Cloprostenol administration in the first week postpartum reduces expression of oxytocin receptors in the endometrium in Holstein-Zebu cows / Administração de cloprostenol na primeira semana pós-parto pode reduzir a expressão de receptores de ocitocina no endométrio de vacas mestiças Holandês-Zebu

The present study investigated the hormonal profile and expression of prostaglandin F2α (PGF2α), oxytocin and estrogen receptors in uterine tissues of postpartum cows treated with cloprostenol. Twenty Holstein-Zebu crossbred cows were treated with saline solution (treatment CONT) or cloprostenol (treatment CLO), both administered two and five days postpartum. Blood samples were collected on days two, seven, 14, 21 and 28 postpartum for progesterone, PGF2α metabolite (PGFM) and estradiol determination, and endometrial biopsy was performed in order to quantify the expression of oxytocin receptor (OXTR), prostaglandin F receptor (PTGFR) and estrogen receptor 1 (ERS1) genes. In the CLO treatment, expression of OXTR was reduced (P<0.05) but no difference (P>0.05) between treatments was found for PTGFR and ERS1 expression. Estrogen concentrations increased progressively until day 14 (P<0.05) and the highest OXTR expression and lowest PTGFR expression were observed on day 14 (P<0.05) in both treatments. Serum PGFM concentrations were high throughout the experiment. In conclusion, cloprostenol administration at days two and five of postpartum seems to reduce OXTR expression in the endometrium in crossbred cows.(AU)

O presente estudo avaliou o perfil hormonal e a expressão gênica de receptores de prostaglandina F2α (PGF2α), ocitocina e estrógeno no endométrio de vacas pós-parto tratadas com cloprostenol. Vinte vacas mestiças Holandês-Zebu foram tratadas com solução salina (tratamento CONT, n = 10) ou cloprostenol (tratamento CLO, n = 10), ambos administrados dois e cinco dias após o parto. Amostras de sangue foram coletadas nos dias dois, sete, 14, 21 e 28 pós-parto para mensuração de progesterona, de metabólito de PGF2α (PGFM) e de estradiol, e foram obtidas biópsias endometriais para quantificar a expressão de PTGFR, OXTR e ESR1. No tratamento CLO, a expressão gênica de receptores de ocitocina foi menor (P<0,05). As concentrações de estrógeno aumentaram progressivamente até o dia 14 (P<0,05). A maior expressão de OXTR foi observada no dia 14 (P<0,05). A expressão de ESR1 foi semelhante entre os tratamentos (P>0,05). Os níveis de PGFM foram altos durante todo o estudo. Conclui-se que a administração de cloprostenol nos dias dois e cinco pós-parto parece diminuir a expressão de OXTR no endométrio em vacas mestiças.(AU)

Cloprostenol administration in the first week postpartum reduces expression of oxytocin receptors in the endometrium in Holstein-Zebu cows / Administração de cloprostenol na primeira semana pós-parto pode reduzir a expressão de receptores de ocitocina no endométrio de vacas mestiças Holandês-Zebu

The present study investigated the hormonal profile and expression of prostaglandin F2α (PGF2α), oxytocin and estrogen receptors in uterine tissues of postpartum cows treated with cloprostenol. Twenty Holstein-Zebu crossbred cows were treated with saline solution (treatment CONT) or cloprostenol (treatment CLO), both administered two and five days postpartum. Blood samples were collected on days two, seven, 14, 21 and 28 postpartum for progesterone, PGF2α metabolite (PGFM) and estradiol determination, and endometrial biopsy was performed in order to quantify the expression of oxytocin receptor (OXTR), prostaglandin F receptor (PTGFR) and estrogen receptor 1 (ERS1) genes. In the CLO treatment, expression of OXTR was reduced (P<0.05) but no difference (P>0.05) between treatments was found for PTGFR and ERS1 expression. Estrogen concentrations increased progressively until day 14 (P<0.05) and the highest OXTR expression and lowest PTGFR expression were observed on day 14 (P<0.05) in both treatments. Serum PGFM concentrations were high throughout the experiment. In conclusion, cloprostenol administration at days two and five of postpartum seems to reduce OXTR expression in the endometrium in crossbred cows.(AU)

O presente estudo avaliou o perfil hormonal e a expressão gênica de receptores de prostaglandina F2α (PGF2α), ocitocina e estrógeno no endométrio de vacas pós-parto tratadas com cloprostenol. Vinte vacas mestiças Holandês-Zebu foram tratadas com solução salina (tratamento CONT, n = 10) ou cloprostenol (tratamento CLO, n = 10), ambos administrados dois e cinco dias após o parto. Amostras de sangue foram coletadas nos dias dois, sete, 14, 21 e 28 pós-parto para mensuração de progesterona, de metabólito de PGF2α (PGFM) e de estradiol, e foram obtidas biópsias endometriais para quantificar a expressão de PTGFR, OXTR e ESR1. No tratamento CLO, a expressão gênica de receptores de ocitocina foi menor (P<0,05). As concentrações de estrógeno aumentaram progressivamente até o dia 14 (P<0,05). A maior expressão de OXTR foi observada no dia 14 (P<0,05). A expressão de ESR1 foi semelhante entre os tratamentos (P>0,05). Os níveis de PGFM foram altos durante todo o estudo. Conclui-se que a administração de cloprostenol nos dias dois e cinco pós-parto parece diminuir a expressão de OXTR no endométrio em vacas mestiças.(AU)

Role of estrogen and progesterone receptors in neonatal uterine cell proliferation in the mouse

The major endocrine regulators of the female reproductive tract are 17β-estradiol (E2) and progesterone (P4). This review discusses our recent work related to the roles of E2 and P4 and their receptors, estrogen receptor 1 (ESR1) and progesterone receptor (PR), respectively, in the neonatal uterus. Neonatal uterine cells in mice are mitogenically responsive to estrogens, but neonatal ovariectomy does not inhibit pre-weaning uterine cell proliferation, indicating that this process does not require endogenous estrogens. Neonatal terine cell proliferation could result from ligand-independent growth factor activation of ESR1, or be independent of ESR1 neonatally despite its obligatory role in adult uterine epithelial proliferation. To determine the role of ESR1 in uterine development, we analyzed cell proliferation and uterine gland development (adenogenesis) in wild-type (WT) and Esr1 knockout (Esr1KO) mice postnatally. Our results indicate that pre-weaning uterine cell proliferation and adenogenesis are independent of ESR1, but these processes become dependent on E2/ESR1 signaling for maintenance and further proliferation and uterine growth during puberty. How pre-weaning uterine cell proliferation and adenogenesis occur independently of E2/ESR1 signaling remains unknown, but ligand-independent activation of ESR1 is not involved in this process. The synthetic glucocorticoid dexamethasone (Dex) inhibits luminal epithelial (LE) proliferation in neonatal mouse uteri, but it has been unclear whether Dex effects were mediated by glucocorticoid receptor (GR) and/or PR. We have used PR knockout (PRKO) mice to test whether PR is required for Dex inhibition of LE proliferation. Our results indicate that maximal inhibitory Dex effects on uterine LE proliferation require PR, possibly reflecting Dex crosstalk with PR.

Role of estrogen and progesterone receptors in neonatal uterine cell proliferation in the mouse

The major endocrine regulators of the female reproductive tract are 17β-estradiol (E2) and progesterone (P4). This review discusses our recent work related to the roles of E2 and P4 and their receptors, estrogen receptor 1 (ESR1) and progesterone receptor (PR), respectively, in the neonatal uterus. Neonatal uterine cells in mice are mitogenically responsive to estrogens, but neonatal ovariectomy does not inhibit pre-weaning uterine cell proliferation, indicating that this process does not require endogenous estrogens. Neonatal terine cell proliferation could result from ligand-independent growth factor activation of ESR1, or be independent of ESR1 neonatally despite its obligatory role in adult uterine epithelial proliferation. To determine the role of ESR1 in uterine development, we analyzed cell proliferation and uterine gland development (adenogenesis) in wild-type (WT) and Esr1 knockout (Esr1KO) mice postnatally. Our results indicate that pre-weaning uterine cell proliferation and adenogenesis are independent of ESR1, but these processes become dependent on E2/ESR1 signaling for maintenance and further proliferation and uterine growth during puberty. How pre-weaning uterine cell proliferation and adenogenesis occur independently of E2/ESR1 signaling remains unknown, but ligand-independent activation of ESR1 is not involved in this process. The synthetic glucocorticoid dexamethasone (Dex) inhibits luminal epithelial (LE) proliferation in neonatal mouse uteri, but it has been unclear whether Dex effects were mediated by glucocorticoid receptor (GR) and/or PR. We have used PR knockout (PRKO) mice to test whether PR is required for Dex inhibition of LE proliferation. Our results indicate that maximal inhibitory Dex effects on uterine LE proliferation require PR, possibly reflecting Dex crosstalk with PR. (AU)

Mecanismo de ação do 17-estradiol no corpo lúteo de cadelas não prenhes

O corpo lúteo (CL) canino é responsável pela síntese de E2 durante o diestro. O E2 atua de forma autócrina e/ou parácrina sobre essa glândula. O mecanismo de atuação do E2 depende da razão entre receptor alfa (ER) e receptor beta (ER). A ligação ao ER tem efeito proliferativo e ao ER antiproliferativo. O objetivo deste trabalho foi entender a sinalização mediada pelo ER e ER no processo de formação e regressão do CL. CLs foram obtidos de cadelas não prenhes (n=30) nos dias 10, 20, 30, 40, 50 e 60 (n=5/grupo) após a ovulação (po). No dia da ovariossalpingohisterectomia foi colhido sangue para mensuração das concentrações de P4 e E2. Dezoito CLs (n=3/grupo) foram submetidos ao sequenciamento de RNA (RNAseq). Os genes diferencialmente expressos (DE) identificados pelo RNAseq foram submetidos ao software oPOSSUM3 para identificação das regiões de ligação mais representadas nos fatores de transcrição relacionados aos genes do ER (ESR1) e ER (ESR2). A análise de expressão temporal revelou a presença de 5.116 diferencialmente expressos (DE) em pelo menos uma comparação e 1.106 não foram anotados ainda no genoma canino, destes genes DE 295 apresentavam regiões de ligação de fatores de transcrição em comum com ESR1 e ESR2. Dez genes DE foram selecionados para validação dos resultados de RNAseq através do qPCR; usando o GAPDH como gene de referência. Desses genes, quatro apresentaram regiões em comum com ESR2 (LEF-1; PAPPA; NDGR2 ATP1A1) e um com ESR1 (CAV1) e os demais controlam proliferação celular (CTNNB1; CCND1; IGFBP 3, 4 e 5). A expressão proteíca de PAPPA e IGFBP 3, 4 e 5 (componentes do sistema IGF) foi avaliada também por imunohistoquímica. Durante a primeira metade do diestro, nossos resultados indicam que a sinalização mediada pelo E2 ocorre via interação do ER com CAV-1 (sinalização não genômica), ativando as vias de sinalização IGF e WNT/catenina, regulando o processo de proliferação celular. Enquanto na segunda metade, o ER regularia a expressão gênica de NDGR2 e ATP1A1, contribuindo para a regressão do CL. Assim nos resultados sugerem que o E2 atue tanto como um fator luteotrófico e quanto regulador da regressão do CL canino.

The canine corpus luteum (CL) is responsible for E2 synthesis during diestrus, which acts in an autocrine and / or paracrine manner in this gland. The E2 mechanism of action depends on the ratio between alpha (ER) and beta (ER) receptor The binding to ER has a proliferative effect whereas to ER an antiproliferative. The objective of this study was to understand the signaling mediated by ER and ER in the formation and regression of the CL. CLs were obtained from non-pregnant bitches (n = 30) on days 10, 20, 30, 40, 50 and 60 (n = 5 / group) post-ovulation (po). On the day of ovariohysterectomy blood was collected for measurement of P4 and E2 concentrations. Eighteen CLs (n = 3 / group) were subjected to RNA sequencing (RNA-Seq). Genes differentially expressed (DE) identified by RNA-Seq were submitted to oPOSSUM3 software for detection of over-represented transcription factors binding sites (TFBS) related to the ER (ESR1) and ER (ESR2) genes. The temporal expression analysis revealed the presence of 5116 differentially expressed (DE) genes in at least one comparison, and 1106 genes, which have not been recorded to the canine genome yet From these, 295 genes showed TFBS in common with ESR1 and ESR2. Ten DE genes were selected to validate the results of RNA-Seq by qPCR; using GAPDH as reference gene. Of these genes, four had TFBS in common with ESR2 (LEF-1; PAPPA; NDGR2; ATP1A1) and one with ESR1 (CAV1) and others genes were chosen because they control cell proliferation (CTNNB1; CCND1, IGFBP 3, 4 and 5). Protein expression of the IGF related genes (PAPPA and IGFBP 3, 4 and 5) was also evaluated by immunohistochemistry. During the first half of diestrus, it appears that E2 mediated signaling occurs via interaction of ER with CAV-1 (non-genomic signaling), activating signaling pathways of IGF and WNT / catenin, then regulating the cell proliferation process. Whereas in the second half, ER appears to regulate NDGR2 and ATP1A1 gene expression, contributing to the regression of the CL. Thus our results suggest that E2 may act as a luteotrophic and also a luteolytic factor in the canine CL.