RESUMO
INTRODUCTION: Despite thromboprophylaxis, women with antiphospholipid syndrome (APS) face high-risk pregnancies due to proinflammatory and prothrombotic states. This highlights the need for new monitoring and prognostic tools. Recent insights into the pathophysiological role of neutrophil activation and extracellular trap (NET) formation in this syndrome led to the exploration of plasma cell-free DNA (cfDNA), a derivative of NETosis, as a promising biomarker. MATERIALS AND METHODS: cfDNA was isolated and quantified from plasma samples of healthy pregnant women (control group, HC) and women with APS (APS group). We assessed the physiological variability of cfDNA across the three trimesters in HC. Levels of cfDNA were compared between APS and HC by gestational trimester. ROC curve analysis was performed to evaluate the efficacy of cfDNA levels for classifying APS patients. Furthermore, cfDNA levels in pregnant women with APS with obstetric complications were compared to those from uncomplicated pregnancies. RESULTS: Among HC, cfDNA significantly increased in the third trimester compared to the first and second. Elevated cfDNA levels in APS compared to HC were observed in the first and second trimesters. First-trimester cfDNA levels demonstrated the highest classification ability to discriminate between APS and HC patients (AUC: 0.906). Among APS, those with complicated pregnancies (fetal growth restriction, preeclampsia, placenta accreta) exhibited significantly elevated cfDNA levels in the second trimester. CONCLUSIONS: Elevated levels of cfDNA in pregnant women with APS, particularly among those with obstetric complications, supports further investigation into the potential of cfDNA as a valuable tool in the obstetric management of women with APS.
Assuntos
Síndrome Antifosfolipídica , Ácidos Nucleicos Livres , Gravidez de Alto Risco , Humanos , Feminino , Gravidez , Síndrome Antifosfolipídica/sangue , Síndrome Antifosfolipídica/complicações , Ácidos Nucleicos Livres/sangue , Adulto , Gravidez de Alto Risco/sangue , Biomarcadores/sangue , Complicações na Gravidez/sangueRESUMO
The use of plasma cell-free DNA (cfDNA) as a useful biomarker in obstetric clinical practice has been delayed due to the lack of reliable quantification protocols. We developed a protocol to quantify plasma cfDNA using an internal standard strategy to overcome difficulties posed by low levels and high fragmentation of cfDNA. cfDNA was isolated from plasma samples of non-pregnant (NP, n = 26) and pregnant (P, n = 26) women using a commercial kit and several elution volumes were evaluated. qPCR parameters were optimized for cfDNA quantification, and several quantities of a recombinant standard were evaluated as internal standard. Absolute quantification was performed using a standard curve and the quality of the complete method was evaluated. cfDNA was eluted in a 50-µl volume, actin-ß (ACTB) was selected as the target gene, and qPCR parameters were optimized. The ACTB standard was constructed and 1000 copies were selected as internal standard. The standard curve showed R2 = 0.993 and E = 109.7%, and the linear dynamic range was defined between 102 and 106 ACTB copies/tube. Repeatability and reproducibility in terms of CV were 19% and up to 49.5% for ACTB copies per milliliter of plasma, respectively. The range of cfDNA levels was 428-18,851 copies/mL in NP women and 4031-2,019,363 copies/mL in P women, showing significant differences between the groups. We recommend the application of internal standard strategy for a reliable plasma cfDNA quantification. This methodology holds great potential for a future application in the obstetric field.
Assuntos
Ácidos Nucleicos Livres , Gestantes , Humanos , Feminino , Gravidez , Reprodutibilidade dos Testes , Ácidos Nucleicos Livres/genética , BiomarcadoresRESUMO
Environmental exposure to agrochemicals during early stages of development can induce subtle alterations that could permanently affect normal physiology. Previously, we reported that in ovo exposure to atrazine (ATZ) disrupts testicular histoarchitecture in postnatal caimans (Caiman latirostris). To assess whether such alterations are the result of disruption of gonadal developmental programming, this study aimed to evaluate the expression of histofunctional biomarkers (VASA, ER, PR, PCNA, and aromatase) and genes involved in gonadal development and differentiation (amh, sox-9, sf-1 and cyp19-a1) in the gonads of male and female caiman embryos and to assess the effect of ATZ exposure on these biomarkers and genes in the gonads of male embryos. Our results suggest that amh, aromatase and sox-9 play a role in sex determination and gonadal differentiation. In male caiman embryos, ATZ exposure increased aromatase expression and altered the temporal expression pattern of amh and sox-9 evidencing an ATZ-induced disruption of gonadal developmental programming. Since the effects of ATZ are consistent across all vertebrate classes, the ATZ-mediated disruptive effects here observed could be present in other vertebrate species.
Assuntos
Jacarés e Crocodilos , Atrazina , Animais , Feminino , Masculino , Atrazina/metabolismo , Aromatase/metabolismo , Gônadas , TestículoRESUMO
Agrochemicals or pesticides are compounds widely used to prevent, destroy or mitigate pests such as insects, rodents, herbs and weeds. However, most of them also act as environmental estrogens, anti-estrogens and/or antiandrogenic chemicals. In addition, both herbicides (such as glyphosate and paraquat) and insecticides (such as pyrethroids, organophosphates, neonicotinoids and rotenone) have been shown to exert significant adverse effects on hippocampal neurogenesis. These effects are particularly important because neurogenesis dysregulation could be associated with cognitive decline and neuropathologies such as Alzheimer's disease. This review focuses on the most commonly used agrochemicals in Argentina and their effects on the hippocampal neurogenesis of mammals. It also discusses the disruption of hormone synthesis and action as a possible mechanism through which these chemical compounds could alter the brain functions. Finally, we propose some lines of research to study the potential endocrine mechanisms involved in the effects of agrochemicals on human health and biodiversity.
Assuntos
Agroquímicos/toxicidade , Neurogênese/efeitos dos fármacos , Animais , Sistema Endócrino/efeitos dos fármacos , Herbicidas/toxicidade , Humanos , Inseticidas/toxicidade , Praguicidas/toxicidadeRESUMO
Ghrelin is a potent orexigenic peptide hormone that acts through the growth hormone secretagogue receptor (GHSR), a G protein-coupled receptor highly expressed in the hypothalamus. In vitro studies have shown that GHSR displays a high constitutive activity, whose physiological relevance is uncertain. As GHSR gene expression in the hypothalamus is known to increase in fasting conditions, we tested the hypothesis that constitutive GHSR activity at the hypothalamic level drives the fasting-induced hyperphagia. We found that refed wild-type (WT) mice displayed a robust hyperphagia that continued for 5 days after refeeding and changed their food intake daily pattern. Fasted WT mice showed an increase in plasma ghrelin levels, as well as in GHSR expression levels and ghrelin binding sites in the hypothalamic arcuate nucleus. When fasting-refeeding responses were evaluated in ghrelin- or GHSR-deficient mice, only the latter displayed an â¼15% smaller hyperphagia, compared with WT mice. Finally, fasting-induced hyperphagia of WT mice was significantly smaller in mice centrally treated with the GHSR inverse agonist K-(D-1-Nal)-FwLL-NH2, compared with mice treated with vehicle, whereas it was unaffected in mice centrally treated with the GHSR antagonists D-Lys3-growth hormone-releasing peptide 6 or JMV2959. Taken together, genetic models and pharmacological results support the notion that constitutive GHSR activity modulates the magnitude of the compensatory hyperphagia triggered by fasting. Thus, the hypothalamic GHSR signaling system could affect the set point of daily food intake, independently of plasma ghrelin levels, in situations of negative energy balance.
Assuntos
Jejum/fisiologia , Grelina/fisiologia , Hiperfagia/etiologia , Receptores de Grelina/fisiologia , Animais , Ingestão de Alimentos/fisiologia , Grelina/metabolismo , Hiperfagia/genética , Hiperfagia/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Grelina/genética , Receptores de Grelina/metabolismo , Transdução de Sinais/genéticaRESUMO
The development of the mammary gland is a hormone-regulated event. Several factors can dysregulate its growth and make the gland more susceptible to cellular transformation. Among these factors, perinatal exposure to xenoestrogens and hormone replacement therapy has been associated with increased risk of developing breast cancer. Here, we assessed the effects induced by estrogen replacement therapy (ERT) in ovariectomized (OVX) middle-aged rats and whether perinatal exposure to diethylstilbestrol (DES) or bisphenol A (BPA) modified these effects in the mammary gland. Pregnant rats were orally exposed to vehicle, 5 µg DES/kg/day, or 0.5 or 50 µg BPA/kg/day from gestational day 9 until weaning. Then, 12-month-old offspring were OVX and treated with 17ß-estradiol for 3 months. Morphological changes and the percentage of epithelial cells that proliferated or expressed estrogen receptor alpha (ESR1) and progesterone receptor (PR) were analyzed in mammary gland samples of 15-month-old animals. ERT induced lobuloalveolar hyperplasia and ductal cysts in the mammary gland of middle-aged rats, associated with a higher proliferation index of epithelial cells. Perinatal exposure to DES followed by ERT increased the number of cysts and induced the formation of fibroadenoma and ductal carcinoma in situ, without modifying the expression of ESR1 or PR. Also, after 3 months of ERT, BPA-exposed rats had a higher incidence of ductal hyperplasia and atypical lobular hyperplasia than animals under ERT alone. In conclusion, perinatal exposure to xenoestrogens increases the susceptibility of the mammary gland to develop cysts and hyperplastic lesions when confronted with ERT later in life.
Assuntos
Compostos Benzidrílicos/efeitos adversos , Cisto Mamário/induzido quimicamente , Carcinoma Intraductal não Infiltrante/induzido quimicamente , Dietilestilbestrol/efeitos adversos , Estradiol/efeitos adversos , Glândulas Mamárias Animais/efeitos dos fármacos , Fenóis/efeitos adversos , Administração Oral , Animais , Compostos Benzidrílicos/administração & dosagem , Cisto Mamário/veterinária , Carcinoma Intraductal não Infiltrante/veterinária , Proliferação de Células/efeitos dos fármacos , Dietilestilbestrol/administração & dosagem , Estradiol/administração & dosagem , Terapia de Reposição de Estrogênios/efeitos adversos , Terapia de Reposição de Estrogênios/métodos , Feminino , Ovariectomia , Fenóis/administração & dosagem , Gravidez , Efeitos Tardios da Exposição Pré-Natal , Ratos , Fatores de RiscoRESUMO
Glyphosate is the active ingredient of several herbicide formulations. Different reports suggest that glyphosate-based herbicides (GBHs) may act as endocrine disruptors. We evaluated the potential estrogenic effects of a GBH formulation using the uterotrophic assay. Adult ovariectomized rats were sc injected for 3 consecutive days with: saline solution (vehicle control), 2.10-5 g E2 /kg/day (uterotrophic dose; UE2 ), 2.10-7 g E2 /kg/day (nonuterotrophic dose; NUE2 ), or 0.5, 5, or 50 mg GBH/kg/day of the. Twenty-four hours after the last injection, the uterus was removed and weighed and processed for histopathology and mRNA extraction. Epithelial cell proliferation and height and expression of estrogen-responsive genes were evaluated (estrogen receptors, ERα and ERß; progesterone receptor, PR; complement 3, C3). Uterine weight and epithelial proliferation were not affected by GBH. However, the luminal epithelial cell height increased at GBH0.5. ERα mRNA was downregulated by all GBH doses and E2 groups, whereas PR and C3 mRNA were diminished by GBH0.5. GBH5-, GBH50-, and UE2 -treated rats showed downregulated ERα protein expression in luminal epithelial cells, while the receptor was upregulated in the stroma. GBH upregulated ERß (GBH0.5-50) and PR (GBH5) expressions in glandular epithelial cells, similar effect to that of NUE2 group. These results indicate that, although the uterine weight was not affected, GBH modulates the expression of estrogen-sensitive genes. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1191-1201, 2017.
Assuntos
Glicina/análogos & derivados , Herbicidas/toxicidade , Útero/efeitos dos fármacos , Animais , Animais Endogâmicos , Estradiol/fisiologia , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/genética , Receptor beta de Estrogênio/metabolismo , Feminino , Expressão Gênica/efeitos dos fármacos , Glicina/toxicidade , Tamanho do Órgão/efeitos dos fármacos , Ovariectomia , Ratos , Ratos Wistar , Receptores de Progesterona/genética , Receptores de Progesterona/metabolismo , Útero/metabolismo , Útero/patologia , GlifosatoRESUMO
With the aim to analyze whether bisphenol A (BPA) modifies ß-Casein (ß-Cas) synthesis and transcriptional regulation in perinatally exposed animals, here, pregnant F0 rats were orally exposed to 0, 0.6 or 52 µg BPA/kg/day from gestation day 9 until weaning. Then, F1 females were bred and mammary glands were obtained on lactation day 2. Perinatal BPA exposure decreased ß-Cas expression without modifying the activation of prolactin receptor. It also decreased the expression of glucocorticoid receptor in BPA52-exposed dams and ß1 and α6 integrins as well as dystroglycan in both BPA groups. In addition, BPA exposure altered the expression of histone-modifying enzymes and induced histone modifications and DNA methylation in the promoter, enhancer and exon VII of the ß-Cas gene. An impaired crosstalk between the extracellular matrix and lactogenic hormone signaling pathways and epigenetic modifications of the ß-Cas gene could be the molecular mechanisms by which BPA decreased ß-Cas expression.
Assuntos
Compostos Benzidrílicos/toxicidade , Caseínas/genética , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Glândulas Mamárias Animais/metabolismo , Fenóis/toxicidade , Efeitos Tardios da Exposição Pré-Natal/genética , Transcrição Gênica/efeitos dos fármacos , Animais , Caseínas/metabolismo , Comunicação Celular/efeitos dos fármacos , Metilação de DNA/genética , Elementos Facilitadores Genéticos/genética , Éxons/genética , Feminino , Histonas/metabolismo , Lactação/genética , Laminina/metabolismo , Gravidez , Efeitos Tardios da Exposição Pré-Natal/patologia , Regiões Promotoras Genéticas , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Ratos Wistar , Receptores de Glucocorticoides/metabolismo , Receptores de Laminina/metabolismo , Receptores da Prolactina/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genéticaRESUMO
Removing dietary phyto-oestrogens in adult male rats causes obesity and diabetes. As whey proteins have been reported to reduce food intake and improve glucose homoeostasis, we investigated whether they could attenuate susceptibility to obesity and diabetes due to phyto-oestrogen deprivation. To this end, thirty male Wistar rats were fed a high-phyto-oestrogen (HP) or a phyto-oestrogen-free (PF) diet for 10 weeks; six rats from each group were killed. The remaining HP animals (six animals) continued receiving the HP diet for 6 weeks. The remaining PF rats (twelve rats) were divided in two groups: one was given the PF diet and the other a variation of the PF diet plus whey protein (PF-W). Body weight, food intake and adipose tissue weights were recorded. Hypothalamic mRNA expressions of orexigenic (neuropeptide Y, agouti-related protein (AgRP)) and anorexigenic (pro-opiomelanocortin (POMC), cocaine-amphetamine-related transcript (CART)) neuropeptides were quantified by real-time PCR. Serum glucose, insulin and total thyroxine (T4), thyroid-stimulating hormone, testosterone and oestradiol were assessed. After 10 weeks of PF diet, increased body weight, adiposity and energy intake, with up-regulation of AgRP and down-regulation of POMC', were observed. Longer treatment exacerbated these results, increased total T4 levels, reduced oestradiol levels and impaired glucose homoeostasis. PF-W reduced energy intake and increased POMC expression; however, body weight and adiposity remained unchanged. PF-W could not prevent the hormonal changes or the high circulating glucose levels induced by phyto-oestrogen deprivation, but reduced fasting insulin. These data demonstrate that, although 6 weeks of whey administration could not prevent obesity in phyto-oestrogen-deprived rats, the reduction in energy intake and circulating insulin could be beneficial with longer treatments.
Assuntos
Ingestão de Energia/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Hipotálamo/metabolismo , Obesidade , Proteínas do Soro do Leite/farmacologia , Ração Animal/análise , Animais , Glicemia , Dieta , Suplementos Nutricionais , Masculino , Fitoestrógenos/administração & dosagem , Fitoestrógenos/farmacologia , RNA/genética , RNA/metabolismo , Ratos , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Neonatal exposure to a low dose of endosulfan may disrupt the expression of Wnt7a and ß-catenin during uterine development leading to the failure of uterine functional differentiation during implantation. New-born female Wistar rats were treated with vehicle, endosulfan (600 µg/kg/d, E600) or diethylstilbestrol (0.2 µg/kg/d, DES) on postnatal days (PNDs) 1, 3, 5 and 7. Subsequently, uterine histomorphology and the protein expression of Wnt7a and ß-catenin were evaluated on PND8, PND21 and gestational day (GD) 5 (pre-implantation period). In the E600 rats, Wnt7a and ß-catenin protein expression was increased in the epithelium on PND8, and Wnt7a expression was decreased in the endometrial glands on PND21. On GD5, the number of uterine glands was decreased in the E600-and DES-treated rats. In addition, Wnt7a expression was decreased in all uterine compartments, and ß-catenin expression was increased in the luminal and glandular epithelia of the E600-and DES-treated rats. Disruption of Wnt7a and ß-catenin uterine expression in the prepubertal and adult females altered the uterine preparation for embryo implantation, which could be associated with the subfertility triggered by endosulfan.
Assuntos
Endossulfano/efeitos adversos , Proteínas Proto-Oncogênicas/metabolismo , Útero/efeitos dos fármacos , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Animais , Animais Recém-Nascidos , Diferenciação Celular/efeitos dos fármacos , Dietilestilbestrol/toxicidade , Endossulfano/administração & dosagem , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Ratos , Ratos Wistar , Útero/crescimento & desenvolvimento , Útero/metabolismoRESUMO
Here we assessed the effects of perinatal exposure to bisphenol A (BPA) on the uterine response to 17ß-estradiol (E2) in aged rats. Pregnant rats were orally exposed to 0.5 or 50 µg BPA/kg/day from gestational day 9 until weaning. On postnatal day (PND) 360, the rats were ovariectomized and treated with E2 for three months. The uterine tissue of BPA50 and BPA0.5 rats showed increased density of glands with squamous metaplasia (GSM) and glands with daughter glands respectively. Wnt7a expression was lower in GSM of BPA50 rats than in controls. The expression of estrogen receptor 1 (ESR1) and its 5'- untranslated exons ESR1-O and ESR1-OT was lower in BPA50 rats. Both doses of BPA modified the expression of coactivator proteins and epigenetic regulatory enzymes. Thus, perinatal BPA-exposed rats showed different glandular abnormalities associated with deregulated expression of E2-target genes. Different mechanisms would be involved depending on the BPA dose administered.
Assuntos
Compostos Benzidrílicos/toxicidade , Disruptores Endócrinos/toxicidade , Estradiol/farmacologia , Fenóis/toxicidade , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Animais , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Masculino , Coativadores de Receptor Nuclear/genética , Coativadores de Receptor Nuclear/metabolismo , Especificidade de Órgãos , Gravidez , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Ratos Wistar , Testículo/metabolismo , Útero/metabolismo , Proteínas Wnt/genética , Proteínas Wnt/metabolismoRESUMO
We assessed the long-term effect of perinatal exposure to bisphenol A (BPA) on the rat uterus and the uterine response to estrogen (E2) replacement therapy. BPA (0.5 or 50µg/kg/day) was administered in the drinking water from gestational day 9 until weaning. We studied the uterus of female offspring on postnatal day (PND) 90 and 360, and the uterine E2 response on PND460 (PND460-E2). On PND90, BPA-exposed rats showed altered glandular proliferation and α-actin expression. On PND360, BPA exposure increased the incidence of abnormalities in the luminal and glandular epithelium. On PND460-E2, the multiplicity of glands with squamous metaplasia increased in BPA50 while the incidence of glands with daughter glands increased in BPA0.5. The expression of steroid receptors, p63 and IGF-I was modified in BPA-exposed rats on PND460-E2. The long-lasting effects of perinatal exposure to BPA included induction of abnormalities in uterine tissue and altered response to E2 replacement therapy.
Assuntos
Compostos Benzidrílicos/toxicidade , Estradiol/farmacologia , Fenóis/toxicidade , Útero/efeitos dos fármacos , Útero/fisiologia , Animais , Apoptose , Atrofia , Compostos Benzidrílicos/administração & dosagem , Proliferação de Células/efeitos dos fármacos , Feminino , Idade Gestacional , Lactação , Ovariectomia , Fenóis/administração & dosagem , Gravidez , Ratos , Ratos Wistar , Receptores de Estrogênio/análise , Receptores de Progesterona/análise , Útero/patologiaRESUMO
We investigated whether neonatal exposure to low doses of endosulfan affects fertility and uterine functional differentiation at pre-implantation in rats. Newborn female rats received the vehicle, 0.2 µg/kg/d of diethylstilbestrol (DES), 6 µg/kg/d of endosulfan (Endo6) or 600 µg/kg/d of endosulfan (Endo600) on postnatal days (PND) 1, 3, 5, and 7. On PND90, the rats were mated to evaluate their reproductive performance on gestational day (GD) 19 and their ovarian steroid serum levels, endometrial proliferation and implantation-associated proteins on GD5. DES and endosulfan decreased the pregnancy rate and the number of implantation sites. On GD5, DES and endosulfan did not change the serum levels of 17ß-estradiol (E2) and progesterone (P); the endometrial proliferation decreased, which was associated with silencing of Hoxa10 in the Endo600-treated rats. Both doses of endosulfan increased the progesterone receptor (PR) expression, whereas the higher dose led additionally to an increase in estrogen receptor alpha (ERα). In the Endo600-treated rats, the down-regulation of Hoxa10 was associated with a deregulation of the steroid receptor coregulators. Alterations in endometrial proliferation and the endocrine pathway of Hoxa10/steroid receptors/coregulators might be the mechanism of endosulfan-induced implantation failure.
Assuntos
Implantação do Embrião/efeitos dos fármacos , Endossulfano/administração & dosagem , Fertilidade/efeitos dos fármacos , Inseticidas/administração & dosagem , Útero/efeitos dos fármacos , Animais , Animais Recém-Nascidos , Carcinógenos/administração & dosagem , Carcinógenos/toxicidade , Proliferação de Células/efeitos dos fármacos , Dietilestilbestrol/administração & dosagem , Dietilestilbestrol/toxicidade , Relação Dose-Resposta a Droga , Endossulfano/toxicidade , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Inseticidas/toxicidade , Gravidez , Taxa de Gravidez , Ratos , Ratos Wistar , Esteroides/sangue , Útero/crescimento & desenvolvimentoRESUMO
Successful implantation is the result of complex molecular interactions between the hormonally primed uterus and a mature blastocyst. This very carefully synchronized interplay of hormonal signals and feedback loops is potentially vulnerable to chemicals such as endocrine disruptors that may disrupt endocrine signaling. Bisphenol A (BPA) is one of the highest-volume chemicals produced worldwide. This chapter describes the effects of brief postnatal exposure to BPA on female reproductive performance and specifically on the uterine adaptations during the preimplantation period. We propose that an early alteration in Hoxa10 gene expression affects the functional differentiation of the preimplantation uterus as part of an altered endocrine signal transduction pathway. These molecular alterations could explain, at least in part, the adverse effects of BPA on uterine implantation. Exposure to endocrine disruptors, such as BPA, could contribute to the impaired female fertility noted over the past decades.
Assuntos
Compostos Benzidrílicos/toxicidade , Implantação do Embrião/efeitos dos fármacos , Perda do Embrião/induzido quimicamente , Disruptores Endócrinos/toxicidade , Exposição Ambiental/efeitos adversos , Poluentes Ambientais/toxicidade , Fenóis/toxicidade , Útero/efeitos dos fármacos , Animais , Animais Recém-Nascidos , Desenvolvimento Infantil/efeitos dos fármacos , Perda do Embrião/patologia , Perda do Embrião/fisiopatologia , Estrogênios não Esteroides/toxicidade , Feminino , Humanos , Recém-Nascido , Infertilidade Feminina/etiologia , Gravidez , Útero/patologiaRESUMO
Caiman latirostris is a reptilian species that exhibits temperature-dependent sex determination (TSD). Male-to-female sex reversal can be achieved after in ovo estrogen/xenoestrogen exposure. This is known as hormone-dependent sex determination (HSD). The amh, sox9 and sf-1 genes are involved in sex determination, sex differentiation, and steroidogenesis. The aims of this study were: (a) to establish the expression patterns of amh, sox9 and sf-1 mRNA in the gonad-adrenal-mesonephros (GAM) complexes of neonatal TSD-male and TSD-female caimans, (b) to compare the expression of these genes between TSD-females and HSD-females (born from E2-exposed eggs incubated at the male-producing temperature) and (c) to evaluate whether in ovo exposure to a low dose of E2 or bisphenol A (BPA) or to a high dose of endosulfan (END) modifies amh, sox9 or sf-1 mRNA expressions in neonatal males. The mRNA expressions of amh, sox9 and sf-1 in GAM complexes from TSD-males and TSD-females and from HSD-females were quantitatively compared by RT-PCR. A sexually dimorphic pattern of amh and sox9 mRNA expression was found, with a higher expression in TSD-males than in TSD-females. sf-1 mRNA did not differ between TSD-males and TSD-females. HSD-females exhibited a higher expression of sox9 than TSD-females. In males, increased mRNA expression of sex-determining genes was observed after in ovo exposure to END. E2 decreased sox9 but increased sf-1 mRNA expression. Changes induced by BPA were evident although not significant. These results provide new insights into the potential mechanisms that lead to the gonadal histo-functional alterations observed in caimans exposed to contaminated environments.
Assuntos
Jacarés e Crocodilos/metabolismo , Disruptores Endócrinos/toxicidade , RNA Mensageiro/genética , Fatores de Transcrição SOX9/genética , Fator Esteroidogênico 1/genética , Animais , Compostos Benzidrílicos/toxicidade , Endossulfano/toxicidade , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/genética , Masculino , Fenóis/toxicidade , Testículo/efeitos dos fármacos , Testículo/metabolismoRESUMO
Endocrine disrupters have been associated with reproductive pathologies such as infertility and gynecological tumors. Using a rat model of early postnatal exposure to bisphenol A (BPA), we evaluated the long-term effects on 1) female reproductive performance, 2) uterine homeobox A10 (Hoxa10) and Hoxa10-target gene expression, and 3) ovarian steroid levels and uterine estrogen receptor α and progesterone (P) receptor expression. Newborn female rats received vehicle, BPA.05 (0.05 mg/kg · d), BPA20 (20 mg/kg · d), diethylstilbestrol.2 (0.2 µg/kg · d), or diethylstilbestrol 20 (20 µg/kg · d) on postnatal d 1, 3, 5, and 7. A significant decrease in the number of implantation sites was assessed in the xenoestrogen-exposed females. To address the molecular effects of postnatal xenoestrogen exposure on the pregnant uterus, we evaluated the expression of implantation-associated genes on d 5 of pregnancy (preimplantation uterus). All xenoestrogen-treated rats showed a lower expression of Hoxa10. In the same animals, two Hoxa10-downstream genes were misregulated in the uterus. ß(3) Integrin, which is up-regulated by Hoxa10 in controls, was decreased, whereas empty spiracles homolog 2, which is down-regulated by Hoxa10, was increased. Furthermore a clear down-regulation of estrogen receptor α and P receptor expression was detected without changes in estradiol and P serum levels. The early exposure to BPA produced a lower number of implantation sites in association with a defective uterine environment during the preimplantation period. Alterations in the endocrine-regulated Hoxa10 gene pathways (steroid receptors--Hoxa10--ß(3) integrin/empty spiracles homolog 2) could explain, at least in part, the BPA effects on the implantation process.
Assuntos
Implantação do Embrião/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Fenóis/toxicidade , Útero/fisiologia , Animais , Animais Recém-Nascidos , Compostos Benzidrílicos , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Proteínas Homeobox A10 , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Integrina beta3/genética , Integrina beta3/metabolismo , Masculino , Ovário/metabolismo , Gravidez , Ratos , Ratos Endogâmicos , Receptores de Progesterona/genética , Receptores de Progesterona/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Hormonally controlled vascular changes play a key role in endometrial development and in the differentiation process necessary for implantation. Vascular endothelial growth factor (VEGF) has emerged as one of the central regulators of the uterine vasculature. Hormonal perturbations during neonatal development may alter sex steroid-dependent regulation of VEGF and may ultimately affect fertility later in life. The aim of this study was to determine whether neonatal exposure to the environmental estrogenic chemical bisphenol A (BPA) affects the adult rat uterine response to hormonal stimuli. Newborn female rats were given s.c. injections of vehicle, BPA (0.05 mg/kg per day or 20 mg/kg per day) or diethylstilbestrol (0.2 microg/kg per day) on Postnatal Days 1, 3, 5, and 7. To evaluate the long-term effects, rats were ovariectomized at Postnatal Day 80 and submitted to hormonal replacement. Rats neonatally exposed to xenoestrogens showed a decreased induction of uterine endothelial proliferation and a decreased Vegf mRNA expression in response to ovarian steroid treatment. Also, although the estrogen receptor alpha (ESR1) expression was lower in subepithelial cells than in controls, a higher expression of silencing mediator of retinoic acid and thyroid hormone receptor (NCOR1, also known as SMRT) corepressor was evidenced in the same compartment. The results indicate that disturbed Vegf expression in BPA rats could be the result of changes in endocrine pathways, such as an altered induction of ESR1 and/or NCOR1 expression. Because of the importance of VEGF in the implantation process, our data suggest that neonatal BPA exposure might have negative consequences on female fertility.
Assuntos
Disruptores Endócrinos/toxicidade , Endotélio Vascular/efeitos dos fármacos , Estrogênios não Esteroides/toxicidade , Fenóis/toxicidade , Útero/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Animais Recém-Nascidos , Compostos Benzidrílicos , Proliferação de Células/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Estradiol/metabolismo , Receptor alfa de Estrogênio/metabolismo , Feminino , Neovascularização Fisiológica/efeitos dos fármacos , Ovariectomia , Progesterona/metabolismo , Ratos , Ratos Endogâmicos , Receptores de Progesterona/metabolismo , Útero/irrigação sanguínea , Útero/metabolismoRESUMO
The estrogenic activity of environmentally relevant doses of endosulfan was investigated using an animal model. Ovariectomized adult rats were injected once a day for 3 days with sesame oil (control), 0.02mg/kg/day 17beta-estradiol (an uterotrophic dose; UE(2)), 0.0002mg/kg/day 17beta-estradiol (a non-uterotrophic dose; NUE(2)), or 0.006, 0.06, 0.6 or 6mg/kg/day endosulfan. After 24h of treatment, the uteri were weighed (uterotrophic assay) and the luminal epithelial cell height (LECH) and progesterone receptor (PR), and estrogen receptor alpha (ERalpha) protein levels were measured. PR, ERalpha, and complement factor-3 (C3) mRNAs were evaluated using real-time PCR. Uterine weight and LECH were only increased in UE(2)-treated rats. PR, ERalpha and C3 expression levels were modified in most of the endosulfan-treated groups, showing an identical pattern of expression to the NUE(2)-group. Our results show that the pesticide endosulfan mimics non-uterotrophic E(2) actions, strengthening the hypothesis that endosulfan is a widespread xenoestrogen.
Assuntos
Endossulfano/toxicidade , Estradiol/farmacologia , Estrogênios não Esteroides/toxicidade , Expressão Gênica/efeitos dos fármacos , Útero/efeitos dos fármacos , Animais , Tamanho Celular/efeitos dos fármacos , Complemento C3/genética , Relação Dose-Resposta a Droga , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Receptor alfa de Estrogênio/genética , Feminino , Imuno-Histoquímica , Tamanho do Órgão/efeitos dos fármacos , RNA Mensageiro/genética , Ratos , Ratos Wistar , Receptores de Progesterona/genética , Útero/patologiaRESUMO
Morphoregulator genes like members of the Hox gene family regulate uterine development and are associated with endocrine-related processes such as endometrial proliferation and differentiation in the adult uterus. Exposure to neonatal endocrine disruptors could affect signaling events governed by Hox genes, altering the developmental trajectory of the uterus with lasting consequences. We investigated whether neonatal exposure to bisphenol A (BPA) alters Hoxa10 and Hoxa11 mRNA uterine expression shortly after treatment as well as in the adult. Moreover, we studied whether xenoestrogen exposure may affect the adult uterine response to hormonal stimuli. Newborn females received vehicle, 0.05 mg/kg.d BPA, 20 mg/kg*d BPA, or diethylstilbestrol (0.2 microg/kg*d) on postnatal d 1, 3, 5, and 7). At postnatal d 8, real time RT-PCR assays showed a decrease in Hoxa10 and Hoxa11 expression in all xenoestrogen-treated groups. To evaluate the long-term effects, we used adult ovariectomized rats with hormonal replacement. The subepithelial stroma in BPA- and diethylstilbestrol-treated animals showed an impaired proliferative response to steroid treatment associated with a silencing of Hoxa10 but not associated with changes in the methylation pattern of the Hoxa10 promoter. BPA animals showed that the Hoxa10 reduction was accompanied by an increased stromal expression of the silencing mediator for retinoic acid and thyroid hormone receptor. The spatial coexpression of steroid receptors Hoxa10 and silencing mediator for retinoic acid and thyroid hormone receptor was established using immunofluorescence. Our data indicate that postnatal BPA exposure affects the steroid hormone-responsiveness of uterine stroma in adulthood. Whether this impaired hormonal response is associated with effects on uterine receptivity and decidualization is currently under investigation.
Assuntos
Hormônios Esteroides Gonadais/farmacologia , Fenóis/toxicidade , Útero/efeitos dos fármacos , Útero/crescimento & desenvolvimento , Algoritmos , Animais , Animais Recém-Nascidos , Compostos Benzidrílicos , Proliferação de Células/efeitos dos fármacos , Dietilestilbestrol/farmacologia , Dietilestilbestrol/toxicidade , Estrogênios não Esteroides/farmacologia , Estrogênios não Esteroides/toxicidade , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Hormônios Esteroides Gonadais/metabolismo , Proteínas Homeobox A10 , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Modelos Biológicos , Ovário/efeitos dos fármacos , Ovário/crescimento & desenvolvimento , Ovário/metabolismo , Fenóis/farmacologia , Ratos , Ratos Wistar , Células Estromais/efeitos dos fármacos , Células Estromais/fisiologia , Útero/metabolismo , Útero/fisiopatologiaRESUMO
The xenoestrogen bisphenol A (BPA) is commonly ingested by humans. We examined the effects of neonatal exposure to low versus high doses of BPA over the control of estrogen receptor alpha (ERalpha) expression in the preoptic area (POA) of prepubertal female rats. Pups received s.c. injections every 48 h of BPA (high dose, 20 mg/kg and low dose, 0.05 mg/kg) or diethylstilbestrol (DES, 0.02 mg/kg) from postnatal day (PND) 1 to PND7 and were killed at PND8 or PND21. Relative expression of ERalpha transcripts containing alternative 5'-untranslated regions OS, ON, O, OT, and E1 in POA were evaluated by RT-PCR. Methylation status of ERalpha promoters was determined by bisulfited DNA restriction analysis and ERalpha protein by immunohistochemistry. In PND8, the high dose of BPA and DES diminished total ERalpha mRNA levels, mediated by the decreased expression of ERalpha-O and ERalpha-OT variants. In contrast, the low dose of BPA augmented total ERalpha mRNA by increasing the expression of the ERalpha-E1 variant. In PND21, both BPA doses increased total ERalpha mRNA by means of the augmented expression of ERalpha-O and ERalpha-OT variants. In PND21, the methylation status of the ERalpha promoters and the circulating levels of estradiol were similar in all experimental groups. At PND8 and PND21, DES and the high dose of BPA decreased, while the low dose of BPA increased ERalpha protein in the POA. These findings show that neonatal BPA exposure alters the abundance of hypothalamic ERalpha transcript variants and protein in a dose-dependent manner.