RESUMO
The developing uterus is highly sensitive to a brief exposure to different substances, in particular those with endocrine-disrupting activity. Thus, exposure to environmental, nutritional, chemical, and other xenobiotic factors affecting signaling events during critical organizational periods can alter the normal course of uterine development with lasting consequences. In this chapter, we provide an experimental protocol to evaluate the development of the rat uterus as a toxicity biomarker at two different developmental time points: (1) the neonatal period, on postnatal day (PND) 8, and (2) the prepubertal period, on PND21. In this experimental approach, we propose to assess: (1) uterine morphology and cytodifferentiation, (2) uterine cell proliferation, and (3) the expression of proteins involved in uterine organogenetic differentiation. All these morphological and molecular markers are useful tools to determine the consequences of exposure to toxicants with the potential to disrupt the uterine development.
Assuntos
Testes de Toxicidade , Útero/efeitos dos fármacos , Fatores Etários , Animais , Animais Recém-Nascidos , Biomarcadores/metabolismo , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Feminino , Imuno-Histoquímica , Microscopia , Organogênese/efeitos dos fármacos , Gravidez , Ratos , Ratos Wistar , Útero/metabolismo , Útero/patologiaRESUMO
Organ cell diversity depends on binary cell-fate decisions mediated by the Notch signalling pathway during development and tissue homeostasis. A clear example is the series of binary cell-fate decisions that take place during asymmetric cell divisions that give rise to the sensory organs of Drosophila melanogaster. The regulated trafficking of Sanpodo, a transmembrane protein that potentiates receptor activity, plays a pivotal role in this process. Membrane lipids can regulate many signalling pathways by affecting receptor and ligand trafficking. It remains unknown, however, whether phosphatidic acid regulates Notch-mediated binary cell-fate decisions during asymmetric cell divisions, and what are the cellular mechanisms involved. Here we show that increased phosphatidic acid derived from Phospholipase D leads to defects in binary cell-fate decisions that are compatible with ectopic Notch activation in precursor cells, where it is normally inactive. Null mutants of numb or the α-subunit of Adaptor Protein complex-2 enhance dominantly this phenotype while removing a copy of Notch or sanpodo suppresses it. In vivo analyses show that Sanpodo localization decreases at acidic compartments, associated with increased internalization of Notch. We propose that Phospholipase D-derived phosphatidic acid promotes ectopic Notch signalling by increasing receptor endocytosis and inhibiting Sanpodo trafficking towards acidic endosomes.
Assuntos
Proteínas de Drosophila/metabolismo , Drosophila/genética , Mecanorreceptores/fisiologia , Organogênese/efeitos dos fármacos , Organogênese/genética , Ácidos Fosfatídicos/farmacologia , Transporte Proteico/genética , Receptores Notch/metabolismo , Transdução de Sinais/efeitos dos fármacos , Complexo 2 de Proteínas Adaptadoras/fisiologia , Animais , Divisão Celular Assimétrica , Drosophila/citologia , Drosophila/embriologia , Proteínas de Drosophila/fisiologia , Endocitose/fisiologia , Endossomos/metabolismo , Feminino , Hormônios Juvenis/fisiologia , Proteínas dos Microfilamentos/metabolismoRESUMO
The aim of this study was to evaluate the impact of prenatal testosterone exposure on prostate development in male and female neonatal gerbils. Pregnant females were exposed to subcutaneous injections of testosterone cypionate (500 µg/animal) at gestational days 20 and 22. Male and female pups were then euthanized at postnatal day 1. Morphological analysis showed that females were severely affected by androgen exposure. We also observed that male and female urogenital sinus (UGS) responded differentially to testosterone treatment, demonstrating heterogeneous immunostaining for the androgen receptor (AR), estrogen receptor alpha (ERα), and proliferating cell nuclear antigen (PCNA). Smooth muscle α-actin (α-SMA) analysis showed that testosterone delays the myodifferentiation, allowing buds to reach the ectopic mesenchymes of the female UGS. Our data showed that abnormal testosterone exposure disrupted prostate organogenesis, altered the expression patterns of important markers, and demonstrated that female UGS was particularly influenced by androgen exposure during a critical window in the developmental period.
Assuntos
Organogênese/efeitos dos fármacos , Próstata/crescimento & desenvolvimento , Testosterona/farmacologia , Animais , Receptor alfa de Estrogênio/metabolismo , Feminino , Gerbillinae , Imageamento Tridimensional , Masculino , Antígeno Nuclear de Célula em Proliferação/metabolismo , Próstata/anatomia & histologia , Próstata/diagnóstico por imagem , Próstata/efeitos dos fármacos , Receptores Androgênicos/metabolismo , Testosterona/sangueRESUMO
Finasteride is a drug that is widely used in the treatment of benign prostatic hyperplasia, hair loss and even as a chemotherapeutic agent in the treatment of prostatic adenocarcinoma. However, its use is known to cause several side effects in adults and it can also cause changes in the embryonic development of the male prostate, which is a cause for concern given the possibility of the accumulation of finasteride in the environment. Nevertheless, no studies have investigated the effects of finasteride on the development of the prostate in females, which occurs in several species of mammals. To evaluate the effects of intrauterine exposure to finasteride (500µgkg-1 day-1) on postnatal prostate development in the Mongolian gerbil in the present study, we used immunohistochemistry, immunofluorescence, serological analysis and three-dimensional reconstruction techniques. Differences were observed in the effects of finasteride on periductal smooth muscle and cell proliferation between the sexes, as well as intersex differences in the presence of the androgen receptor, which was elevated in males, and the oestrogen receptor ERα, which was increased in females. Together, the data indicate that the female prostate has its own hormone dynamics and that there are sex-specific differences in the way in which the female prostate reacts to prenatal exposure to finasteride.
Assuntos
Finasterida/farmacologia , Gerbillinae/crescimento & desenvolvimento , Efeitos Tardios da Exposição Pré-Natal , Próstata/efeitos dos fármacos , Próstata/crescimento & desenvolvimento , Animais , Feminino , Imuno-Histoquímica , Masculino , Organogênese/efeitos dos fármacos , Gravidez , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Efeitos Tardios da Exposição Pré-Natal/fisiopatologia , Efeitos Tardios da Exposição Pré-Natal/veterinária , Próstata/metabolismo , Receptores Androgênicos/efeitos dos fármacos , Receptores Androgênicos/metabolismo , Receptores de Estrogênio/efeitos dos fármacos , Receptores de Estrogênio/metabolismo , Caracteres SexuaisRESUMO
Perigestational alcohol consumption up to early organogenesis can produce abnormal maternal vascularization via altered decidual VEGF/receptor expression. CF-1 female mice were administered with 10% ethanol in drinking water for 17 days prior to and up to day 10 of gestation. Control females received water without ethanol. Treated females had reduced frequency of implantation sites with expanded vascular lumen (P < 0.05), α-SMA-immunoreactive spiral arteries in proximal mesometrial decidua, reduced PCNA-positive endothelial cells (P < 0.01) and diminished uterine NK cell numbers (P < 0.05) in proximal decidua compared to controls. The VEGF expression (laser capture microscopy, RT-PCR, western blot and immunohistochemistry) was reduced in decidual tissue after perigestational alcohol consumption (P < 0.05). The uNK-DBA+ cells of treated females had reduced VEGF immunoexpression compared to controls (P < 0.01). Very low decidual and endothelial cell KDR immunoreactivity and reduced decidual gene and protein KDR expression was found in treated females compared to controls (P < 0.001). Instead, strong FLT-1 immunoexpression was detected in decidual and uNK cells (P < 0.05) in the proximal decidua from treated females compared to controls. In conclusion, perigestational alcohol ingestion induces the reduction of lumen expansion of spiral arteries, concomitant with reduced endothelial cell proliferation and uNK cell population, and uncompleted remodeling of the artery smooth muscle. These effects were supported by low decidual VEGF and KDR gene and protein expression and increased FLT-1 expression, suggesting that VEGF and KDR reduction may contribute, in part, to mechanisms involved in deficient decidual angiogenesis after perigestational alcohol consumption in mouse.
Assuntos
Consumo de Bebidas Alcoólicas/efeitos adversos , Decídua/irrigação sanguínea , Endotélio Vascular/patologia , Exposição Materna/efeitos adversos , Neovascularização Patológica/patologia , Organogênese/efeitos dos fármacos , Efeitos Tardios da Exposição Pré-Natal/patologia , Animais , Células Cultivadas , Decídua/efeitos dos fármacos , Decídua/metabolismo , Decídua/patologia , Implantação do Embrião/efeitos dos fármacos , Embrião de Mamíferos/irrigação sanguínea , Embrião de Mamíferos/efeitos dos fármacos , Embrião de Mamíferos/metabolismo , Embrião de Mamíferos/patologia , Desenvolvimento Embrionário , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Feminino , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/metabolismo , Células Matadoras Naturais/patologia , Camundongos , Neovascularização Patológica/induzido quimicamente , Gravidez , Efeitos Tardios da Exposição Pré-Natal/etiologia , Útero/efeitos dos fármacos , Útero/metabolismo , Útero/patologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
Yerba mate (Ilex paraguariensis A. St.-Hil.; Aquifoliaceae) is a popular tonic and stimulant beverage that is widely consumed in different South American countries. Estimates indicate the consumption of >1 L per day in southern Brazil and Uruguay. Despite its relatively high consumption, data on reproductive toxicity during critical periods of gestation remain unclear. Thus, we evaluated the effects of an aqueous extract of I. paraguariensis leaves ("chimarrão" [IPC]) at two critical periods of gestation in Wistar rats: preimplantation embryonic stage and fetal organogenesis. Pregnant Wistar rats were orally treated with IPC (3, 30, and 300 mg/kg) from days 1 to 7 or 8 to 21 of pregnancy. The respective control groups received vehicle. During treatment, clinical signs of maternal toxicity, maternal body weight, and food and water intake were monitored. The rats were killed on days 8 and 20 of pregnancy, and the following parameters were evaluated: weight of the maternal uterus, weight of the liver, weight of the kidneys, weight of the spleen, total embryo implantation, preimplantation loss, the mean of live fetuses, the percentage of dead fetuses, fetus weight, and fetal malformation. The aqueous extract of the leaves of I. paraguariensis L. did not present any deleterious effects on preimplantation embryos or the organogenesis of offspring from female Wistar rats. These safety data provide evidence that IPC may be safe for consumption during gestation.
Assuntos
Desenvolvimento Fetal/efeitos dos fármacos , Feto/efeitos dos fármacos , Ilex paraguariensis/toxicidade , Reprodução/efeitos dos fármacos , Chás de Ervas/toxicidade , Animais , Contraindicações de Medicamentos , Feminino , Organogênese/efeitos dos fármacos , Folhas de Planta , Gravidez , Ratos Wistar , América do SulRESUMO
Ganciclovir (GCV) has been implicated in the development of testicular alterations. Exposure on gestational day (GD) 10 in rats induced permanent effects, including focal reduction or absence of germ cells (Sertoli cell-only tubules). Because the timing of exposure can be critical for testicular effects, we exposed rat dams to 300 mg/kg GCV (3 100 mg/kg subcutaneous injections) on GD10, 14 and 19, when germ cells have high rates of migration, proliferation and are mitotically quiescent, respectively. Males exposed to GCV in utero on GD10 and 14 were evaluated for androgenization markers, serum and fecal androgens, and testicular histomorphometry at adulthood. Double-labeling immunofluorescence for DAZL and Ki67 were used to assess gonocytes number and the proliferative activity of germ and somatic cells in fetal testes on GD15 and 20, ie, 24 h after GCV exposure. Adult rats exposed on GD14 showed delayed puberty onset, despite normal androgen levels. Also, there was a 50% reduction in testicular weight and about 30% of seminiferous tubules lacking germ cells. Effects on GD10 animals were less pronounced. In the fetal testis, the number of gonocytes was reduced by 50% in rats exposed on GD14, but normal in GD19 fetuses. GCV also reduced Sertoli cell proliferation immunolabeling in GD19 fetuses and Sertoli cell number in adults. In conclusion, GCV toxicity on germ cells seems to be linked to their proliferation rate and GD14 is a critical window in rats, when GCV exposure causes an acute massive loss of germ cells that persists until adulthood.
Assuntos
Antivirais/administração & dosagem , Ganciclovir/administração & dosagem , Organogênese/efeitos dos fármacos , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Maturidade Sexual/efeitos dos fármacos , Testículo/efeitos dos fármacos , Animais , Antivirais/toxicidade , Proliferação de Células/efeitos dos fármacos , Feminino , Ganciclovir/toxicidade , Células Germinativas/efeitos dos fármacos , Células Germinativas/patologia , Idade Gestacional , Masculino , Gravidez , Efeitos Tardios da Exposição Pré-Natal/patologia , Ratos , Ratos Wistar , Testículo/embriologia , Testículo/crescimento & desenvolvimento , Fatores de TempoRESUMO
Endocrine disrupting compounds (EDCs) have the potential to alter fish reproduction at various levels of organization. The aim of this study was to assess the impact of a natural environment with heavily anthropogenic influence on the physiological processes involved in reproduction in the freshwater fish lambari (Astyanax fasciatus) using different biomarkers. Adult males and females were collected in different seasons from two distinct sites in the same watershed: Ponte Nova Reservoir (PN) considered a pristine or small anthropogenic influence reference point; and Billings Reservoir (Bil), subjected to a large anthropogenic impact. Biological indices, such as hepatosomatic index and gonadosomatic index (GSI), gonadal histomorphology, fecundity, and biomarkers such as plasma levels of estradiol (E2) as well as hepatic gene expression of its alfa nuclear receptor (ERα), were analyzed. Hepatic vitellogenin (VTG) gene expression was evaluated in both sexes, as an indicator of xenoestrogen exposure. Females collected at PN presented a typical annual variation reflected in GSI, whereas for those sampled at Bil the index did not change through the seasons. The higher concentration of E2 in males collected at Bil during spring/2013, together with the detection of VTG gene expression, suggest the presence of EDCs in the water. These EDCs may have also influenced fecundity of females from Bil, which was higher during winter and spring/2013. Gene expression of ERα and ovarian morphology did not differ between fish from both sites. Water conditions from Bil reservoir impacted by anthropic activity clearly interfered mainly with biomarkers of biological effect such as plasma E2 levels and absolute and relative fecundity, but also altered biomarkers of exposure as VTG gene expression. These facts support the notion that waterborne EDCs are capable of causing estrogenic activity in A. fasciatus.
Assuntos
Characidae/metabolismo , Disruptores Endócrinos/toxicidade , Monitoramento Ambiental/métodos , Gônadas/efeitos dos fármacos , Organogênese/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade , Animais , Biomarcadores/metabolismo , Brasil , Characidae/crescimento & desenvolvimento , Disruptores Endócrinos/análise , Estradiol/metabolismo , Feminino , Fertilidade/efeitos dos fármacos , Água Doce/química , Expressão Gênica/efeitos dos fármacos , Gônadas/crescimento & desenvolvimento , Gônadas/patologia , Masculino , Reprodução/efeitos dos fármacos , Estações do Ano , Vitelogeninas/metabolismo , Poluentes Químicos da Água/análiseRESUMO
Maytenus ilicifolia Mart. ex Reissek is a plant commonly used in folklore medicine in the management of gastric diseases in South America. This study explores the effects of a supratherapeutic dose of aqueous and ethanol extracts of M. ilicifolia (1360 mg/kg) on fertility and neurobehavioral status in male and pregnant rats. A battery of sensory-motor developmental endpoints was carried out to assess impairments on pups of dams orally treated with the aqueous extract of M. ilicifolia during the organogenesis period of pregnancy (GD 9 through GD 14). The neuromotor maturation reflexes and physical developments of the offspring were not significantly different between the groups (p < 0.05). Also, the hippocampal morphology revealed no indices of cell loss in the CA1, CA2, CA3 and CA4 areas. As second protocol, some fertility aspects were investigated in young post pubertal male Wistar rats treated with the ethanol extract for 30 days. The semen quality and testicular tissue morphology of male rats treated with the ethanol extract of M. ilicifolia remained unaffected upon treatment. Thus, the results indicate that the high-dose of M. ilicifolia extracts have no neurotoxic potential on offspring and seem not to affect the sperm quality of male rats.
Assuntos
Comportamento Animal/efeitos dos fármacos , Fertilidade/efeitos dos fármacos , Maytenus/química , Medicina Tradicional/efeitos adversos , Extratos Vegetais/efeitos adversos , Gastropatias/tratamento farmacológico , Animais , Etanol/química , Feminino , Masculino , Organogênese/efeitos dos fármacos , Gravidez , Ratos , Ratos Wistar , Análise do Sêmen , América do Sul , Testículo/efeitos dos fármacos , Água/químicaRESUMO
Perigestational alcohol consumption by CF-1 mouse, from before mating up to the period of embryo organogenesis, leads to retarded early embryo development and neural tube defects. Here, we addressed if perigestational alcohol ingestion up to Day 10 of pregnancy induces oxidative stress and changes in macromolecules and organ tissues of early organogenic embryos. Adult CF-1 female mice were administered 10% ethanol in their drinking water for 17 days prior to mating and until Day 10 of gestation, whereas control females were administered ethanol-free water. Our results demonstrated significantly reduced Catalase abundance and activity and increased glutathione content in the embryos of ethanol-treated females. The nitrite level was significantly reduced, but TBARS (thiobarbituric acid reactive substances) content, an index of lipid peroxidation, did not change. Embryos derived from ethanol-treated females also showed higher abundance of 3-nitrotyrosine (3-NT)-containing proteins in all tissues, compared to the control group. Apoptosis was significantly increased in the ectoderm and mesoderm, but not in the heart-although this organ did contain more cleaved Caspase-3-positive cardiomyocytes per area of ventricular myocardium than controls. In sum, moderate perigestational alcohol ingestion up to Day 10 of gestation in mice induces oxidative stress by altering radical nitrogen species and antioxidant enzymatic and non-enzymatic mechanisms in embryos. Further, generalized protein nitration, due to unbalanced nitric oxide levels associated with tissue-specific apoptosis, was detected in embryos, suggesting that oxidative mechanisms may play an important role in the perigestational alcohol-induced malformation of organogenic embryos exposed to ethanol.