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1.
PLoS One ; 15(8): e0237430, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32841307

RESUMO

BACKGROUND & AIMS: Given ongoing challenges in non-invasive non-alcoholic liver disease (NAFLD) diagnosis, we sought to validate an ALT-based NAFLD phenotype using measures readily available in electronic health records (EHRs) and population-based studies by leveraging the clinical and genetic data in the Million Veteran Program (MVP), a multi-ethnic mega-biobank of US Veterans. METHODS: MVP participants with alanine aminotransferases (ALT) >40 units/L for men and >30 units/L for women without other causes of liver disease were compared to controls with normal ALT. Genetic variants spanning eight NAFLD risk or ALT-associated loci (LYPLAL1, GCKR, HSD17B13, TRIB1, PPP1R3B, ERLIN1, TM6SF2, PNPLA3) were tested for NAFLD associations with sensitivity analyses adjusting for metabolic risk factors and alcohol consumption. A manual EHR review assessed performance characteristics of the NAFLD phenotype with imaging and biopsy data as gold standards. Genetic associations with advanced fibrosis were explored using FIB4, NAFLD Fibrosis Score and platelet counts. RESULTS: Among 322,259 MVP participants, 19% met non-invasive criteria for NAFLD. Trans-ethnic meta-analysis replicated associations with previously reported genetic variants in all but LYPLAL1 and GCKR loci (P<6x10-3), without attenuation when adjusted for metabolic risk factors and alcohol consumption. At the previously reported LYPLAL1 locus, the established genetic variant did not appear to be associated with NAFLD, however the regional association plot showed a significant association with NAFLD 279kb downstream. In the EHR validation, the ALT-based NAFLD phenotype yielded a positive predictive value 0.89 and 0.84 for liver biopsy and abdominal imaging, respectively (inter-rater reliability (Cohen's kappa = 0.98)). HSD17B13 and PNPLA3 loci were associated with advanced fibrosis. CONCLUSIONS: We validate a simple, non-invasive ALT-based NAFLD phenotype using EHR data by leveraging previously established NAFLD risk-associated genetic polymorphisms.


Assuntos
Alanina Transaminase/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , 17-Hidroxiesteroide Desidrogenases/genética , Abdome/diagnóstico por imagem , Proteínas Adaptadoras de Transdução de Sinal/genética , Idoso , Alanina Transaminase/genética , Registros Eletrônicos de Saúde , Feminino , Loci Gênicos , Predisposição Genética para Doença , Variação Genética , Humanos , Lipase/genética , Fígado/patologia , Lisofosfolipase/genética , Masculino , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Hepatopatia Gordurosa não Alcoólica/etnologia , Hepatopatia Gordurosa não Alcoólica/genética , Fenótipo , Fatores de Risco , Veteranos
2.
Sci Rep ; 10(1): 4040, 2020 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-32132633

RESUMO

Flaviviridae infections represent a major global health burden. By deciphering mechanistic aspects of hepatitis C virus (HCV)-host interactions, one could discover common strategy for inhibiting the replication of related flaviviruses. By elucidating the HCV interactome, we identified the 17-beta-hydroxysteroid dehydrogenase type 12 (HSD17B12) as a human hub of the very-long-chain fatty acid (VLCFA) synthesis pathway and core interactor. Here we show that HSD17B12 knockdown (KD) impairs HCV replication and reduces virion production. Mechanistically, depletion of HSD17B12 induces alterations in VLCFA-containing lipid species and a drastic reduction of lipid droplets (LDs) that play a critical role in virus assembly. Oleic acid supplementation rescues viral RNA replication and production of infectious particles in HSD17B12 depleted cells, supporting a specific role of VLCFA in HCV life cycle. Furthermore, the small-molecule HSD17B12 inhibitor, INH-12, significantly reduces replication and infectious particle production of HCV as well as dengue virus and Zika virus revealing a conserved requirement across Flaviviridae virus family. Overall, the data provide a strong rationale for the advanced evaluation of HSD17B12 inhibition as a promising broad-spectrum antiviral strategy for the treatment of Flaviviridae infections.


Assuntos
17-Hidroxiesteroide Desidrogenases/metabolismo , Hepacivirus/fisiologia , Hepatite C/enzimologia , Ácido Oleico/farmacologia , Replicação Viral/efeitos dos fármacos , 17-Hidroxiesteroide Desidrogenases/genética , Animais , Chlorocebus aethiops , Células HeLa , Células Hep G2 , Hepatite C/genética , Humanos , Células Vero , Replicação Viral/genética
3.
J Steroid Biochem Mol Biol ; 199: 105586, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-31926269

RESUMO

Recent studies have shown that an adrenal steroid 11ß-hydroxy-4-androstene-3,17-dione serves as the precursor to androgens, 11-ketotestosterone and 11-ketodihydrotestosterone (11KDHT). The biosynthetic pathways include the reduction of 3- and 17-keto groups of the androgen precursors 11-keto-C19-steroids, which has been reported to be mediated by three human enzymes; aldo-keto reductase (AKR)1C2, AKR1C3 and 17ß-hydroxysteroid dehydrogenase (HSD) type-3. To explore the contribution of the enzymes in the reductive metabolism, we kinetically compared the substrate specificity for 11-keto-C19-steroids among purified recombinant preparations of four AKRs (1C1, 1C2,1C3 and 1C4) and DHRS11, which shows 17ß-HSD activity. Although AKR1C1 did not reduce the 11-keto-C19-steroids, AKR1C3 and DHRS11 reduced 17-keto groups of 11-keto-4-androstene-3,17-dione, 11-keto-5α-androstane-3,17-dione (11K-Adione) and 11-ketoandrosterone with Km values of 5-28 µM. The 3-keto groups of 11KDHT and 11K-Adione were reduced by AKR1C4 (Km 1 µM) more efficiently than by AKR1C2 (Km 5 and 8 µM, respectively). GC/MS analysis of the products showed that DHRS11 acts as 17ß-HSD, and that AKR1C2 and AKR1C4 are predominantly 3α-HSDs, but formed a minor 3ß-metabolite from 11KDHT. Since DHRS11 was thus newly identified as 11-keto-C19-steroid reductase, we also investigated its substrate-binding mode by molecular docking and site-directed mutagenesis of Thr163 and Val200, and found the following structural features: 1). There is a space that accommodates the 11-keto group of the 11-keto-C19-steroids in the substrate-binding site. 2) Val200 is a critical determinant for exhibiting the strict 17ß-HSD activity of the enzyme, because the Val200Leu mutation resulted in both significant impairment of the 17ß-HSD activity and emergence of 3ß-HSD activity towards 5α-androstanes including 11KDHT.


Assuntos
17-Hidroxiesteroide Desidrogenases/química , 20-Hidroxiesteroide Desidrogenases/química , Aldo-Ceto Redutases/química , Esteroides/biossíntese , 17-Hidroxiesteroide Desidrogenases/genética , 17-Hidroxiesteroide Desidrogenases/metabolismo , 20-Hidroxiesteroide Desidrogenases/genética , 20-Hidroxiesteroide Desidrogenases/metabolismo , Membro C3 da Família 1 de alfa-Ceto Redutase/química , Membro C3 da Família 1 de alfa-Ceto Redutase/genética , Membro C3 da Família 1 de alfa-Ceto Redutase/metabolismo , Aldo-Ceto Redutases/genética , Aldo-Ceto Redutases/metabolismo , Androgênios/biossíntese , Androgênios/química , Vias Biossintéticas/genética , Humanos , Simulação de Acoplamento Molecular , Oxirredutases/química , Oxirredutases/genética , Oxirredutases/metabolismo , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Esteroides/química , Especificidade por Substrato , Testosterona/análogos & derivados , Testosterona/metabolismo
4.
J Steroid Biochem Mol Biol ; 199: 105597, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-31958634

RESUMO

Sex steroid hormones play important roles in fish sex differentiation, gonadal development and secondary sexual characteristics. Olive flounder Paralichthys olivaceus is a valuable commercial marine fish species and has marked sexual dimorphism. However, the mechanisms of action of sex hormones in flounder sex are still unclear. In this study, a total of ten Hsd17b family genes, including Hsd17b3, -4, -7, -8, -9, -10, -12a, -12b, -14 and -15, were identified in the flounder, which encoded critical enzymes acting on sex steroid synthesis and metabolism. Hsd17b genes were distributed on eight chromosomes. Hsd17b12a and -12b were located on chromosomes 19 and 7, respectively. It was speculated that these two genes were just highly similar rather than different transcripts derived from the same gene. According to the results of domain and motif analyses, they all belonged to the SDR superfamily and contained conserved Hsd17b motifs TGxxxGxG, PGxxxT, NNAG and YxxxK. Analysis of amino acid sequences predicted that Hsd17b1, -4, -7, -12a and -14 were hydrophilic proteins. The stability of Hsd17b1, -3 and -12b proteins was predicted to be low. The various Hsd17b family genes differed in tissue expression pattern, and Hsd17b10, -12a and -12b were highly expressed in the flounder ovary. Moreover, throughout gonadal development, Hsd17b3 was highly expressed in the testis, and Hsd17b1, -12a and -12b were highly expressed in the ovary, suggesting that they might play an important role in testosterone synthesis in the testis or estrogen synthesis in the ovary. Activities of Hsd17b3 at stages I-V were all significantly higher in the testis than in the ovary (P < 0.05, P < 0.01). Transfection analysis in HEK293T cells showed that Hsd17b1 and -3 were located in both the cytoplasm and nucleus. Additionally, after challenging fish with tamoxifen, Hsd17b3 expression level in the testis decreased significantly (P < 0.01), and in the ovary no significant change was observed. Moreover, the expression of Hsd17b1 in the ovary was significantly upregulated after injection with flutamide (P < 0.05). These findings introduce the characteristics of the flounder Hsd17b in subfamily, which contribute to our understanding of the regulation of sex steroid hormone synthesis in fish gonadal development.


Assuntos
17-Hidroxiesteroide Desidrogenases/genética , Proteínas de Peixes/genética , Linguado/genética , Hormônios Esteroides Gonadais/genética , 17-Hidroxiesteroide Desidrogenases/química , Sequência de Aminoácidos/genética , Animais , Feminino , Regulação da Expressão Gênica no Desenvolvimento/genética , Hormônios Esteroides Gonadais/biossíntese , Gônadas/crescimento & desenvolvimento , Gônadas/metabolismo , Masculino , Família Multigênica/genética , Ovário/crescimento & desenvolvimento , Ovário/metabolismo , Caracteres Sexuais , Testículo/crescimento & desenvolvimento , Testículo/metabolismo
5.
J Steroid Biochem Mol Biol ; 196: 105494, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31610224

RESUMO

In spite of the significant progress of estrogen-dependent breast cancer (BC) treatment, aromatase inhibitor resistance is a major problem limiting the clinical benefit of this frontier endocrine-therapy. The aim of this study was to determine the differential expression of steroid-converting enzymes between tumor and adjacent normal tissues, as well as their correlation in modulating intratumoral steroid-hormone levels in post-menopausal estrogen-dependent BC. RNA sequencing dataset (n = 1097) of The-Cancer-Genome-Atlas (Breast Invasive Carcinoma) retrieved through the data portal of Genomic Data Commons was used for differential expressions and expression correlation analyses by Mann-Whitney U and Spearman's rank test, respectively. The results showed significant up-regulation of 17ß-HSD7 (2.50-fold, p < 0.0001) in BC, supporting its effect in sex-hormone control. Besides, suppression of 11ß-HSD1 expression (-8.29-fold, p < 0.0001) and elevation of 11ß-HSD2 expression (2.04-fold, p < 0.0001) provide a low glucocorticoid environment diminishing BC anti-proliferation. Furthermore, 3α-HSDs were down-regulated (-1.59-fold, p < 0.01; -8.18-fold, p < 0.0001; -33.96-fold, p < 0.0001; -31.85-fold, p < 0.0001 for type 1-4, respectively), while 5α-reductases were up-regulated (1.41-fold, p < 0.0001; 2.85-fold, p < 0.0001; 1.70-fold, p < 0.0001 for type 1-3, respectively) in BC, reducing cell proliferation suppressers 4-pregnenes, increasing cell proliferation stimulators 5α-pregnanes. Expression analysis indicates significant correlations between 11ß-HSD1 with 3α-HSD4 (r = 0.605, p < 0.0001) and 3α-HSD3 (r = 0.537, p < 0.0001). Significant expression correlations between 3α-HSDs were also observed. Our results systematically present the regulation of steroid-converting enzymes and their roles in modulating the intratumoral steroid-hormone levels in BC with a vivid 3D-schema, supporting novel therapy targeting the reductive 17ß-HSD7 and proposing a new combined therapy targeting 11ß-HSD2 and 17ß-HSD7.


Assuntos
Neoplasias da Mama/genética , Carcinoma Ductal de Mama/genética , Sistema Enzimático do Citocromo P-450/genética , Hormônios Esteroides Gonadais/genética , Receptores Citoplasmáticos e Nucleares/genética , 17-Hidroxiesteroide Desidrogenases/genética , 17-Hidroxiesteroide Desidrogenases/metabolismo , Neoplasias da Mama/epidemiologia , Neoplasias da Mama/metabolismo , Carcinoma Ductal de Mama/epidemiologia , Carcinoma Ductal de Mama/metabolismo , Estudos de Coortes , Sistema Enzimático do Citocromo P-450/metabolismo , Bases de Dados Factuais/estatística & dados numéricos , Estradiol/farmacologia , Feminino , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Hormônios Esteroides Gonadais/metabolismo , Humanos , Setor Público/estatística & dados numéricos , Receptores Citoplasmáticos e Nucleares/metabolismo
6.
J Steroid Biochem Mol Biol ; 196: 105493, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31614207

RESUMO

17ß-Hydroxysteroid dehydrogenases (17ß-HSDs) catalyze the reduction of 17-ketosteroids and the oxidation of 17ß-hydroxysteroids to regulate the production of androgens and estrogens. Among them, 17ß-HSD type 3 (HSD17B3) is expressed almost exclusively in testicular Leydig cells and contributes to development of male sexual characteristics by converting androstenedione (A4) to testosterone (T). Mutations of HSD17B3 genes cause a 46,XY disorder of sexual development (46,XY DSD) as a result of low T production. Therefore, the evaluation of 17ß-HSD3 enzymatic activity is important for understanding and diagnosing this disorder. We adapted a method that easily evaluates enzymatic activity of 17ß-HSD3 by quantifying the conversion from A4 to T using androgen receptor (AR)-mediated transactivation. HEK293 cells were transduced to express human HSD17B3, and incubated medium containing A4. Depending on the incubation time with HSD17B3-expressing cells, the culture media progressively increased luciferase activities in CV-1 cells, transfected with the AR expression vector and androgen-responsive reporter. Culture medium from HSD17B1 and HSD17B5-expressing cells also increased the luciferase activities. This system is also applicable to detect the conversion of 11-ketoandrostenedione to 11-ketotestosterone by HSD17B3. Establishment of HEK293 cells expressing various missense mutations in the HSD17B3 gene associated with 46,XY DSD revealed that this system is effective to evaluate the enzymatic activities of mutant proteins.


Assuntos
17-Hidroxiesteroide Desidrogenases/metabolismo , Receptores Androgênicos/fisiologia , Ativação Transcricional/genética , 17-Hidroxiesteroide Desidrogenases/genética , Animais , Células Cultivadas , Transtorno 46,XY do Desenvolvimento Sexual/genética , Transtorno 46,XY do Desenvolvimento Sexual/metabolismo , Ativação Enzimática/genética , Indução Enzimática/genética , Células HEK293 , Humanos , Mutação de Sentido Incorreto/fisiologia , Transfecção
7.
Nat Rev Gastroenterol Hepatol ; 17(1): 40-52, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31641249

RESUMO

Nonalcoholic fatty liver disease (NAFLD) affects around a quarter of the global population, paralleling worldwide increases in obesity and metabolic syndrome. NAFLD arises in the context of systemic metabolic dysfunction that concomitantly amplifies the risk of cardiovascular disease and diabetes. These interrelated conditions have long been recognized to have a heritable component, and advances using unbiased association studies followed by functional characterization have created a paradigm for unravelling the genetic architecture of these conditions. A novel perspective is to characterize the shared genetic basis of NAFLD and other related disorders. This information on shared genetic risks and their biological overlap should in future enable the development of precision medicine approaches through better patient stratification, and enable the identification of preventive and therapeutic strategies. In this Review, we discuss current knowledge of the genetic basis of NAFLD and of possible pleiotropy between NAFLD and other liver diseases as well as other related metabolic disorders. We also discuss evidence of causality in NAFLD and other related diseases and the translational significance of such evidence, and future challenges from the study of genetic pleiotropy.


Assuntos
Hepatopatia Gordurosa não Alcoólica/genética , 17-Hidroxiesteroide Desidrogenases/genética , Aciltransferases/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Carcinoma Hepatocelular/genética , Causalidade , Progressão da Doença , Pleiotropia Genética , Predisposição Genética para Doença , Humanos , Lipase/genética , Cirrose Hepática Alcoólica/genética , Neoplasias Hepáticas/genética , Proteínas de Membrana/genética , Análise da Randomização Mendeliana , Síndrome Metabólica/genética , Biologia de Sistemas
9.
Hepatology ; 71(1): 56-66, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31155741

RESUMO

A common loss-of-function variant in HSD17B13 (rs72613567:TA) was recently found to protect from chronic liver disease. Whether the variant confers protection from specific risk factors for liver disease is unclear. We tested the association of rs72613567 with plasma levels of alanine transaminase (ALT) and clinical liver disease and mortality in 111,612 individuals from the Danish general population, including 497 with cirrhosis and 113 with hepatocellular carcinoma. HSD17B13 rs72613567:TA was associated with stepwise lower levels of plasma ALT of up to 1.3 U/L in TA/TA homozygotes versus T/T homozygotes. For each TA-allele, the risk of cirrhosis and hepatocellular carcinoma was reduced by 15% and 28%, respectively. In prospective analyses, the TA-allele was associated with up to 33% lower rates of liver-related mortality in the general population, and with up to 49% reduced liver-related mortality in patients with cirrhosis. The ALT-lowering effect of rs72613567:TA was amplified by increasing adiposity, alcohol consumption, and genetic risk of fatty liver disease. The TA-allele was associated with only marginally lower ALT in lean nondrinkers with low genetic risk of hepatic steatosis. In contrast, compared with T/T homozygotes, TA/TA homozygotes had 12% to 18% lower plasma ALT among the most obese, in heavy drinkers, and in individuals carrying three or four steatogenic alleles in patatin-like phospholipase domain-containing protein 3 (PNPLA3) and transmembrane 6 superfamily 2 (TM6SF2). Conclusion: High risk of fatty liver disease amplifies the ALT-lowering effect of HSD17B13 rs72613567:TA in the Danish general population.


Assuntos
17-Hidroxiesteroide Desidrogenases/genética , Alanina Transaminase/sangue , Carcinoma Hepatocelular/epidemiologia , Fígado Gorduroso/epidemiologia , Fígado Gorduroso/genética , Cirrose Hepática/epidemiologia , Neoplasias Hepáticas/complicações , Neoplasias Hepáticas/epidemiologia , Idoso , Carcinoma Hepatocelular/complicações , Dinamarca , Fígado Gorduroso/complicações , Feminino , Variação Genética , Humanos , Cirrose Hepática/complicações , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Medição de Risco
10.
Cell Mol Life Sci ; 77(6): 1153-1175, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31302749

RESUMO

Metabolic reprogramming of tumor cells involves upregulation of fatty acid (FA) synthesis to support high bioenergetic demands and membrane synthesis. This has been shown for cytosolic synthesis of FAs with up to 16 carbon atoms. Synthesis of long-chain fatty acids (LCFAs), including ω-6 and ω-3 polyunsaturated FAs, takes place at the endoplasmic reticulum. Despite increasing evidence for an important role of LCFAs in cancer, the impact of their synthesis in cancer cell growth has scarcely been studied. Here, we demonstrated that silencing of 17ß-hydroxysteroid dehydrogenase type 12 (17ß-HSD12), essentially catalyzing the 3-ketoacyl-CoA reduction step in LCFA production, modulates proliferation and migration of breast cancer cells in a cell line-dependent manner. Increased proliferation and migration after 17ß-HSD12 knockdown were partly mediated by metabolism of arachidonic acid towards COX2 and CYP1B1-derived eicosanoids. Decreased proliferation was rescued by increased glucose concentration and was preceded by reduced ATP production through oxidative phosphorylation and spare respiratory capacity. In addition, 17ß-HSD12 silencing was accompanied by alterations in unfolded protein response, including a decrease in CHOP expression and increase in eIF2α activation and the folding chaperone ERp44. Our study highlights the significance of LCFA biosynthesis for tumor cell physiology and unveils unknown aspects of breast cancer cell heterogeneity.


Assuntos
17-Hidroxiesteroide Desidrogenases/metabolismo , Neoplasias da Mama/metabolismo , 17-Hidroxiesteroide Desidrogenases/genética , Acil Coenzima A/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Ácidos Graxos/metabolismo , Feminino , Inativação Gênica , Humanos , Lipogênese , Células MCF-7
11.
Int J Mol Sci ; 21(24)2020 Dec 20.
Artigo em Inglês | MEDLINE | ID: mdl-33419257

RESUMO

In early stages of Alzheimer's disease (AD), amyloid beta (Aß) accumulates in the mitochondrial matrix and interacts with mitochondrial proteins, such as cyclophilin D (cypD) and 17ß-hydroxysteroid dehydrogenase 10 (17ß-HSD10). Multiple processes associated with AD such as increased production or oligomerization of Aß affect these interactions and disbalance the equilibrium between the biomolecules, which contributes to mitochondrial dysfunction. Here, we investigate the effect of the ionic environment on the interactions of Aß (Aß1-40, Aß1-42) with cypD and 17ß-HSD10 using a surface plasmon resonance (SPR) biosensor. We show that changes in concentrations of K+ and Mg2+ significantly affect the interactions and may increase the binding efficiency between the biomolecules by up to 35% and 65% for the interactions with Aß1-40 and Aß1-42, respectively, in comparison with the physiological state. We also demonstrate that while the binding of Aß1-40 to cypD and 17ß-HSD10 takes place preferentially around the physiological concentrations of ions, decreased concentrations of K+ and increased concentrations of Mg2+ promote the interaction of both mitochondrial proteins with Aß1-42. These results suggest that the ionic environment represents an important factor that should be considered in the investigation of biomolecular interactions taking place in the mitochondrial matrix under physiological as well as AD-associated conditions.


Assuntos
Doença de Alzheimer/genética , Peptídeos beta-Amiloides/química , Técnicas Biossensoriais/métodos , Ressonância de Plasmônio de Superfície/métodos , 17-Hidroxiesteroide Desidrogenases/química , 17-Hidroxiesteroide Desidrogenases/genética , Doença de Alzheimer/diagnóstico , Doença de Alzheimer/patologia , Ciclofilina D/química , Ciclofilina D/genética , Humanos , Íons/química , Mitocôndrias/química , Proteínas Mitocondriais/química , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genética
12.
Reprod Biol Endocrinol ; 17(1): 111, 2019 Dec 26.
Artigo em Inglês | MEDLINE | ID: mdl-31878927

RESUMO

BACKGROUND: Previous studies of expression profiles of major endometrial effectors of steroid physiology in endometriosis have yielded markedly conflicting conclusions, presumably because the relative effects of type of endometriosis, fertility history and menstrual cycle phases on the measured variables were not considered. In the present study, endometrial mRNA and protein levels of several effectors of steroid biosynthesis and action in patients with stage III-IV ovarian endometriosis (OE) with known fertility and menstrual cycle histories were compared with the levels in control endometrium to test this concept. METHODS: Endometrial samples were collected from patients without endometriosis (n = 32) or OE stages III-IV (n = 52) with known fertility and cycle histories. qRT-PCR and immunoblotting experiments were performed to measure levels of NR5A1, STAR, CYP19A1, HSD17Bs, ESRs and PGR transcripts and proteins, respectively. Tissue concentrations of steroids (P4, T, E1 and E2) were measured using ELISAs. RESULTS: The levels of expression of aromatase and ERß were lower (P < 0.0001) and 17ß-HSD1 (P < 0.0001) and PRA (P < 0.01) were higher in OE endometrium. Lower aromatase levels and higher 17ß-HSD1 levels were detected in fertile (aromatase: P < 0.05; 17ß-HSD1: P < 0.0001) and infertile (aromatase: P < 0.0001; 17ß-HSD1: P < 0.0001) OE endometrium than in the matched control tissues. Both proliferative (PP) and secretory (SP) phase OE samples expressed aromatase (P < 0.0001) and ERß (PP: P < 0.001; SP: P < 0.01) at lower levels and 17ß-HSD1 (P < 0.0001) and PRA (PP: P < 0.01; SP: P < 0.0001) at higher levels than matched controls. Higher 17ß-HSD1 (P < 0.01) and E2 (P < 0.05) levels and a lower (P < 0.01) PRB/PRA ratio was observed in infertile secretory phase OE endometrium than in control. CONCLUSIONS: We report that dysregulated expression of 17ß-HSD1 and PGR resulting in hyperestrogenism and progesterone resistance during the secretory phase of the menstrual cycle, rather than an anomaly in aromatase expression, was the hallmark of eutopic endometrium from infertile OE patients. Furthermore, the results provide proof of concept that the fertility and menstrual cycle histories exerted relatively different effects on steroid physiology in the endometrium from OE patients compared with the control subjects.


Assuntos
Endometriose/metabolismo , Endométrio/metabolismo , Hormônios Esteroides Gonadais/metabolismo , Doenças Ovarianas/metabolismo , Receptores de Esteroides/metabolismo , 17-Hidroxiesteroide Desidrogenases/análise , 17-Hidroxiesteroide Desidrogenases/genética , Adolescente , Adulto , Aromatase/análise , Aromatase/genética , Endométrio/química , Estradiol/análise , Feminino , Expressão Gênica , Humanos , Infertilidade Feminina/metabolismo , Ciclo Menstrual , Progesterona/análise , Receptores Estrogênicos/análise , Receptores de Progesterona/análise , Receptores de Esteroides/genética , Adulto Jovem
13.
J Steroid Biochem Mol Biol ; 195: 105483, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31550505

RESUMO

Hydroxysteroid 17-Beta Dehydrogenase 3 (Hsd17b3), primarily expressed in Leydig cells (LCs) of the mammalian testes, is essential for testosterone biosynthesis and male fertility. The aim of our study was to profile the expression, splice variants (SV) and novel insertion/deletion (indel) of Hsd17b3 in boars. Quantitative analysis showed that the expression level of Hsd17b3 in the testis was significantly highest. Among different testicular cell types, the Hsd17b3 mRNA expression level of LCs was significantly higher than that of SSCs (spermatogonial stem cells) and SCs (Sertoli cells). Furthermore, the SV was firstly identified in pigs and it was highly expressed in LCs comparing with SSCs and SCs. In addition, two mutations were identified in pig Hsd17b3 gene promotor and intron, respectively, which were associated with male reproductive traits (P <  0.05). In conclusion, both transcripts of Hsd17b3 gene were highly expressed in pig testes and LCs; the two novel indel variants of Hsd17b3 gene can be used as potential DNA makers for the marker-assisted selection in pigs. All these findings would enrich the study of Hsd17b3 gene in pig genetic breeding.


Assuntos
17-Hidroxiesteroide Desidrogenases/genética , Células Intersticiais do Testículo/metabolismo , Reprodução/genética , Sus scrofa/genética , Células-Tronco Germinativas Adultas/metabolismo , Processamento Alternativo , Animais , Variação Genética , Mutação INDEL , Masculino , Regiões Promotoras Genéticas , Células de Sertoli/metabolismo
14.
Anim Genet ; 50(6): 705-711, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31476086

RESUMO

The genetic background of disorders of sex development (DSD) in dogs with a normal male sex chromosome set (78,XY) is poorly described. In this study, we present for the first time, an analysis of six genes of the testosterone pathway, encoding enzymes (CYP17A1, HSD3B2, HSD17B3, SRD5A2) and transcription factors (NR5A1, AR). The entire coding sequence and flanking regions of the introns, 5'-UTR and 3'-UTR were analyzed in five DSD dogs (78,XY, SRY-positive) with ambiguous external genitalia and in 15 control dogs. A homozygous deletion of 2 bp in exon 2 of HSD17B3 (hydroxysteroid 17-beta dehydrogenase 3) was found in a Dachshund dog with enlarged clitoris, vulva and abdominal gonads and decreased serum testosterone level. In silico analysis revealed that this deleterious variant causes truncation of the encoded polypeptide (from 306 to 65 amino acids) and deprivation of the active site of the encoded enzyme. Genotyping of 23 control Dachshund dogs showed a normal homozygous genotype. Thus, we assumed that the 2-bp deletion is the causative variant. Moreover, 24 SNPs (four in CYP17A1, three in HSD3B2, six in HSD17B3, five in SRD5A2, one in AR and five in NR5A1), two intronic indels (one in HSD3B2 and one in SRD5A2) and two microsatellite polymorphisms in exon 1 of AR were found. Six SNPs appeared to be novel. No association with DSD phenotype was observed. Identification of the first case of DSD in domestic animals caused by a deleterious variant of a gene involved in testosterone synthesis showed that these genes are important candidates in such studies.


Assuntos
Transtornos do Desenvolvimento Sexual/veterinária , Doenças do Cão/genética , Testosterona/biossíntese , 17-Hidroxiesteroide Desidrogenases/genética , Animais , Códon de Terminação , Transtornos do Desenvolvimento Sexual/genética , Transtornos do Desenvolvimento Sexual/patologia , Cães , Feminino , Deleção de Genes , Genitália/patologia , Masculino , Testosterona/sangue
15.
J Gastrointestin Liver Dis ; 28(3): 297-302, 2019 Sep 12.
Artigo em Inglês | MEDLINE | ID: mdl-31517326

RESUMO

BACKGROUND AND AIMS: Two single nucleotide polymorphisms (SNPs) in SERPINA1 (Pi*Z rs28929474 and Pi*S rs17580) are risk factors for developing liver cirrhosis. A recent study identified a common SNP in HSD17B13 (rs72613567) that conferred protection from chronic liver disease. The aim of the present study was to test these associations in a cohort of Lithuanian patients with liver fibrosis or cirrhosis. METHODS: The study included 302 patients with cirrhosis, 127 patients with liver fibrosis (METAVIR stages I-III) and 548 controls, all from Lithuania. SNPs were genotyped by quantitative PCR, using TaqMan allelic discrimination assays. Adjusted p value of ≤ 0.016 was considered significant. RESULTS: Genotype distributions of SERPINA1 and HSD17B13 SNPs were in Hardy-Weinberg equilibrium. SERPINA1 Pi*Z was not associated with liver fibrosis or cirrhosis. HSD17B13 rs10433937 (in high linkage disequilibrium with rs72613567; r 2 =0.96) also showed no overall association with liver disease, but the GG- genotype was associated with reduced risk of liver fibrosis (aOR 0.37, p=0.03). SERPINA1 Pi*S was associated with higher risk of developing hepatic fibrosis (aOR 3.42, p=0.001) and cirrhosis (aOR 2.59, p=0.02). CONCLUSIONS: We found that SERPINA1 Pi*S variant conferred an increased risk of developing liver fibrosis, while SERPINA1 Pi*Z and HSD17B13 rs10433937 were not associated with liver fibrosis or cirrhosis of different aetiology.


Assuntos
17-Hidroxiesteroide Desidrogenases/genética , Cirrose Hepática/genética , Polimorfismo de Nucleotídeo Único , alfa 1-Antitripsina/genética , Adulto , Idoso , Alelos , Estudos de Casos e Controles , Estudos de Coortes , Feminino , Predisposição Genética para Doença , Genótipo , Humanos , Cirrose Hepática/etiologia , Masculino , Pessoa de Meia-Idade , Fatores de Risco
16.
PLoS One ; 14(7): e0220492, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31348804

RESUMO

Rhodococcus ruber Chol-4 is a potent steroid degrader that has a great potential as a biotechnological tool. As proof of concept, this work presents testosterone production from 4-androstene-3,17-dione by tailoring innate catabolic enzymes of the steroid catabolism inside the strain. A R. ruber quadruple mutant was constructed in order to avoid the breakage of the steroid nucleus. At the same time, an inducible expression vector for this strain was developed. The 17-ketoreductase gene from the fungus Cochliobolus lunatus was cloned and overexpressed in this vector. The engineered strain was able to produce testosterone from 4-androstene-3,17-dione using glucose for cofactor regeneration with a molar conversion of 61%. It is important to note that 91% of the testosterone was secreted outside the cell after 3 days of cell biotransformation. The results support the idea that Rhodococcus ruber Chol-4 can be metabolically engineered and can be used for the production of steroid intermediates.


Assuntos
17-Hidroxiesteroide Desidrogenases/metabolismo , Proteínas de Bactérias/metabolismo , Engenharia Metabólica/métodos , Rhodococcus/genética , Rhodococcus/metabolismo , Testosterona/metabolismo , 17-Hidroxiesteroide Desidrogenases/genética , Proteínas de Bactérias/genética , Biotransformação , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Rhodococcus/crescimento & desenvolvimento
17.
J Strength Cond Res ; 33(9): 2344-2351, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31343553

RESUMO

Pickering, C, Suraci, B, Semenova, EA, Boulygina, EA, Kostryukova, ES, Kulemin, NA, Borisov, OV, Khabibova, SA, Larin, AK, Pavlenko, AV, Lyubaeva, EV, Popov, DV, Lysenko, EA, Vepkhvadze, TF, Lednev, EM, Leonska-Duniec, A, Pajak, B, Chycki, J, Moska, W, Lulinska-Kuklik, E, Dornowski, M, Maszczyk, A, Bradley, B, Kana-ah, A, Cieszczyk, P, Generozov, EV, and Ahmetov, II. A genome-wide association study of sprint performance in elite youth football players. J Strength Cond Res 33(9): 2344-2351, 2019-Sprint speed is an important component of football performance, with teams often placing a high value on sprint and acceleration ability. The aim of this study was to undertake the first genome-wide association study to identify genetic variants associated with sprint test performance in elite youth football players and to further validate the obtained results in additional studies. Using micro-array data (600 K-1.14 M single nucleotide polymorphisms [SNPs]) of 1,206 subjects, we identified 12 SNPs with suggestive significance after passing replication criteria. The polymorphism rs55743914 located in the PTPRK gene was found as the most significant for 5-m sprint test (p = 7.7 × 10). Seven of the discovered SNPs were also associated with sprint test performance in a cohort of 126 Polish women, and 4 were associated with power athlete status in a cohort of 399 elite Russian athletes. Six SNPs were associated with muscle fiber type in a cohort of 96 Russian subjects. We also examined genotype distributions and possible associations for 16 SNPs previously linked with sprint performance. Four SNPs (AGT rs699, HSD17B14 rs7247312, IGF2 rs680, and IL6 rs1800795) were associated with sprint test performance in this cohort. In addition, the G alleles of 2 SNPs in ADRB2 (rs1042713 & rs1042714) were significantly over-represented in these players compared with British and European controls. These results suggest that there is a genetic influence on sprint test performance in footballers, and identifies some of the genetic variants that help explain this influence.


Assuntos
Desempenho Atlético/fisiologia , Grupo com Ancestrais do Continente Europeu/genética , Corrida/fisiologia , Futebol/fisiologia , 17-Hidroxiesteroide Desidrogenases/genética , Aceleração , Adolescente , Alelos , Angiotensinogênio/genética , Criança , Estudos de Coortes , Feminino , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Fator de Crescimento Insulin-Like II/genética , Interleucina-6/genética , Masculino , Polônia , Polimorfismo de Nucleotídeo Único , Proteínas Tirosina Fosfatases Classe 2 Semelhantes a Receptores/genética , Receptores Adrenérgicos beta 2/genética , Federação Russa , Reino Unido , Adulto Jovem
18.
J Steroid Biochem Mol Biol ; 193: 105411, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31207361

RESUMO

Reductive 17ß-hydroxysteroid dehydrogenases (17ß-HSDs) and 11ß-hydroxysteroid dehydrogenase 2 (11ß-HSD2) play crucial roles in respectively regulating steroids and glucocorticoids for the progression of hormone-dependent breast cancer. Most studies focused on the function and individual regulation of these enzymes. However, mutual regulation of these enzymes and the induced modulation on the estrogen and androgen receptors for breast cancer promotion are not yet clear. In this study, MCF-7 and T47D cells were treated with inhibitors of 17ß-HSD1, 17ß-HSD7, aromatase or steroid sulfatase (STS), then mRNA levels of 17ß-HSD7, STS, 11ß-HSD 2, estrogen receptors α (ERα) and androgen receptor (AR) were determined by Q-PCR. ER negative cell line MDA-MB-231 was used as a negative control. Our results demonstrate that 17ß-HSD7, STS and 11ß-HSD2 are all regulated by the same estrogen estradiol via ERα. When the gene of ERα (ESR1) was knocked down, there was no longer significant mutual regulation of these enzymes. Our results demonstrate that important steroidogenic enzymes transcriptionally regulated by ERα, can be mutually closely correlated. Inhibition of one of them can reduce the expression of another, thereby amplifying the role of the inhibition. Furthermore, inhibition of 17ß-HSD7 increases the expression of AR gene which is considered as a marker for better prognosis in ER + breast cancer, while maintaining ERα level. Thus, our mechanistic finding provides a base for further improving the endocrine therapy of ER + breast cancer, e.g., for selecting the target steroid enzymes, and for the combined targeting of human 17ß-HSD7 and ERα.


Assuntos
17-Hidroxiesteroide Desidrogenases/antagonistas & inibidores , Neoplasias da Mama/genética , Receptor alfa de Estrogênio/genética , Receptores Androgênicos/genética , 11-beta-Hidroxiesteroide Desidrogenase Tipo 2/antagonistas & inibidores , 17-Hidroxiesteroide Desidrogenases/genética , Inibidores da Aromatase/farmacologia , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Di-Hidrotestosterona/metabolismo , Estradiol/metabolismo , Feminino , Humanos , Esteril-Sulfatase/antagonistas & inibidores , Esteril-Sulfatase/genética , Esteril-Sulfatase/metabolismo
19.
J Cell Physiol ; 234(12): 22819-22832, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31124138

RESUMO

CREBZF, including the two isoforms SMILE (long isoform of CREBZF) and Zhangfei (short isoform of CREBZF), has been identified as a novel transcriptional coregulator of a variety of nuclear receptors. Our previous studies found that SMILE is expressed in the mouse uterine luminal and glandular epithelium and is upregulated by estrogen. In the present study, CREBZF was age-dependently and -specifically expressed in mouse interstitial Leydig cells during sexual maturation. The expression pattern of CREBZF exhibited an age-related increase, and SMILE was the dominant isoform in the mouse testis. Although hCG did not affect CREBZF expression, CREBZF silencing significantly inhibited hCG-stimulated testosterone production in primary Leydig cells and MLTC-1 cells. Meanwhile, the serum concentration of testosterone was significantly decreased after microinjection of lentiviral-mediated shRNA-CREBZF into the mature mouse testis. In addition, CREBZF silencing markedly decreased P450c17, 17ß-HSD, and 3ß-HSD expression following hCG stimulation in primary Leydig cells, and this inhibitory effect was obviously reversed by overexpression of CREBZF. Furthermore, CREBZF significantly upregulated the mRNA levels of Nr4a1 and Nr5a1, which are the essential orphan nuclear receptors for steroidogenic gene expression. Together our data indicate that CREBZF promotes hCG-induced testosterone production in mouse Leydig cells by affecting Nr4a1 and Nr5a1 expression levels and subsequently increasing the expression of steroidogenic genes such as 3ß-HSD, 17ß-HSD, and P450c17, suggesting a potential important role of CREBZF in testicular testosterone synthesis.


Assuntos
Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Células Intersticiais do Testículo/metabolismo , Testosterona/biossíntese , 17-Hidroxiesteroide Desidrogenases/genética , 17-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Fatores de Transcrição de Zíper de Leucina Básica/genética , Linhagem Celular , Gonadotropina Coriônica/farmacologia , Regulação Enzimológica da Expressão Gênica , Células Intersticiais do Testículo/efeitos dos fármacos , Masculino , Camundongos , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/genética , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Cultura Primária de Células , Progesterona Redutase/genética , Progesterona Redutase/metabolismo , Maturidade Sexual , Esteroide 17-alfa-Hidroxilase/genética , Esteroide 17-alfa-Hidroxilase/metabolismo , Esteroide Isomerases/genética , Esteroide Isomerases/metabolismo , Fator Esteroidogênico 1/genética , Fator Esteroidogênico 1/metabolismo
20.
Toxicol In Vitro ; 60: 383-388, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31132478

RESUMO

Polybrominated Diphenyl Ethers (PBDEs) have been extensively applied as flame retardants in different polymeric materials since the 1970s, which have become a group of long-lasting environmental pollutants. They have been reported from previous studies to accumulate and then disrupt the endocrine system in humans. However, the mechanisms are still little known. In the present study, mouse Leydig tumor cells were utilized to investigate steroidogenic activity influenced by deca-brominated diphenyl ether (BDE-209). Our data showed that BDE-209 did not change intracellular cAMP level in the presence of human Chorionic Gonadotropin (hCG), cholera toxin (CT), and forskolin, which indicated that reduction of progesterone may not be related to the hCG-cAMP signal pathway in MLTC-1 cells. Furthermore, the reduction of progesterone generation was not shifted by 8-Br-cAMP, an analog of cAMP, indicating that BDE-209 may inhibit post-cAMP sites. In addition, mRNA expression levels of P450 side-chain cleavage enzyme (P450scc) and 3ß-hydroxysteroid dehydrogenase (3ß-HSD) presented a concentration-dependent decrease. In conclusion, this study suggested that BDE-209 may attenuate the progesterone secretion mainly through lowering the expression of these two enzymes.


Assuntos
Retardadores de Chama/toxicidade , Éteres Difenil Halogenados/toxicidade , Tumor de Células de Leydig/metabolismo , Progesterona/antagonistas & inibidores , 17-Hidroxiesteroide Desidrogenases/genética , 17-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Enzima de Clivagem da Cadeia Lateral do Colesterol/genética , Enzima de Clivagem da Cadeia Lateral do Colesterol/metabolismo , AMP Cíclico/metabolismo , Tumor de Células de Leydig/genética , Camundongos , Progesterona/metabolismo , RNA Mensageiro/metabolismo
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