RESUMO
We previously reported that exposure during gestation and lactation to a low dose of glyphosate-based herbicide (GBH) reduced the area and perimeter of male offspring mammary gland at postnatal day 60 (PND60), whereas a higher dose increased the longitudinal growth of the gland. Here, our aim was to assess whether perinatal exposure to GBH exhibits endocrine disruptive action in male mammary gland at an early time point (pre-puberty), which could be related to the changes observed after puberty. We also wanted to explore whether an early evaluation of the male rat mammary gland is appropriate to assess exposure to potential endocrine disrupting chemicals (EDCs). Pregnant rats were orally exposed, through the diet, to vehicle (saline solution), 3.5 or 350 mg/kg/day of GBH from gestational day 9 until weaning. At PND21, the male offspring were euthanized, and mammary gland samples were collected. The histology and proliferation index of the mammary glands were evaluated, and the mRNA expression of estrogen (ESR1) and androgen (AR) receptors, cyclin D1 (Ccnd1), amphiregulin (Areg), insulin-like growth factor 1 (IGF1), epidermal growth factor receptor (EGFR) and IGF1 receptor (IGF1R) were assessed. Moreover, the phosphorylated-Erk1/2 (p-ERK1/2) protein expression was determined. No differences were observed in mammary epithelial structures and AR expression between experimental groups; however, the proliferation index was reduced in GBH3.5-exposed males. This result was associated with decreased ESR1, Ccnd1, Areg, IGF1, EGFR and IGF1R mRNA expressions, as well as reduced p-Erk1/2 protein expression in these animals. ESR1, Ccnd1, IGF1R and EGFR expressions were also reduced in GBH350-exposed males. In conclusion, the mammary gland development of pre-pubertal male rats is affected by perinatal exposure to GBH. Although further studies are still needed to understand the molecular mechanisms involved in GBH350 exposure, the present results may explain the alterations observed in mammary gland growth of post-pubertal males exposed to low doses of GBH. Our results also suggest that early evaluation of the male rat mammary gland is useful in assessing exposure to potential EDCs. However, analysis of EDCs effects at later time points should not be excluded.
Assuntos
Disruptores Endócrinos/toxicidade , Glicina/análogos & derivados , Herbicidas/toxicidade , Glândulas Mamárias Animais/crescimento & desenvolvimento , Actinas/metabolismo , Animais , Feminino , Glicina/toxicidade , Peptídeos e Proteínas de Sinalização Intercelular/biossíntese , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Glândulas Mamárias Animais/efeitos dos fármacos , Gravidez , Efeitos Tardios da Exposição Pré-Natal/patologia , Ratos , Ratos Wistar , Receptores de Estrogênio/efeitos dos fármacos , Receptores de Estrogênio/genética , Receptores de Fatores de Crescimento/biossíntese , Receptores de Esteroides/biossíntese , GlifosatoRESUMO
INTRODUCTION: The present study investigates, in 23-day-old and adult male rats, the effect of severe food restriction in utero on blood pressure (BP), and its association with nephron structure and function changes, angiotensin II (AT1R/AT2R), glucocorticoid (GR) and mineralocorticoid (MR) receptor expression. MATERIALS AND METHODS: The daily food supply to pregnant rats was measured and one group (n=15) received normal quantity of food (NF) while the other received 50% of that (FR50%) (n=15). Kidneys were processed to AT1R, AT2R, MR, and GR immunolocalization and for western blotting analysis. The renal function was estimated by creatinine and lithium clearances in 12-week-old offspring. RESULTS: By stereological analyses, FR50% offspring present a reduction of nephron numbers (35%) with unchanged renal volume. Expression of AT1R and AT2R was significantly decreased in FR50% while the expression of GR and MR increased in FR50%. We also verified a pronounced decrease in urinary sodium excretion accompanied by increased BP in 12-week-old FR50% offspring. CONCLUSION: The current data suggest that changes in renal function are conducive to excess sodium tubule reabsorption, and this might potentiate the programming of adult hypertension. It is plausible to arise in the current study an association between decreasing natriuresis, reciprocal changes in renal AngII and steroid receptors with the hypertension development found in FR50% compared with age-matched NF offspring.
Assuntos
Restrição Calórica , Desenvolvimento Fetal/fisiologia , Rim/embriologia , Receptores de Esteroides/biossíntese , Sódio/urina , Animais , Peso ao Nascer , Pressão Sanguínea , Creatinina/urina , Feminino , Feto , Rim/crescimento & desenvolvimento , Testes de Função Renal , Glomérulos Renais/embriologia , Glomérulos Renais/crescimento & desenvolvimento , Masculino , Gravidez , Ratos , Ratos Wistar , Receptor Tipo 2 de Angiotensina/biossíntese , Receptor Tipo 2 de Angiotensina/genética , Receptores de Glucocorticoides/biossíntese , Receptores de Mineralocorticoides/biossínteseRESUMO
OBJECTIVE: To evaluate the effect of melatonin both on the ovaries of pinealectomized female rats through histomorphometric analysis and on steroid receptors, proliferating cell nuclear antigen (PCNA), and vascular endothelial growth factor (VEGF) expression. DESIGN: Experimental study. SETTING: Federal University of São Paulo, Brazil. ANIMAL(S): Forty female rats. INTERVENTION(S): Forty rats were divided equally into four groups: GI-vehicle without surgery; GII--surgery without removal of the pineal gland (sham); GIII--pinealectomized with vehicle; and GIV--pinealectomized with melatonin treatment. After treatment for 3 consecutive months, the animals were killed and their ovaries removed for analysis. MAIN OUTCOME MEASURE(S): Estrogen and progesterone receptors, histologic and immunohistochemical analysis. RESULT(S): The GIII samples presented signals of proliferation on ovarian surface epithelium and interstitial cells as well as high expressions of PCNA and VEGF in those structures compared with GI, GII, and GIV. Also, the levels of progesterone receptor (fmol/g) in ovaries of GIII (250.6 ± 32.4) were significantly lower than in those of GI (429.0 ± 23,8), GII (442.3 ± 30.2), and GIV (564.1 ± 78.7). The levels of progesterone in GIII were superior to those in GI, GII, and GIV. CONCLUSION(S): Our findings suggest that melatonin may attenuate proliferation in ovarian structures and increase the number of luteal bodies as well as the levels of progesterone receptor.
Assuntos
Melatonina/fisiologia , Ovário/metabolismo , Glândula Pineal/cirurgia , Antígeno Nuclear de Célula em Proliferação/biossíntese , Receptores de Esteroides/biossíntese , Fator A de Crescimento do Endotélio Vascular/biossíntese , Animais , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Feminino , Regulação da Expressão Gênica , Ovário/citologia , Glândula Pineal/metabolismo , Ligação Proteica/genética , Ratos , Ratos Wistar , Receptores de Esteroides/genética , Transdução de Sinais/genética , Fator A de Crescimento do Endotélio Vascular/genéticaRESUMO
Fine particulate air pollutants, mainly their organic fraction, have been demonstrated to be associated with cardiovascular and respiratory health problems. Puerto Rico has been reported to have the highest prevalence of pulmonary diseases (e.g., asthma) in the United States. The aim of this study was to assess, for the first time, the immunological response of human bronchial epithelial cells (BEAS-2B) to organic extracts isolated from airborne particulate matter (PM(2.5)) in Puerto Rico. Organic extracts from PM(2.5) collected throughout an 8-month period (2000-2001) were pooled (composite) in order to perform chemical analysis and biological activity testing. BEAS-2B cells were exposed to PM(2.5) organic extract to assess cytotoxicity, levels of cytokines and relative gene expression of MHC-II, hPXR and CYP3A5. Our findings show that organic PM(2.5) consist of toxic as well as bioactive components that can regulate the secretion of cytokines in BEAS-2B, which could modulate inflammatory response in the lung. Trace element analyses confirmed the presence of metals in organic extracts highlighting the relative high abundance of Cu and Zn in polar organic extracts. Polar organic extracts exhibited dose-dependant toxicity and were found to significantly induce the release of interleukin 6 (IL-6), IL-1beta and IL-7 while significantly inhibiting the secretion of IL-8, G-CSF and MCP-1. Moreover, MHC-II transcriptional activity was up-regulated after 24 h of exposure, whereas PXR and CYP3A5 were down-regulated. This research provides a new insight into the effects of PM(2.5) organic fractions on specific effectors and their possible role in the development of respiratory inflammatory diseases in Puerto Rico.
Assuntos
Poluentes Ocupacionais do Ar/toxicidade , Biomarcadores/análise , Brônquios/patologia , Células Epiteliais/imunologia , Células Epiteliais/patologia , Material Particulado/toxicidade , Poluentes Ocupacionais do Ar/análise , Brônquios/citologia , Brônquios/imunologia , Linhagem Celular , Citocromo P-450 CYP3A/biossíntese , Citocinas/análise , Monitoramento Ambiental , Genes MHC da Classe II/efeitos dos fármacos , Antígeno HLA-DR2/biossíntese , Humanos , Material Particulado/análise , Receptor de Pregnano X , Porto Rico , RNA Mensageiro/biossíntese , RNA Mensageiro/isolamento & purificação , Receptores de Esteroides/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Supressão Genética/efeitos dos fármacos , Oligoelementos/toxicidade , Ativação TranscricionalRESUMO
OBJECTIVE: To evaluate gene and protein expression of steroid receptors, nuclear receptor coregulators, and uterine receptivity markers in midsecretory phase endometria from untreated women with polycystic ovary syndrome (PCOS). DESIGN: Case-control study. SETTING: Hospital research unit. PATIENT(S): Eight patients with PCOS and eight fertile women of similar age to those with PCOS. INTERVENTION(S): Endometrial samples were obtained from women with PCOS (PCOSE) and normal (NE) women during the midsecretory phase of the menstrual cycle. MAIN OUTCOME MEASURE(S): Expression studies (immunohistochemistry, reverse transcription-polymerase chain reaction [RT-PCR] and Western blot). RESULT(S): Endometria from PCOS exhibit higher levels of messenger RNA (mRNA) and protein for estrogen receptor alpha and coactivators than NE. Epithelial cells had a greater expression of progesterone receptor in PCOSE, whereas, no differences were observed in gene and protein expression of the nuclear corepressor (NcoR) and the antiadhesion molecule mucin type-1 (MUC-1) between PCOSE and NE. Immunodetection for the coactivator ARA70 was higher in PCOSE than in NE; in contrast, expression of beta3-integrin in epithelia was lower in PCOSE than in control endometria. CONCLUSION(S): The higher response to steroid hormones of endometria from untreated PCOS-women induces diminished expression of beta3 integrin, which partially explain implantation failure in PCOS patients.
Assuntos
Implantação do Embrião/fisiologia , Endométrio/metabolismo , Síndrome do Ovário Policístico/metabolismo , Receptores de Esteroides/biossíntese , Fatores de Transcrição/biossíntese , Adulto , Western Blotting , Estudos de Casos e Controles , Endométrio/química , Feminino , Humanos , Integrina beta3/biossíntese , Integrina beta3/genética , Síndrome do Ovário Policístico/fisiopatologia , RNA Mensageiro/biossíntese , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Útero/química , Útero/metabolismoRESUMO
The aim of this study was to examine, using semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) the changes in mRNA expression of the two estrogen receptor (ER) subtypes, ERalpha and ERbeta, prolactin receptor long and short form, and progesterone (Pg) receptor (PgR), in liver and mammary gland during gestation, early lactation, and weaning in both hyperthyroid (HT) and normal rats. Pregnancy increased long prolactin receptors (PRL-R(L)) and ERalpha mRNAs in liver and PRL-R(I) in mammary gland. Lactation decreased PRL-R(L) in liver and ERbeta and PgR in mammary gland. HT decreased PRL-R(L), at the end of pregnancy (G21), ERalpha (in G21 and L1) in liver and PRL-R(L) in L1 as well as short prolactin receptors (PRL-R(S)) (G7, L1) and ERbeta (G7, G14, L4) in mammary gland. In conclusion, our data indicated that (1) PRL-R1 and ERalpha expression levels are differentially regulated in the liver, and PgR and ERbeta in mammary gland during pregnancy and lactation (2) ERbeta is variably expressed depending on the state of thyroid hormones, however the ERalpha gene expression remained constant in mammary gland. (3) PRL-R1 mRNA expression is highly induced in the mammary gland during late pregnancy and abruptly declines on the first day of lactation for the HT rats.
Assuntos
Hipertireoidismo/metabolismo , Fígado/metabolismo , Glândulas Mamárias Animais/metabolismo , Prenhez/metabolismo , Receptores da Prolactina/biossíntese , Receptores de Esteroides/biossíntese , Hormônios Tireóideos/fisiologia , Animais , Receptor alfa de Estrogênio/biossíntese , Receptor alfa de Estrogênio/genética , Receptor beta de Estrogênio/biossíntese , Receptor beta de Estrogênio/genética , Feminino , Regulação da Expressão Gênica , Hipertireoidismo/genética , Lactação , Período Pós-Parto/genética , Período Pós-Parto/metabolismo , Gravidez , Prenhez/genética , RNA/biossíntese , RNA/genética , Ratos , Ratos Wistar , Receptores de Progesterona/biossíntese , Receptores de Progesterona/genética , Receptores da Prolactina/genética , Receptores de Esteroides/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
NOR-1 is an orphan member of the nuclear receptor superfamily, which includes a group of transcription factors involved in the response to steroids, fatty acids, retinoic acids, and other lipophilic molecules. The NOR-1 subfamily (NR4), composed also of Nurr1 and Nurr77, has been implicated in cell proliferation, differentiation, apoptosis, chondrosarcomas, inflammation, and atherogenesis. The NOR-1 receptor is an orphan ligand receptor which acts over gene transactivation. No ligands, if such in fact exist, are known for this receptor. Recently, the three-dimensional structure of the homolog receptor Nurr1 has been solved using protein crystallography techniques. Surprisingly, the structure does not present either a typical cavity for ligand binding or a classical co-factor binding site in the ligand binding domain (LBD). To allow for structural studies of other members of NR4 subfamily, we have subcloned, overexpressed in Escherichia coli cells, purified, and characterized the rat orphan nuclear receptor NOR-1 LBD domain. We obtained NOR-1 LBD at a high degree of purity and with an overall yield of 3 mg/L of culture media. CD spectroscopic analysis shows a high alpha-helical secondary structure content (52%), similar to that of Nurr 1 LBD three-dimensional structure. Thermal denaturation monitored by UV absorption and CD spectroscopy suggests proper folding of recombinant NOR-1 LBD.
Assuntos
Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/isolamento & purificação , Receptores Citoplasmáticos e Nucleares/química , Receptores Citoplasmáticos e Nucleares/isolamento & purificação , Receptores de Esteroides/química , Receptores de Esteroides/isolamento & purificação , Fatores de Transcrição/química , Fatores de Transcrição/isolamento & purificação , Sequência de Aminoácidos , Animais , Sítios de Ligação , Dicroísmo Circular , Proteínas de Ligação a DNA/biossíntese , Eletroforese em Gel de Poliacrilamida , Escherichia coli/metabolismo , Vetores Genéticos , Ligantes , Espectrometria de Massas , Dados de Sequência Molecular , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares , Ligação Proteica , Conformação Proteica , Dobramento de Proteína , Estrutura Terciária de Proteína , Ratos , Receptores Citoplasmáticos e Nucleares/biossíntese , Receptores de Esteroides/biossíntese , Proteínas Recombinantes/química , Análise de Sequência de Proteína , Fatores de Tempo , Fatores de Transcrição/biossíntese , Raios UltravioletaRESUMO
OBJECTIVE: To evaluate the expression of steroid hormone receptors, proteins related to apoptosis, and cell proliferation in endometria from women with polycystic ovary syndrome (PCOS). DESIGN: Case-control study. SETTING: Hospital research unit. PATIENT(S): Eight women with PCOS and 12 fertile healthy women of similar age to those with PCOS. INTERVENTION(S): Endometrial samples were obtained from women with PCOS (PCOSE) and normal (NE) women during the proliferative phase of the menstrual cycle. MAIN OUTCOME MEASURE(S): Expression studies (immunohistochemistry and reverse transcription-polymerase chain reaction) and DNA fragmentation [TdT-mediated dUTP nick end labeling (TUNEL)]. RESULT(S): In stroma, protein expression of estrogen receptor alpha, Bcl-2, and Bax was higher in PCOSE than in NE; epithelial cells had a greater expression of androgen receptor in the nucleus and a lower expression in the cytoplasm of PCOSE. Cell proliferation was higher in the epithelia of NE, while the expression of caspase-3 and DNA fragmentation was similar in both groups. The bax mRNA expression was higher in PCOSE, and bcl-2 mRNA expression was similar between groups. A higher bcl-2/bax relative ratio in PCOSE was observed. CONCLUSION(S): An alternating expression of proteins related to cell survival in endometria from PCOSE may potentially be associated with the disruption of their endometrial cell cycle.
Assuntos
Apoptose/fisiologia , Endométrio/metabolismo , Regulação da Expressão Gênica/fisiologia , Síndrome do Ovário Policístico/metabolismo , Receptores de Esteroides/biossíntese , Adulto , Estudos de Casos e Controles , Caspase 3 , Caspases/biossíntese , Caspases/genética , Endométrio/patologia , Receptor alfa de Estrogênio , Receptor beta de Estrogênio , Feminino , Humanos , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Antígeno Ki-67/biossíntese , Antígeno Ki-67/genética , Síndrome do Ovário Policístico/patologia , Proteínas Proto-Oncogênicas/biossíntese , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Proteínas Proto-Oncogênicas c-bcl-2/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Receptores Androgênicos/biossíntese , Receptores Androgênicos/genética , Receptores de Estrogênio/biossíntese , Receptores de Estrogênio/genética , Receptores de Progesterona/biossíntese , Receptores de Progesterona/genética , Receptores de Esteroides/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteína X Associada a bcl-2RESUMO
Os autores fazem uma revisäo sobre a influência que o EGF e seu receptor (EGF-R) exercem no tecido endometrial, enfocando sua possível regulaçäo pelos esteróides e seus receptores e sua importância nos processos de transformaçäo secretora e de decidualizaçäo.