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
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
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
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.
Assuntos
Consumo de Bebidas Alcoólicas/efeitos adversos , Apoptose/efeitos dos fármacos , Embrião de Mamíferos/efeitos dos fármacos , Etanol/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Efeitos Tardios da Exposição Pré-Natal , Consumo de Bebidas Alcoólicas/genética , Consumo de Bebidas Alcoólicas/metabolismo , Consumo de Bebidas Alcoólicas/patologia , Animais , Animais não Endogâmicos , Antioxidantes/metabolismo , Dano ao DNA/efeitos dos fármacos , Embrião de Mamíferos/metabolismo , Desenvolvimento Embrionário/efeitos dos fármacos , Desenvolvimento Embrionário/genética , Etanol/efeitos adversos , Feminino , Camundongos , Organogênese/efeitos dos fármacos , Gravidez , Efeitos Tardios da Exposição Pré-Natal/genética , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Efeitos Tardios da Exposição Pré-Natal/patologiaRESUMO
Studies about toxicological potential of usnic acid are limited. This way, the vast majority of data available in the literature are related only to biological activities. This is the first study that aimed to evaluate the oral toxicity of usnic acid during the period of organogenesis. Females rats were distributed in the control groups, treated I and II, at doses of 15 and 25 mg/kg, administered by gavage during the 6° to 15° days of pregnancy. After 20 days the fetuses were removed and analyzed. A reduction in weight gain during pregnancy, increased resorption, reduction in the number of viable fetuses, and their body weight were observed. Morphological changes in the litter were visualized as exposure of the eye and atrophy of the limbs at the dose of 25 mg/kg. Histological analysis of the liver of the fetus showed reduction in the number of megakaryocytes between experimental groups and increase in the number of hepatocytes in a dose of 25 mg/kg. The experimental model used in this study reveals teratogenic effect of usnic acid in the period of organogenesis. Since this achievement, the importance of evaluating the toxic effects of natural substances is imperative, in order to elucidate the care in their indication as drug.
Assuntos
Benzofuranos/administração & dosagem , Organogênese/efeitos dos fármacos , Teratogênese/efeitos dos fármacos , Animais , Ascomicetos/química , Benzofuranos/química , Peso Corporal/efeitos dos fármacos , Feminino , Desenvolvimento Fetal/efeitos dos fármacos , Feto/efeitos dos fármacos , Masculino , Tamanho do Órgão/efeitos dos fármacos , Gravidez , RatosRESUMO
The Wolffian duct (WD) undergoes morphological changes induced by androgens to form the epididymis, which is an organ essential for sperm maturation. Androgen action in WD epithelium involves paracrine factors of mesenchymal origin that function by still poorly understood mechanisms. Here we studied the antimicrobial ß-defensin SPAG11C as a new player in duct morphogenesis, localized prenatally in the WD mesenchyme. Organotypic culture of rat WDs and tissues from Androgen Receptor (AR) knockout mice (ARKO) were used. Our results show that androgen/AR signaling differentially regulated SPAG11C expression at mRNA and protein levels in the developing WD. WDs incubated with recombinant human SPAG11C were shorter and less coiled as a result of reduced epithelial cell proliferation, but not increased apoptosis. Our results suggested ß-defensin SPAG11C as an androgen-target required for WD morphogenesis. This highlights the multifunctional repertoire of the ß-defensin protein family and their potential contribution to the in utero environment that determines male reproductive success.
Assuntos
Androgênios/farmacologia , Anti-Infecciosos/farmacologia , Morfogênese/efeitos dos fármacos , Ductos Mesonéfricos/efeitos dos fármacos , beta-Defensinas/farmacologia , Animais , Antígenos de Superfície/metabolismo , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Epididimo/efeitos dos fármacos , Epididimo/metabolismo , Epitélio/efeitos dos fármacos , Epitélio/metabolismo , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Humanos , Masculino , Camundongos , Camundongos Knockout , Organogênese/efeitos dos fármacos , Ratos , Ratos Wistar , Receptores Androgênicos/metabolismo , Ductos Mesonéfricos/metabolismoRESUMO
The aim of this study was to evaluate the toxicity of Stryphnodendron fissuratum pods in guinea pigs (Cavia porcellus) and test the hypothesis that this plant has teratogenic effects. Thus, sixteen guinea pigs were randomly divided into four groups of four animals each. Groups 10, 20 and 40 consisted of guinea pigs that received commercial food that contained crushed pods of S. fissuratum at concentrations of 10, 20 and 40 g/kg, respectively, during the period of organogenesis. Control group consisted of guinea pigs under the same management conditions that did not receive crushed pods of S. fissuratum in their food. In all experimental groups, the main clinical signs of poisoning consisted of anorexia, prostration, absence of vocalizations, alopecia, diarrhea, and abortions within the adult guinea pigs. Those that did not abort gave birth to weak, malnourished pups, some of which had fetal malformations. The main teratogenic changes consisted of eventration, arthrogryposis, amelia of the forelimbs, anophthalmia, microphthalmia, anotia and agnathia. The reductions in the number of offspring and the malformations observed in the experimental groups suggest that S. fissuratum affects fetal development and is teratogenic.
Assuntos
Anormalidades Induzidas por Medicamentos , Fabaceae/toxicidade , Desenvolvimento Fetal/efeitos dos fármacos , Cobaias/embriologia , Exposição Materna , Teratogênicos/toxicidade , Animais , Feminino , Organogênese/efeitos dos fármacos , Gravidez , Distribuição AleatóriaRESUMO
High levels of homocysteine (Hcy) are related to an increased risk of the occurrence of congenital anomalies, including limb defects. However, few evaluations about how toxic levels of Hcy affect limb development have been reported. We investigated whether Hcy can affect the cell cycle proteins and proteins involved in mesenchymal cell differentiation during limb development, in a chicken embryo model. Embryos were treated with 20 µmol d-l Hcy/50 µl saline at embryonic day 2 and analyzed at embryonic day 6. Untreated control embryos received exclusively 50 µl saline solution. To identify cells in proliferation and cell cycle proteins, as well as Pax1/9 and Sox9 proteins, we performed immunolocalization and flow cytometry analyses using the antibodies anti-phosphohistone H3, anti-p53, anti-p21, anti-proliferating cell nuclear antigen, anti-Pax1, anti-Pax9 and anti-Sox9. No significant differences in cell proliferation were observed between Hcy-treated and untreated embryos. We observed a decrease of the proliferating cell nuclear antigen and p21 proteins, both involved in the G1 phase of cell cycle progression. On the other hand, in mesenchymal cells of the limbs, Hcy induces an increase of p53 protein, which can be activated by DNA damage. In cell differentiation, Hcy induced an increase mainly of Pax9 and Sox9 proteins. Our data indicate that the treatment with Hcy changes the mesenchymal cell dynamics during limb development, but does not change the morphology of the cartilage molds. These findings provide information to understand better the cellular basis of the toxicity of Hcy on chondrogenesis during limb development.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Condrogênese/efeitos dos fármacos , Homocisteína/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Organogênese/efeitos dos fármacos , Animais , Embrião de Galinha , Dano ao DNA , Extremidades/embriologia , Células-Tronco Mesenquimais/metabolismo , Fator de Transcrição PAX9/genética , Fator de Transcrição PAX9/metabolismo , Fatores de Transcrição SOX9/genética , Fatores de Transcrição SOX9/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
Tebuconazole is a synthetic agrochemical action systemic antifungal triazole. The maternal exposure to triazoles, during pregnancy, can cause adverse effects on maternal-placental-fetal united, resulting in alterations in the development of the fetus, thus it becomes essential to characterize these effects. This study aimed to analyze behavioral aspects of rats and physical development, sensory and motor litter. We used pregnant wistar rats treated orally with Tebuconazole doses (20 and 50 mg / kg / day) and rats in the Control Group with distilled water during the organogenic period (6th to 15th). After birth, there was the maternal reproductive performance: time of pregnancy, number of live pups, stillbirths, congenital malformations and the weight of the litter. Observed maternal behavior and lactation. Litters were weighed, daily from the 2nd to the 9th day after birth and the 10th dpn to 2 th of lactation, the individual weight. Also pups evaluated in the physical, sensoral development and motor activity in the open field. The results showed signs of acute toxicity in pregnant rats treated with 50 mg / kg / day occurring decrease in body weight gain, reducing the number of corpora lutea and maternal deaths as compared to controls and to the group 20mg/kg / days. In litters of rats exposed to doses of 20 and 50mg/kg/day there was a delay in the development of sensory and motor activity. Treatment with Tebuconazole, in subtoxic doses caused maternal toxicity, adverse effects on reproductive performance and fetotoxic.
O Tebuconazol é um agroquímico sintético de ação sistêmica, antifúngico do grupo dos triazóis. A exposição aos triazóis, durante o período gestacional, pode causar efeitos adversos na unidade materno-placentária-fetal, resultando em alterações no desenvolvimento do concepto, dessa forma torna-se imprescindível a caracterização desses efeitos. Objetivou-se analisar aspectos comportamentais das ratas e o desenvolvimento físico, sensorial e motor das ninhadas. Utilizou-se ratas Wistar prenhes, tratadas por via oral com o Tebuconazol nas doses (20 e 50 mg/kg/dia) e as ratas do Grupo Controle com água destilada, durante o período organogênico (6º ao 15º dia). Após o nascimento, observou-se a performance reprodutiva materna: tempo de prenhez, número de filhotes vivos, natimortos, malformações e ou anomalias e o peso total da ninhada. Observou o comportamento materno e a lactação. Pesaram-se as ninhadas diariamente, do 2º até o 9º dia-pós-nascimento e a partir do 10º dpn ao 21º dia de lactação verificou-se o peso individual. Também avaliou-se nos filhotes o desenvolvimento físico, a análise sensorial e a atividade motora em campo aberto. Os resultados mostraram sinais de intoxicação aguda nas ratas prenhes tratadas com a dose de 50 mg/kg/dia ocorrendo diminuição do ganho de peso corporal, redução do número de corpos lúteos e mortes maternas, quando comparados aos controles e ao grupo...
Assuntos
Animais , Ratos , Feto/anormalidades , Organogênese/efeitos dos fármacos , Ratos Wistar , Toxicidade , Triazóis/efeitos adversosRESUMO
Tebuconazole is a synthetic agrochemical action systemic antifungal triazole. The maternal exposure to triazoles, during pregnancy, can cause adverse effects on maternal-placental-fetal united, resulting in alterations in the development of the fetus, thus it becomes essential to characterize these effects. This study aimed to analyze behavioral aspects of rats and physical development, sensory and motor litter. We used pregnant wistar rats treated orally with Tebuconazole doses (20 and 50 mg / kg / day) and rats in the Control Group with distilled water during the organogenic period (6th to 15th). After birth, there was the maternal reproductive performance: time of pregnancy, number of live pups, stillbirths, congenital malformations and the weight of the litter. Observed maternal behavior and lactation. Litters were weighed, daily from the 2nd to the 9th day after birth and the 10th dpn to 2 th of lactation, the individual weight. Also pups evaluated in the physical, sensoral development and motor activity in the open field. The results showed signs of acute toxicity in pregnant rats treated with 50 mg / kg / day occurring decrease in body weight gain, reducing the number of corpora lutea and maternal deaths as compared to controls and to the group 20mg/kg / days. In litters of rats exposed to doses of 20 and 50mg/kg/day there was a delay in the development of sensory and motor activity. Treatment with Tebuconazole, in subtoxic doses caused maternal toxicity, adverse effects on reproductive performance and fetotoxic.(AU)
O Tebuconazol é um agroquímico sintético de ação sistêmica, antifúngico do grupo dos triazóis. A exposição aos triazóis, durante o período gestacional, pode causar efeitos adversos na unidade materno-placentária-fetal, resultando em alterações no desenvolvimento do concepto, dessa forma torna-se imprescindível a caracterização desses efeitos. Objetivou-se analisar aspectos comportamentais das ratas e o desenvolvimento físico, sensorial e motor das ninhadas. Utilizou-se ratas Wistar prenhes, tratadas por via oral com o Tebuconazol nas doses (20 e 50 mg/kg/dia) e as ratas do Grupo Controle com água destilada, durante o período organogênico (6º ao 15º dia). Após o nascimento, observou-se a performance reprodutiva materna: tempo de prenhez, número de filhotes vivos, natimortos, malformações e ou anomalias e o peso total da ninhada. Observou o comportamento materno e a lactação. Pesaram-se as ninhadas diariamente, do 2º até o 9º dia-pós-nascimento e a partir do 10º dpn ao 21º dia de lactação verificou-se o peso individual. Também avaliou-se nos filhotes o desenvolvimento físico, a análise sensorial e a atividade motora em campo aberto. Os resultados mostraram sinais de intoxicação aguda nas ratas prenhes tratadas com a dose de 50 mg/kg/dia ocorrendo diminuição do ganho de peso corporal, redução do número de corpos lúteos e mortes maternas, quando comparados aos controles e ao grupo...(AU)
Assuntos
Animais , Ratos , Ratos Wistar , Toxicidade , Organogênese/efeitos dos fármacos , Triazóis/efeitos adversos , Feto/anormalidadesRESUMO
The potential of oral exposure to calcium and magnesium citrate, a natural product obtained from dolomite, to initiate teratogenesis was analyzed in Wistar rats. Animals received calcium and magnesium citrate oral doses of 250, 500 and 1000 mg/kg during the period of gestation from day 6 to 17 post conception. Maternal, embryo and fetal toxicity was evaluated. Calcium and magnesium citrate exposure did not produce maternal toxicity assessed by clinical observations, body weight gain, food intake, hematology, biochemical parameters and necropsy finding. Signs of embryo-fetal toxicity were not observed. Skeletal and visceral malformations were seen occasionally in all drug-treated and control groups. Skeletal and visceral variations were similar in control and drug-treated groups except for incomplete ossification rib. These finding was spontaneous and unrelated to the drug. In conclusion, in this study we found that the oral exposure to rats of up to 1000 mg/kg of calcium and magnesium citrate during organogenesis did not induce significant maternal and embryo-fetal toxicity. The experimentally derived NOAEL for developmental toxicity was 1000 mg/kg.
Assuntos
Citrato de Cálcio/toxicidade , Ácido Cítrico/toxicidade , Exposição Materna/efeitos adversos , Organogênese/efeitos dos fármacos , Compostos Organometálicos/toxicidade , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Animais , Carbonato de Cálcio/química , Citrato de Cálcio/isolamento & purificação , Ácido Cítrico/isolamento & purificação , Suplementos Nutricionais , Relação Dose-Resposta a Droga , Desenvolvimento Embrionário/efeitos dos fármacos , Determinação de Ponto Final , Feminino , Desenvolvimento Fetal/efeitos dos fármacos , Magnésio/química , Anormalidades Musculoesqueléticas/induzido quimicamente , Anormalidades Musculoesqueléticas/embriologia , Nível de Efeito Adverso não Observado , Compostos Organometálicos/isolamento & purificação , Gravidez , Ratos , Ratos Wistar , Costelas/anormalidades , Costelas/efeitos dos fármacos , Costelas/embriologiaRESUMO
Scientific assessment of harmful interactions of chemicals over the entire reproductive cycle are divided into three segments based on the period: from premating and mating to implantation (I), from implantation to major organogenesis (II), and late pregnancy and postnatal development (III). We combined the segments I and II to assess Plathymenia reticulata aqueous extract safety. In order to investigate reproductive toxicity (segment I), pregnant rats received orally 0.5 or 1.0 g/kg of extract, daily, during 18 days. These concentrations were determined by a preliminary in vitro LD50 test in CHO-k1 cells. A control group received deionized water. The offspring was removed at the 19th day, by caesarean, and a teratology study (segment II) was carried out. The corpora lutea, implants, resorptions, live, and dead fetuses were then counted. Placenta and fetuses were weighted. External and visceral morphology were provided by the fixation of fetuses in Bouin, whereas skeletal analysis was carried out on the diaphanizated ones. The increase in the weights of placenta and fetuses was the only abnormality observed. Since there was no sign of alteration on reproduction parameters at our experimental conditions, we conclude that P. reticulata aqueous extract is safe at 0.5 to 1.0 g/kg and is not considered teratogenic.
Assuntos
Feto/efeitos dos fármacos , Organogênese/efeitos dos fármacos , Extratos Vegetais/administração & dosagem , Animais , Relação Dose-Resposta a Droga , Fabaceae/química , Feminino , Humanos , Extratos Vegetais/química , Gravidez , Ratos , Teratogênicos/química , Teratogênicos/farmacologia , Água/químicaRESUMO
Although some studies have pointed out to embryo/fetal toxicity, knowledge about the potential toxicity of the fungicide epoxiconazole is still limited. Once the results of these previous studies have raised some concern, this study studied the effects of epoxiconazole maternal exposure on the physical endpoints in the development of rat pups. To accomplish that, the effects of epoxiconazole (50.0, 100.0, and 150.0 mg/kg) were examined when rats were exposed at two different developmental stages: during the first 6 days of pregnancy or in the organogenesis period (6-15 days). After parturition, pups were tested for growth and maturational milestones. Maternal exposure to the fungicide, independently of phase, resulted in significantly early mean time to vaginal opening and delayed time to testes descent in pups. Weight gain rate in pups and their mothers was not affected for the tested exposure period. The findings of this study emphasize that epoxiconazole maternal exposure may lead to alterations in developmental patterns in nursing pups, consistent with the known influence of epoxiconazole on steroid hormone synthesis.
Assuntos
Compostos de Epóxi/toxicidade , Fungicidas Industriais/toxicidade , Organogênese/efeitos dos fármacos , Efeitos Tardios da Exposição Pré-Natal , Reprodução/efeitos dos fármacos , Triazóis/toxicidade , Animais , Animais Recém-Nascidos/crescimento & desenvolvimento , Relação Dose-Resposta a Droga , Disruptores Endócrinos/toxicidade , Feminino , Lactação , Modelos Lineares , Masculino , Gravidez , Ratos , Ratos WistarRESUMO
Proteolysis carried out by different proteases control cellular processes during development and regeneration. Here we investigated the function of the proteasome and other proteases in the process of intestinal regeneration using as a model the sea cucumber Holothuria glaberrima. This echinoderm possesses the ability to regenerate its viscera after a process of evisceration. Enzymatic activity assays showed that intestinal extracts at different stages of regeneration possessed chymotrypsin-like activity. This activity was inhibited by i) MG132, a reversible inhibitor of chymotrypsin and peptidylglutamyl peptidase hydrolase (PGPH) activities of the proteasome, ii) E64d, a permeable inhibitor of cysteine proteases and iii) TPCK, a serine chymotrypsin inhibitor, but not by epoxomicin, an irreversible and potent inhibitor of all enzymatic activities of the proteasome. To elucidate the role which these proteases might play during intestinal regeneration, we carried out in vivo experiments injecting MG132, E64d and TPCK into regenerating animals. The results showed effects on the size of the regenerating intestine, cell proliferation and collagen degradation. These findings suggest that proteolysis by several proteases is important in the regulation of intestinal regeneration in H. glaberrima.