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1.
Endocrinology ; 163(1)2022 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-34734245

RESUMO

Polycyclic aromatic hydrocarbons like benzo[a]pyrene (BaP) are generated during incomplete combustion of organic materials. Prior research has demonstrated that BaP is a prenatal ovarian toxicant and carcinogen. However, the metabolic pathways active in the embryo and its developing gonads and the mechanisms by which prenatal exposure to BaP predisposes to ovarian tumors later in life remain to be fully elucidated. To address these data gaps, we orally dosed pregnant female mice with BaP from embryonic day (E) 6.5 to E11.5 (0, 0.2, or 2 mg/kg/day) for metabolite measurement or E9.5 to E11.5 (0 or 3.33 mg/kg/day) for embryonic gonad RNA sequencing. Embryos were harvested at E13.5 for both experiments. The sum of BaP metabolite concentrations increased significantly with dose in the embryos and placentas, and concentrations were significantly higher in female than male embryos and in embryos than placentas. RNA sequencing revealed that enzymes involved in metabolic activation of BaP are expressed at moderate to high levels in embryonic gonads and that greater transcriptomic changes occurred in the ovaries in response to BaP than in the testes. We identified 490 differentially expressed genes (DEGs) with false discovery rate P-values < 0.05 when comparing BaP-exposed to control ovaries but no statistically significant DEGs between BaP-exposed and control testes. Genes related to monocyte/macrophage recruitment and activity, prolactin family genes, and several keratin genes were among the most upregulated genes in the BaP-exposed ovaries. Results show that developing ovaries are more sensitive than testes to prenatal BaP exposure, which may be related to higher concentrations of BaP metabolites in female embryos.


Assuntos
Benzo(a)pireno/metabolismo , Gônadas/metabolismo , Placenta/metabolismo , Prenhez , Transcriptoma , Animais , Cromatografia Líquida de Alta Pressão , Biologia Computacional , Feminino , Inflamação , Queratinas/metabolismo , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/metabolismo , Ovário/metabolismo , Gravidez , RNA-Seq , Fatores Sexuais , Testículo/metabolismo , Fatores de Tempo
2.
PLoS One ; 16(10): e0257161, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34614002

RESUMO

The current study used RNA sequencing to determine transcriptional profiles of equine endometrium collected 14, 22, and 28 days after ovulation from pregnant mares. In addition, the transcriptomes of endometrial samples obtained 20 days after ovulation from pregnant mares, and from non-pregnant mares which displayed and failed to display extended luteal function following the administration of oxytocin, were determined and compared in order to delineate genes whose expressions depend on the presence of the conceptus as opposed to elevated progesterone alone. A mere fifty-five transcripts were differentially expressed between samples collected from mares at Day 22 and Day 28 of pregnancy. This likely reflects the longer-term exposure to a relatively constant, progesterone-dominated environment with little change in factors secreted by the conceptus that would affect endometrial gene expression. The complement system was amongst the canonical pathways significantly enriched in transcripts differentially expressed between Day 14 and Day 22/28 of pregnancy. The expression of complement components 7 and 8 was confirmed using in situ hybridization. The expression of SERPING1, an inhibitor of the complement system, was confirmed by immunohistochemistry. In line with the resumed capacity of the endometrium to produce prostaglandin, prostaglandin G/H synthase 1 was expressed at higher levels at Days 22 and 28 than at Day 14 of pregnancy. Our data suggest that this up-regulation is enhanced by the presence of the conceptus; samples obtained from mares at Day 20 of pregnancy had significantly higher levels of prostaglandin G/H synthase 1 transcript than mares with extended luteal function.


Assuntos
Endométrio/metabolismo , Cavalos/genética , Ocitocina/farmacologia , Prenhez , Transcriptoma , Animais , Corpo Lúteo/efeitos dos fármacos , Corpo Lúteo/fisiologia , Feminino , Cavalos/fisiologia , Ovulação/efeitos dos fármacos , Ocitocina/administração & dosagem , Gravidez , Transcriptoma/efeitos dos fármacos
3.
Theriogenology ; 176: 183-187, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34624812

RESUMO

The aim of the study was to compare three methods of reducing twin pregnancy in mares to maintain a single pregnancy. As multiple pregnancies in mare are always undesirable, early ultrasound diagnosis makes possible management of twin pregnancies and extra embryo removal. In years 2010-2018, 16494 mares were sonographically tested for early pregnancy, finding 868 cases of twins (471 bilateral and 397 unilateral). 260 mares with a confirmed bilateral tween pregnancy were subjected to manual crushing of one embryo and administration of flunixin at a dose of 1.1 mg/kg BW. 186 mares were subjected only to the embryo crushing procedure. 25 mares from this group were on a restrictive diet. In the unilateral twin pregnancy mare group, 62 were subjected to manual embryo reduction with simultaneous treatment with flunixin, 60 had only manual embryonic vesicle crush and 210 had a restrictive diet. Determination of success, measured as the development of a single pregnancy, were monitored 2 weeks after the procedure, between the 50th and 60th day of pregnancy and after the 90th day of pregnancy. In general, warm-blooded mares were more prone to a twin pregnancy, and at the same time, all the procedures used to reduce it to a single pregnancy caused a greater risk of losing both embryos than in the case of cold-blooded mares. The beneficial effect of administering flunixin after manual removal of one embryo on the maintenance of the other has been experimentally proven in both unilateral and bilateral twin pregnancy.


Assuntos
Redução de Gravidez Multifetal , Prenhez , Animais , Embrião de Mamíferos , Feminino , Cavalos , Gravidez , Redução de Gravidez Multifetal/veterinária , Gravidez Múltipla , Estudos Retrospectivos , Ultrassonografia
4.
Arq. Ciênc. Vet. Zool. UNIPAR (Online) ; 24(2, cont.): e2408, jul-dez. 2021. ilus, graf
Artigo em Português | ID: biblio-1352315

RESUMO

A inseminação artificial em tempo fixo (IATF) é uma biotécnica com inúmeras vantagens para a produção de bovinos. Para a realização da IATF, as fêmeas bovinas devem receber um protocolo farmacológico que induz a ovulação de forma sincronizada. Esse protocolo inicia com a inserção de um dispositivo impregnado com progesterona que deve permanecer no fundo vaginal da fêmea por tempo determinado. O presente relato tem por objetivo descrever a remoção de um implante impregnado com progesterona que foi encontrado na cavidade abdominal de uma novilha. Durante um atendimento reprodutivo de rotina em uma propriedade rural, uma novilha da raça Jersey, recém-adquirida, com histórico de ter sido submetida à IATF, foi avaliada para diagnóstico de gestação. No exame a novilha não estava gestante e apresentava um cisto folicular ovariano. A novilha foi submetida novamente à IATF em que foi inserido e removido um implante, com isso o cisto regrediu e ao inseminar a novilha não foi possível passar a cérvix com o aplicador. Vinte dias após a inseminação a novilha manifestou estro e ao ser avaliada para nova inseminação, foi identificado um implante solto na cavidade abdominal da novilha. Foi realizada uma laparotomia pelo flanco esquerdo e o implante foi removido. Após a recuperação da cirurgia a novilha foi inseminada e novamente e ficou gestante. Os protocolos de IATF devem ser realizados por profissionais capacitados para evitar problemas como a presente situação em que um implante foi encontrado na cavidade abdominal de uma novilha.(AU)


The fixed-time artificial insemination (FTAI) is a biotechnique with numerous advantages for cattle production. In order to perform the FTAI, the cows must receive a pharmacological protocol to induce ovulation in a synchronized way. This protocol begins with the insertion of a device impregnated with progesterone that must remain in the vaginal fundus of the cow for a specific period of time. This report aims at describing the removal of a progesterone impregnated implant that was found in the abdominal cavity of a heifer. During a routine reproductive service in a rural property, a recently acquired Jersey heifer, with history of having been submitted to FTAI, was evaluated for pregnancy diagnosis. At the exam, it was observed that the heifer was not pregnant and presented an ovarian follicular cyst. The heifer was then submitted to FTAI again, where an implant was inserted and removed. This made the cyst to recede, and upon subsequent insemination, the applicator was not able to pass the cervix. Twenty days after insemination, the heifer was in estrus and when it was evaluated for insemination, a loose implant was identified in the abdominal cavity of the heifer. A laparotomy was performed through the left flank and the implant was removed. After recovery from the surgery, the heifer was inseminated again and became pregnant. The FTAI protocols should only be carried out by trained professionals to avoid problems such as the one presented herein, where an implant was found in the abdominal cavity of a heifer.(AU)


La inseminación artificial a tiempo fijo (FTAI) es una biotécnica con numerosas ventajas para la producción de ganado. Para realizar FTAI, las hembras bovinas deben recibir un protocolo farmacológico que induzca la ovulación de forma sincronizada. Este protocolo comienza con la inserción de un dispositivo impregnado con progesterona que debe permanecer en el fondo vaginal de la hembra durante un tiempo específico. Este informe tiene como objetivo describir la extracción de un implante impregnado de progesterona que se encontró en la cavidad abdominal de una novilla. Durante la atención reproductiva de rutina en una propiedad rural, se evaluó el diagnóstico de preñez de una novilla de Jersey recién adquirida con antecedentes de someterse a FTAI. En el examen, la novilla no estaba embarazada y tenía un quiste folicular ovárico. La novilla fue nuevamente sometida a FTAI, en la cual se insertó y removió un implante, con esto el quiste retrocedió y al inseminar a la novilla no fue posible pasar el cérvix con el aplicador. Veinte días después de la inseminación, la vaquilla manifestó estro y cuando se evaluó para una nueva inseminación, se identificó un implante suelto en la cavidad abdominal de la vaquilla. Se realizó una laparotomía a través del flanco izquierdo y se extrajo el implante. Después de recuperarse de la cirugía, la novilla fue inseminada y nuevamente quedó embarazada. Los protocolos FTAI deben ser realizados por profesionales capacitados para evitar problemas como la situación actual donde se encontró un implante en la cavidad abdominal de una novilla.(AU)


Assuntos
Animais , Bovinos , Ovulação , Progesterona , Prenhez , Inseminação Artificial , Colo do Útero , Corpos Estranhos , Estro
5.
Int J Mol Sci ; 22(18)2021 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-34575877

RESUMO

Embryo manipulation is a requisite step in assisted reproductive technology (ART). Therefore, it is of great necessity to appraise the safety of ART and investigate the long-term effect, including lipid metabolism, on ART-conceived offspring. Augmenting our ART rabbit model to investigate lipid metabolic outcomes in offspring longitudinally, we detected variations in hepatic DNA methylation ART offspring in the F3 generation for embryonic exposure (multiple ovulation, vitrification and embryo transfer). Through adult liver metabolomics and proteomics, we identified changes mainly related to lipid metabolism (e.g., polyunsaturated fatty acids, steroids, steroid hormone). We also found that DNA methylation analysis was linked to changes in lipid metabolism and apoptosis genes. Nevertheless, these differences did not apparently alter the general health status. Thus, our findings suggest that ART is likely to be a player in embryo epigenetic events related to hepatic homeostasis alteration in adulthood.


Assuntos
Metilação de DNA , Transferência Embrionária , Epigenômica , Fígado/embriologia , Técnicas de Reprodução Assistida , Animais , Embrião de Mamíferos/metabolismo , Epigênese Genética , Feminino , Genoma , Humanos , Metabolismo dos Lipídeos , Fígado/metabolismo , Masculino , Metaboloma , Gravidez , Prenhez , Proteoma , Proteômica/métodos , Coelhos , Reprodução , Esteroides/biossíntese , Vitrificação
6.
Sci Rep ; 11(1): 16569, 2021 08 16.
Artigo em Inglês | MEDLINE | ID: mdl-34400677

RESUMO

Maternal immune adaptation to accommodate pregnancy depends on sufficient availability of regulatory T (Treg) cells to enable embryo implantation. Toll-like receptor 4 is implicated as a key upstream driver of a controlled inflammatory response, elicited by signals in male partner seminal fluid, to initiate expansion of the maternal Treg cell pool after mating. Here, we report that mice with null mutation in Tlr4 (Tlr4-/-) exhibit impaired reproductive outcomes after allogeneic mating, with reduced pregnancy rate, elevated mid-gestation fetal loss, and fetal growth restriction, compared to Tlr4+/+ wild-type controls. To investigate the effects of TLR4 deficiency on early events of maternal immune adaptation, TLR4-regulated cytokines and immune regulatory microRNAs were measured in the uterus at 8 h post-mating by qPCR, and Treg cells in uterus-draining lymph nodes were evaluated by flow cytometry on day 3.5 post-coitum. Ptgs2 encoding prostaglandin-endoperoxide synthase 2, cytokines Csf2, Il6, Lif, and Tnf, chemokines Ccl2, Cxcl1, Cxcl2, and Cxcl10, and microRNAs miR-155, miR-146a, and miR-223 were induced by mating in wild-type mice, but not, or to a lesser extent, in Tlr4-/- mice. CD4+ T cells were expanded after mating in Tlr4+/+ but not Tlr4-/- mice, with failure to expand peripheral CD25+FOXP3+ NRP1- or thymic CD25+FOXP3+ NRP1+ Treg cell populations, and fewer Treg cells expressed Ki67 proliferation marker and suppressive function marker CTLA4. We conclude that TLR4 is an essential mediator of the inflammation-like response in the pre-implantation uterus that induces generation of Treg cells to support robust pregnancy tolerance and ensure optimal fetal growth and survival.


Assuntos
Retardo do Crescimento Fetal/imunologia , Reabsorção do Feto/imunologia , Prenhez/imunologia , Receptor 4 Toll-Like/deficiência , Animais , Quimiotaxia de Leucócito , Ciclo-Oxigenase 2/biossíntese , Ciclo-Oxigenase 2/genética , Citocinas/biossíntese , Citocinas/genética , Feminino , Retardo do Crescimento Fetal/genética , Reabsorção do Feto/genética , Idade Gestacional , Mutação com Perda de Função , Linfonodos/imunologia , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , MicroRNAs/biossíntese , MicroRNAs/genética , Tamanho do Órgão , Placenta/anatomia & histologia , Gravidez , Resultado da Gravidez , Taxa de Gravidez , Sêmen/imunologia , Subpopulações de Linfócitos T/imunologia , Linfócitos T Reguladores/imunologia , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologia , Útero/metabolismo
7.
Biomed Res Int ; 2021: 6681491, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34195279

RESUMO

Background: Although hypothyroidism during pregnancy may develop grave outcomes for both mothers and offspring, management of which is still a challenge due to the insufficient understanding of this disease. The close correlation between hypothyroidism and preeclampsia is well documented, suggesting that preeclampsia is a potential risk factor for the development of maternal hypothyroidism. However, the exact role of preeclampsia in gestational hypothyroidism is still obscure. Objective: In this study, we explored the possible mechanisms of the effect of preeclampsia on thyroid function of maternal rats. Methods: Thirty pregnant rats were randomly divided into normal pregnancy control (NOP), preeclampsia (PE), and preeclampsia supplemented with amlodipine besylate (PEAml). NG-Nitro-L-arginine-methyl ester was used to induce preeclamptic symptoms. On gestational day 21, rats were sacrificed, and then, the ultrastructure of the thyroid gland, type 1 iodothyronine deiodinase (Dio1) expression, and serum-free thyroxine (FT4), free triiodothyronine (FT3), and thyroid stimulation hormones (TSH) were assessed. Results: Compared to NOP rats, results of PE rats showed that thyroid follicular cells' ultrastructure was damaged; both hepatic Dio1 mRNA and protein levels were decreased. Interestingly, these changes were ameliorated in PEAml rats. Additionally, FT4, FT3, and TSH levels have no significant differences among groups. Conclusion: These findings indicated that preeclampsia could disrupt synthesis, secretion, and metabolism function of thyroid hormones by damaging thyroid follicular cells and interfering Dio1 expression.


Assuntos
Hipotireoidismo/metabolismo , Iodeto Peroxidase/metabolismo , Fígado/enzimologia , Pré-Eclâmpsia/metabolismo , Glândula Tireoide/ultraestrutura , Hormônios Tireóideos/sangue , Animais , Feminino , Hipotireoidismo/complicações , NG-Nitroarginina Metil Éster/metabolismo , Gravidez , Prenhez , Ratos , Ratos Sprague-Dawley , Fatores de Risco , Tireotropina/sangue , Tiroxina/sangue , Fatores de Tempo , Tri-Iodotironina/sangue
8.
J Anim Sci ; 99(8)2021 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-34240172

RESUMO

Modulation of the immune system is known to be important for successful pregnancy but how immune function might differ between the lymph nodes draining the reproductive tract and peripheral lymph nodes is not well understood. Additionally, if immune system changes in response to the presence of an embryo during early pregnancy, and if this response differs in local versus peripheral immune tissue, has not been well characterized. To address these questions, we examined expression of genes important for immune function using NanoString technology in the ampulla and isthmus of the oviduct, endometrium, lymph nodes draining the reproductive tract (lumbo-aortic and medial iliac) as well as a peripheral lymph node (axillary), the spleen, and circulating immune cells from ewes on day 5 of the estrous cycle or pregnancy. Concentrations of estradiol and progesterone in plasma were also determined. Principal component analysis revealed separation of the local from the peripheral lymph nodes (MANOVA P = 3.245e-08, R2 = 0.3) as well as separation of tissues from pregnant and nonpregnant animals [lymph nodes (MANOVA P = 2.337e-09, R2 = 0.5), reproductive tissues (MANOVA P = 2.417e-14, R2 = 0.47)]. Nine genes were differentially (FDR < 0.10) expressed between lymph node types, with clear difference in expression of these genes between the lumbo-aortic and axillary lymph nodes. Expression of these genes in the medial iliac lymph node was not consistently different to either the axillary or the lumbo-aortic lymph node. Expression of IL10RB was increased (FDR < 0.05) by 24% in the reproductive tissue of the pregnant animals compared to nonpregnant animals. Analysis of gene categories revealed that expression of genes of the T-cell receptor pathway in reproductive tract tissues was associated (P < 0.05) with pregnancy status. In conclusion, assessment of gene expression of reproductive and immune tissue provides evidence for a specialization of the local immune system around the reproductive tract potentially important for successful establishment of pregnancy. Additionally, differences in gene expression patterns in reproductive tissue from pregnant and nonpregnant animals could be discerned as early as day 5 of pregnancy. This was found to be associated with expression of genes important for T-cell function and thus highlights the important role of these cells in early pregnancy.


Assuntos
Prenhez , Progesterona , Animais , Endométrio , Ciclo Estral , Feminino , Expressão Gênica , Sistema Imunitário , Linfonodos , Gravidez , Ovinos
9.
Pol J Vet Sci ; 24(2): 183-190, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-34250776

RESUMO

The aim of this study was to evaluate the possibility to predict outcomes of artificial insemi- nation (AI) in dairy cows based on in-line milk progesterone (P4) concentration. The research was carried out on the herd of loose housing 245 dairy cows of 2-4 lactations, with average milk yielding 11.000 kg per cow. Milk sampling, measuring, and recording of milk P4 concentration was carried out using the Herd Navigator (HN). The grouping was performed according to the following three indices: the first by reproductive condition - pregnant or not pregnant after AI, the second by P4 concentration from day 20 before AI to day 20 after AI, and the third by P4 concentration at AI time. There was a significant difference in P4 concentration in the group of pregnant cows from day 15 to day 9 before AI, and it was by 18.3% higher com- pared to that in the group of non-pregnant cows in the said period (p⟨0.01). The milk P4 concen- trations began to differ mostly from day 10 after AI. At that time, the average P4 concentration in the group of pregnant dairy cows was by 36.8% higher compared to that in the group of non-pregnant cows (p⟨0.01). A statistically significant difference between the ratio of the cows with high, medium, and low P4 concentration on days 20-16 before AI (p⟨0.01) was determined. The highest number of cows with up to 2-3 ng/ml P4 concentration became pregnant at the AI time. In-line milk P4 records captured on day 10-15 before AI can be used to predict the proper for reproduction period. By P4 concentrations on day10 after AI, the ratio of pregnant cows in herd can be assessed.


Assuntos
Bovinos , Inseminação Artificial/veterinária , Leite/química , Prenhez , Progesterona/química , Progesterona/metabolismo , Animais , Feminino , Masculino , Gravidez , Prenhez/fisiologia
10.
Reprod Domest Anim ; 56(10): 1315-1328, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34310786

RESUMO

Infrared thermography is a non-invasive technique which allows to distinguish between pregnant and non-pregnant animals. Detecting accurate body surface temperatures can be challenging due to external factors altering thermograph measurements. This study aimed to determine the associations between the ambient temperature, the hair coat features and the temperatures of mares' abdomens. It compared pregnant and non-pregnant mares throughout 11 months. The research was carried out on 40 Konik Polski mares, which were divided into pregnant and non-pregnant groups. The temperature (Tmax, maximal; Taver, average; Tmin, minimal) of the mares' abdomen was evaluated in two regions of interest: the whole area of the lateral surface of the mares' abdomen (Px1) and the flank area of the lateral surface of mares' abdomen (Px2). During the increasing period, the slopes in the linear regression equation did not differ significantly for ambient (Tamb) and surface temperatures in both groups. In the decreasing period, the slopes did not differ significantly for Tamb and Tmax in the non-pregnant group. They also did not differ for Tamb and Taver in Px1 and Tamb and Tmin in Px1 in both pregnant and non-pregnant groups respectively. Other slopes varied significantly (p < .001). There was no evidence of parallel changes in hair coat features and measured temperatures. The flank area appears more suitable for thermal imaging in pregnant mares due to the seasonal fluctuations in hair coat lengths.


Assuntos
Abdome/diagnóstico por imagem , Cavalos/fisiologia , Testes de Gravidez/veterinária , Prenhez , Termografia/veterinária , Pelo Animal , Animais , Feminino , Gravidez , Testes de Gravidez/métodos , Temperatura , Termografia/métodos
11.
Int J Mol Sci ; 22(13)2021 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-34201586

RESUMO

The molecular mechanism underlying embryonic implantation is vital to understand the correct communications between endometrium and developing conceptus during early stages of pregnancy. This study's objective was to determine molecular changes in the uterine endometrial proteome during the preimplantation and peri-implantation between 9 days (9D), 12 days (12D), and 16 days (16D) of pregnant Polish Large White (PLW) gilts. 2DE-MALDI-TOF/TOF and ClueGOTM approaches were employed to analyse the biological networks and molecular changes in porcine endometrial proteome during maternal recognition of pregnancy. A total of sixteen differentially expressed proteins (DEPs) were identified using 2-DE gels and MALDI-TOF/TOF mass spectrometry. Comparison between 9D and 12D of pregnancy identified APOA1, CAPZB, LDHB, CCT5, ANXA4, CFB, TTR upregulated DEPs, and ANXA5, SMS downregulated DEPs. Comparison between 9D and 16D of pregnancy identified HP, APOA1, ACTB, CCT5, ANXA4, CFB upregulated DEPs and ANXA5, SMS, LDHB, ACTR3, HP, ENO3, OAT downregulated DEPs. However, a comparison between 12D and 16D of pregnancy identified HP, ACTB upregulated DEPs, and CRYM, ANXA4, ANXA5, CAPZB, LDHB, ACTR3, CCT5, ENO3, OAT, TTR down-regulated DEPs. Outcomes of this study revealed key proteins and their interactions with metabolic pathways involved in the recognition and establishment of early pregnancy in PLW gilts.


Assuntos
Implantação do Embrião/fisiologia , Endométrio/metabolismo , Prenhez/metabolismo , Proteínas/metabolismo , Animais , Feminino , Gravidez , Proteínas/análise , Reprodutibilidade dos Testes , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Suínos
12.
Gene ; 801: 145855, 2021 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-34293448

RESUMO

Selecting stably expressed reference genes which are not affected by physiological or pathophysiological conditions is crucial for reliable quantification in gene expression studies. This study examined the expression stability of a panel of twelve reference genes in tissues from the female mouse reproductive axis and the uterus. Gene expression studies were carried out using reverse transcriptase quantitative polymerase chain reaction (RT-qPCR). cDNA was synthesised from RNA extracted from hypothalami, pituitaries, ovaries and uteri of female mice at ages representing weaning, puberty and adulthood as well as pregnancy (13 ± 1 days post-coitus) (n = a minimum of 3 at each age and at pregnancy). The reference genes examined included 18 s, Actb, Atp5b, B2m, Canx, Cyc1, Eif4a2, Gapdh, Rpl13a, Sdha, Ubc and Ywhaz. The RT-qPCR raw data were imported into the qBASE+ software to analyse the expression stability using GeNorm. These data were also subsequently analysed using other software packages (Delta CT, Normfinder, BestKeeper). A comprehensive ranking was conducted considering all stability rankings generated from the different software analyses. B2m and Eif4a2 deviated from the acceptable range for amplification efficiency and therefore were excluded from the further analyses. The stability of the reference genes is influenced by the software used for the analysis with BestKeeper providing markedly different results than the other analyses. GeNorm analysis of tissues taken at different ages but not including pregnant animals, indicated that the expression of the reference genes is tissue specific with the most stable genes being: in the hypothalamus, Canx and Actb; in the pituitary, Sdha and Cyc1; in the ovary, 18s, Sdha and Ubc; and in the uterus, Ywhaz, Cyc1, Atp5b, 18s and Rpl13a. The optimal number of reference genes to be used was determined to be 2 in the first three tissues while in the uterus, the V-score generated by the GeNorm analysis was higher than 0.15 suggesting that 3 or more genes should be used for normalisation. Inclusion of tissues from pregnant mice changed the reference genes identified as being the most stable: Ubc and Sdha were the most stable genes in the hypothalamus, pituitary and the ovary. The addition of pregnant tissue had no effect on the stability of the genes in uterus (Ywhaz, Cyc1, Atp5b, 18s and Rpl13a). Identification of these stable reference genes will be of use to those interested in studying female fertility and researchers should be alert to the effects of pregnancy on reference gene stability. This study also signifies the importance of re-examining reference gene stability if the experimental conditions are changed, as shown with the introduction of pregnancy as a new factor in this research.


Assuntos
Prenhez/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/normas , Animais , Feminino , Hipotálamo/fisiologia , Camundongos Endogâmicos C57BL , Ovário/fisiologia , Hipófise/fisiologia , Gravidez , Padrões de Referência , Reprodutibilidade dos Testes , Útero/fisiologia
13.
Gene ; 801: 145831, 2021 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-34274485

RESUMO

The main strategy for preventing porcine reproductive and respiratory syndrome (PRRS) is vaccination. However, current commercial porcine reproductive and respiratory syndrome virus (PRRSV) vaccines have limited effectiveness and may even cause infections in pigs. The identification of stable molecular markers associated with immune responses to PRRSV vaccination in pigs provides a new approach for PRRS prevention. DNA methylation, the most stable epigenetic molecular marker related to PRRSV vaccination, has not been investigated. In the current research, we used whole genome bisulfite sequencing (WGBS) to investigate DNA methylation in pregnant sows that received PRRSV vaccination and their piglets with high and low PRRSV-specific antibody levels. By performing methylation data analysis and basing on our previous transcriptomic studies, we identified several differentially methylated genes (DMGs) that are involved in the pathways of inflammatory and immune responses. Among the DMGs, ISG15, MX1, SERPINE1, GNG11 and IFIT3 were common hub genes in the two generations. MX1 and GNG11 were located in quantitative trait loci related with PRRSV antibody titer and PRRSV susceptibility, respectively. These results suggest that PRRSV vaccination in sows induces DNA methylation changes in genes and DNA methylation changes occur through intergenerational transmission. The novel DNA methylation markers and target genes observed in our study provide new insights into the molecular mechanisms of immune responses to PRRSV vaccination across two pig generations.


Assuntos
Anticorpos Antivirais/sangue , Metilação de DNA , Síndrome Respiratória e Reprodutiva Suína/genética , Vacinas Virais/imunologia , Animais , Animais Recém-Nascidos/imunologia , Animais Recém-Nascidos/virologia , Anticorpos Antivirais/genética , Feminino , Regulação da Expressão Gênica , Ontologia Genética , Transmissão Vertical de Doenças Infecciosas , Síndrome Respiratória e Reprodutiva Suína/imunologia , Síndrome Respiratória e Reprodutiva Suína/transmissão , Gravidez , Prenhez , Mapas de Interação de Proteínas/genética , Mapas de Interação de Proteínas/imunologia , Locos de Características Quantitativas , Suínos
14.
Int J Mol Sci ; 22(10)2021 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-34070219

RESUMO

Age-associated decline in oocyte quality is one of the dominant factors of low fertility. Aging alters several key processes, such as telomere lengthening, cell senescence, and cellular longevity of granulosa cells surrounding oocyte. To investigate the age-dependent molecular changes, we examined the expression, localization, and correlation of telomerase reverse transcriptase (TERT) and ß-Klotho (KLB) in bovine granulosa cells, oocytes, and early embryos during the aging process. Herein, cumulus-oocyte complexes (COCs) obtained from aged cows (>120 months) via ovum pick-up (OPU) showed reduced expression of ß-Klotho and its co-receptor fibroblast growth factor receptor 1 (FGFR1). TERT plasmid injection into pronuclear zygotes not only markedly enhanced day-8 blastocysts' development competence (39.1 ± 0.8%) compared to the control (31.1 ± 0.5%) and D-galactose (17.9 ± 1.0%) treatment groups but also enhanced KLB and FGFR1 expression. In addition, plasmid-injected zygotes displayed a considerable enhancement in blastocyst quality and implantation potential. Cycloastragenol (CAG), an extract of saponins, stimulates telomerase enzymes and enhances KLB expression and alleviates age-related deterioration in cultured primary bovine granulosa cells. In conclusion, telomerase activation or constitutive expression will increase KLB expression and activate the FGFR1/ß-Klotho pathway in bovine granulosa cells and early embryos, inhibiting age-related malfunctioning.


Assuntos
Blastocisto/metabolismo , Bovinos/embriologia , Bovinos/genética , Proteínas de Membrana/genética , Prenhez/genética , Telomerase/genética , Envelhecimento/genética , Envelhecimento/fisiologia , Animais , Bovinos/fisiologia , Células Cultivadas , Fase de Clivagem do Zigoto/metabolismo , Implantação do Embrião/genética , Implantação do Embrião/fisiologia , Desenvolvimento Embrionário/genética , Desenvolvimento Embrionário/fisiologia , Feminino , Expressão Gênica , Células da Granulosa/metabolismo , Proteínas de Membrana/metabolismo , Oócitos/crescimento & desenvolvimento , Oócitos/metabolismo , Gravidez , Prenhez/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética
15.
Anim Sci J ; 92(1): e13555, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-33938089

RESUMO

Dairy cattle management lacks consideration of fetal breed, the effect of which on fetal growth and nutrition are unclear. We investigated blood parameters in 12 late-pregnant Holstein heifers with similar (Holstein, n = 5) or different (Japanese Black [n = 4] or F1 cross [n = 3]; Holstein × Japanese Black) fetus breeds and in their umbilical cords and calves. Samples were obtained from dams 1 week before calving (-1 week) and immediately after calving, from the umbilical vein at calving, and from calves immediately after birth. Dams with beef fetuses had higher serum glucose levels (-1 week; p < .05) than those with Holstein fetuses. Plasma total amino acid, total essential amino acid, total nonessential amino acid, and other amino acid concentrations were lower in the umbilical veins of dams with calves of the beef breeds than in those of the Holstein breeds (p < .05). Furthermore, serum glucose and plasma amino acid levels were lower in the beef calves than in the Holstein calves (p < .05). Overall, nutrient supply from dams to beef fetuses was lower than that to Holstein fetuses. Our findings may facilitate feeding management of dairy cattle pregnant with beef breeds for appropriate fetal growth and nutrition.


Assuntos
Fenômenos Fisiológicos da Nutrição Animal , Animais Recém-Nascidos/sangue , Bovinos/sangue , Feto/metabolismo , Estado Nutricional , Período Pós-Parto/sangue , Prenhez/sangue , Veias Umbilicais/metabolismo , Aminoácidos/sangue , Animais , Glicemia/metabolismo , Bovinos/embriologia , Bovinos/metabolismo , Feminino , Troca Materno-Fetal , Gravidez
16.
Anim Reprod Sci ; 229: 106758, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33964763

RESUMO

The side of ovulation (left ovary, LO; right ovary, RO) and side of the next ovulation were compared between (1) beginning and end of an interovulatory interval (IOI) and beginning and end of consecutive sets of two and three IOI (n = 900 IOI), (2) beginning and end of the IOI for two and three follicular waves per IOI (n = 1300), and (3) beginning of pregnancy and first postpartum ovulation (n = 793). Pairs of sides of ovulation were designated LL (LO and LO), RR, LR, and RL. The frequency of ovulation pairs for two ends of an IOI was not different from two ends of two or three consecutive IOI indicating that differences between LO and RO were more likely inherent than from factors that developed in each IOI. For each end of an IOI or two consecutive IOI, the least frequency (P < 0.05) was for LL (16 %) with no differences among RR, LR, and RL (28 % for each). Frequencies between ipsilateral (LL, RR) and contralateral (LR, RL) ovulations pairs were not different for two-wave IOI (48 % compared with 52 %) but differed (P < 0.0001) for three-wave IOI (32 % compared with 68 %) and for pregnancy/postpartum (34 % compared with 66 %). In pregnancy/postpartum, each pair was different (P < 0.05) from each other: LL (13 %), RR (21 %), LR (30 %), RL (36 %). The lesser frequency for LL than for any of the others for an IOI, consecutive IOI, and pregnancy/postpartum indicated a ubiquity of the small propensity for LO ovulation.


Assuntos
Bovinos/fisiologia , Ovário/fisiologia , Ovulação/fisiologia , Prenhez , Animais , Feminino , Gravidez , Prenhez/fisiologia
17.
Arq. bras. med. vet. zootec. (Online) ; 73(3): 653-657, May-June 2021. ilus
Artigo em Inglês | ID: biblio-1278359

RESUMO

Squamous metaplasia of the rete ovarii is an ovarian pathologic change characterized by replacement of the normal single layered cuboidal epithelium of the rete ovarii by a stratified squamous keratinized epithelium. Uterus and ovaries from a local slaughterhouse pregnant crossbreed cow were evaluated through ultrasound, macroscopically and histologically. Grossly, there were multiple cysts in both ovaries, which were histologically characterized as rete ovarii cysts with squamous metaplasia and intraluminal accumulation of keratinized material. Squamous metaplasia of the rete ovarii has been previously reported in cows, however this is the first report of this condition in a pregnant animal, demonstrating that this ovarian change is compatible with pregnancy.(AU)


A metaplasia escamosa da rete ovarii é uma patologia ovariana caracterizada pela substituição do epitélio simples cuboidal normal da rete ovarii por um epitélio estratificado escamoso queratinizado. Útero e ovários de uma vaca mestiça gestante, proveniente de abatedouro, foram avaliados por ultrassonografia, macroscopia e histologia. Verificaram-se vários cistos em ambos os ovários, histologicamente caracterizados como cistos de rete ovarii com metaplasia escamosa, com acúmulo intraluminal de material queratinizado. Metaplasia escamosa da rete ovarii foi relatada anteriormente em vacas, porém este é o primeiro relato em que essa alteração ovariana é compatível com manutenção da ciclicidde ovariana e gestação na vaca.(AU)


Assuntos
Animais , Feminino , Gravidez , Bovinos , Ovário/patologia , Teratoma/veterinária , Prenhez/fisiologia , Ciclo Estral/fisiologia , Cisto Epidérmico/veterinária , Epitélio/patologia , Metaplasia/veterinária
18.
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
19.
J Am Heart Assoc ; 10(8): e019922, 2021 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-33843249

RESUMO

Background Antenatal intrauterine fetal hypoxia is a common pregnancy complication that has profound adverse effects on an individual's vascular health later in life. Pulmonary arteries are sensitive to hypoxia, but adverse effects of antenatal hypoxia on pulmonary vasoreactivities in the offspring remain unknown. This study aimed to determine the effects and related mechanisms of antenatal hypoxia on pulmonary artery functions in adult male offspring. Methods and Results Pregnant Sprague-Dawley rats were housed in a normoxic or hypoxic (10.5% O2) chamber from gestation days 10 to 20. Male offspring were euthanized at 16 weeks old (adult offspring). Pulmonary arteries were collected for vascular function, electrophysiology, target gene expression, and promoter methylation studies. In pulmonary artery rings, contractions to serotonin hydrochloride, angiotensin II, or phenylephrine were reduced in the antenatal hypoxic offspring, which resulted from inactivated L-type Ca2+ channels. In pulmonary artery smooth muscle cells, the basal whole-cell Ca2+ currents, as well as vasoconstrictor-induced Ca2+ transients were significantly reduced in antenatal hypoxic offspring. In addition, increased promoter methylations within L-type Ca2+ channel subunit alpha1 C were compatible with its reduced expressions. Conclusions This study indicated that antenatal hypoxia programmed long-lasting vascular hypocontractility in the male offspring that is linked to decreases of L-type Ca2+ channel subunit alpha1 C in the pulmonary arteries. Antenatal hypoxia resulted in pulmonary artery adverse outcomes in postnatal offspring, was strongly associated with reprogrammed L-type Ca2+ channel subunit alpha1 C expression via a DNA methylation-mediated epigenetic mechanism, advancing understanding toward the effect of antenatal hypoxia in early life on long-term vascular health.


Assuntos
Canais de Cálcio Tipo L/genética , Regulação para Baixo , Regulação da Expressão Gênica , Hipóxia/genética , Prenhez , Artéria Pulmonar/fisiopatologia , Vasoconstrição/fisiologia , Animais , Pressão Sanguínea/fisiologia , Canais de Cálcio Tipo L/biossíntese , Células Cultivadas , Feminino , Hipóxia/metabolismo , Hipóxia/fisiopatologia , Masculino , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/patologia , Gravidez , Artéria Pulmonar/metabolismo , Artéria Pulmonar/patologia , RNA/genética , Ratos , Ratos Sprague-Dawley
20.
Sci Rep ; 11(1): 8698, 2021 04 22.
Artigo em Inglês | MEDLINE | ID: mdl-33888798

RESUMO

Visfatin appears to be an energy sensor involved in the regulation of female fertility, which creates a hormonal link integrating the control of energy homeostasis and reproduction. This study evaluates the expression levels of visfatin gene and protein in selected areas of the porcine hypothalamus responsible for gonadotropin-releasing hormone synthesis: the mediobasal hypothalamus (MBH) and preoptic area (POA), and visfatin concentrations in the blood plasma. The tissue samples were harvested from gilts on days 2-3, 10-12, 14-16, and 17-19 of the estrous cycle, and on days 10-11, 12-13, 15-16, 27-28 of pregnancy. Visfatin was localized in the cytoplasm and nucleus of cells creating both studied hypothalamic structures. The study demonstrated that visfatin gene and protein expression in MBH and POA depends on hormonal status related to the phase of the estrous cycle or early pregnancy. Blood plasma concentrations of visfatin during the estrous cycle were higher on days 2-3 in relation to other studied phases of the cycle, while during early pregnancy, the highest visfatin contents were observed on days 12-13. This study demonstrated visfatin expression in the porcine hypothalamus and its dependence on the hormonal milieu related to the estrous cycle and early pregnancy.


Assuntos
Estro , Hipotálamo/metabolismo , Nicotinamida Fosforribosiltransferase/sangue , Prenhez/sangue , Animais , Feminino , Nicotinamida Fosforribosiltransferase/genética , Nicotinamida Fosforribosiltransferase/metabolismo , Gravidez
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