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1.
Elife ; 102021 05 24.
Artigo em Inglês | MEDLINE | ID: mdl-34029184

RESUMO

Receptor-interacting serine/threonine-protein kinase 3 (RIPK3) normally signals to necroptosis by phosphorylating MLKL. We report here that when the cellular RIPK3 chaperone Hsp90/CDC37 level is low, RIPK3 also signals to apoptosis. The apoptotic function of RIPK3 requires phosphorylation of the serine 165/threonine 166 sites on its kinase activation loop, resulting in inactivation of RIPK3 kinase activity while gaining the ability to recruit RIPK1, FADD, and caspase-8 to form a cytosolic caspase-activating complex, thereby triggering apoptosis. We found that PGF2α induces RIPK3 expression in luteal granulosa cells in the ovary to cause luteal regression through this RIPK3-mediated apoptosis pathway. Mice carrying homozygous phosphorylation-resistant RIPK3 S165A/T166A knockin mutations failed to respond to PGF2α but retained pro-necroptotic function, whereas mice with phospho-mimicking S165D/T166E homozygous knock-in mutation underwent spontaneous apoptosis in multiple RIPK3-expressing tissues and died shortly after birth. Thus, RIPK3 signals to either necroptosis or apoptosis depending on its serine 165/threonine 166 phosphorylation status.


Assuntos
Apoptose , Corpo Lúteo/enzimologia , Dinoprosta/metabolismo , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Animais , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Chaperoninas/genética , Chaperoninas/metabolismo , Corpo Lúteo/patologia , Proteína de Domínio de Morte Associada a Fas/genética , Proteína de Domínio de Morte Associada a Fas/metabolismo , Feminino , Células HEK293 , Células HT29 , Células HeLa , Humanos , Células MCF-7 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Mutação , Fosforilação , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Proteína Serina-Treonina Quinases de Interação com Receptores/genética , Transdução de Sinais
2.
Int J Mol Sci ; 22(7)2021 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-33915762

RESUMO

The corpus luteum (CL) is a temporary endocrine gland vital for pregnancy establishment and maintenance. Estradiol-17ß (E2) is the major embryonic signal in pigs supporting the CL's function. The mechanisms of the luteoprotective action of E2 are still unclear. The present study aimed to determine the effect of E2 on luteal expression of factors involved in CL function. An in vivo model of intrauterine E2 infusions was applied. Gilts on day 12 of pregnancy and the estrous cycle were used as referential groups. Concentrations of E2 and progesterone were elevated in CLs of gilts receiving E2 infusions, compared to placebo-treated gilts. Estradiol-17ß stimulated luteal expression of DNA-methyltransferase 1 (DNMT1), but decreased expression of DNMT3B gene and protein, as well as DNMT3A protein. Similar results for DNMT3A and 3B were observed in CLs on day 12 of pregnancy compared to day 12 of the estrous cycle. Intrauterine infusions of E2 altered luteal expression of the genes involved in CL function: PTGFR, PTGES, STAR, HSD17B1, CYP19A1, and PGRMC1. Our findings indicate a role for E2 in expression regulation of factors related to CL function and a novel potential for E2 to regulate DNA methylation as putative physiological mechanisms controlling luteal gene expression.


Assuntos
Corpo Lúteo/enzimologia , DNA (Citosina-5-)-Metiltransferases/metabolismo , Estradiol/fisiologia , Prenhez/metabolismo , Animais , Estradiol/administração & dosagem , Feminino , Gravidez , Progesterona/metabolismo , Suínos
3.
Sci Rep ; 10(1): 3681, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-32111948

RESUMO

Corpora lutea (CL) are transient endocrine glands supporting pregnancy by progesterone production. They develop at the site of ovulation from the remaining follicle, are highly metabolically active and undergo distinct, transformative processes during their lifetime. In contrast to other species, CL of lynxes do not regress at the end of cycle, but remain functionally active (persist) for years. Reactive oxygen species (ROS) and anti-oxidative enzymes are described to be important for the functionality of CL. We examined ten anti-oxidative enzymes in fresh and persistent CL of lynxes as well as in domestic cat CL of different luteal stages. The gene expression profiles, especially those of SOD1 and SOD2, showed some remarkable differences between CL stages during non-pregnant and pregnant cycles of domestic cats and between fresh and persistent CL of lynxes. Lynx gene expression profiles of SODs were confirmed by western blot analysis, immunohistochemistry and activity assays. SOD2 was characterized by a conspicuous high expression and enzyme activity exclusively in persistent CL. We suggest that SOD2 is required to detoxify potential elevated superoxide anion levels by producing H2O2 in the physiologically persistent CL. This product might also act as a signaling molecule, securing the CL from apoptosis and insuring long-term luteal cell survival.


Assuntos
Antioxidantes/metabolismo , Corpo Lúteo/enzimologia , Lynx/fisiologia , Superóxido Dismutase/metabolismo , Animais , Gatos , Feminino , Masculino , Gravidez
4.
Reprod Fertil Dev ; 28(5): 565-73, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25194502

RESUMO

There is considerable evidence of the neuroendocrine control involved in luteal regression in the rat. In addition, circulating prolactin (PRL), which increases during the night before parturition, may gain access to the coeliac ganglion (CG), indirectly impacting the physiology of the ovary because of the known connection between the CG and the ovary via the superior ovarian nerve (SON). In this work we investigated in the CG-SON-ovary system and whether PRL added to the CG has an impact, indirectly via the SON, on luteal regression on Day 21 of pregnancy. The system was incubated without (control) or with PRL added to the CG. We measured the ovarian release of progesterone (P), oestradiol and prostaglandin F2 alpha (PGF2α) by radioimmunoassay, and nitrites (NO) by the Griess method. Luteal mRNA expression of 3ß-hydroxysteroid dehydrogenase (3ß-HSD), 20α-HSD, aromatase, inducible nitric oxide synthase (iNOS) and apoptosis regulatory factors was analysed by reverse transcription-polymerase chain reaction. P release, the expression of Bcl-2 and the Bcl-2:Bax ratio was lower than control preparations, while the expression of 20α-HSD and the release of NO and PGF2α were higher in the experimental group. In conclusion, PRL acts at the CG and, by a neural pathway, modulates luteal function at the end of pregnancy.


Assuntos
Corpo Lúteo/inervação , Gânglios Simpáticos/efeitos dos fármacos , Luteólise/efeitos dos fármacos , Ovário/inervação , Prolactina/farmacologia , 20-alfa-Hidroxiesteroide Desidrogenase/genética , 20-alfa-Hidroxiesteroide Desidrogenase/metabolismo , 3-Hidroxiesteroide Desidrogenases/genética , 3-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Aromatase/genética , Aromatase/metabolismo , Corpo Lúteo/enzimologia , Corpo Lúteo/patologia , Dinoprosta/metabolismo , Estradiol/metabolismo , Feminino , Gânglios Simpáticos/fisiologia , Idade Gestacional , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Nitritos/metabolismo , Ovário/metabolismo , Gravidez , Progesterona/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ratos Sprague-Dawley , Fatores de Tempo , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismo
5.
J Steroid Biochem Mol Biol ; 154: 75-84, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26170243

RESUMO

European lynx species demonstrate an atypical ovarian cycle compared to other felids. The physiological persistence of corpora lutea (CLs), reflected in constantly elevated progesterone (P4) concentrations in serum, is thought to ensure a seasonal monooestrus. Moreover, the coexistence of CLs from a recent ovulation (freshCLs) and persistent CLs from previous years (perCLs) on the same ovary has been proven. We assume that perCLs in lynxes occur due to fundamentally different mechanisms of luteal regression. Our study presents a detailed analysis of steroidogenic enzymes and steroids in fresh and perCLs obtained from Iberian lynxes during metoestrus, and in perCLs obtained from Eurasian lynxes during prooestrus. By quantitative PCR we measured relative mRNA amounts of steroidogenic acute regulatory protein (STAR), cytochrome P450 oxidases (CYPs), hydroxysteroid dehydrogenases (HSDs) and a steroid reductase (SRD). Protein expression in CLs was investigated for CYP11A1, CYP17A1, CYP19A1 and HSD3B. Additionally, the intraluteal and serum steroid content was determined. During metoestrus, mRNA amounts of STAR, CYP11A1, CYP19A1, HSD17B7 and SRD5A1 were significantly higher in perCLs compared to freshCLs. Protein of CYP11A1 was detected independently of the CL age in metoestrus, but expression was less evident in prooestrous perCLs. The protein signal of CYP17A1 was strong in freshCLs and perCLs of metoestrus, but weak at prooestrus. The presence of CYP19A1 protein was confirmed in each stage of the CL. These findings contribute to the hypothesis that CLs from previous years might support freshly developed CLs for pregnancy maintenance. However, initiation of ovulation might require a functional down-regulation of perCLs prior to breeding. It is noteworthy that the HSD3B1 mRNA amount was significantly elevated in fresh compared to perCLs (metoestrus). Accordingly, HSD3B protein was substantially present in freshCLs, whereas signals were literally absent in all perCLs. Elevated expression of HSD3B coincided with high intraluteal oestrogen concentrations in freshCLs; however, the enzyme pattern was less concordant with intraluteal P4 and androgen concentrations. Serum P4 concentrations of Iberian lynxes were constant between prooestrus and prolonged dioestrus. Moreover, constantly high serum oestrogen concentrations were measured during pro-, met- and prolonged dioestrus. The physiology of exceptionally high serum oestrogen concentrations outside the breeding season of lynxes merits further investigation. In conclusion our study supports the concept that the unique reproductive strategy of lynxes is directly linked to sustained intraluteal steroid biogenesis in persistent CLs.


Assuntos
Corpo Lúteo/enzimologia , Sistema Enzimático do Citocromo P-450/metabolismo , Estro , Hidroxiesteroide Desidrogenases/metabolismo , Lynx/fisiologia , Animais , Feminino
6.
J Reprod Dev ; 60(2): 150-4, 2014 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-24492656

RESUMO

Regression of the corpus luteum (CL) is characterized by a decay in progesterone (P4) production (functional luteolysis) and disappearance of luteal tissues (structural luteolysis). In mares, structural luteolysis is thought to be caused by apoptosis of luteal cells, but functional luteolysis is poorly understood. 20α-hydroxysteroid dehydrogenase (20α-HSD) catabolizes P4 into its biologically inactive form, 20α-hydroxyprogesterone (20α-OHP). In mares, aldo-keto reductase (AKR) 1C23, which is a member of the AKR superfamily, has 20α-HSD activity. To clarify whether AKR1C23 is associated with functional luteolysis in mares, we investigated the expression of AKR1C23 in the CL in different luteal phases. The luteal P4 concentration and levels of 3ß-hydroxysteroid dehydrogenase (3ß-HSD) mRNA were higher in the mid luteal phase than in the late and regressed luteal phases (P<0.05), but the level of 3ß-HSD protein was higher in the late luteal phase than in the regressed luteal phase (P<0.05). The luteal 20α-OHP concentration and the level of AKR1C23 mRNA were higher in the late luteal phase than in the early and mid luteal phases (P<0.05), and the level of AKR1C23 protein was also highest in the late luteal phase. Taken together, these findings suggest that metabolism of P4 by AKR1C23 is one of the processes contributing to functional luteolysis in mares.


Assuntos
Aldeído Redutase/biossíntese , Corpo Lúteo/enzimologia , Cavalos/metabolismo , Fase Luteal/metabolismo , Luteólise/fisiologia , 20-alfa-Di-Hidroprogesterona/biossíntese , 20-alfa-Di-Hidroprogesterona/genética , 3-Hidroxiesteroide Desidrogenases/biossíntese , 3-Hidroxiesteroide Desidrogenases/genética , Aldeído Redutase/genética , Aldo-Ceto Redutases , Animais , Western Blotting , Feminino , Regulação Enzimológica da Expressão Gênica , Progesterona/biossíntese , Progesterona/genética , RNA/química , RNA/genética , Reação em Cadeia da Polimerase em Tempo Real/veterinária
7.
Reprod Biol Endocrinol ; 11: 111, 2013 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-24330451

RESUMO

BACKGROUND: During female reproductive cycles, a rapid fall in circulating progesterone (P4) levels is one of the earliest events that occur during induced luteolysis in mammals. In rodents, it is well recognized that during luteolysis, P4 is catabolized to its inactive metabolite, 20alpha-hydroxyprogesterone (20alpha-OHP) by the action of 20alpha-hydroxysteroid dehydrogenase (20alpha-HSD) enzyme and involves transcription factor, Nur77. Studies have been carried out to examine expression of 20alpha-HSD and its activity in the corpus luteum (CL) of buffalo cow. METHODS: The expression of 20alpha-HSD across different bovine tissues along with CL was examined by qPCR analysis. Circulating P4 levels were monitored before and during PGF2alpha treatment. Expression of 20alpha-HSD and Nur77 mRNA was determined in CL at different time points post PGF2alpha treatment in buffalo cows. The chromatographic separation of P4 and its metabolite, 20alpha-OHP, in rat and buffalo cow serum samples were performed on reverse phase HPLC system. To further support the findings, 20alpha-HSD enzyme activity was quantitated in cytosolic fraction of CL of both rat and buffalo cow. RESULTS: Circulating P4 concentration declined rapidly in response to PGF2alpha treatment. HPLC analysis of serum samples did not reveal changes in circulating 20alpha-OHP levels in buffalo cows but serum from pseudo pregnant rats receiving PGF2alpha treatment showed an increased 20alpha-OHP level at 24 h post treatment with accompanying decrease in P4 concentration. qPCR expression of 20alpha-HSD in CL from control and PGF2alpha-treated buffalo cows showed higher expression at 3 and 18 h post treatment, but its specific activity was not altered at different time points post PGF2alpha treatment. The Nur77 expression increased several fold 3 h post PGF2alpha treatment similar to the increased expression observed in the PGF2alpha-treated pseudo pregnant rats which perhaps suggest initiation of activation of apoptotic pathways in response to PGF2alpha treatment. CONCLUSIONS: The results taken together suggest that synthesis of P4 appears to be primarily affected by PGF2alpha treatment in buffalo cows in contrast to increased metabolism of P4 in rodents.


Assuntos
20-Hidroxiesteroide Desidrogenases/metabolismo , Búfalos/metabolismo , Corpo Lúteo/enzimologia , Dinoprosta/farmacologia , Progesterona/sangue , 20-Hidroxiesteroide Desidrogenases/sangue , Animais , Cromatografia Líquida de Alta Pressão , Feminino , Reação em Cadeia da Polimerase , Gravidez , Ratos
8.
Cytokine ; 64(1): 18-21, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23941776

RESUMO

Normal reproductive function involves the expression of inflammatory mediators. Regarding the corpus luteum (CL), cytokines promote the cross-talk between immune, vascular and steroidogenic cells, among others. Moreover, TNF, IFNG and FASL were shown to regulate equine CL establishment and regression. We hypothesized that cytokines action on equine CL may be mediated by nitric oxide (NO), through the regulation of endothelial NO synthase (eNOS) expression. TNF increased eNOS mRNA level and NO metabolite (nitrite) production during CL growth. Cytokines combined action (TNF+IFNG+FASL) promoted eNOS protein upregulation in mid-CL and nitrite production in mid and late-CL. However, in late-CL, TNF alone decreased nitrite secretion. These results indicate that in equine CL, cytokines TNF, IFNG and FASL regulate NO activity, via eNOS expression modulation.


Assuntos
Corpo Lúteo/metabolismo , Proteína Ligante Fas/metabolismo , Interferon gama/metabolismo , Óxido Nítrico/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Animais , Células Cultivadas , Corpo Lúteo/enzimologia , Corpo Lúteo/crescimento & desenvolvimento , Feminino , Cavalos , Óxido Nítrico Sintase Tipo III/genética , Óxido Nítrico Sintase Tipo III/metabolismo , Nitritos/metabolismo , RNA Mensageiro/biossíntese , Regulação para Cima
9.
Mol Cell Endocrinol ; 377(1-2): 103-11, 2013 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-23856004

RESUMO

We examined the effects of LPA on TNFα and IFNγ - induced decrease of P4 synthesis and on the cytokine - induced apoptosis of the cultured luteal cells. In the steroidogenic luteal cells LPA reversed the inhibitory effect of TNFα and IFNγ on P4 synthesis and also inhibited the stimulatory effects of TNFα and IFNγ on the expression of Bax, TNFR1, Fas and FasL as well as caspase 3 activity. These results suggest that TNFα and IFNγ cannot induce apoptosis in the presence of LPA, which orientates the steroidogenic luteal cells towards the survival state. In conclusion our results indicate that LPA supports P4 synthesis and action in the bovine CL.


Assuntos
Corpo Lúteo/efeitos dos fármacos , Corpo Lúteo/metabolismo , Interferon gama/farmacologia , Lisofosfolipídeos/farmacologia , Fator de Necrose Tumoral alfa/farmacologia , Animais , Apoptose/efeitos dos fármacos , Caspase 3/genética , Caspase 3/metabolismo , Caspase 8/genética , Caspase 8/metabolismo , Bovinos , Separação Celular , Células Cultivadas , Corpo Lúteo/citologia , Corpo Lúteo/enzimologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Progesterona/biossíntese , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Tipo I de Fatores de Necrose Tumoral/genética , Receptores Tipo I de Fatores de Necrose Tumoral/metabolismo , Receptores Tipo II do Fator de Necrose Tumoral/genética , Receptores Tipo II do Fator de Necrose Tumoral/metabolismo , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo
10.
Anim Reprod Sci ; 140(3-4): 138-46, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23896394

RESUMO

In the present paper, cellular composition of buffalo corpus luteum (CL) with its functional characterization based on 3ß-HSD and progesterone secretory ability at different stages of estrous cycle and pregnancy was studied. Buffalo uteri along with ovaries bearing CL were collected from the local slaughter house. These were classified into different stages of estrous cycle (Stage I, II, III and IV) and pregnancy (Stage I, II and III) based on morphological appearance of CL, surface follicles on the ovary and crown rump length of conceptus. Luteal cell population, progesterone content and steroidogenic properties were studied by dispersion of luteal cells using collagenase type I enzyme, RIA and 3ß-HSD activity, respectively. Large luteal cells (LLC) appeared as polyhedral or spherical in shape with a centrally placed large round nucleus and an abundance of cytoplasmic lipid droplets. However, small luteal cells (SLC) appeared to be spindle shaped with an eccentrically placed irregular nucleus and there was paucity of cytoplasmic lipid droplets. The size of SLC (range 12-23µm) and LLC (range 25-55µm) increased (P<0.01) with the advancement of stage of estrous cycle and pregnancy. The mean progesterone concentration per gram and per CL increased (P<0.01) from Stage I to III of estrous cycle with maximum concentration at Stage III of estrous cycle and pregnancy. The progesterone concentration decreased at Stage IV (day 17-20) of estrous cycle coinciding with CL regression. Total luteal cell number (LLC and SLC) also increased (P<0.01) from Stage I to III of estrous cycle and decreased (P<0.05), thereafter, at Stage IV indicating degeneration of luteal cells and regression of the CL. Total luteal cell population during pregnancy also increased (P<0.01) from Stage I to II and thereafter decreased (P>0.05) indicating cessation of mitosis. Increased (P<0.05) large luteal cell numbers from Stage I to III of estrous cycle and pregnancy coincided with the increased progesterone secretion and 3ß-HSD activity of CL. Thus, proportionate increases of large compared with small luteal cells were primarily responsible for increased progesterone secretion during the advanced stages of the estrous cycle and pregnancy. Total luteal cells and progesterone content per CL during the mid-luteal stage in buffalo as observed in the present study seem to be less than with cattle suggesting inherent luteal deficiency.


Assuntos
Búfalos/fisiologia , Corpo Lúteo/fisiologia , Ciclo Estral/fisiologia , Ovário/fisiologia , 3-Hidroxiesteroide Desidrogenases/análise , 3-Hidroxiesteroide Desidrogenases/fisiologia , Animais , Corpo Lúteo/citologia , Corpo Lúteo/enzimologia , Feminino , Histocitoquímica/veterinária , Células Lúteas/citologia , Células Lúteas/enzimologia , Células Lúteas/fisiologia , Ovário/citologia , Gravidez , Progesterona/análise , Progesterona/fisiologia
11.
Folia Morphol (Warsz) ; 72(1): 36-40, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23749709

RESUMO

The aim of this study was to evaluate the occurrence and the activity of prostaglandin-endoperoxide synthase 1 (PTGS1), PTGS2, and endothelial, neuronal, and inducible nitric oxide synthase (e-, n-, and iNOS) in early, mid, late, and regressive corpora lutea (CL) of bovines during diestrus. PTGS1 immunoreactivity was localised mainly in the cytoplasm of small luteal cells, whereas PTGS2 was detected in the cytoplasm of large luteal cells during early, mid, and late stages. The immunoexpression of all NOS isoforms was observed in the nuclei of luteal cells in the CL stages examined. PTGS1 enzyme activity was higher in late CL and lower in regressive ones; PTGS2 increased from early to late CL and lowered in regressive ones. Constitutive NOS enzymatic activity (eNOS plus nNOS) was higher in late CL and lower in regressive ones; iNOS was lower in regressive CL. These results support the idea that PTGSs and NOSs regulate the bovine CL life span mainly during the transition from the luteotrophic to the luteolytic phase.


Assuntos
Corpo Lúteo/enzimologia , Ciclo-Oxigenase 1/metabolismo , Ciclo-Oxigenase 2/metabolismo , Diestro/metabolismo , Óxido Nítrico Sintase/metabolismo , Animais , Bovinos , Feminino , Imuno-Histoquímica , Óxido Nítrico Sintase Tipo I/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo
12.
Biol Reprod ; 89(1): 2, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23677986

RESUMO

There is no distinct explanation of the mechanism for the prepartal prostaglandin F2alpha (PGF2alpha) increase in pregnant dogs. Although the PGF2alpha-synthase (PGFS [AKR1C3]) mRNA expression and localization profiles have been previously investigated in canine utero/placental compartments, the availability and biochemical activity of the PGFS (AKR1C3) protein remain unknown. In order to better understand the regulation of canine uterine PGF2alpha availability and eventual prepartum release in luteolytic amounts in dogs, canine-specific PGFS (AKR1C3) and 15-hydroxyprostaglandin dehydrogenase (HPGD) antibodies were generated and used to characterize the expression, cellular localization, and biochemical properties of PGFS (AKR1C3) and HPGD in the utero/placental compartments and corpus luteum throughout pregnancy and at prepartum luteolysis. PGFS (AKR1C3) expression was weak or absent in luteal samples. Uterine PGFS (AKR1C3) was up-regulated postimplantation and declined prepartum. The utero/placental expression of PGFS (AKR1C3) was identified in the superficial uterine glands throughout gestation and in the trophoblast cells within the feto-maternal contact zone during placentation, suggesting a possible role for PGFS (AKR1C3) in the trophoblast invasion. Utero-placental HPGD was up-regulated until postimplantation, lower at midgestation, and greatly suppressed at prepartum. Expression was routinely identified in the endometrial surface and glandular epithelia, and positive signals were also observed in the trophoblast cells at the feto-maternal contact zone. The biochemical activity of recombinant PGFS (AKR1C3) and HPGD was confirmed after its expression in a heterologous system. The colocalization of HPGD with PGFS (AKR1C3) expression suggests a modulatory role for HPGD as a gatekeeper of the supply of prostaglandin in the pregnant canine uterus.


Assuntos
Corpo Lúteo/enzimologia , Dinoprosta/biossíntese , Hidroxiprostaglandina Desidrogenases/biossíntese , Placenta/enzimologia , Prenhez/metabolismo , Animais , Chlorocebus aethiops , Dinoprosta/genética , Dinoprosta/fisiologia , Cães , Feminino , Hidroxiprostaglandina Desidrogenases/genética , Hidroxiprostaglandina Desidrogenases/fisiologia , Gravidez , Útero/enzimologia , Células Vero
13.
Anim Reprod Sci ; 138(1-2): 118-32, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23517855

RESUMO

The study was designed to examine the aglepristone (RU534) mechanisms affecting the corpora lutea (CL) lifespan in pseudopregnant rabbits. Aglepristone (10 mg/kg b.w.) was injected subcutaneously twice at either early- or mid-luteal phase (Days 3 and 4, or Days 8 and 9, respectively) after induction of ovulation with GnRH (Day 0). Corpora lutea and uteri, explanted at days 6 and 11, were evaluated for immunohistochemistry and Western blotting of progesterone (PR) and estrogen (ER) receptors, cyclooxygenase 1 (COX1), COX2, and PGE2-9-ketoreductase (PGE2-9-K) enzymatic activities, and progesterone, PGF2α, and PGE2 in vitro synthesis. Independent of luteal stage, aglepristone prolonged the functional luteal phase by 3 Days over that of controls as assessed by blood progesterone profiles. Aglepristone decreased protein for ER during both luteal-stages in CL and uteri. Progesterone receptor protein was decreased by RU354 at Days 6 in the uterus and at Days 11 in CL, whereas RU534 increased PR at Days 11 in uteri. In the CL, RU534 enhanced progesterone production at Days 6 and 11, whereas it decreased PGF2α and increased PGE2 at Day 11. In the uteri, RU534 decreased PGF2α and increased PGE2 synthesis at both days. COX2 and PGE2-9K activities were decreased by RU534 in the CL at Day 11, whereas in the uteri COX2 increased and PGE2-9-K decreased at Days 6 and 11. In conclusion, these data on aglepristone effects suggest that progesterone has a regulatory role on luteal function through direct and uterine-mediated mechanisms in pseudopregnant rabbits.


Assuntos
Corpo Lúteo/metabolismo , Estrenos/farmacologia , Pseudogravidez/metabolismo , Coelhos/metabolismo , Útero/metabolismo , Animais , Western Blotting/veterinária , Corpo Lúteo/enzimologia , Ciclo-Oxigenase 1/análise , Ciclo-Oxigenase 2/análise , Dinoprosta/análise , Dinoprostona/análise , Feminino , Hidroxiprostaglandina Desidrogenases/análise , Imuno-Histoquímica/veterinária , Fase Luteal/fisiologia , Progesterona/sangue , Receptores de Estrogênio/análise , Receptores de Progesterona/análise , Estatísticas não Paramétricas , Útero/enzimologia
14.
Biol Reprod ; 88(3): 62, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23365414

RESUMO

The expression of peroxisome proliferator-activated receptor gamma (PPARgamma) and its role in corpora lutea (CL) function were studied in pseudopregnant rabbits. Corpora lutea were collected at an early stage (Day 4), midstage (Day 9), and late stage (Day 13) of pseudopregnancy. Immunohistochemistry found evidence for the presence of PPARgamma in the perinuclear cytoplasm and nucleus of all the luteal cells; immunoreactivity decreased from the early to the late stage, with immunonegativity of the nuclei of late stage CL. PPARgamma mRNA transcript was expressed in all the luteal stages with the lowest level in the late stage. In CL cultured in vitro, the PPARgamma agonist (15-deoxy delta12,14 prostaglandin J2 [15d-PGJ2], 200 nM) increased and the antagonist (T0070907, 50 nM) decreased progesterone secretion at early and midluteal stages, whereas 15d-PGJ2 reduced and T0070907 increased PGF2alpha at the same stages. Prostaglandin-endoperoxide synthase 2 (PTGS2) activity was reduced by 15d-PGJ2 and increased by T0070907 in CL of early and midluteal stages. Conversely, 15d-PGJ2 increased and T0070907 reduced 3beta-hydroxysteroid dehydrogenase (3beta-HSD) activity in early and midluteal stage CL. PGE2 in vitro secretion as well as PTGS1 and 20alpha-HSD enzymatic activities were not affected by 15d-PGJ2 and T0070907 in any CL types. These results indicate that PPARgamma plays a luteotropic role in pseudopregnant rabbits, through PTGS2 down-regulation and 3beta-HSD up-regulation, with a consequent PGF2alpha decrease and progesterone increase.


Assuntos
Corpo Lúteo/enzimologia , PPAR gama/metabolismo , Pseudogravidez/metabolismo , Animais , Ciclo-Oxigenase 1/metabolismo , Ciclo-Oxigenase 2/metabolismo , Dinoprosta/metabolismo , Dinoprostona/metabolismo , Feminino , Expressão Gênica , Hidroxiprostaglandina Desidrogenases/metabolismo , Imuno-Histoquímica , Progesterona/metabolismo , Progesterona Redutase/metabolismo , RNA Mensageiro/metabolismo , Coelhos
15.
Reprod Domest Anim ; 48(1): 85-9, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22612285

RESUMO

In most mammals, the corpus luteum (CL) and placenta are the major sources of progesterone. The goat pregnancy depends on the presence of CL after mid-gestation, while sheep pregnancy does not. The expression and distribution of P450-aromatase (P450-Aro) mRNA throughout gestation has not been investigated in the goat CL and partially in the sheep CL. The present research was designed to characterize the expression of P450-Aro mRNA in small ruminant CL with emphasis in the goat. For this purpose, ovaries from Criollo goats and Pelibuey sheeps were analysed using in situ reverse transcription-polymerase chain reaction (RT-PCR) for the histological detection of P450-Aro transcripts. In addition, P450-Aro expression was determined by in vitro RT-PCR. In situ RT-PCR studies showed that the goat and sheep CL were rich in cells positive for P450-Aro mRNA. We have also found in vitro RT-PCR expression of P450-Aro mRNA in goat CL at 1, 3 and 4 months of gestation. This study shows that the goat CL expresses P450-Aro mRNA along gestation, suggesting that this structure is capable to produce oestrogens up to the end of gestation.


Assuntos
Aromatase/metabolismo , Corpo Lúteo/enzimologia , Cabras/fisiologia , Prenhez , Animais , Feminino , Regulação Enzimológica da Expressão Gênica , Células da Granulosa/enzimologia , Células da Granulosa/metabolismo , Gravidez , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Ovinos/fisiologia
16.
Reprod Biomed Online ; 25(6): 551-60, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23063822

RESUMO

Progesterone produced by the corpus luteum (CL) regulates the synthesis of various endometrial proteins required for embryonic implantation and development. Compromised CL progesterone production is a potential risk factor for prenatal development. Reactive oxygen species (ROS) play diverse roles in mammalian reproductive biology. ROS-induced oxidative damage and subsequent adverse developmental outcomes constitute important issues in reproductive medicine. The CL is considered to be highly exposed to locally produced ROS due to its high blood vasculature and steroidogenic activity. ROS-induced apoptotic cell death is involved in the mechanisms of CL regression that occurs at the end of the non-fertile cycle. Luteal ROS production and propagation depend upon several regulating factors, including luteal antioxidants, steroid hormones and cytokines, and their crosstalk. However, it is unknown which of these factors have the greatest contribution to the maintenance of CL integrity and function during the oestrous/menstrual cycle. There is evidence to suggest that antioxidants play important roles in CL rescue from luteolysis when pregnancy ensues. As luteal phase defect impacts fertility by preventing implantation and early conceptus development in livestock and humans, this review attempts to address the importance of ROS-scavenging antioxidant enzymes in the control of mammalian CL function and integrity. The corpus luteum (CL) is a transient endocrine organ that develops after ovulation from the ovulated follicle during each reproductive cycle. The main function of the CL is the production and secretion of progesterone which is necessary for embryonic implantation and development. Compromised CL progesterone production is a potential risk factor for prenatal development and pregnancy outcomes. Reactive oxygen species (ROS), which are natural by-products of cellular respiration and metabolism, play diverse roles in mammalian reproductive biology. ROS-induced oxidative damage and subsequent development of adverse pregnancy outcomes constitute important issues in reproductive medicine. Before the end of the first trimester, a high rate of human and animal conceptions end in spontaneous abortion and most of these losses occur at the time of implantation in association with ROS-induced oxidative damage. Every cell in the body is normally able to defend itself against the oxidative damage caused by the ROS. The cellular antioxidant enzymes constitute the first line of defence against the toxic effects of ROS. The CL is considered to be highly exposed to locally produced ROS due to its high blood vasculature and metabolic activity. There is now evidence to suggest that cellular antioxidants play important roles in CL rescue from regression when pregnancy ensues. As defective CL function impacts fertility by preventing implantation and early conceptus development in livestock and humans, this review attempts to address the importance of antioxidant enzymes in the control of mammalian CL function and integrity.


Assuntos
Manutenção do Corpo Lúteo/metabolismo , Corpo Lúteo/enzimologia , Estresse Oxidativo , Oxirredutases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Animais , Apoptose , Hipóxia Celular , Corpo Lúteo/irrigação sanguínea , Corpo Lúteo/metabolismo , Feminino , Humanos , Gravidez , Progesterona/metabolismo
17.
Microsc Res Tech ; 75(12): 1682-90, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22865504

RESUMO

Immunopresence, gene expression, and enzymatic activity of cyclooxygenase 1 (COX1), COX2, PGE2-9-ketoreductase (PGE2-9-K), endothelial (eNOS), and inducible nitric oxide synthases (iNOS), and hormone in vitro production were examined in early, mid, late, and regressive buffalo corpora lutea (CL). COX1 immunosignals were detected in the cytoplasm of small luteal cells, COX2 in large luteal cells, and PGE2-9-K in all luteal cells. COX2 and PGE2-9-K immunosignals were greater in late CL. Immunopresence of both NOS types were evidenced in the nuclei and cytoplasm of all luteal cells, as well as in the nuclei of endothelial cells, during all stages studied. The eNOS and iNOS immunosignals increased during the early stage. COX1 transcripts were lower in late and regressive CL, COX2 in late, PGE2-9-K higher in regressive, and iNOS higher in early and lower in regressive CL. COX1 enzymatic activity was lower in regressive CL, COX2 increased in mid and late stages, and PGE2-9-K was higher in late CL. Endothelial NOS activity was higher during mid and late stages and lower in regressive, whereas iNOS was greater in late and lower in early. Progesterone in vitro release was higher in mid and lower in late phase, while PGF2α synthesis was higher in late CL and lower in regressive, and PGE2 was higher during regressive stage. These results support the idea that COX, NOS, and PGE2-9-K regulate buffalo CL life span. In particular, regressive CL seems involved in the development of the contralateral early CL, through the production of the luteotrophic PGE2.


Assuntos
Búfalos/fisiologia , Corpo Lúteo/enzimologia , Diestro , Expressão Gênica , Hidroxiprostaglandina Desidrogenases/análise , Óxido Nítrico Sintase/análise , Prostaglandina-Endoperóxido Sintases/análise , Animais , Núcleo Celular/química , Citoplasma/química , Feminino , Perfilação da Expressão Gênica , Hidroxiprostaglandina Desidrogenases/genética , Imuno-Histoquímica , Microscopia , Óxido Nítrico Sintase/genética , Prostaglandina-Endoperóxido Sintases/genética
18.
Gene ; 503(2): 222-8, 2012 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-22575614

RESUMO

Neuron-specific enolase (NSE) is the key molecular marker for diffuse neuroendocrine system (DNES) cells, its expression in the pregnant corpus luteum (CL) of dairy goats was studied by the immunofluorescence method and the ultra structural characteristics of luteal cells were detected by the electron microscopy to identify the existence of DNES cells in the pregnant CL of dairy goats. Besides, the coding sequence of dairy goats NSE gene was cloned and its biological information was analyzed. Results revealed that NSE immunopositive cells exhibited widespread cytoplasmic staining throughout the whole pregnant CL. In addition, these cells showed typical characteristics of DNES cells in the electron microscopy. These results suggested that many DNES cells exist in the pregnant CL of dairy goats. Meanwhile, we identified the coding sequence of dairy goats NSE (GenBank Accession No. JN887466). Its nucleotide sequence homology was found to be 97.9%, 89.3%, 90% and 92.6%, respectively, compared with that of Bos taurus, Rattus norvegicus, Mus musculus and Homo sapiens, while the amino acid sequence homology was 99.1%, 97%, 97.2% and 98.2% respectively. These results first showed that the functional amino acids coded by the NSE gene were highly conserved in Caprine, B. taurus, R. norvegicus, M. musculus and H. sapiens. It was implied that the gene NSE in dairy goats had close homology to that of NSE of other species. Our findings demonstrated the possible existence of DNES cells in pregnant CL, providing new clue for further understanding of interactions between the neuroendocrine and reproductive systems. Characterization of gene sequence of dairy goats NSE will enable us to synthesize interference RNA for further study on the role of NSE on the formation, function and apoptosis of pregnant CL in dairy goats.


Assuntos
Corpo Lúteo/enzimologia , Cabras/genética , Fosfopiruvato Hidratase/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Corpo Lúteo/ultraestrutura , Feminino , Genótipo , Cabras/metabolismo , Dados de Sequência Molecular , Sistemas Neurossecretores/citologia , Gravidez , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos
19.
Reprod Sci ; 19(4): 416-22, 2012 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-22101240

RESUMO

There is evidence suggesting that estradiol (E(2)) regulates the physiology of the ovary and the sympathetic neurons associated with the reproductive function. The objective of this study was to investigate the effect of E(2) on the function of late pregnant rat ovaries, acting either directly on the ovarian tissue or indirectly via the superior ovarian nerve (SON) from the celiac ganglion (CG). We used in vitro ovary (OV) or ex vivo CG-SON-OV incubation systems from day 21 pregnant rats. Various concentrations of E(2 )were added to the incubation media of either the OV alone or the ganglion compartment of the CG-SON-OV system. In both experimental schemes, we measured the concentration of progesterone in the OV incubation media by radioimmunoassay at different times. Luteal messenger RNA (mRNA) expression of 3ß-hydroxysteroid dehydrogenase (3ß-HSD) and 20α-hydroxysteroid dehydrogenase (20α-HSD) enzymes, respectively, involved in progesterone synthesis and catabolism, and of antiapoptotic B-cell lymphoma 2 (Bcl-2) and proapoptotic Bcl-2-associated X protein (Bax), were measured by reverse transcriptase-polymerase chain reaction (RT-PCR) at the end of the incubation period. Estradiol added directly to the OV incubation or to the CG of the CG-SON-OV system caused a decline in the concentration of progesterone accumulated in the incubation media. In addition, E(2), when added to the OV incubation, decreased the expression of 3ß-HSD and the ratio of Bcl-2/Bax. We conclude that through a direct effect on the OV, E(2) favors luteal regression at the end of pregnancy in rats, in association with neural modulation from the CG via the SON.


Assuntos
Corpo Lúteo/efeitos dos fármacos , Estradiol/farmacologia , Gânglios Simpáticos/efeitos dos fármacos , Luteólise/efeitos dos fármacos , Ovário/efeitos dos fármacos , Progesterona/metabolismo , 20-alfa-Hidroxiesteroide Desidrogenase/genética , 20-alfa-Hidroxiesteroide Desidrogenase/metabolismo , 3-Hidroxiesteroide Desidrogenases/genética , 3-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Corpo Lúteo/enzimologia , Corpo Lúteo/inervação , Corpo Lúteo/fisiologia , Feminino , Gânglios Simpáticos/enzimologia , Gânglios Simpáticos/fisiologia , Técnicas In Vitro , Luteólise/fisiologia , Ovário/enzimologia , Ovário/inervação , Ovário/fisiologia , Gravidez , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , RNA Mensageiro/química , RNA Mensageiro/genética , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismo
20.
J Anim Sci ; 89(5): 1338-46, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21521814

RESUMO

In the present study, we performed quantitative reverse-transcription PCR (qPCR) to examine changes in gene expression of prolactin receptor (long form: l-PRLR; short form: s-PRLR) and 20α-hydroxysteroid dehydrogenase (20α-HSD; EC 1.1.1.149) in the bovine corpus luteum (CL) throughout the estrous cycle and pregnancy. Western blotting was used to determine protein abundance. Bovine CL were collected and luteal stages (n = 6/stage) were classified by macroscopic observation as early (d 1 to 4 after ovulation), mid (d 5 to 10), late (d 11 to 17), and regressing (d 18 to 20). A CL of pregnancy (n = 6) was determined by the presence of conceptus (d 28 to term). The mRNA for both forms of PRLR were expressed at all the luteal stages. Expression of s-PRLR and l-PRLR mRNA was less (P < 0.01) during early and regressing luteal stages compared with mid and late stages. Expression of s-PRLR mRNA in CL of pregnancy was greater (P < 0.01) than early, mid, and regressing CL and did not differ from late luteal stage expression. A greater (P < 0.01) expression of l-PRLR mRNA was observed in pregnant vs. early and regressing CL. In addition, qPCR showed the presence of 20α-HSD mRNA during all luteal stages of the estrous cycle, with the greatest (P < 0.01) expression observed in the regressing luteal stage. Western blotting revealed protein abundance of both PRLR isoforms during all luteal stages and pregnancy, with a predominance of the s-PRLR protein. Densitometry analysis indicated that protein abundances of s-PRLR were greater (P < 0.05) than l-PRLR during early, mid, and late luteal stages and did not differ during the regressing luteal stage. Protein abundances of 20α-HSD were least (P < 0.05) during the early luteal stage. In conclusion, results of the current study suggest a possible involvement of PRLR, especially s-PRLR, in the regulation of progesterone secretion and metabolism during the bovine estrous cycle and pregnancy.


Assuntos
Bovinos/fisiologia , Corpo Lúteo/metabolismo , Ciclo Estral/fisiologia , Progesterona/metabolismo , Receptores da Prolactina/biossíntese , 20-alfa-Hidroxiesteroide Desidrogenase/biossíntese , 20-alfa-Hidroxiesteroide Desidrogenase/genética , Animais , Western Blotting/veterinária , Corpo Lúteo/enzimologia , Feminino , Regulação Enzimológica da Expressão Gênica , Gravidez , Isoformas de Proteínas , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Receptores da Prolactina/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária
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