Resumo
O conhecimento do processo de luteólise dentro da fisiologia reprodutiva se torna importante para que ocorra a compreensão do reconhecimento materno da gestação e seja possível formular estratégias anti-luteolíticas que visem manter esta fase conferindo aumentos na eficiência reprodutiva e produtiva. Durante o processo de luteólise, o tecido luteal sofre mudanças bruscas na capacidade esteroidogênica, vascularização e remodelamento, resultando em substituição da glândula por tecido conjuntivo, sendo a PGF2α o principal determinante na indução espontânea da luteólise em bovinos. Simultaneamente, ocorre um íntimo processo entre as relações anatômicas que compreende o ovário, útero e suas vascularizações. A exposição do útero à interação coordenada entre P4, E2 e ocitocina aliada à expressão de seus receptores são essenciais para a secreção pulsátil de PGF2α pelo endométrio a qual induzirá a luteólise. Nesta regressão do corpo lúteo (CL), estão envolvidos ainda os hormônios (LH, prolactina, cortisol) e várias substâncias vasoativas (óxido nítrico, VEGF, EG-VEGF, bFGF, END-1, AngII). A concentração de PGF2α pode ser representada pela mensuração sanguínea da concentração de seu metabólito 13,14-dihidro-15-ceto- PGF2α (PGFM) sendo a pulsatilidade da secreção de PGF2α durante os períodos pré-luteolítico, luteolítico e pós-luteolítico mensurável pelo aumento na concentração de PGFM. Neste artigo de revisão, será possível compreender e discutir os principais aspectos endócrinos e moleculares que ocorrem com as mudanças morfofuncionais no CL durante o processo de luteólise fisiológica em fêmeas bovinas.
The knowledge of the luteolysis process within reproductive physiology becomes important to occur the understanding of maternal recognition of gestation being possible to formulate anti-luteolytic strategies aimed at maintaining this phase conferring increases in reproductive and productive efficiency. During the luteolysis process, luteal tissue undergoes abrupt changes in steroidogenic capacity, vascularization and remodeling, resulting in replacement of the gland by connective tissue, and PGF2α is the main determinant in the spontaneous induction of luteolysis in cattle. Simultaneously, an intimate process occurs between the anatomical relationships that comprise the ovary, uterus and its vascularizations. The exposure of the uterus to the coordinated interaction between P4, E2 and oxytocin, together with the expression of its receptors, are essential for the pulsating secretion of PGF2α by the endometrium, which will induce luteolysis. In this regression of the corpus luteum (CL), hormones (LH, prolactin, cortisol) and various vasoactive substances (nitric oxide, VEGF, EG-VEGF, bFGF, END-1, AngII) are also involved. The concentration of PGF2α can be represented by the blood measurement of the concentration of its metabolite 13,14-dihydro-15-keto-PGF2α (PGFM),being the pulsatility of PGF2α secretion during the pre-luteolytic, luteolytic and post-luteolytic measured by the increased concentration of PGFM. In this review article, it will be possible to understand and discuss the main endocrine and molecular aspects that occur with the morphofunctional changes in CL during the physiological luteolysis process in bovine females.
Assuntos
Feminino , Animais , Bovinos , Corpo Lúteo/fisiologia , Dinoprosta/análise , Luteólise/fisiologia , Fenômenos Reprodutivos FisiológicosResumo
O conhecimento do processo de luteólise dentro da fisiologia reprodutiva se torna importante para que ocorra a compreensão do reconhecimento materno da gestação e seja possível formular estratégias anti-luteolíticas que visem manter esta fase conferindo aumentos na eficiência reprodutiva e produtiva. Durante o processo de luteólise, o tecido luteal sofre mudanças bruscas na capacidade esteroidogênica, vascularização e remodelamento, resultando em substituição da glândula por tecido conjuntivo, sendo a PGF2α o principal determinante na indução espontânea da luteólise em bovinos. Simultaneamente, ocorre um íntimo processo entre as relações anatômicas que compreende o ovário, útero e suas vascularizações. A exposição do útero à interação coordenada entre P4, E2 e ocitocina aliada à expressão de seus receptores são essenciais para a secreção pulsátil de PGF2α pelo endométrio a qual induzirá a luteólise. Nesta regressão do corpo lúteo (CL), estão envolvidos ainda os hormônios (LH, prolactina, cortisol) e várias substâncias vasoativas (óxido nítrico, VEGF, EG-VEGF, bFGF, END-1, AngII). A concentração de PGF2α pode ser representada pela mensuração sanguínea da concentração de seu metabólito 13,14-dihidro-15-ceto- PGF2α (PGFM) sendo a pulsatilidade da secreção de PGF2α durante os períodos pré-luteolítico, luteolítico e pós-luteolítico mensurável pelo aumento na concentração de PGFM. Neste artigo de revisão, será possível compreender e discutir os principais aspectos endócrinos e moleculares que ocorrem com as mudanças morfofuncionais no CL durante o processo de luteólise fisiológica em fêmeas bovinas.(AU)
The knowledge of the luteolysis process within reproductive physiology becomes important to occur the understanding of maternal recognition of gestation being possible to formulate anti-luteolytic strategies aimed at maintaining this phase conferring increases in reproductive and productive efficiency. During the luteolysis process, luteal tissue undergoes abrupt changes in steroidogenic capacity, vascularization and remodeling, resulting in replacement of the gland by connective tissue, and PGF2α is the main determinant in the spontaneous induction of luteolysis in cattle. Simultaneously, an intimate process occurs between the anatomical relationships that comprise the ovary, uterus and its vascularizations. The exposure of the uterus to the coordinated interaction between P4, E2 and oxytocin, together with the expression of its receptors, are essential for the pulsating secretion of PGF2α by the endometrium, which will induce luteolysis. In this regression of the corpus luteum (CL), hormones (LH, prolactin, cortisol) and various vasoactive substances (nitric oxide, VEGF, EG-VEGF, bFGF, END-1, AngII) are also involved. The concentration of PGF2α can be represented by the blood measurement of the concentration of its metabolite 13,14-dihydro-15-keto-PGF2α (PGFM),being the pulsatility of PGF2α secretion during the pre-luteolytic, luteolytic and post-luteolytic measured by the increased concentration of PGFM. In this review article, it will be possible to understand and discuss the main endocrine and molecular aspects that occur with the morphofunctional changes in CL during the physiological luteolysis process in bovine females.(AU)
Assuntos
Animais , Feminino , Bovinos , Corpo Lúteo/fisiologia , Luteólise/fisiologia , Dinoprosta/análise , Fenômenos Reprodutivos FisiológicosResumo
In vitro embryo production has the potential to produce more offspring from genetically valuable animals than standard MO ET as it is capable of avoiding most of the causes of failure in MOET (poor response to superovulation, poor fertilization and premature luteolysis). It also allows repeating collection in the donor animals more often and more times in their reproductive life. However, consistent results are not easy to obtain when conducting large scale programs, mainly due to variability associated with the in vitro fertilization results. Cloning by somatic cell nuclear transfer has been used by a few groups to successfully produce genetic copies of individuals of high genetic merit, but remains to be a very inefficient reproduction technology even in the hand s of those that have been successful in producing multiple live clones. This review provides a collection of tips and points to consider when planning in vitro embryo production and cloning programs in sheep and goats.ien
Assuntos
Animais , Fertilização , Genética/tendências , Luteólise/fisiologia , Técnicas de Cultura Embrionária/instrumentação , Cabras/classificação , Clonagem de Organismos , Ovinos/classificaçãoResumo
In vitro embryo production has the potential to produce more offspring from genetically valuable animals than standard MO ET as it is capable of avoiding most of the causes of failure in MOET (poor response to superovulation, poor fertilization and premature luteolysis). It also allows repeating collection in the donor animals more often and more times in their reproductive life. However, consistent results are not easy to obtain when conducting large scale programs, mainly due to variability associated with the in vitro fertilization results. Cloning by somatic cell nuclear transfer has been used by a few groups to successfully produce genetic copies of individuals of high genetic merit, but remains to be a very inefficient reproduction technology even in the hand s of those that have been successful in producing multiple live clones. This review provides a collection of tips and points to consider when planning in vitro embryo production and cloning programs in sheep and goats.(AU)ien
Assuntos
Animais , Genética/tendências , Técnicas de Cultura Embrionária/instrumentação , Fertilização , Luteólise/fisiologia , Ovinos/classificação , Cabras/classificação , Clonagem de OrganismosResumo
Foi avaliado o efeito da aplicação de um luteolítico em dois momentos distintos sobre a taxa de concepção na inseminação artificial em tempo fixo (IATF) de vacas com cria ao pé, além de verificar a eficácia de implantes com 1 grama de P4 de primeiro e segundo uso. Foram utilizadas 210 vacas de corte lactantes da raça Montana divididas em dois lotes de 105 vacas, comparando oito grupos distribuídos nos lotes. O lote D6,5 foi tratado no dia zero com 2mg de benzoato de estradiol i.m. (BE, Estrogin®) juntamente com implante intravaginal de progesterona de 1º uso (G1- Sincrogest®, n=29; G2- Primer®, n=26) e de 2º uso (G3- Sincrogest®, n=26; G4- Primer®, n=24), no dia 6,5 foi aplicado 150mcg i.m. de Cloprostenol Sódico (Sincrocio®), sendo a retirada do implante de P4 no dia 8. Dia 9, ocorreu a aplicação de 1mg de BE i.m. e no dia 10 a tarde a IATF. O lote D8 iniciou o protocolo de IATF e foi tratado da mesma forma, à exceção da aplicação do luteolítico que ocorreu no oitavo dia. Os grupos foram formados: implante de 1º uso (G5- Sincrogest®, n=27; G6- Primer®, n=28) e de 2º uso (G7-Sincrogest®, n=26; G8- Primer®, n=24). As taxas de prenhez dos grupos do lote D6,5 foram de 55,17%(G1), 69,23%(G2), 57,67%(G3), 70,83% (G4) (p=0,263). No lote D8 os grupos G5, G6, G7, G8 deste lote apresentaram 44,44%, 67,86%, 46,15%, e 58,33% de prenhez (p=0,573). As taxas de prenhez para os lotes D6,5 e D8 foram, de 62,86% e 54,29%. Não houve diferença nas taxas de concepção a IATF entre os lotes (p= 0,262). Os implantes de primeiro e segundo uso Sincrogest® e Primer® foram efetivos para obter-se uma taxa de concepção em torno de 50% em vacas de corte lactantes. A antecipação da aplicação do Cloprostenol Sódico (D6,5) não foi efetiva para aumentar as taxas de concepção a IATF e prenhez ao final da estação de monta. A idade, condição corporal e os dias pós-parto não influenciaram na taxa de concepção a IATF e taxa de prenhez ao final da estação de monta.(AU)
The effect of luteolytic application was evaluated in two distinct moments about the range of pregnancy in the artificial insemination in fixed time of post partum cows, besides checking the efficacy of implants with 1 gram of progesterone of first and second use. 210 post partum cows of Montana breed were used divided into two different lots of 105 cows, comparing 8 groups. The lot D6,5 was treated on the day zero with 2mg of estradiol benzoate i.m. (BE, Estrogin®) together with the intravaginal implant of progesterone of first use (G1- Sincrogest®, n=29; G2- Primer®, n=26) and of second use (G3- Sincrogest®, n=26; G4- Primer®, n=24), on the day 6,5 150mcg i.m of Cloprostenol (Sincrocio®), the removal of the implant of P4 on the day 8. On the day 9, there was an application 1mg of BE i.m and on the day 10 in the afternoon the TAI. The lot D8 was treated on the same way, except for the application of luteolytic, that occurred on the eighth day. The groups were formed this way: first use (G5- Sincrogest®, n=27; G6- Primer®, n=28) and second use (G7-Sincrogest®, n=26; G8- Primer®, n=24). The ranges of pregnancy of the groups in the lot D6,5 were 55,17%(G1), 69,23%(G2), 57,67%(G3), 70,83%(G4) (p=0,263). In the lot D8 the groups G1, G2, G3, G4 of this lot presented 44,44%, 67,86%, 46,15%, e 58,33% of pregnancy (p=0,573). The ranges of pregnancy for the lots D6,5 and D8 were 62,86% e 54,29%. There was no difference in the ranges of pregnancy TAI between the lots (p= 0,262). The implants of first and second use Sincrogest ® and Primer ® were effective to obtain a pregnancy rate around 50% in lactating beef cows. The anticipation of applying Cloprostenol Sodic (D6, 5) was not effective to increase the conception rate to TAI and pregnancy rate at the end of the breeding season. The age, body condition and days postpartum did not influence the conception rate to TAI and pregnancy rate at the end of the breeding season.(AU)
Assuntos
Animais , Feminino , Progesterona/análise , Bovinos/embriologia , Bovinos/fisiologia , Inseminação Artificial/métodos , Luteólise/fisiologia , Período Pós-PartoResumo
The effect of prostaglandin F2α (PGF) infusion for 3 h into the jugular vein on progesterone concentrations was studied in 24 Holstein heifers.Plasma concentrations of PGF were assessed by assay of 13,14-dihydro-15-keto-PGF (PGFM). The 3 h of PGF infusion were used to approximate the duration of the major concentrations of PGFM in a natural pulse. During infusion of 5, 10, or 15 mg/3 h, the concentrations of PGFM greatly exceeded the peak of a natural pulse. Plasma concentrations of progesterone decreased (P < 0.05) in the three PGF-treated groups by Hour 1 (Hour 0 = beginning of infusion). Progesterone increased between Hours 1 and 2, but the increase was significant (P < 0.03) only in the 5-mg group. Concentrations decreased more gradually between Hours 2 and 6 than between Hours 0 and 1 with no differences among PGF groups. The percentage of CL area with color-Doppler signals of blood flow were elevated similarly in the three PGF-treated groups at Hours 1 to 3 and by Hour 5 decreased to below the percentage at Hour 0. In a second experiment, approximating a natural PGFM pulse by intravenous infusion of PGF at a dose of 0.7 mg/3 h did not affect plasma progesterone concentrations. Results indicatedthat intravenous infusion of PGF for 3 h decreased theprogesterone concentration when the total dose was equivalent to doses that have been shown to be completely luteolytic when given as a single systemic injection. However, intravenous infusion of a dose of PGF that approximately simulated a natural PGFM pulse did not effect progesterone concentration.(AU)
Assuntos
Animais , Veias Jugulares/anatomia & histologia , Luteólise/fisiologia , Corpo Lúteo/anatomia & histologia , Gado/classificação , Prostaglandinas/análiseResumo
The effect of prostaglandin F2α (PGF) infusion for 3 h into the jugular vein on progesterone concentrations was studied in 24 Holstein heifers.Plasma concentrations of PGF were assessed by assay of 13,14-dihydro-15-keto-PGF (PGFM). The 3 h of PGF infusion were used to approximate the duration of the major concentrations of PGFM in a natural pulse. During infusion of 5, 10, or 15 mg/3 h, the concentrations of PGFM greatly exceeded the peak of a natural pulse. Plasma concentrations of progesterone decreased (P < 0.05) in the three PGF-treated groups by Hour 1 (Hour 0 = beginning of infusion). Progesterone increased between Hours 1 and 2, but the increase was significant (P < 0.03) only in the 5-mg group. Concentrations decreased more gradually between Hours 2 and 6 than between Hours 0 and 1 with no differences among PGF groups. The percentage of CL area with color-Doppler signals of blood flow were elevated similarly in the three PGF-treated groups at Hours 1 to 3 and by Hour 5 decreased to below the percentage at Hour 0. In a second experiment, approximating a natural PGFM pulse by intravenous infusion of PGF at a dose of 0.7 mg/3 h did not affect plasma progesterone concentrations. Results indicatedthat intravenous infusion of PGF for 3 h decreased theprogesterone concentration when the total dose was equivalent to doses that have been shown to be completely luteolytic when given as a single systemic injection. However, intravenous infusion of a dose of PGF that approximately simulated a natural PGFM pulse did not effect progesterone concentration.
Assuntos
Animais , Corpo Lúteo/anatomia & histologia , Luteólise/fisiologia , Veias Jugulares/anatomia & histologia , Gado/classificação , Prostaglandinas/análiseResumo
The corpus luteum (CL) undergoes drastic changes in its function and structure during the estrous cycle. To secrete a sufficient amount of progesterone (P4) to ensure the occurrence of pregnancy in a cow with a body weight greater than 500 kg, the bovine CL weighs 5-8 g which is 2-3 thousand times heavier than rat CL. If pregnancy does not occur successfully, rapid luteolysis is caused by prostaglandin F2α (PGF2α) that is released from the endometrium around days 17-19 of the estrous cycle in the cow. Thus, it is clear that the bovine CL lifespan is controlled by well-coordinated mechanisms. As the CL matures, the steroidogenic cells establish contact with many capillary vessels, so that the CL is composed of a large number of vascular endothelial cells that can account for up to 50% of all cells in the bovine CL. Also, luteal endothelial cells secrete several vasoactive substances such as PGF2α, nitric oxide, endothelin-1 and angiotensin II that regulate blood flow as well as P4 secretion in an autocrine/paracrine manner within the CL. Therefore, blood vessels and endothelial cells within the CL have an essential role in luteal function in the cow, suggesting that the study of vasoactive molecules from the CL is of great importance to give an insight into systems which regulate luteolysis locally. In the present review, we describe novel concepts on the luteolytic mechanisms in the cow, with emphasis on luteal blood flow and vasoactive molecules.
Assuntos
Feminino , Animais , Bovinos , Ciclo Estral/fisiologia , Corpo Lúteo/crescimento & desenvolvimento , Luteólise/fisiologia , Dinoprosta/efeitos adversos , Endotelina-1/efeitos adversosResumo
The corpus luteum (CL) undergoes drastic changes in its function and structure during the estrous cycle. To secrete a sufficient amount of progesterone (P4) to ensure the occurrence of pregnancy in a cow with a body weight greater than 500 kg, the bovine CL weighs 5-8 g which is 2-3 thousand times heavier than rat CL. If pregnancy does not occur successfully, rapid luteolysis is caused by prostaglandin F2α (PGF2α) that is released from the endometrium around days 17-19 of the estrous cycle in the cow. Thus, it is clear that the bovine CL lifespan is controlled by well-coordinated mechanisms. As the CL matures, the steroidogenic cells establish contact with many capillary vessels, so that the CL is composed of a large number of vascular endothelial cells that can account for up to 50% of all cells in the bovine CL. Also, luteal endothelial cells secrete several vasoactive substances such as PGF2α, nitric oxide, endothelin-1 and angiotensin II that regulate blood flow as well as P4 secretion in an autocrine/paracrine manner within the CL. Therefore, blood vessels and endothelial cells within the CL have an essential role in luteal function in the cow, suggesting that the study of vasoactive molecules from the CL is of great importance to give an insight into systems which regulate luteolysis locally. In the present review, we describe novel concepts on the luteolytic mechanisms in the cow, with emphasis on luteal blood flow and vasoactive molecules.(AU)
Assuntos
Animais , Feminino , Bovinos , Luteólise/fisiologia , Corpo Lúteo/crescimento & desenvolvimento , Ciclo Estral/fisiologia , Dinoprosta/efeitos adversos , Endotelina-1/efeitos adversosResumo
The luteinizing hormone (LH) receptor is fundamental for the regulation of the corpus luteum (CL) in women and non-human primates. Its ligands, LH and human chorionic gonadotropin (hCG), have key roles in the regulation of tissue and vascular remodeling associated with luteal formation and regression. However this remodeling involves the regulation of cells that do not express LH receptors including endothelial cells, pericytes, fibroblasts and macrophages. We have taken a candidate molecule approach to identify important LH/hCG-regulated paracrine molecules and their receptors in CL and assess the effects of their manipulation in vivo and in vitro. Vascular endothelial growth factor (VEGF) acts on endothelial cells and is a major paracrine regulator of luteal angiogenesis and vasculature maintenance. Luteolysis is associated with increased SLIT/ROBO, connective tissue growth factor (CTGF) and matrix metalloproteinase (MMP) expression in luteal fibroblasts. Investigation of the inhibition of these changes by hCG has identified activin A as a novel paracrine luteolysin and locally generated cortisol as a novel paracrine luteotropin. The molecular regulation of luteal function in the primate is complex and the paracrine regulation of luteal function is still not fully understood. Locally, the luteolytic activities of SLIT/ROBO and activin-A are inhibited by hCG and the luteotropic activities of VEGF and cortisol are stimulated by hCG.
Assuntos
Feminino , Animais , Fator A de Crescimento do Endotélio Vascular/fisiologia , Gonadotropina Coriônica/efeitos adversos , Gravidez/fisiologia , Luteinização/fisiologia , Receptores do LH/metabolismo , Hormônio Luteinizante/fisiologia , Luteólise/fisiologia , Neovascularização Fisiológica/fisiologiaResumo
The luteinizing hormone (LH) receptor is fundamental for the regulation of the corpus luteum (CL) in women and non-human primates. Its ligands, LH and human chorionic gonadotropin (hCG), have key roles in the regulation of tissue and vascular remodeling associated with luteal formation and regression. However this remodeling involves the regulation of cells that do not express LH receptors including endothelial cells, pericytes, fibroblasts and macrophages. We have taken a candidate molecule approach to identify important LH/hCG-regulated paracrine molecules and their receptors in CL and assess the effects of their manipulation in vivo and in vitro. Vascular endothelial growth factor (VEGF) acts on endothelial cells and is a major paracrine regulator of luteal angiogenesis and vasculature maintenance. Luteolysis is associated with increased SLIT/ROBO, connective tissue growth factor (CTGF) and matrix metalloproteinase (MMP) expression in luteal fibroblasts. Investigation of the inhibition of these changes by hCG has identified activin A as a novel paracrine luteolysin and locally generated cortisol as a novel paracrine luteotropin. The molecular regulation of luteal function in the primate is complex and the paracrine regulation of luteal function is still not fully understood. Locally, the luteolytic activities of SLIT/ROBO and activin-A are inhibited by hCG and the luteotropic activities of VEGF and cortisol are stimulated by hCG.(AU)
Assuntos
Animais , Feminino , Receptores do LH/metabolismo , Luteinização/fisiologia , Gonadotropina Coriônica/efeitos adversos , Fator A de Crescimento do Endotélio Vascular/fisiologia , Gravidez/fisiologia , Hormônio Luteinizante/fisiologia , Luteólise/fisiologia , Neovascularização Fisiológica/fisiologiaResumo
Durante o período crítico do reconhecimento materno, compreendidoentre o 15º e 19º dias da gestação, o concepto deve sintetizarcompetentemente moléculas capazes de bloquear a síntese deprostaglandina F2α (PGF2α) e a luteólise. Em bovinos, a principalmacromolécula protéica envolvida em tal bloqueio é o interferon-tau(IFN-τ). Durante o período crítico, falhas neste reconhecimentodeterminam à mortalidade embrionária em até 40% das fêmeasinseminadas. Informações sobre o IFN-τ em animais Bos taurus indicus,ainda são restritas. Este estudo objetivou uma avaliação quantitativado IFN-τ durante o período crítico do reconhecimento materno, emlavados uterinos obtidos por sonda de Foley (dias 14, 16 e 18 pósestro) ou post-mortem (dia 18 pós-estro). Para tanto, foram utilizadasfêmeas multíparas azebuadas (Bos taurus indicus), cíclicas ou prenhes,nos dias 14, 16 e 18 pós-estro. Para a obtenção dos lavados, os úterosforam infundidos com solução de Ringer Simples. Os lavados foramconcentrados por ultra-filtração e liofilizados. As macromoléculasprotéicas foram separadas por Eletroforese Unidimensional SDSPAGE, em gel com 15% de poliacrilamida. A quantificação doIFN-τ nos lavados uterinos foi realizada por Western-Blotting edensitometria. Tanto nos lavados obtidos por sonda de Foley quantonos post-mortem foi possível observar bandas de proteínas queapresentaram reação cruzada com os anticorpos utilizados no WesternBlotting. O IFN-τ foi detectado apenas nos lavados uterinospost-mortem de vacas prenhes (P<0,05). A densidade óptica não foiafetada pelo dia do período crítico, estado (cíclico ou prenhe) ouinteração dia x estado. Nos lavados post-mortem não houve efeito depeso do concepto ou concentração de progesterona plasmática no diado lavado na densidade da banda protéica referente ao IFN-τ .(AU)
During the critical period of the maternal recognition, which occursbetween days 15 and 19 of pregnancy, the conceptus must competentlysynthesize molecules capable of blocking the synthesis ofprostaglandin F2α (PGF2α) and luteolysis. In cattle, the majormacromolecule involved in suck blockage is the protein interferontau (IFN-τ). During the critical period, failures in the recognition ofpregnancy determine embryonic mortality on up to 40% ofinseminated cows. Data about IFN-τ in Bos taurus indicus are stillscarce. Objective of this study was to quantitatively evaluate the presenceof IFN-τ during the critical period for maternal recognition ofpregnancy in uterine flushings obtained in vivo by Foley catheter (Days14, 16 and 18 post estrus) or post-mortem (Day 18 post estrus).Multiparous, cyclic or pregnant zebu cows (Bos taurus indicus) on days14, 16 and 18 post estrus were used for in vivo or post mortem uterineflushing collection. In both cases, a Ringer solution was used to washthe uterus of cows. Uterine flushings were concentrated by ultrafiltration and lyophilized. Proteins were separated by one-dimensionalelectrophoresis (SDS-PAGE) in a 15% polyacrilamide gel. Interferontau quantification in uterine flushings was performed by westernblotting and densitometry. Non-specific protein bands were observedin both in vivo and post mortem uterine flushings. Interferon-tau wasdetected only in uterine flushings obtained from pregnant cowspost-mortem (P<0.05). Optical density of protein bands was not affectedby the day of the critical period, state (cyclic or pregnant) or interactionday x state. There was no effect of the conceptus weight orprogesterone concentration on the day of uterine flushing collectionin the optical density of the IFN-τ protein band.(AU)
Assuntos
Animais , Feminino , Prenhez , Prostaglandinas/efeitos adversos , Luteólise/fisiologia , Interferons/efeitos adversos , Interferons/análise , Mortalidade Fetal , BovinosResumo
Com o intuito de investigar se a hipóxia representa um dos desencadeadores da regressão luteínica em cadelas não prenhes, o presente estudo foi delineado para analisar a expressão do fator transcricional indutível por hipóxia HIF1A e a de seus genes-alvo relacionados à angiogênese, como o fator de crescimento endotelial vascular (VEGFA) e à captação de glicose, como as proteínas transportadoras facilitadoras GLUT1/SLC2A1 e GLUT4/SLC2A4 no corpo lúteo canino ao longo do diestro (dias 10 a 70 após a ovulação). Para tal, utilizou-se imuno-histoquímica e western blotting para localizar e quantificar as proteínas do HIF1A, GLUT1 e GLUT4 e PCR em tempo real para quantificar a expressão do RNAm de HIF1A, SLC2A1, SLC2A4, VEGFA, FLT1 e KDR. Além disso, células luteínicas nas fases inicial (dia 10), média (dia 30) e final (dia 60) foram submetidas ao tratamento com cloreto de cobalto (CoCl2) a 500 µM para avaliar os efeitos da hipóxia sobre a expressão gênica dos fatores acima citados, bem como sobre a produção de progesterona e 17β-estradiol. Nossos resultados demonstraram que o HIF1A é expresso pelo corpo lúteo canino de maneira tempo-dependente ao longo do diestro, e que a expressão de seu RNAm está diretamente correlacionada a expressão gênica de SLC2A1, SLC2A4, VEGFA, FLT1 e KDR e com as concentrações de progesterona periférica. No cultivo primário de células luteínicas, a hipóxia induzida pelo CoCl2 diminuiu a produção de progesterona e de 17β-estradiol e estimulou significativamente a expressão de HIF1A, SLC2A1, SLC2A4 e VEGFA. Esses resultados sugerem que o HIF1A constitui um dos fatores regulatórios da função do corpo lúteo canino participando da modulação de processos como esteroidogêne, angiogênese e da captação de glicose, atuando como fator luteolítico
This study was designed to investigate if hypoxia is one of the triggers of luteal regression in non-pregnant bitches. For that, we analyzed the hypoxia- inducible factor (HIF1A) expression as well as the expression of its target genes related to angiogenesis (vascular endothelial growth factor VEGFA) and to glucose uptake (glucose transporters GLUT/SLC2A 1 and 4) in canine corpus luteum throughout diestrus (days 10 to 70 after ovulation). We used immunohistochemistry and western blotting to localize and quantify the protein expression of HIF1A, GLUT1 and GLUT4, respectively, and real time PCR to analyze HIF1A, SLC2A1, SLC2A4, VEGFA, FLT1 and KDR gene expression. Moreover, luteal cells from early (day 10), mid (day 30) and late luteal phase (day 60) were submitted to 500 µM cobalt chloride (CoCl2) treatment to verify hypoxia effects on gene expression of the above cited genes and on progesterone and 17β-estradiol production. Our results showed that luteal cells expressed HIF1A in a time-dependent manner over diestrus and that its expression was directly correlated to both SLC2A1, SLC2A4, VEGFA, FLT1 and KDR gene expression and progesterone production. The protein expression of the studied genes also changed over diestrus and was correlated with the respective gene expression. In primary luteal cells culture, cobalt chloride-induced hypoxia downregulated progesterone and 17β-estradiol production, but upregulated HIF1A, SLC2A1, SLC2A4 and VEGFA gene expression. These findings suggest that HIF1A is one of the factors regulating canine luteal function by modulating important process as steroidogenesis, angiogenesis and glucose uptake, acting as a pro-survival factor
Assuntos
Feminino , Animais , Gravidez , Cães , Corpo Lúteo/embriologia , Hipóxia/diagnóstico , Hipóxia/embriologia , Hipóxia/veterinária , Luteólise/fisiologia , DiestroResumo
Com o intuito de investigar se a hipóxia representa um dos desencadeadores da regressão luteínica em cadelas não prenhes, o presente estudo foi delineado para analisar a expressão do fator transcricional indutível por hipóxia HIF1A e a de seus genes-alvo relacionados à angiogênese, como o fator de crescimento endotelial vascular (VEGFA) e à captação de glicose, como as proteínas transportadoras facilitadoras GLUT1/SLC2A1 e GLUT4/SLC2A4 no corpo lúteo canino ao longo do diestro (dias 10 a 70 após a ovulação). Para tal, utilizou-se imuno-histoquímica e western blotting para localizar e quantificar as proteínas do HIF1A, GLUT1 e GLUT4 e PCR em tempo real para quantificar a expressão do RNAm de HIF1A, SLC2A1, SLC2A4, VEGFA, FLT1 e KDR. Além disso, células luteínicas nas fases inicial (dia 10), média (dia 30) e final (dia 60) foram submetidas ao tratamento com cloreto de cobalto (CoCl2) a 500 µM para avaliar os efeitos da hipóxia sobre a expressão gênica dos fatores acima citados, bem como sobre a produção de progesterona e 17β-estradiol. Nossos resultados demonstraram que o HIF1A é expresso pelo corpo lúteo canino de maneira tempo-dependente ao longo do diestro, e que a expressão de seu RNAm está diretamente correlacionada a expressão gênica de SLC2A1, SLC2A4, VEGFA, FLT1 e KDR e com as concentrações de progesterona periférica. No cultivo primário de células luteínicas, a hipóxia induzida pelo CoCl2 diminuiu a produção de progesterona e de 17β-estradiol e estimulou significativamente a expressão de HIF1A, SLC2A1, SLC2A4 e VEGFA. Esses resultados sugerem que o HIF1A constitui um dos fatores regulatórios da função do corpo lúteo canino participando da modulação de processos como esteroidogêne, angiogênese e da captação de glicose, atuando como fator luteolítico (AU)
This study was designed to investigate if hypoxia is one of the triggers of luteal regression in non-pregnant bitches. For that, we analyzed the hypoxia- inducible factor (HIF1A) expression as well as the expression of its target genes related to angiogenesis (vascular endothelial growth factor VEGFA) and to glucose uptake (glucose transporters GLUT/SLC2A 1 and 4) in canine corpus luteum throughout diestrus (days 10 to 70 after ovulation). We used immunohistochemistry and western blotting to localize and quantify the protein expression of HIF1A, GLUT1 and GLUT4, respectively, and real time PCR to analyze HIF1A, SLC2A1, SLC2A4, VEGFA, FLT1 and KDR gene expression. Moreover, luteal cells from early (day 10), mid (day 30) and late luteal phase (day 60) were submitted to 500 µM cobalt chloride (CoCl2) treatment to verify hypoxia effects on gene expression of the above cited genes and on progesterone and 17β-estradiol production. Our results showed that luteal cells expressed HIF1A in a time-dependent manner over diestrus and that its expression was directly correlated to both SLC2A1, SLC2A4, VEGFA, FLT1 and KDR gene expression and progesterone production. The protein expression of the studied genes also changed over diestrus and was correlated with the respective gene expression. In primary luteal cells culture, cobalt chloride-induced hypoxia downregulated progesterone and 17β-estradiol production, but upregulated HIF1A, SLC2A1, SLC2A4 and VEGFA gene expression. These findings suggest that HIF1A is one of the factors regulating canine luteal function by modulating important process as steroidogenesis, angiogenesis and glucose uptake, acting as a pro-survival factor (AU)
Assuntos
Animais , Feminino , Gravidez , Cães , Corpo Lúteo/embriologia , Hipóxia/diagnóstico , Hipóxia/embriologia , Hipóxia/veterinária , Luteólise/fisiologia , DiestroResumo
O corpo lúteo (CL) é uma estrutura endócrina transitória formada por células luteais esteroidogênicas pequenas (SLC) e grandes (LLC), que associadas aos fibroblastos e a uma ampla rede de capilares constituem uma estrutura especializada na síntese de progesterona (P4). De maneira geral, para a síntese de P4 nas células luteais esteroidogênicas, o colesterol se liga a receptores específicos na membrana celular e é transportado ao citosol. Posteriormente, o colesterol dirige-se a membrana mitocondrial interna e por ação da enzima P450scc transforma-se em pregnenolona. No retículo endoplasmático liso a pregnenolona é convertida a P4 pela enzima 3β-hidroxiesteróide deidrogenase (3β-HSD). A proteína de regulação aguda da esteroidogênese (StAR), o receptor benzodiazepínico tipo periférico (PBR) e a endozepina participam no transporte do colesterol para os diferentes compartimentos mitocondriais. Assim, supõe-se que a capacidade de síntese de P4 no CL esteja relacionada à concentração celular de receptores que captam o colesterol, às enzimas P450scc e 3β-HSD, e às proteínas celulares transportadoras de colesterol. Na espécie bovina, as LLC são responsáveis por mais de 80% da produção deste hormônio no CL. A menor concentração de proteínas transportadoras de colesterol na mitocôndria parecem limitar a síntese de P4 nas SLC. A P4 favorece um meio ambiente uterino apropriado para o desenvolvimento do(s) concepto(s), dependendo da espécie. Na maioria das espécies, a ausência da fertilização ou a incapacidade do concepto em sinalizar sua existência no útero estabelecem a luteólise. Tal evento fisiológico caracteriza-se pela regressão funcional e estrutural do(s) corpo(s) lúteo(s). Para o estabelecimento e a manutenção da prenhez torna-se necessário o bloqueio da luteólise por mecanismos que diferem entre as espécies. Em primatas e equídeos esse reconhecimento ocorre pela secreção de gonadotrofinas específicas e em ruminantes pela presença de fatores anti-luteolíticos.(AU)
The corpus luteum (CL) is a transitory endocrinal structure formed by steroidogenic small luteal cells (SLC) and large luteal cells (LLC) that associated with fibroblast and a wide web of capillaries form a structure specialized in synthesis of progesterone (P4) In general, for the synthesis of P4 in the steroidogenic luteal cells, cholesterol joints specific receptors on the cellular membrane and is transported to the cytosol. Later, cholesterol goes to an internal mitochondrial membrane and by the action of the enzyme P450scc is transformed into pregnenolone. In the smooth endoplasmic reticulum pregnenolone is converted to P4 by the enzyme 3β-hydroxysteroid dehydrogenase (3 β -HSD). The steroidogenic acute regulatory protein (StAR), the peripheral benzodiazepine receptor (PBR) and endozepine participate in the transport of cholesterol to the different mitochondrial compartments. Therefore, it is supposed that the capacity of synthesis of P4 in the CL is related to the cellular concentration of receptors that catch cholesterol, to the enzymes P450scc and 3β-HSD and to the cholesterol cellular transport proteins. In bovine, the LLC are responsible for more than 80 of P4 production in the CL. The lowest concentration of cholesterol transport proteins in the mitochondria seems to limit the synthesis of P4 in the SLC. P4 supports a proper uterine environment for the development of conceptuses, depending on the specie. In most species, the lack of fertilization or the conceptus incapacity to signalize its presence in the uterus establishes luteolysis. This physiolocal event is characterized by the functional and structural regression of the CL. For the establishment and maintenance of pregnancy it is necessary to block luteolysis through different mechanisms among species. In primates and equids it occurs by the secretion of specific gonadotropins and in ruminants by antiluteolytic factors. (AU)