RESUMO
Many sex differences in liver gene expression originate in the brain, depend on GH secretion and may underlie sex disparities in hepatic disease. Because epigenetic mechanisms may contribute, we studied promoter methylation and microRNA abundance in the liver, associated with expression of sexual dimorphic genes in mice with selective disruption of the dopamine D2 receptor in neurons (neuroDrd2KO), which decreases hypothalamic Ghrh, pituitary GH, and serum IGFI and in neonatally androgenized female mice which have increased pituitary GH content and serum IGFI. We evaluated mRNA levels of the female predominant genes prolactin receptor (Prlr), alcohol dehydrogenase 1 (Adh1), Cyp2a4, and hepatocyte nuclear transcription factor 6 (Hnf6) and the male predominant gene, Cyp7b1. Female predominant genes had higher mRNA levels compared to males, but lower methylation was only detected in the Prlr and Cyp2a4 female promoters. In neuroDrd2KO mice, sexual dimorphism was lost for all genes; the upregulation (feminization) of Prlr and Cyp2a4 in males correlated with decreased methylation of their promoters, and the downregulation (masculinization) of Hnf-6 mRNA in females correlated inversely with its promoter methylation. Neonatal androgenization of females evoked a loss of sexual dimorphism only for the female predominant Hnf6 and Adh1 genes, but no differences in promoter methylation were found. Finally, mmu-miR-155-5p, predicted to target Cyp7b1 expression, was lower in males in association with higher Cyp7b1 mRNA levels compared to females and was not modified in neuroDrd2KO or TP mice. Our results suggest specific regulation of gene sexually dimorphic expression in the liver by methylation or miRNAs.
Assuntos
Álcool Desidrogenase/genética , Hidrocarboneto de Aril Hidroxilases/genética , Família 2 do Citocromo P450/genética , Família 7 do Citocromo P450/genética , Hormônio do Crescimento/farmacologia , Fator 6 Nuclear de Hepatócito/genética , Receptores da Prolactina/genética , Esteroide Hidroxilases/genética , Álcool Desidrogenase/metabolismo , Animais , Hidrocarboneto de Aril Hidroxilases/metabolismo , Família 2 do Citocromo P450/metabolismo , Família 7 do Citocromo P450/metabolismo , Epigênese Genética/efeitos dos fármacos , Epigênese Genética/fisiologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Hormônio do Crescimento/metabolismo , Fator 6 Nuclear de Hepatócito/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Regiões Promotoras Genéticas/genética , Receptores da Prolactina/metabolismo , Caracteres Sexuais , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Esteroide Hidroxilases/metabolismoRESUMO
The fields of pharmacogenetics and pharmacogenomics have become increasingly promising regarding the clinical application of genetic data to aid in prevention of adverse reactions. Specific screening tests can predict which animals express modified proteins or genetic sequences responsible for adverse effects associated with a drug. Among the genetic variations that have been investigated in dogs, the multidrug resistance gene (MDR) is the best studied. However, other genes such as CYP1A2 and CYP2B11 control the protein syntheses involved in the metabolism of many drugs. In the present study, the MDR-1, CYP1A2 and CYP2B11 genes were examined to identify SNP polymorphisms associated with these genes in the following four canine breeds: Uruguayan Cimarron, Border Collie, Labrador Retriever and German Shepherd. The results revealed that several SNPs of the CYP1A2 and CYP2B11 genes are potential targets for drug sensitivity investigations.
Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Hidrocarboneto de Aril Hidroxilases/genética , Citocromo P-450 CYP1A2/genética , Cães/genética , Polimorfismo de Nucleotídeo Único , Esteroide Hidroxilases/genética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Animais , Hidrocarboneto de Aril Hidroxilases/metabolismo , Citocromo P-450 CYP1A2/metabolismo , Família 2 do Citocromo P450 , Cães/metabolismo , Esteroide Hidroxilases/metabolismoRESUMO
Placenta is an important source of endocrine and immunological factors. During pregnancy, calcitriol, the active metabolite of vitamin D, is also metabolized by decidua and placental tissue by means of CYP27B1 and CYP24A1 for synthesis and inactivation of calcitriol respectively. Calcitriol production is regulated by several factors in a tissue-specific manner. However, the association of pro-inflammatory cytokines on calcitriol metabolism has not been studied in human placenta. The aim of the present study was to investigate the effects of TNF-α, INF-γ, IL-6 and IL-1ß upon CYP27B1 and CYP24A1 gene expression in primary cultures of human placental cells. Placentas were obtained immediately after delivery by cesarean section from normotensive women. Cytokine effects upon mRNA of CYPs in enriched trophoblastic cell preparations were evaluated by using qPCR. The results showed that incubation of trophoblasts in the presence of each cytokine resulted in a significant increase of both CYPs expression. Interestingly, TNF-α increased significantly the ratio of CYP24A1/CYP27B1 gene expression, while IFN-γ preferentially induced CYP27B1, whereas IL-1ß and IL-6 stimulated gene expression of both CYPs in the same proportion. The results suggest that cytokines among other factors regulate calcitriol metabolism in human placenta; specifically, INF-γ may contribute to calcitriol production while TNF-α favors its catabolism.
Assuntos
25-Hidroxivitamina D3 1-alfa-Hidroxilase/metabolismo , Interferon gama/farmacologia , Interleucina-1beta/farmacologia , Interleucina-6/farmacologia , Esteroide Hidroxilases/metabolismo , Trofoblastos/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , 25-Hidroxivitamina D3 1-alfa-Hidroxilase/genética , Antivirais/farmacologia , Células Cultivadas , Ensaio de Imunoadsorção Enzimática , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Placenta/citologia , Placenta/efeitos dos fármacos , Placenta/metabolismo , Gravidez , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Recombinantes/genética , Proteínas Recombinantes/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Esteroide Hidroxilases/genética , Trofoblastos/citologia , Trofoblastos/efeitos dos fármacos , Vitamina D3 24-HidroxilaseRESUMO
Neonatal androgenization masculinizes the GH axis and thus may impact on liver gene regulation. Neonatal testosterone administration to female mice decreased (defeminized) female predominant GH-dependent liver gene expression (Hnf6, Adh1, Prlr, Cyp3a41) and did not modify male predominant genes (Cyp7b1, Cyp4a12, Slp). Female predominance of Cis mRNA, an inhibitor of episodic GH signaling pathway, was unaltered. At birth, Cyp7b1 promoter exhibited a higher methylation status in female livers, while the Hnf6 promoter was equally methylated in both sexes; no differences in gene expression were detected at this age. In adulthood, consistent with sex specific predominance, lower methylation status was determined for the Cyp7b1 promoter in males, and for the Hnf6 promoter in females, and this last difference was prevented by neonatal androgenization. Therefore, early steroid treatment or eventually endocrine disruptor exposure may alter methylation status and sexual dimorphic expression of liver genes, and consequently modify liver physiology in females.
Assuntos
Androgênios/farmacologia , Hormônio do Crescimento/genética , Fator 6 Nuclear de Hepatócito/genética , Fígado/efeitos dos fármacos , Esteroide Hidroxilases/genética , Testosterona/farmacologia , Animais , Animais Recém-Nascidos , Família 7 do Citocromo P450 , Metilação de DNA/efeitos dos fármacos , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Hormônio do Crescimento/metabolismo , Fator 6 Nuclear de Hepatócito/metabolismo , Fígado/crescimento & desenvolvimento , Fígado/metabolismo , Masculino , Camundongos , Regiões Promotoras Genéticas/efeitos dos fármacos , Caracteres Sexuais , Transdução de Sinais , Esteroide Hidroxilases/metabolismo , Virilismo/genéticaRESUMO
A novel mutation in CYP24A1 provides insight into idiopathic infantile hypercalcemia. In this report of 3 brothers, in twins supplemented with vitamin D (1900 IU/d), only the twin homozygous for CYP24A1 exhibited idiopathic infantile hypercalcemia. A subsequently affected younger brother given vitamin D 400 IU/d was not hypercalcemic.
Assuntos
Suplementos Nutricionais , Hipercalcemia/genética , Doenças do Recém-Nascido/genética , Erros Inatos do Metabolismo/genética , Mutação , Esteroide Hidroxilases/genética , Vitamina D/uso terapêutico , Alelos , Doenças em Gêmeos , Éxons , Feminino , Homozigoto , Humanos , Lactente , Masculino , Hormônio Paratireóideo/metabolismo , Linhagem , Reação em Cadeia da Polimerase/métodos , Análise de Sequência de DNA , Irmãos , Vitamina D3 24-HidroxilaseRESUMO
In order to better understand vitamin D3 in cattle metabolism, we quantified 1alpha-HYD and 24-HYD gene expression. In the kidneys of 35 male Nellore cattle, these were divided into a control group and two treatment groups (2 x 10(6) international units of vitamin D3 administered for 2 or 8 consecutive days pre-slaughter). Vitamin D3 supplementation resulted in a significant increase in 1alpha-HYD gene expression; however, significantly increased 24-HYD gene expression was only detected in cattle that had 8 days of supplementation. The finding of upregulation of 24-HYD due to vitamin D supplementation is in line with the expected rise in 24,25-di-hydroxy-vitamin D3 synthesis observed when plasma vitamin D3 concentrations are high, stimulating excretion by the organism. On the other hand, upregulation of 1alpha-HYD was unexpected, since vitamin D3 supplementation has been reported to impact these two genes in opposite directions. We conclude that vitamin D3 metabolism in these animals is more complex than previously reported.
Assuntos
25-Hidroxivitamina D3 1-alfa-Hidroxilase/biossíntese , Bovinos/metabolismo , Colecalciferol/farmacologia , Rim/metabolismo , Esteroide Hidroxilases/biossíntese , 25-Hidroxivitamina D3 1-alfa-Hidroxilase/genética , Animais , Cálcio/sangue , Suplementos Nutricionais , Exposição Ambiental , Expressão Gênica , Gliceraldeído 3-Fosfato Desidrogenase (NADP+)/biossíntese , Masculino , Carne , Fator 1 de Elongação de Peptídeos/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Proteínas Ribossômicas/biossíntese , Esteroide Hidroxilases/genética , Luz Solar , Vitamina D3 24-HidroxilaseRESUMO
The hypothalamic-growth hormone (GH)-liver axis represents a new concept in endocrine regulation of drug toxicity. Preponderant sex differences are found in liver gene expression, mostly dependent on the sexually dimorphic pattern of GH secretion which is set during the neonatal period by gonadal steroids. We tested if GH-dependent sexually dimorphic liver enzymes and proteins was perturbed by neonatal Bisphenol A (BPA) treatment in female rats. Female rats were sc injected with BPA (50 or 500 µg/50 µl) or castor oil vehicle from postnatal day 1 to 10. At five months serum prolactin, pituitary GH, and serum and liver insulin growth factor-I (IGF-I) were measured by RIA. Major urinary proteins (MUPs) were determined by electrophoresis. Liver Cyp2c11, Cyp2c12, Adh1, Hnf6, and Prlr mRNA levels were determined by real time PCR. Pituitary GH content and liver IGF-I concentration were increased by neonatal BPA treatment, indicating partial masculinization of the GH axis in treated females. GH-dependent female predominant liver enzyme genes (Cyp2c12 and Adh1) and a transcription factor (Hnf6) were downregulated or defeminized, while there were no changes in a male predominant gene (Cyp2c11) or protein (MUP). Our findings indicate that perinatal exposure to BPA may compromise the sexually dimorphic capacity of the liver to metabolize drugs and steroids.
Assuntos
Disruptores Endócrinos/toxicidade , Estrogênios não Esteroides/toxicidade , Hormônio do Crescimento/metabolismo , Fígado/efeitos dos fármacos , Fenóis/toxicidade , Hipófise/efeitos dos fármacos , Fatores Etários , Envelhecimento/genética , Envelhecimento/metabolismo , Álcool Desidrogenase/genética , Animais , Animais Recém-Nascidos , Hidrocarboneto de Aril Hidroxilases/genética , Compostos Benzidrílicos , Família 2 do Citocromo P450 , Esquema de Medicação , Eletroforese em Gel de Poliacrilamida , Disruptores Endócrinos/administração & dosagem , Estrogênios não Esteroides/administração & dosagem , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Fator 6 Nuclear de Hepatócito/genética , Injeções Subcutâneas , Fator de Crescimento Insulin-Like I/metabolismo , Fígado/metabolismo , Masculino , Fenóis/administração & dosagem , Hipófise/metabolismo , Prolactina/sangue , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Receptores da Prolactina/genética , Caracteres Sexuais , Fatores Sexuais , Esteroide 16-alfa-Hidroxilase/genética , Esteroide Hidroxilases/genéticaRESUMO
Primary hepatocytes are widely used in investigating drug metabolism and its toxicological effects. N-Nitrosodiethylamine (NDEA)-induced genotoxicity and cytotoxicity was used in primary cultures of female rat hepatocytes in the presence of phenobarbital (PB). PB pre-treatment (1mM) increased the number of necrotic (2-fold) and apoptotic cells (4-fold) after NDEA treatment (0.21-105 µg/mL). The mitotic indices and the number of micronucleated cells decreased, thus suggesting cytotoxicity. An increased number of chromosomal aberrations were observed after pre-treatment with PB. NDEA-treatment (0.21-21 µg/mL) induced expression of the CYP2B1 and CYP2B2 mRNA and PB treatment alone induced ~6-fold and ~2-fold increases of CYP2B1 and CYP2B2 mRNA, respectively. NDEA treatment following PB exposure increased CYP2B1 mRNA expression under all tested concentrations and also increased CYP2B2 expression at 21 and 105 µg/mL. Our data suggest that the alteration of CYP2B1/2 expression by PB increased the cytotoxicity and genotoxicity of NDEA leading to the final genotoxic metabolite.
Assuntos
Carcinógenos/toxicidade , Sistema Enzimático do Citocromo P-450/biossíntese , Dietilnitrosamina/toxicidade , Hepatócitos/efeitos dos fármacos , Mutagênicos/toxicidade , Fenobarbital/toxicidade , Regulação para Cima/efeitos dos fármacos , Animais , Hidrocarboneto de Aril Hidroxilases/biossíntese , Hidrocarboneto de Aril Hidroxilases/genética , Morte Celular/efeitos dos fármacos , Células Cultivadas , Aberrações Cromossômicas/efeitos dos fármacos , Cocarcinogênese , Citocromo P-450 CYP2B1/biossíntese , Citocromo P-450 CYP2B1/genética , Sistema Enzimático do Citocromo P-450/genética , Indução Enzimática/efeitos dos fármacos , Feminino , Hepatócitos/patologia , Micronúcleos com Defeito Cromossômico/efeitos dos fármacos , Índice Mitótico , RNA Mensageiro/metabolismo , Ratos , Esteroide Hidroxilases/biossíntese , Esteroide Hidroxilases/genéticaRESUMO
Calcitriol, the hormonal form of vitamin D(3), exerts immunomodulatory effects through the vitamin D(3) receptor (VDR) and increases prolactin (PRL) expression in the pituitary and decidua. Nevertheless, the effects of calcitriol upon lymphocyte PRL have not been evaluated. Therefore, we investigated calcitriol effects upon PRL in resting and phytohemagglutinin-activated human peripheral blood mononuclear cells (PBMNC) and Jurkat T lymphoma cells. Immunoblots showed constitutive expression of the 50-kDa VDR species in activated PBMNC and Jurkat cells, while a 75-kDa species was recognized in both resting and activated-PBMNC. Only in resting PBMNC calcitriol significantly stimulated PRL expression in a dose-dependent manner. The positive control CYP24A1, a highly VDR-responsive gene, was stimulated by calcitriol, effect that was stronger in resting than in activated-PBMNC (P<0.05), and without effect in Jurkat cells. Calcitriol upregulation of PRL and CYP24A1 was significantly inhibited by the VDR antagonist TEI-9647. EMSA showed that resting PBMNC contain a protein that binds to DR3-type VDRE. Cell activation reduced basal CYP24A1 while induced CYP27B1, VDR and pregnane X receptor (PXR) expression. In summary, calcitriol stimulated PRL and CYP24A1 gene expression in quiescent lymphocytes through a VDR-mediated mechanism. Our results suggest that the 75-kDa VDR species could be participating in calcitriol-mediated effects, and that activation induces factors such as PXR that restrain VDR transcriptional processes. This study supports the presence of a functional VDR in quiescent lymphocytes, providing evidence to reevaluate the VDR paradigm that assumes that lymphocytes respond to calcitriol only after activation. Altogether, our results offer new insights into the mechanisms whereby PRL is regulated in immune cells.
Assuntos
Calcitriol/fisiologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/fisiologia , Prolactina/metabolismo , Adulto , Animais , Calcitriol/farmacologia , Expressão Gênica/efeitos dos fármacos , Humanos , Leucócitos Mononucleares/citologia , Masculino , Receptor de Pregnano X , Prolactina/genética , Receptores de Calcitriol/metabolismo , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Esteroide Hidroxilases/genética , Esteroide Hidroxilases/metabolismo , Vitamina D3 24-Hidroxilase , Vitaminas/farmacologiaRESUMO
We tested the hypothesis that polymorphisms in cytochrome P450c17alpha (CYP17), aromatase (CYP19), 17beta-hydroxysteroid dehydrogenase type I (17beta-HSD1) and sex hormone-binding globulin (SHBG) genes may modify the association between isoflavone intake and breast cancer risk. We conducted hospital-based, case-control studies in Nagano, Japan and Sao Paulo, Brazil. A total of 846 pairs (388 Japanese, 79 Japanese Brazilians, and 379 non-Japanese Brazilians) completed validated food frequency questionnaires. Four single nucleotide polymorphisms (SNPs) in CYP17 (rs743572), CYP19 (rs10046), 17beta-HSD1 (rs605059), and SHBG (rs6259) genes were genotyped. We found no association between the 4 SNPs and breast cancer risk. In combination analyses of isoflavone intake and SNPs, an inverse association between intake and risk was limited to women with at least one A allele of the rs605059 polymorphism for all 3 populations, albeit without statistical significance. For the rs6259 polymorphism, the inverse association was limited to postmenopausal Japanese with the GG genotype (odds ratio [OR] for highest vs. lowest tertile = 0.50, 95% confidence interval [CI] = 0.29-0.87; P for trend < 0.01), and to non-Japanese Brazilians with at least one A allele (OR for consumers vs. nonconsumer = 0.21, 95% CI = 0.06-0.77). We found no remarkable difference for the rs743572 and rs10046 polymorphisms. Our findings suggest that polymorphisms in the 17beta-HSD1 and SHBG genes may modify the association between isoflavone intake and breast cancer risk.