP pesticide in rainbow trout RTG-2 cells.


Assuntos
Proteínas Quinases Dependentes de AMP Cíclico/efeitos dos fármacos , Estradiol/biossíntese , Inseticidas/toxicidade , Monocrotofós/toxicidade , Transdução de Sinais/efeitos dos fármacos , Testosterona/biossíntese , Animais , Aromatase/biossíntese , Aromatase/efeitos dos fármacos , Linhagem Celular , Enzima de Clivagem da Cadeia Lateral do Colesterol/biossíntese , Enzima de Clivagem da Cadeia Lateral do Colesterol/efeitos dos fármacos , Disruptores Endócrinos/toxicidade , Gônadas/citologia , Gônadas/efeitos dos fármacos , Gônadas/metabolismo , Oncorhynchus mykiss/metabolismo , Oncorhynchus mykiss/fisiologia , Reação em Cadeia da Polimerase em Tempo Real , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroide 17-alfa-Hidroxilase/efeitos dos fármacos
12.
Toxicology ; 322: 14-22, 2014 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-24810611

RESUMO

In this study we tried to answer a question which component of Halowax 1051 is responsible for, observed in previously published study, androgenic effects of the mixture, and whether it is possible to draw conclusions about the action of mixtures by examining the effect of an indicator congener. Ovarian follicles were incubated with individual congeners of an artificial mixture for 6-24h. At the end of the incubation period, media were collected for determination of progesterone (P4), androstenedione (A4), testosterone (T) and estradiol (E2) levels by enzyme immunoassay, and follicles were retained for an examination of aryl hydrocarbon receptor (AHR), cytochrome p450 enzymes (CYP1A1, CYP17, CYP19), and 17ß-hydroxysteroid dehydrogenase (17ß-HSD) protein expression by Western blotting. CN73 in dose 50pg/ml after 6h had no effect and decreased AHR expression after 24h, while at dose 400pg/ml increased AHR protein expression after 6h of exposure which remained elevated after 24h. CN74 and CN75 at both concentrations tested (25 and 50pg/ml) stimulated AHR protein expression after 6h and decreased it after 24h of exposure. Individual congeners induced a rapid increase in CYP1A1 protein expression, with a rank order of efficacy of CN73>CN74=CN75. All congeners increased P4/A4 and T/E2 secretion ratios in association with a decrease in the A4/T ratio, pointing to androgenic and anti-estrogenic properties of PCNs in ovarian follicles. The most potent congener in this context was CN73. The effects of mixtures were comparable to those of CN74 and CN75, and were not as strong as those observed for CN73. Collectively, these data suggest antagonistic actions of single congeners in a mixture, indicating that the actions of a mixture cannot be predicted based on the actions of individual congeners.


Assuntos
Citocromo P-450 CYP1A1/biossíntese , Hidrocarbonetos Clorados/toxicidade , Naftalenos/toxicidade , Folículo Ovariano/metabolismo , Receptores de Hidrocarboneto Arílico/biossíntese , Esteroides/metabolismo , 17-Hidroxiesteroide Desidrogenases/biossíntese , Animais , Aromatase/biossíntese , Western Blotting , Ciclo Estral/efeitos dos fármacos , Feminino , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Hidrocarbonetos Clorados/química , Naftalenos/química , Folículo Ovariano/efeitos dos fármacos , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroides/biossíntese , Suínos
13.
Mol Cell Endocrinol ; 387(1-2): 35-43, 2014 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-24576611

RESUMO

The basis for the pattern of adrenal androgen production in the chimpanzee, which resembles that of humans, is poorly defined. We characterized the developmental zonation and expression of elements of the androgen biosynthetic pathway in the chimpanzee adrenal. The newborn adrenal contained a broad fetal zone (FZ) expressing CYP17, SULT2A1, and Cytochrome B5 (CB5) but not HSD3B; the outer cortex expressed HSD3B but not SULT2A1 or CB5. During infancy, the FZ involuted and the HSD3B-expressing outer cortex broadened. By 3years of age, a thin layer of cells that expressed CB5, SULT2A1, and CYP17 adjoined the medulla and likely represented the zona reticularis; the outer cortex consisted of distinct zonae fasiculata and glomerulosa. Thereafter, the zona reticularis broadened as also occurs in the human. The adult chimpanzee adrenal displayed other human-like characteristics: intramedullary clusters of reticularis-like cells and also a cortical cuff of zona fasiculata-like cells adjoining the central vein.


Assuntos
Androgênios/biossíntese , Zona Fasciculada/crescimento & desenvolvimento , Zona Glomerulosa/crescimento & desenvolvimento , Zona Reticular/crescimento & desenvolvimento , Animais , Citocromos b5/biossíntese , Desidroepiandrosterona/biossíntese , Sulfato de Desidroepiandrosterona/sangue , Feminino , Masculino , Pan troglodytes , Esteroide 17-alfa-Hidroxilase/biossíntese , Sulfotransferases/biossíntese , Zona Fasciculada/anatomia & histologia , Zona Fasciculada/metabolismo , Zona Glomerulosa/anatomia & histologia , Zona Glomerulosa/metabolismo , Zona Reticular/anatomia & histologia , Zona Reticular/metabolismo
14.
J Biol Chem ; 288(46): 33387-97, 2013 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-24097974

RESUMO

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.


Assuntos
Mutação em Linhagem Germinativa , Proteínas Nucleares/metabolismo , Neoplasias Ovarianas/metabolismo , Células Tecais/metabolismo , Transativadores/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Antígenos CD/biossíntese , Antígenos CD/genética , Antígenos de Neoplasias/biossíntese , Antígenos de Neoplasias/genética , Linhagem Celular , Feminino , Regulação Enzimológica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Proteínas Nucleares/genética , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroide 17-alfa-Hidroxilase/genética , Células Estromais/metabolismo , Células Estromais/patologia , Células Tecais/patologia , Transativadores/genética , Proteínas Supressoras de Tumor/genética
15.
Andrology ; 1(5): 772-8, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23843177

RESUMO

We analysed an outbreed mouse line which was selected for the phenotype 'high fertility' for 158 generations. During this selection period the mouse strain increased the number of offspring per litter from 10.4 to 17.1 and the total litter weight up to ~160%. In this study, we initially characterize the reproductive phenotype of high fertility males. Surprisingly, male bucks of the fertility line (FL1) show reduced percentage of motile and progressive motile spermatozoa; however, other sperm motility characteristics (e.g. velocity parameters) are improved compared with an unselected control line. Cytometrical investigation of the testicular cell-type composition indicated a significant increased concentration of diploid cells by a concomitant reduction in haploid cells in the testicular parenchyma of FL1. Furthermore, total testosterone concentrations in blood are dramatically increased in FL1 (>20 ng/mL). In line with increased testosterone levels, we observed increased expression rates of steroidogenic key enzymes Cyp11 and Cyp17 from FL1 testis samples. These data indicate that FL1 males have a manifest 'high fertility phenotype'. Diallelic crosses imply that male-only contribution largely determines the reproductive outcome in cross-breeding experiments. FL1 therefore is a promising model for future investigations on male factor (in)fertility. Our observation might also offer valuable cues for human reproductive medicine.


Assuntos
Fertilidade/genética , Tamanho da Ninhada de Vivíparos/genética , Animais , Cruzamento , Expressão Gênica , Masculino , Camundongos , Modelos Animais , Motilidade dos Espermatozoides , Espermatozoides/fisiologia , Esteroide 17-alfa-Hidroxilase/biossíntese , Testículo/enzimologia , Testículo/metabolismo , Testosterona/sangue
16.
J Endocrinol ; 218(1): 117-24, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23606751

RESUMO

Gastric parietal cells synthesize and secrete estradiol-17ß (E2) into gastric veins joining the portal vein, and a large amount of gastric E2 first binds to its receptors in the liver. However, the role of the gastric E2 is not entirely clear during postnatal development. The objective of this study was to reveal the onset of aromatase and other steroid-synthesizing enzymes in the gastric mucosa; to determine the period of rising E2 levels in the portal vein; and to further understand the relationship between gastric E2 and liver estrogen receptor α (ERα). The immunoblot bands and the immunohistochemistry of gastric mucosa revealed that aromatase protein began to express itself at 20 days and then increased in the levels of aromatase protein from 20 days onward. Expression of mRNAs for gastric aromatase (Cyp19a1) and other steroid-synthesizing enzymes, 17α-Hydroxylase (Cyp17a1) and 17ß-hydroxysteroid dehydrogenase (HSD17b3), also increased similar to the increment of aromatase protein. Portal venous E2 levels were elevated after 20 days and increased remarkably between 23 and 30 days, similar to gastric aromatase mRNA levels. The E2 level was approximately three times higher at 40 days than that at 20 days. The liver weight and Esr1 levels began to increase after 20 days and the increment was positively correlated with the change of portal venous E2 levels. These findings suggest that some changes may occur around 20 days to regulate the gastric E2 synthesis and secretion.


Assuntos
Aromatase/metabolismo , Estradiol/sangue , Mucosa Gástrica/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Sistema Porta/crescimento & desenvolvimento , 17-Hidroxiesteroide Desidrogenases/biossíntese , 17-Hidroxiesteroide Desidrogenases/genética , 17-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Aromatase/biossíntese , Aromatase/genética , Western Blotting , Estradiol/metabolismo , Receptor alfa de Estrogênio/biossíntese , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Mucosa Gástrica/citologia , Mucosa Gástrica/crescimento & desenvolvimento , Imuno-Histoquímica , Fígado/crescimento & desenvolvimento , Fígado/metabolismo , Masculino , Tamanho do Órgão , Veia Porta , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroide 17-alfa-Hidroxilase/genética , Esteroide 17-alfa-Hidroxilase/metabolismo
17.
J Cell Physiol ; 228(5): 1120-6, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23065845

RESUMO

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.


Assuntos
3-Hidroxiesteroide Desidrogenases , Enzima de Clivagem da Cadeia Lateral do Colesterol , Glicosídeos/administração & dosagem , Pregnanos/administração & dosagem , Esteroide 11-beta-Hidroxilase , Esteroide 17-alfa-Hidroxilase , 3-Hidroxiesteroide Desidrogenases/antagonistas & inibidores , 3-Hidroxiesteroide Desidrogenases/biossíntese , Corticosteroides/metabolismo , Androstenodiona/análogos & derivados , Animais , Linhagem Celular Tumoral , Enzima de Clivagem da Cadeia Lateral do Colesterol/antagonistas & inibidores , Enzima de Clivagem da Cadeia Lateral do Colesterol/biossíntese , Corticosterona/biossíntese , Corticosterona/metabolismo , Cortisona/metabolismo , Humanos , Progesterona/análogos & derivados , Progesterona/biossíntese , Esteroide 11-beta-Hidroxilase/antagonistas & inibidores , Esteroide 11-beta-Hidroxilase/biossíntese , Esteroide 17-alfa-Hidroxilase/antagonistas & inibidores , Esteroide 17-alfa-Hidroxilase/biossíntese , Relação Estrutura-Atividade
18.
Endocrine ; 43(1): 184-90, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22798247

RESUMO

The objective of the present study was to characterize the effect of insulin plus hCG on the expression of steroidogenic enzymes (P450scc and CYP17) in polycystic ovaries of rats. Changes in estrous cycle, ovarian morphology, hormonal levels, and protein levels by immunohistochemistry and western-blot were determined. Rats treated with insulin plus hCG displayed abnormal estrous cycles with increasing androgen biosynthesis. Meanwhile, insulin plus hCG resulted in multiple large cysts with diminished granulosa layers and increased thecal layers and stromal-interstitial tissue. Moreover, there was an increase in the expression of P450scc and CYP17 in thecal and stromal cells in our PCOS rat model compared with control rats. These results indicate that administration of insulin with hCG can synergistically result in endogenous hyperandrogenism which may partially upregulate the expression of steroidogenic enzymes in ovarian tissue.


Assuntos
Enzima de Clivagem da Cadeia Lateral do Colesterol/biossíntese , Modelos Animais de Doenças , Indução Enzimática , Hiperandrogenismo/etiologia , Ovário/enzimologia , Síndrome do Ovário Policístico/metabolismo , Esteroide 17-alfa-Hidroxilase/biossíntese , Androstenodiona/sangue , Animais , Western Blotting , Enzima de Clivagem da Cadeia Lateral do Colesterol/metabolismo , Feminino , Imuno-Histoquímica , Ovário/metabolismo , Ovário/patologia , Síndrome do Ovário Policístico/enzimologia , Síndrome do Ovário Policístico/patologia , Síndrome do Ovário Policístico/fisiopatologia , Ratos , Ratos Sprague-Dawley , Esteroide 17-alfa-Hidroxilase/metabolismo , Células Estromais/enzimologia , Células Estromais/metabolismo , Células Estromais/patologia , Testosterona/sangue , Células Tecais/enzimologia , Células Tecais/metabolismo , Células Tecais/patologia , Regulação para Cima
19.
Gene ; 512(2): 444-9, 2013 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-23124045

RESUMO

Cytochrome P450c17 (CYP17, 17α-hydroxylase/17, 20-lyase) plays a critical role in the production of androgens and estrogens in vertebrates. We isolated the full length cDNAs of P450c17-I and P450c17-II from Sebastes schlegeli. The cDNA sequences of P450c17-I and P450c17-II encoded 515 and 533 amino acid residues respectively. The putative P450c17-I and P450c17-II enzymes of Korean rockfish share high sequence identity with that of Japanese flounder (92% and 81%) respectively. Our current study describes that P450c17s of Korean rockfish are mainly expressed in gonads, head kidney and kidney by RT-PCR. Quantitative real-time PCR showed that the expression patterns of Korean rockfish P450c17s were developmental stage-dependency. In addition, the testosterone (T) and gonadosomatic index (GSI) levels further support the important role of P450c17-I during shift in steroidogenesis. Taken together, this study provides information about the Korean rockfish P450c17s characterization and mRNA expression as such helps in further understanding of its function in gonadal development.


Assuntos
Regulação Enzimológica da Expressão Gênica/fisiologia , Perciformes/metabolismo , Esteroide 17-alfa-Hidroxilase/biossíntese , Animais , Clonagem Molecular , DNA Complementar/genética , Proteínas de Peixes , Peixes , Isoenzimas/biossíntese , Isoenzimas/genética , Especificidade de Órgãos/fisiologia , Perciformes/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Esteroide 17-alfa-Hidroxilase/genética
20.
Cell Tissue Res ; 350(3): 513-23, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23053053

RESUMO

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.


Assuntos
Enzima de Clivagem da Cadeia Lateral do Colesterol/biossíntese , Hormônio Liberador de Gonadotropina/agonistas , Nafarelina/análogos & derivados , Fosfoproteínas/biossíntese , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroides/biossíntese , Testículo/fisiologia , Animais , Enzima de Clivagem da Cadeia Lateral do Colesterol/genética , Cães , Regulação para Baixo/efeitos dos fármacos , Implantes de Medicamento , Hormônio Liberador de Gonadotropina/genética , Hormônio Liberador de Gonadotropina/metabolismo , Masculino , Nafarelina/administração & dosagem , Fosfoproteínas/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Esteroide 17-alfa-Hidroxilase/genética , Testículo/efeitos dos fármacos , Testículo/metabolismo
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