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2.
Theriogenology ; 141: 48-53, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31518728

RESUMO

An efficient system to collect large numbers of vital zygotes is a pre-requisite for application of zygote genome-editing technology, including development of efficient models for xenotransplantation using pigs. Owing to the sub-optimal in vitro production of zygotes in pigs, efficient collection of in vivo developed zygotes is required. Timing of ovulation is a key factor to sustain efficiency since the interval between pronuclear formation and the first division is very short in pigs. The weaning-to-estrus interval can, due to its inverse relation with length of estrus and time of ovulation, interfere with ovulation and make it asynchronous, which reduces the probability of obtaining zygotes. This retrospective study compared the effects of three weaning-to-estrus intervals of 3, 4 or 5 days on zygote collection efficiency in a total of 17 trials over a 3-year period including 223 sows. Donor sows in groups of 10-15 animals were super-ovulated with eCG 24 h after weaning and those in estrus at 48-72 h post-eCG were immediately treated with hCG, followed by insemination 6 and 24 h thereafter. Collected structures during laparotomy on Day 2 (Day 0: onset of estrus) were morphologically evaluated and only those with a single cell and two visible polar bodies were considered as zygotes. Zygotes were injected with CRISPR-Cas9 editor mixture and cultured for 6 days to evaluate their developmental ability against non-injected control zygotes. Of all recovered structures (N = 5,468), 67.4%, 30.8% and 1.8% were zygotes, 2-cell embryos and oocytes-degenerated embryos, respectively. The different weaning-to-estrus intervals did not affect either the percentages of collected zygotes (range: 64.1%-70.0%) or the percentages of sows with zygotes at collection time (range: 69.0%-73.3%). The weaning-to-estrus intervals did not affect the in vitro developmental ability of zygotes. After 24 h of culture, 78.1 ±â€¯2.0% and 95.1 ±â€¯0.6 (P < 0.05) of injected (N = 2,345) and non-injected (N = 335) zygotes, respectively, developed to 2-to-4-cell embryo stage. The total efficiency of the system was 64.1 ±â€¯2.2% and 85.8 ±â€¯1.5% (P < 0.05) for injected and non-injected zygotes, respectively. In conclusion, the results indicate that neither the efficiency of collecting in vivo derived porcine zygotes from superovulated sows nor the zygote ability to develop to blastocyst after cytoplasmic genome-editing injection were affected by a weaning-to-estrus interval between 3-to-5 days.


Assuntos
Gonadotropina Coriônica/farmacologia , Oócitos , Suínos/fisiologia , Animais , Técnicas de Cultura Embrionária/veterinária , Embrião de Mamíferos , Inseminação Artificial/veterinária , Estudos Retrospectivos , Superovulação/efeitos dos fármacos , Suínos/embriologia , Fatores de Tempo , Coleta de Tecidos e Órgãos
3.
Theriogenology ; 141: 168-172, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31542520

RESUMO

This study evaluated embryo production after superovulation (SO) with a reduced number of FSH applications and increased eCG dose in 26 Bos taurus × Bos indicus donors. On Day 0, donors received an intravaginal device (CIDR) with 1.9 g of progesterone plus 2.5 mg of estradiol benzoate and 50 mg of progesterone via IM. On Day 4, donors were randomly allotted to one of three SO treatments: 1) 455 IU of Folltropin +400 IU of eCG (n = 9), 2) 350 IU of Folltropin +600 IU of eCG (n = 9), and 3) 500 IU of Pluset + 600 IU of eCG (n = 8). In treatment 455 IU of Folltropin +400 IU of eCG, donors received eight IM Folltropin injections in decreasing dose 12 h apart from Day 4 to Day 7. On Day 6, at the same time as the Folltropin, donors received via IM 25 mg of dinoprost tromethamine (PGF2a). On Day 7, the CIDR was removed, and together with the Folltropin, donors received 200 IU of eCG via IM. In treatment 350 IU of Folltropin +600 IU of eCG, donors received four IM Folltropin injections in decreasing dose 12 h apart on Days 4 and 5. On Day 6, donors received via IM 600 IU of eCG in the morning and two doses of 25 mg of PGF2a 12 h apart. On Day 7, the CIDR was removed. Donors from treatment 500 IU of Pluset +600 IU of eCG received four IM Pluset injections in decreasing dose 12 h apart on Days 4 and 5. On Day 6, donors received via IM 600 IU of eCG in the morning and two doses of 25 mg of PGF2a 12 h apart. On Day 7, the CIDR was removed. In the morning of Day 8, donors from the three treatments received 0.25 mg of GnRH via IM. Artificial insemination was performed on Day 8 (pm) and Day 9 (am). Embryos were collected on Day 15. Variables evaluated were number of CL before embryo collection, number of structures recovered, transferable embryos, degenerate embryos and unfertilized oocytes, recovery rate, and viability rate. There was no difference in any variable among treatments (P > 0.05). In conclusion, replacement of four Folltropin or Pluset injections from a conventional eight FSH-injection SO protocol, by a single injection of 600 IU of eCG, is a good alternative to reduce donor handling without decreasing yield of transferable embryos.


Assuntos
Bovinos/embriologia , Gonadotropina Coriônica/farmacologia , Hormônio Foliculoestimulante/farmacologia , Superovulação/efeitos dos fármacos , Animais , Gonadotropina Coriônica/administração & dosagem , Técnicas de Cultura Embrionária , Feminino , Fertilização In Vitro , Hormônio Foliculoestimulante/administração & dosagem , Doadores de Tecidos
4.
Anim Reprod Sci ; 208: 106116, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31405452

RESUMO

Cryptorchidism, the failure of one or both testes to descend to an extra-abdominal location during mammalian development, is a common reproductive malady that often requires surgical intervention to remedy. Leydig cells are responsible for producing insulin-like peptide 3 (INSL3), a peptide hormone that is essential for normal testicular descent. The insl3 promoter in Leydig cells can be activated by cAMP through the transcription factor Nur77, which also regulates the promoters of the steroidogenic enzymes, cyp17 and 3ß-hsd. While the mechanism of LH action on testosterone production is well characterized, the effect of LH on insl3 abundance has not been described. The MA-10 Leydig cell line was used to test the hypothesis that abundance of insl3 mRNA is increased by LH/CG via the cAMP pathway. Cells treated with hCG had a transient robust increase in abundance of nur77 mRNA, while insl3 mRNA abundance remained unchanged. Further, while cAMP addition increased nur77 mRNA abundance, it failed to affect insl3 mRNA. Inhibition of LH-receptor-linked signal transduction pathways in the presence of hCG implicated multiple signaling networks in the regulation of both the insl3 and nur77 genes. Treatment with hCG and cAMP also increased abundance of cyp17 mRNA but not 3ß-hsd. Abundance of insl3 mRNA was not affected by hCG, testosterone or the combination of hCG and testosterone. Collectively, these results provide support for the constitutive regulation of insl3 mRNA abundance in the MA-10 Leydig cell line rather than acute regulation by LH/CG and cAMP.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Insulina/metabolismo , Células Intersticiais do Testículo/fisiologia , Proteínas/metabolismo , Animais , Linhagem Celular , Gonadotropina Coriônica/farmacologia , Flavonoides/farmacologia , Regulação da Expressão Gênica/fisiologia , Insulina/genética , Isoquinolinas/farmacologia , Masculino , Camundongos , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/genética , Membro 1 do Grupo A da Subfamília 4 de Receptores Nucleares/metabolismo , Perileno/análogos & derivados , Perileno/farmacologia , Proteínas/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Sulfonamidas/farmacologia , Testosterona/farmacologia , Wortmanina/farmacologia
5.
Anim Reprod Sci ; 208: 106108, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31405462

RESUMO

The objective of this study was to evaluate the effects of plasma progesterone (P4) concentrations during eCG-ovarian follicular superstimulatory treatment performed in early luteal phase and estradiol concentrations during peri-ovulatory period on ovarian response, number and embryo quality. On Day -2, females (n = 75) having a follicle ≥7 mm were treated with GnRH to induce ovulation. On Day 0, females that had ovulations (n = 54) were treated with 1000 IU eCG and were assigned to one of two treatments: (1) intravaginal device (ID) containing 0.5 g P4 (P4 group) and (2) no ID (Control group). On Day 5, females were administered PGF2α and the ID was removed. On Day 7 and 8, females were mated and embryo recovery was performed 7 or 8 days later. Blood samples were collected from Day 0 to 9. Number (± SD) of follicles ≥7 mm on day of mating was greater (P =  0.04) in the control (9.7 ± 4.2) than P4-treated (6.7 ±â€¯4.9) group; number of corpora lutea did not differ (5.5 ±â€¯3.1 and 5.2 ±â€¯3.4 respectively). Ovulation rate was greater (P <  0.01) in the P4-group (77.4%; 130/168) than control group (53.3%; 135/253). Number of embryos with an excellent grade (grade 1) tended to be greater (P =  0.07) in the P4-group (82.4%; 42/51) than control group (65.4%; 36/55). It was concluded that supplementation with exogenous P4 during eCG treatment in early luteal phase inhibits excessive follicular growth, increases ovulation rate and improves embryo quality.


Assuntos
Camelídeos Americanos/fisiologia , Gonadotropina Coriônica/farmacologia , Corpo Lúteo/fisiologia , Progesterona/farmacologia , Superovulação/efeitos dos fármacos , Animais , Camelídeos Americanos/sangue , Gonadotropina Coriônica/administração & dosagem , Corpo Lúteo/efeitos dos fármacos , Feminino , Progestinas/farmacologia , Substâncias para o Controle da Reprodução/farmacologia
6.
Anim Reprod Sci ; 208: 106112, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31405481

RESUMO

The establishment of protocols for the control of the ovarian function of collared peccaries is recommended for the development of assisted reproductive techniques. The goals were to (1) compare a gonadotropin combination with prostaglandin analogue to synchronize timing of onset of estrus among animals, and (2) elucidate the effects of the most desirable protocol for performing an artificial insemination study and macroscopic evaluation of the ovaries. Three of five females treated with a double administration of 120 µg prostaglandin (cloprostenol) at a 9-day interval expressed symptoms of estrus 9 days after the second injection. One female presented estrus after 6 days, whereas other did not respond to the treatment. All females (5/5) treated with a single dose containing 400 IU eCG and 200 IU hCG manifested estrus 6 days after the hormone injection. In a second experiment, ten females that were estrous synchronized using eCG/hCG, were artificially inseminated with fresh semen and monitored for pregnancy every 30 days. Although there was no detection of fetuses by ultrasonic examination, seven females (7/10) had greater than basal progesterone values for 60 days after the treatments were imposed. Ovaries from two females treated with eCG/hCG were collected 6 days post-injection. There was confirmation of an ovarian stimulation as a result of the presence of 88 and 25 antral follicles, as well as three and eight hemorrhagic structures in ovaries of each female, respectively. It, therefore, is proposed that eCG/hCG can be used as an effective treatment for estrous synchronization in collared peccaries.


Assuntos
Artiodáctilos/fisiologia , Gonadotropina Coriônica/farmacologia , Sincronização do Estro/métodos , Animais , Gonadotropina Coriônica/administração & dosagem , Cloprostenol/farmacologia , Relação Dose-Resposta a Droga , Feminino
7.
J Steroid Biochem Mol Biol ; 194: 105460, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31470110

RESUMO

The bile acid receptor Farnesoid-X-Receptor alpha (FXRα), a member of the nuclear receptor superfamily, is well known for its roles in the enterohepatic tract. In addition, FXRα regulates testicular physiology through the control of both endocrine and exocrine functions. The endocrine function of the Leydig cells is mainly controlled by the hypothalamo-pituitary axis viaLH/chorionic gonadotropin (CG). If FXRα was demonstrated to control the expression of the Lhcgr gene, encoding the LH receptor; the impact of the LH/CG signaling on the Fxrα expression has not been defined so far. Here, we demonstrate that hCG increases the Fxrα gene expression through the protein kinase-A signaling pathway. Fxrα is then involved in a negative feedback of steroid synthesis. These data improve our knowledge of the local control of the testicular steroidogenesis with the identification of the link between the hypothalamo-pituitary axis and the FXRα signaling pathway.


Assuntos
Gonadotropina Coriônica/farmacologia , Receptores Citoplasmáticos e Nucleares/genética , Testículo/efeitos dos fármacos , Animais , Linhagem Celular , Masculino , Camundongos Endogâmicos C57BL , Fosfoproteínas/genética , Progesterona/metabolismo , Receptores do LH/genética , Transdução de Sinais/efeitos dos fármacos , Testículo/metabolismo , Testosterona/sangue , Testosterona/metabolismo
8.
Theriogenology ; 138: 39-46, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31284220

RESUMO

Control of ovarian function in cheetahs is sub-optimal, which currently limits the integration of assisted reproductive techniques into the genetic management of that endangered species. The objective of this study was to determine the effect of preemptive progestin treatment on the quality of ovarian responses after exogenous gonadotropin stimulation in cheetahs. Adult females received either 1) 200 IU equine chorionic gonadotropin (eCG) followed with 3,000 IU porcine luteinizing hormone (pLH) (intramuscular route) (n = 5; control group) or 2) similar eCG/pLH administration preceded by a 7-day treatment with oral progestin (0.1 mg/kg altrenogest; ALT group; n = 7). At 42 h post-pLH administration, a series of metrics was assessed via laparoscopy (number of follicles ≥ 2 mm, number of corpora lutea, oviduct and uterine cornua diameter and overall vascularization). Concentrations of fecal estradiol, progesterone and glucocorticoid metabolites (FEM, FPM, and FGM, respectively) were measured by enzyme immunoassay for 3 wk before ALT treatment (Period 1), 7 d during ovarian suppression period (Period 2), throughout eCG/LH treatment and laparoscopy (Period 3), and 6 wk following laparoscopy (Period 4). Overall, nine out of 12 cheetahs (4/5 in control and 5/7 ALT group) had freshly-formed corpora lutea at the time of laparoscopy. Mean follicle and corpora lutea numbers in the control versus ALT group were not different (P > 0.05). Overall measurements and vascularization scores also did not differ (P > 0.05) among groups. FEM average concentrations increased (P ≤ 0.05) in response to eCG for the ALT-treated females between Periods 2 and 3 and were sustained during Period 4. However, FEM average concentrations did not vary (P > 0.05) for control females throughout Periods 1-4. Post-ovulatory FPM average concentrations (Period 4) did not differ (P > 0.05) between the ALT-treated females and controls. FPM average concentration from both groups increased in Period 4 compared to Periods 1-3 (P ≤ 0.05). Females receiving the ALT treatment also had lower (P ≤ 0.05) FGM metabolite average concentrations than control females during ovarian suppression (suggesting adrenal suppression). Collective results suggest that ovarian response to gonadotropin treatment in the cheetah was improved following oral progestin administration due to the normative increase in estradiol following stimulation for these females compared with control. This treatment should lead to more effective timed assisted reproduction procedures for this species.


Assuntos
Acinonyx/fisiologia , Sincronização do Estro/métodos , Gonadotropinas Equinas/farmacologia , Ovário/efeitos dos fármacos , Indução da Ovulação , Progestinas/administração & dosagem , Administração Oral , Animais , Gonadotropina Coriônica/farmacologia , Estradiol/análise , Estradiol/metabolismo , Fezes/química , Feminino , Glucocorticoides/análise , Glucocorticoides/metabolismo , Hormônio Luteinizante/farmacologia , Ovário/fisiologia , Indução da Ovulação/métodos , Indução da Ovulação/veterinária , Progesterona/análise , Progesterona/metabolismo , Progestinas/farmacologia , Resultado do Tratamento , Acetato de Trembolona/administração & dosagem , Acetato de Trembolona/análogos & derivados , Acetato de Trembolona/farmacologia
9.
Medicine (Baltimore) ; 98(27): e16213, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31277129

RESUMO

RATIONALE: The borderline form of empty follicle syndrome (EFS) is a phenomenon where only a few mature or immature oocytes are retrieved despite adequate response to controlled ovarian hyperstimulation (COH). It is a rare phenomenon with an unclear underlying mechanism, and there is currently no effective treatment. PATIENT CONCERNS: The patient received 3 assisted reproductive technology cycles, and although her follicular development and estrogen levels were normal during COH, the outcome with respect to the oocytes obtained was unsatisfactory. DIAGNOSES: Borderline form of EFS. INTERVENTIONS: In the context of undergoing GnRH-antagonist protocol, we implemented a double-trigger with human chorionic gonadotropin (hCG) after 6 hours of gonadotropin-releasing hormone agonist (GnRH-a) administration. OUTCOMES: Eleven oocytes were obtained (M I × 3, M II × 8), which underwent in vitro fertilization (IVF). After 18 hours, 7 oocytes showed normal fertilization, with 2 embryos formed 72 hours later (embryo rating, 6C II × 1, 9C II × 1); the embryos were then frozen. LESSONS: Oocyte maturation and ovulation are time-dependent processes, and that different patients require different lengths/intervals of time for treatment. Therefore, the borderline form of EFS, in general, may be treatable, and our novel trigger method provides a new treatment option for such patients in the future.


Assuntos
Gonadotropina Coriônica/farmacologia , Fármacos para a Fertilidade Feminina/farmacologia , Hormônio Liberador de Gonadotropina/agonistas , Síndrome de Hiperestimulação Ovariana/terapia , Indução da Ovulação/métodos , Adulto , Feminino , Fertilização In Vitro , Humanos , Recuperação de Oócitos , Gravidez , Substâncias para o Controle da Reprodução/farmacologia
10.
Reprod Biol ; 19(2): 210-217, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31262644

RESUMO

Clinical outcomes of fresh embryo transfer in non-hCG triggered in vitro maturation (IVM) cycles are inferior compared to vitrified-warmed embryo transfer. This is a prospective observational pilot study in a consecutive cohort of 31 polycystic ovary syndrome (PCOS) patients and 37 normo-ovulatory egg donors who underwent IVM without fresh embryo transfer between July 2009 and June 2014. All subjects received 150 IU of highly purified menotropin (HP-hMG) daily for three days. On cycle day 6, all patients started transdermal oestradiol (E2) at a daily dose of 9 mg. There was no human chorionic gonadotropin (hCG) trigger before oocyte retrieval (OR). Vaginal micronized progesterone was commenced on the evening after OR, at a daily dose of 600 mg. Additional luteal phase support (LPS) was administered as follows: Group A: no additional LPS; Group B: 1500 IU of hCG administered 4 h after OR and Group C: 5000 IU of hCG administered 4 h after OR + an additional injection of 5000 IU of hCG 1 day before endometrial biopsy. Endometrial biopsy for histology and immunohistochemistry (IHC) was performed on day 5 or 6 after OR. Instead of being downregulated, both PR-B and ERα in endometrial glands and stroma were moderately to strongly expressed in all three protocols, suggesting that the mid-luteal histological signature of endometrial receptivity is deficient in a non-hCG-triggered IVM cycle. Poor clinical outcomes after fresh embryo transfer following IVM are probably related to inappropriate endometrial development which may be linked to the short follicular phase of IVM cycles.


Assuntos
Gonadotropina Coriônica/farmacologia , Endométrio/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Síndrome do Ovário Policístico/metabolismo , Receptores de Esteroides/metabolismo , Adulto , Gonadotropina Coriônica/administração & dosagem , Estudos de Coortes , Estradiol/administração & dosagem , Estradiol/farmacologia , Feminino , Humanos , Técnicas de Maturação in Vitro de Oócitos , Projetos Piloto , Progesterona/administração & dosagem , Progesterona/farmacologia , Estudos Prospectivos , Receptores de Esteroides/genética , Adulto Jovem
11.
Theriogenology ; 138: 145-151, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31352176

RESUMO

Although thyroid hormone (TH) plays important roles in regulating ovarian development, the mechanism are still unclear. Cytochrome P450 lanosterol 14α-demethylase (CYP51) is a key enzyme in sterols and steroids biosynthesis that involved in folliculogenesis and oocyte maturation, which is regulated by follicle stimulating hormone (FSH). However, the effect of TH on CYP51 expression in ovarian cells is unclear. The objective of this study was to determine the effects of TH on CYP51 in rat ovary. Hypothyroidism rats were induced by 6-propyl-2-thiouracil (PTU), genes expressions in ovary were analyzed by Western blot or qRT-PCR. The data showed that CYP51 was significantly decreased in hypothyroidism, which was accompanied by the down-regulation of mRNA level. Meanwhile, similar tendency was also showed in FSHR expression in hypothyroidism. To evaluate the effect of the gonadotropin on CYP51 and FSHR expression in ovarian cells in vivo, hypo rats were injected subcutaneously with equine chorionic gonadotropin (eCG) respectively. The results showed that eCG reversed CYP51 and FSHR expression in hypo group. Moreover, FSH-induced CYP51 expression was meditated by FSHR. In addition, serum concentration of FSH and E2 were also decreased in hypothyroidism, and E2 was up-regulated by eCG treatment. These results indicate that hypothyroidism changes CYP51 and FSHR expression in ovary, which are regulated by gonadotropin. Moreover, genes changes in ovary are at least partially attributed to steroids biosynthesis.


Assuntos
Hipotireoidismo/genética , Ovário/metabolismo , Receptores do FSH/genética , Esterol 14-Desmetilase/genética , Animais , Células Cultivadas , Gonadotropina Coriônica/farmacologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Hormônios Esteroides Gonadais/biossíntese , Gonadotropinas Equinas/farmacologia , Hipotireoidismo/metabolismo , Hipotireoidismo/patologia , Ovário/efeitos dos fármacos , Ovário/patologia , Ratos , Ratos Sprague-Dawley , Receptores do FSH/metabolismo , Esterol 14-Desmetilase/metabolismo , Hormônios Tireóideos/farmacologia
12.
Bull Exp Biol Med ; 167(1): 57-61, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31177451

RESUMO

We studied the effect of estriol, chorionic gonadotropin, oncostatin M, and hormone-cytokine combinations on the expression of recombinase RAG-1 in T-regulatory (Treg) and T helper 17 (Th17) lymphocytes. It was found that estriol in a concentration corresponding to the first trimester of pregnancy increased the level of Treg (CD4+FoxP3+) cells and suppressed the formation of Th17 (CD4+RORC+) lymphocytes. This effect was nor observed after individual administration of chorionic gonadotropin and oncostatin M, but some combinations of the studied hormones with oncostatin M increased the percentage of CD4+FOXP3+ cells. In the presence of oncostatin M, the studied hormones enhanced the expression of RAG-1 in CD4+FoxP3+ cells, but not in CD4+RORC+ cells, thereby initiating of Treg T-cell receptor (TCR) revision. The mechanisms of hormone cytokine control of induction of the immune tolerance during pregnancy are discussed.


Assuntos
Gonadotropina Coriônica/farmacologia , Estriol/farmacologia , Oncostatina M/farmacologia , Recombinases/metabolismo , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/enzimologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/metabolismo , Células Cultivadas , Feminino , Fatores de Transcrição Forkhead/metabolismo , Humanos
13.
Med Sci Monit ; 25: 4377-4383, 2019 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-31187785

RESUMO

BACKGROUND Patients with endometriosis (EMs) are routinely advised to take GnRH-a for 3-6 months to improve the internal reproductive environment, but this may not be necessary. MATERIAL AND METHODS This retrospective study examined the effects of in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) with shortened (n=311) or conventional (n=213) long-term pituitary downregulation in EMs patients between January 2013 and July 2017. RESULTS The 2 groups showed no significant differences in gonadotropin (Gn) dose, number of oocytes retrieved, or miscarriage rate. Follicle-stimulating hormone (FSH), luteinizing hormone (LH), and estradiol (E2) levels on the initiation day and the LH level on human chorionic gonadotropin (hCG) day (1.22±1.39 vs. 0.74±0.55 P=0.0026) were higher in the study group than in the control group. The cumulative live birth rates in the second cycle were 69.13% in the study group (95% confidence interval (CI), 64-74.27%) vs. 68.54% in the control group (95% CI, 62.31-74.78%, P=0.88, respectively). CONCLUSIONS This study showed that the shortened regimen and the ultralong regimen did not produce different pregnancy outcomes after ART, and the single-application, long-term GnRH-a protocol may serve as a cost-effective and safe treatment protocol for EMs patients.


Assuntos
Fertilização In Vitro/métodos , Hormônio Liberador de Gonadotropina/farmacologia , Injeções de Esperma Intracitoplásmicas/métodos , Aborto Espontâneo , Adulto , Gonadotropina Coriônica/farmacologia , Regulação para Baixo , Transferência Embrionária , Endometriose/complicações , Feminino , Humanos , Recuperação de Oócitos/métodos , Oócitos/efeitos dos fármacos , Indução da Ovulação , Hipófise/efeitos dos fármacos , Hipófise/patologia , Gravidez , Resultado da Gravidez , Taxa de Gravidez , Resultado do Tratamento
14.
Theriogenology ; 135: 1-6, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31185424

RESUMO

Superovulation technique is important to improve the efficiency of oocyte and animal production and reduce the number of oocyte donors. Previously, we have reported that the coadministration of inhibin antiserum (IAS) and equine chorionic gonadotropin (eCG) results in the production of >100 oocytes in a 4-week-old female C57BL/6 mice. It is well established that superovulation depends on the age of the female mice. However, detailed data regarding the ovulation of juvenile, mature, and aged female mice following the administration of IAS and eCG as well as the performance of reproductive technologies using oocytes have not yet been investigated. In the present study, we examined the effect of the age of female mice (3-50 weeks old) on the number of ovulated oocytes via the coadministration of IAS and eCG or eCG alone. Treatment with IAS plus eCG produced the maximum number of oocytes at 4 weeks of age. Moreover, IAS plus eCG produced more oocytes than eCG alone in mice aged between 3 and 5 weeks or 7 and 30 weeks. The fertilization and birth rates were similar between the two treatments at any age. Moreover, after vitrifying and warming the embryos, the survival and birth rates of two-cell embryos were similar between the two treatments. Subsequently, we examined the optimal ages of female mice (between 24 and 34 days) to obtain a high and stable number of oocytes. In mice aged between 24 and 32 days, IAS plus eCG induced the production of more eggs than eCG alone. Notably, the coadministration of IAS and eCG in mice aged between 25 and 31 days resulted in stable ovulation and high number of oocytes. Using the tip of the optimal female aged between 25 and 31 days old, we demonstrated an efficient production of embryos and offspring between homozygous knockout males and few females aged 26-28 days via in-vitro fertilization and embryo transfer. In summary, the coadministration of IAS and eCG resulted in a higher number of oocytes in juvenile, mature, and aged female mice. This treatment may be useful for the efficient production of homozygous mutant mice from a limited number of female mice.


Assuntos
Gonadotropina Coriônica/farmacologia , Soros Imunes/farmacologia , Inibinas/antagonistas & inibidores , Superovulação/efeitos dos fármacos , Envelhecimento , Animais , Gonadotropina Coriônica/administração & dosagem , Criopreservação , Quimioterapia Combinada , Técnicas de Cultura Embrionária , Feminino , Fertilização In Vitro , Soros Imunes/administração & dosagem , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Oócitos/fisiologia
15.
Reprod Biol ; 19(2): 173-178, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31151753

RESUMO

The tumor suppressor gene KCTD11 plays a critical role in cell proliferation, differentiation and invasion. The current study investigated the regulation and the spatiotemporal expression pattern of Kctd11 in the rat ovary during the periovulatory period. Ovaries, granulosa cells, or theca-interstitial cells were collected at various times after hCG administration using an established gonadotropin-primed immature rat model that induces follicular development and ovulation. Real-time quantitative PCR analysis revealed that mRNA for Kctd11 was significantly induced both in theca-intersititial and granulosa cells after hCG treatment although their temporal expression patterns differed. In situ hybridization analysis demonstrated that Kctd11 mRNA expression was induced in theca-intersititial cells at 6 h after hCG, and the expression remained elevated until 12 h after hCG. Kctd11 mRNA was stimulated in granulosa cells at 6 h and reached the highest expression at 12 h. There was negligible Kctd11 mRNA signal observed in newly forming corpora lutea. In addition, the data indicate that both the protein kinase A and the protein kinase C pathway regulate the expression of Kctd11 mRNA in granulosa cells. Either forskolin or phorbol 12 myristate 13-acetate can mimic hCG induction of Kctd11 expression. Furthermore, the stimulation of Kctd11 by hCG requires new protein synthesis. Inhibition of progesterone action and the EGF pathway blocked Kctd11 mRNA expression, whereas inhibition of prostaglandin synthesis had no effect. Our finding suggest that the induction of the Kctd11 may be important for theca and granulosa cell differentiation into luteal cells.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Ovário/metabolismo , Ovulação/fisiologia , Transferases/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Animais , Proteínas de Ciclo Celular/genética , Gonadotropina Coriônica/farmacologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Células da Granulosa/fisiologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Células Tecais/fisiologia , Transferases/genética , Proteínas Supressoras de Tumor/genética
16.
Reprod Biol ; 19(2): 179-188, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31151754

RESUMO

Corpus luteum (CL) is an endocrine tissue involved in regulation of reproductive cycle and early pregnancy establishment. In the present study DEAD-box helicase-5 (Ddx5), a member of the DEAD box family of RNA helicases was investigated for its expression, regulation and function in CL of Wistar rats. Ddx5 was expressed in adult rat CL. Primary cell culture from supra-ovulated ovaries were established for in vitro studies. Addition of luteinizing hormone (LH; 100 ng/ml), a luteotrophic factor in primary cell culture, decreased Ddx5 RNA expression (foldchange:0.6 ±â€¯0.075) while prostaglandin alpha (PGF2α; 1µM), a luteolytic factor caused an increase (foldchange:2.4 ±â€¯0.4) compared to control group. Under in vivo conditions, the administration of PGF2α or gonadotropin-releasing hormone antagonist; cetrorelix (CET) caused luteolysis as well as an increase in the protein level of Ddx5 (foldchange:1.9 ±â€¯0.27 and 1.4 ±â€¯0.09 viz.; p < 0.05) in CL of adult rats. LH was administered post CET treatment which suppressed Ddx5 protein expression (foldchange:0.8 ±â€¯0.16; p < 0.05) compared to CET treated group. Further, it was observed that the expression of Ddx5 was upregulated (foldchange:1.5 ±â€¯0.23; p < 0.05) in CL during late pregnancy compared to mid pregnancy concomitant to luteolysis in adult rats. Overall, the results suggest for the first time that Ddx5 is expressed in rat CL and regulated by luteolytic and luteotrophic factors in an inverse fashion. Further, the data significantly correlates ddx5 expression to CL regression suggesting involvement of ddx5 in luteolysis. These results suggest a significant role of Ddx5 in female reproduction biology and warrant in depth examination of the function of Ddx5 in CL.


Assuntos
Cloprostenol/farmacologia , Corpo Lúteo/metabolismo , RNA Helicases DEAD-box/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Hormônio Liberador de Gonadotropina/análogos & derivados , Animais , Gonadotropina Coriônica/farmacologia , RNA Helicases DEAD-box/genética , Feminino , Hormônio Liberador de Gonadotropina/farmacologia , Antagonistas de Hormônios/farmacologia , Luteolíticos/farmacologia , Gravidez , Ratos , Ratos Wistar
17.
Gen Comp Endocrinol ; 281: 83-90, 2019 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-31170402

RESUMO

The function of insulin-like growth factor (Igf) system in ovary has attracted much attention, but the role of Igf binding proteins (Igfbps) in ovary is still largely unknown. In this study, the role of Igfbps in oocyte maturation was investigated in zebrafish. The expression of all eight identified Igfbps except Igfbp6b could be detected in the adult ovary and exhibited differential expression profiles during folliculogenesis. The expression of several Igfbps is dynamically changed during oocyte maturation induced by human chorionic gonadotropin (hCG). By treatment of an Igfbps inhibitor NBI-31772 in vitro, the oocyte maturation could be stimulated in a clear dose-, time- and stage-dependent manner. Such effects were also observed by administration of NBI-31772 in vivo. Igfbps are differentially expressed in both follicular cells and oocytes, but the effect of NBI-31772 could only be found in intact follicles and not in the denuded oocytes. Previous studies have demonstrated that Igf3 is the major Igf member in regulating oocyte maturation of zebrafish. Interestingly, NBI-31772 could increase the effect of Igf3 on oocyte maturation. Furthermore, we found the effect of NBI-31772 on oocyte maturation could be blocked by an Igf type 1 receptor inhibitor BMS-536924 in vitro, suggesting the Igfbps can inhibit the oocyte maturation via Igf/Igf1r pathway. Together, we provided the first evidence in fish that Igfbps inhibit oocyte maturation of zebrafish.


Assuntos
Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/metabolismo , Oócitos/fisiologia , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/metabolismo , Animais , Catecóis/farmacologia , Gonadotropina Coriônica/farmacologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Humanos , Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/genética , Fator de Crescimento Insulin-Like I/metabolismo , Isoquinolinas/farmacologia , Oócitos/metabolismo , Oogênese/efeitos dos fármacos , Folículo Ovariano/citologia , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/metabolismo , Receptor IGF Tipo 1/metabolismo , Transdução de Sinais/efeitos dos fármacos
18.
Mol Med Rep ; 19(6): 5353-5360, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31059097

RESUMO

Ppm1b, a metal­dependent serine/threonine protein phosphatase, catalyzes the dephosphorylation of a variety of phosphorylated proteins. Ppm1b­/­ mouse embryos die at the fertilized oocyte stage, whereas Ppm1b+/­ mice with a C57BL/6 background exhibit no phenotypic abnormalities. Because the C57BL/6 strain produces a limited number of pups, in an attempt to produce Ppm1b­/­ mice, congenic Ppm1b+/­ mice with an ICR background were established, which are more fertile and gave birth to more pups. As a result, however, no Ppm1b­/­ offspring were obtained when pairs of Ppm1b+/­ ICR mice were bred again. Ppm1b+/­ male and female ICR mice were analyzed from the viewpoint of fecundity. The Ppm1b haploinsufficiency had no effect on testicular weight or the number of sperm in male mice. Despite the fact that the levels of Ppm1b protein in the ovaries of sexually mature Ppm1b+/­ mice were decreased compared with those of Ppm1b+/+ mice, there appeared to be no significant difference in the histological appearance of the ovaries, litter sizes or plasma progesterone levels at the estrous stage. When superovulation was induced by stimulation using a hormone treatment, the number of ovulated oocytes were the same for Ppm1b+/­ and Ppm1b+/+ mice at 4 weeks of age when the estrous cycle did not proceed, however, the number of ovulated oocytes was lower in sexually mature Ppm1b+/­ mice at 11 weeks of age compared with Ppm1b+/+ mice in the first and the second superovulation cycles. These collective results suggest that follicle development is excessive in Ppm1b+/­ mice, and that this leads to a partial depletion of matured follicles and a corresponding decrease in the number of ovulated oocytes.


Assuntos
Proteína Fosfatase 2C/genética , Superovulação , Animais , Gonadotropina Coriônica/farmacologia , Feminino , Heterozigoto , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos ICR , Oócitos/citologia , Oócitos/efeitos dos fármacos , Oócitos/metabolismo , Ovário/metabolismo , Gravidez , Progesterona/sangue , Proteína Fosfatase 2C/metabolismo , Superovulação/efeitos dos fármacos
19.
Theriogenology ; 133: 113-118, 2019 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-31082747

RESUMO

This study aimed to evaluate 1) the effect of inserting an intravaginal sponge containing medroxyprogesterone during the late luteal phase on the corpus luteum (CL) function and endogenous production of P4; 2) the effectiveness of two different equine chorionic gonadotrophin (eCG) doses on synchronization of ovulations for a resynchronization treatment; and 3) the inclusion of an early pregnancy diagnosis in an early resynchronization protocol for ovulation in ewes. For all studies, the synchronization protocol was based on a short-term protocol of six days of progestogen treatment plus one dose of prostaglandin F2alpha, one dose of eCG, and gonadorelin acetate after sponge withdrawal. For the first study, the ewes were mated with fertile rams; a second sponge was inserted in half of these ewes 12 days later, and blood samples were collected daily for six days, until sponge withdrawal. For the second study, the ewes were not mated, and received a second sponge during the same period, after which they were divided into three groups according to eCG dose (0, 200, or 300 IU). In the third study, all ewes were artificially inseminated and received the second sponge during the same period. At sponge withdrawal, pregnancy was diagnosed by color Doppler ultrasonography (DUS) of the CL, and only non-pregnant ewes were re-inseminated two days later. In the first study, serum progesterone values were similar regardless of whether an intravaginal sponge had been inserted. In the second study, the ovulation time was more concentrated in those ewes which received 200 IU of eCG. In the third study, there was no difference between the experimental groups (with or without a previous pregnancy diagnosis) in pregnancy rate at the first insemination, accumulated pregnancy rate, and pregnancy loss. The insertion of an intravaginal sponge impregnated with medroxyprogesterone acetate did not affect the endogenous production of P4. The application of 200 IU of eCG provided the best result with regard to the synchronization of ovulations in the resynchronization treatment. Also, the inclusion of an early pregnancy diagnosis with DUS is useful and improves the general results of resynchronization programs, shortening the total working period.


Assuntos
Gonadotropina Coriônica/farmacologia , Sincronização do Estro/métodos , Indução da Ovulação/veterinária , Progestinas/farmacologia , Ovinos/fisiologia , Animais , Feminino , Cavalos , Ovário/diagnóstico por imagem , Gravidez , Taxa de Gravidez , Fatores de Tempo
20.
Mar Biotechnol (NY) ; 21(4): 537-549, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31129797

RESUMO

Oocyte maturation and hydration are regulated by a complex interplay of various hormones and local factors. We have investigated the morphological changes of follicles and serum steroid levels during the HCG (human choionic gonadotophin)-induced oocyte maturation in the orange-spotted grouper. For the first time, a large-scale transcriptomic analysis of follicles during the maturation has been conducted in a fish species which produce pelagic oocytes. Eight cDNA libraries of follicle samples, from full-grown immature follicles to mature follicles, were constructed. A total of 402,530,284 high-quality clean reads were obtained after filtering, 79.66% of which perfectly mapped to the orange-spotted grouper genome. Real-time PCR results of 12 representative genes related to oocyte maturation and hydration verified the reliability of the RNA-seq data. A large number of genes related to oocyte maturation and hydration were identified in the transcriptome dataset. And the transcriptomic analysis revealed the dynamic changes of the steroid synthesis pathway and the pathway of hydration during oocyte maturation. The present study will facilitate future study on the oocyte maturation and hydration in the orange-spotted grouper and other marine pelagic egg spawner.


Assuntos
Proteínas de Peixes/genética , Regulação da Expressão Gênica no Desenvolvimento , Genoma , Oócitos/metabolismo , Perciformes/genética , Transcriptoma , Animais , Gonadotropina Coriônica/farmacologia , Estradiol/sangue , Feminino , Proteínas de Peixes/classificação , Proteínas de Peixes/metabolismo , Perfilação da Expressão Gênica , Biblioteca Gênica , Ontologia Genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Hidroxiprogesteronas/sangue , Anotação de Sequência Molecular , Oócitos/citologia , Oócitos/efeitos dos fármacos , Oócitos/crescimento & desenvolvimento , Folículo Ovariano/citologia , Folículo Ovariano/efeitos dos fármacos , Folículo Ovariano/crescimento & desenvolvimento , Folículo Ovariano/metabolismo , Perciformes/crescimento & desenvolvimento , Perciformes/metabolismo , Progesterona/sangue , Testosterona/sangue
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