WorldWideScience

Sample records for bovine embryo transfer

  1. Nucleolar ultrastructure in bovine nuclear transfer embryos

    DEFF Research Database (Denmark)

    Kaňka, Jiří; Smith, Steven Dale; Soloy, Eva;

    1999-01-01

    in nuclear morphology as a transformation of the nucleolus precursor body into a functional rRNA synthesising nucleolus with a characteristic ultrastructure. We examined nucleolar ultrastructure in bovine in vitro produced (control) embryos and in nuclear transfer embryos reconstructed from a MII phase...... at 1 hr after fusion and, by 3 hr after fusion, it was restored again. At this time, the reticulated fibrillo-granular nucleolus had an almost round shape. The nucleolar precursor body seen in the two-cell stage nuclear transfer embryos consisted of intermingled filamentous components and secondary...... time intervals after fusion. In the two-cell stage nuclear transfer embryo, the originally reticulated nucleolus of the donor blastomere had changed into a typical nucleolar precursor body consisting of a homogeneous fibrillar structure. A primary vacuole appeared in the four-cell stage nuclear...

  2. Factors that affect the reproductive efficiency of the recipient within a bovine embryo transfer program

    Directory of Open Access Journals (Sweden)

    Arturo Duica A.

    2007-12-01

    Full Text Available The embryo transfer is a biotechnological technique that allows increasing the descendant of animals with high genetic value. The positive results, represented in pregnancy after the application of this technique, are affected by some factors that are inherent to the donor, the embryo, the technique, and the recipients which receive a strange embryo in the uterus allowing pregnancy. This review describes some factors affecting the reproductive efficiency of the recipients of bovine embryos within a program of embryo transfer. Its important to evaluate the parameters in this kind of recipients, as race, age, physiological status, health status, weight, reproductive tract integrity and management, and also too monitoring the ovarian structures while the estrus synchronization, and within previous and posterior stages in embryo transfer procedure. Therefore an optimum follicular development will be determinant to corpus luteum formation which generates enough serum progesterone concentrations to offer a right uterine environment allowing the optimum embryo development. Controlling the factors that affect the efficiency of the embryo transfer, it will obtain an increasing of positive results represented in pregnancies and births of individuals come from animals with high genetic value.

  3. Transcriptional reprogramming of gene expression in bovine somatic cell chromatin transfer embryos

    Directory of Open Access Journals (Sweden)

    Page Grier P

    2009-04-01

    Full Text Available Abstract Background Successful reprogramming of a somatic genome to produce a healthy clone by somatic cells nuclear transfer (SCNT is a rare event and the mechanisms involved in this process are poorly defined. When serial or successive rounds of cloning are performed, blastocyst and full term development rates decline even further with the increasing rounds of cloning. Identifying the "cumulative errors" could reveal the epigenetic reprogramming blocks in animal cloning. Results Bovine clones from up to four generations of successive cloning were produced by chromatin transfer (CT. Using Affymetrix bovine microarrays we determined that the transcriptomes of blastocysts derived from the first and the fourth rounds of cloning (CT1 and CT4 respectively have undergone an extensive reprogramming and were more similar to blastocysts derived from in vitro fertilization (IVF than to the donor cells used for the first and the fourth rounds of chromatin transfer (DC1 and DC4 respectively. However a set of transcripts in the cloned embryos showed a misregulated pattern when compared to IVF embryos. Among the genes consistently upregulated in both CT groups compared to the IVF embryos were genes involved in regulation of cytoskeleton and cell shape. Among the genes consistently upregulated in IVF embryos compared to both CT groups were genes involved in chromatin remodelling and stress coping. Conclusion The present study provides a data set that could contribute in our understanding of epigenetic errors in somatic cell chromatin transfer. Identifying "cumulative errors" after serial cloning could reveal some of the epigenetic reprogramming blocks shedding light on the reprogramming process, important for both basic and applied research.

  4. Cloned embryos from semen. Part 2: Intergeneric nuclear transfer of semen-derived eland (Taurotragus oryx) epithelial cells into bovine oocytes

    Science.gov (United States)

    Nel-Themaat, L.; Gomez, M.C.; Pope, C.E.; Lopez, M.; Wirtu, G.; Jenkins, J.A.; Cole, A.; Dresser, B.L.; Bondioli, K.R.; Godke, R.A.

    2008-01-01

    The production of cloned offspring by nuclear transfer (NT) of semen-derived somatic cells holds considerable potential for the incorporation of novel genes into endangered species populations. Because oocytes from endangered species are scarce, domestic species oocytes are often used as cytoplasts for interspecies NT. In the present study, epithelial cells isolated from eland semen were used for intergeneric transfer (IgNT) into enucleated bovine oocytes and compared with bovine NT embryos. Cleavage rates of bovine NT and eland IgNT embryos were similar (80 vs. 83%, respectively; p > 0.05); however, development to the morula and blastocyst stage was higher for bovine NT embryos (38 and 21%, respectively; p bovine NT or eland IgNT cybrids before activation, but in 75 and 70% of bovine NT and eland igNT embryos, respectively, cell-cycle resumption was observed at 16 h postactivation (hpa). For eland IgNT embryos, 13% had ???8 cells at 84 hpa, while 32% of the bovine NT embryos had ???8 cells at the same interval. However, 100 and 66% of bovine NT and eland IgNT embryos, respectively, that had ???8 cells synthesized DNA. From these results we concluded that (1) semen-derived epithelial cell nuclei can interact and be transcriptionally controlled by bovine cytoplast, (2) the first cell-cycle occurred in IgNT embryos, (3) a high frequency of developmental arrest occurs before the eight-cell stage in IgNT embryos, and (4) IgNT embryos that progress through the early cleavage stage arrest can (a) synthesize DNA, (b) progress through subsequent cell cycles, and (c) may have the potential to develop further. ?? 2008 Mary Ann Liebert, Inc.

  5. 牛胚胎移植技术的产业化研究%Study of Industrilization of Bovine Embryo Transferring

    Institute of Scientific and Technical Information of China (English)

    冯建忠; 史远刚; 张秀陶; 梁小军; 薛伟; 扬奇; 孟华; 李毓华

    2001-01-01

    Embryo transfer and relative technology was explicated, including setting up of nucleus herd of cattle by importing frozen embryo, choice of donor, superovalution, cryopreservation of embryo and embryo splitting. This study make a base for the industrilization of bovine embryo transferring.%本文探讨了引进国外优质奶、肉牛冷冻胚胎建立核心群,以及供体牛选择、超数排卵、胚胎冷冻保存和分割等相关技术,为牛胚胎移植产业化奠定基础。

  6. Characteristics of pregnancies and offspring following transfer of bovine in vivo embryos assessed by nanorespirometry

    DEFF Research Database (Denmark)

    Lopes, Ana Sofia; Madsen, S E; Greve, Torben;

    2008-01-01

    It has been speculated whether the metabolism of the pre-implantation embryo may be reflected on the pregnancy and characteristics of the newborn animal. The present study investigated whether respiration rates of individual embryos were correlated with gestation length, type of parturition, birth......, III), stage of development, and diameter and were subsequently transferred individually (n = 43) to synchronized recipients. Gestation length of the recipients (n = 22) was calculated and the type of parturition (no assistance, light traction, heavy traction, or caesarean section) recorded. Sex......, weight, and condition of the calves at birth (weak, normal, or very active) were also assessed. Results were evaluated by chi-square analysis and using a linear mixed model. The pregnancy rate was 60% (26/43), and the respiration rates of individual embryos influenced gestation length as well...

  7. Transcriptional reprogramming of gene expression in bovine somatic cell chromatin transfer embryos

    OpenAIRE

    Page Grier P; Kasinathan Poothappillai; Wang Zhongde; Rodriguez-Osorio Nelida; Robl James M; Memili Erdogan

    2009-01-01

    Abstract Background Successful reprogramming of a somatic genome to produce a healthy clone by somatic cells nuclear transfer (SCNT) is a rare event and the mechanisms involved in this process are poorly defined. When serial or successive rounds of cloning are performed, blastocyst and full term development rates decline even further with the increasing rounds of cloning. Identifying the "cumulative errors" could reveal the epigenetic reprogramming blocks in animal cloning. Results Bovine clo...

  8. Transcriptional Reprogramming of Gene Expression in Bovine Somatic Cell Chromatin Transfer Embryos

    OpenAIRE

    Rodriguez-Osorio, N.; Wang, Zhongde; Page, G. P.; Robl, J M; Memili, E.

    2009-01-01

    Background Successful reprogramming of a somatic genome to produce a healthy clone by somatic cells nuclear transfer (SCNT) is a rare event and the mechanisms involved in this process are poorly defined. When serial or successive rounds of cloning are performed, blastocyst and full term development rates decline even further with the increasing rounds of cloning. Identifying the "cumulative errors" could reveal the epigenetic reprogramming blocks in animal cloning. Results Bovine clones from ...

  9. In vitro development of OPU-derived bovine embryos cultured either individually or in groups with the silk protein sericin and the viability of frozen-thawed embryos after transfer.

    Science.gov (United States)

    Isobe, Tomohiro; Ikebata, Yoshihisa; Do, Lanh Thi Kim; Tanihara, Fuminori; Taniguchi, Masayasu; Otoi, Takeshige

    2015-07-01

    The optimization of single-embryo culture conditions is very important, particularly in the in vitro production of bovine embryos using the ovum pick-up (OPU) procedure. The purpose of this study was to examine the development of embryos derived from oocytes obtained by OPU that were cultured either individually or in groups in medium supplemented with or without sericin and to investigate the viability of the frozen-thawed embryos after a direct transfer. When two-cell-stage embryos were cultured either individually or in groups for 7 days in CR1aa medium supplemented with or without 0.5% sericin, the rates of development to blastocysts and freezable blastocysts were significantly lower for the embryos cultured individually without sericin than for the embryos cultured in groups with or without sericin. Moreover, the rate of development to freezable blastocysts of the embryos cultured individually with sericin was significantly higher than that of the embryos cultured without sericin. When the frozen-thawed embryos were transferred directly to recipients, the rates of pregnancy, abortion, stillbirth and normal calving in the recipients were similar among the groups, irrespective of the culture conditions and sericin supplementation. Our findings indicate that supplementation with sericin during embryo culture improves the quality of the embryos cultured individually but not the viability of the frozen-thawed embryos after transfer to recipients. PMID:25488699

  10. Splitting and biopsy for bovine embryo sexing under field conditions.

    Science.gov (United States)

    Lopes, R F; Forell, F; Oliveira, A T; Rodrigues, J L

    2001-12-01

    Improvements on embryo micromanipulation techniques led to the use of embryo bisection technology in commercial embryo transfer programs, and made possible the direct genetic analysis of preimplantation bovine embryos by biopsy. For example, aspiration and microsection, allow bovine embryos sexing by detection of male-specific Y-chromosome in a sample of embryonic cells. We report on the application of the methodologies of splitting and biopsy of bovine embryos in field conditions, and on the results of embryo sex determination by the polymerase chain reaction (PCR). Pregnancy rates achieved with fresh bisected or biopsied embryos (50 to 60%) were similar to the fresh intact embryos (55 to 61%). The PCR protocol used for embryo sexing showed 92% to 94% of efficiency and 90 to 100% of accuracy. These results demonstrate these procedures are suitable for use in field conditions.

  11. MODELAGEM BIOECONÔMICA DA TRANSFERÊNCIA DE EMBRIÕES EM BOVINOS BIOECONOMIC MODEL IN BOVINE EMBRYO TRANSFER

    Directory of Open Access Journals (Sweden)

    Renato Travassos Beltrame

    2010-04-01

    Full Text Available

    O objetivo deste trabalho foi desenvolver um modelo matemático orientado a eventos de simulação, para auxiliar tomadas de decisão relativas à transferência de embriões em bovinos, considerando-se as dinâmicas de dois componentes da transferência de embriões: receptoras e embriões. Na simulação, não se avaliaram respostas individuais de doadoras a coletas consecutivas e eventos correspondentes na transferência de embriões. Simulou-se o mesmo protocolo para superovulação a todas as doadoras. Receptoras foram sincronizadas simulando-se o uso de prostaglandina. O número de embriões viáveis produzido por doadora e sua variabilidade tiveram como base um processo aleatório de simulação de Monte Carlo, que pressupôs uma distribuição exponencial negativa de densidade de probabilidade. Custos e receitas foram inseridos no modelo por meio de um cenário-base para calcular indicadores econômicos de rentabilidade. A análise sugeriu a impraticabilidade da atividade, se realizada diante do cenário proposto (VPL – R$: 57.596,69. A partir do cenário proposto, o custo médio estimado foi de R$ 1.178,19, e de R$ 980,03, para se obter uma prenhez a partir de uma situação otimizada, sugerida pelo modelo (5/100; 5/190.

    PALAVRAS-CHAVES: Otimização, receptoras, simulação, transferência de embriões, viabilidade econômica.

    A simulation model related to embryo transfer programs in bovine was carried out through a mathematical model directed to events, considering the dynamic of two resources: recipients and embryos. Individual answers of donors to consecutive collections and corresponding events in embryo transfer were not evaluated. The same protocol for superovulation was simulated for all the donor collections, using similar doses of hormones and drugs for all the animals. Recipients were synchronized using prostaglandin. Meantime, the number of viable embryos produced by donor and its variability were based at

  12. Gestaciones producidas con embriones bovinos clonados por transferencia nuclear Pregnancies produced by bovine embryos cloned by nuclear transfer

    Directory of Open Access Journals (Sweden)

    M A Martínez Díaz

    2007-01-01

    Full Text Available El presente estudio comunica la obtención de gestaciones de embriones clonados por transferencia nuclear de células somáticas en bovinos por primera vez en Chile. Ovocitos bovinos obtenidos de ovarios de matadero fueron madurados in vitro y enucleados por micromanipulación. Células donantes de núcleos fueron obtenidas de la oreja de una vaca adulta, cultivadas por 9-14 días y criopreservadas en nitrógeno líquido. Células somáticas confluentes fueron desagregadas e insertadas individualmente en el espacio perivitelino de un ovocito enucleado. Cada par ovocito-célula obtenido fue tratado con dos pulsos eléctricos para inducir su fusión y luego los embriones fueron cultivados por 2 horas y tratados con ionomicina y 6-dimetilaminopurina para su activación. Los embriones fueron cultivados en medio sintético oviductual por 8-9 días hasta el estado de blastocisto. Blastocistos fueron transferidos a vaquillas receptoras 7 a 9 días postcelo. Se realizaron 25 transferencias a vaquillas receptoras y se logró la preñez en 5 (20% de ellas. Dos de éstas abortaron a los 42 días y una tercera a los 120 días. Las dos vaquillas preñadas restantes mantuvieron su gestación (más de 45 días hasta el momento de escribir esta comunicación.With the aim of obtaining pregnancies from nuclear transfer embryos reconstituted with somatic cells and enucleated oocytes, bovine oocytes from slaughterhouse were matured in vitro and enucleated by micromanipulation. Nuclear donor cells were obtained from the ear of an adult cow, cultured for 9 to 14 days and cryopreserved in liquid nitrogen. Confluent cells were inserted individually in the perivitelline space of enucleated oocytes and treated with 2 electric pulses to induce fusion. The reconstituted zygotes were then cultured for 2 hours and treated with ionomycin and 6-dimethylaminopurine for their activation. The embryos were cultured in synthetic oviduct fluid for 8 to 9 days to obtain the

  13. Influence of recipient cytoplasm cell stage on transcription in bovine nucleus transfer embryos

    DEFF Research Database (Denmark)

    Smith, Steven D.; Soloy, Eva; Kanka, Jiri;

    1996-01-01

    . NTE were produced using either a MII phase (nonactivated) cytoplasts at 32 hr of maturation or S-phase (activated) cytoplasts activated with calcium ionophore A23187 and cycloheximide treatment approximately 8 hr prior to fusion with a blastomere from an in-vitro-produced morula stage embryo at 32 hr...

  14. Effect of two activation treatments and age of blastomere karyoplasts on in vitro development of bovine nuclear transfer embryos

    DEFF Research Database (Denmark)

    Booth, P J; Holm, P; Vajta, G;

    2001-01-01

    , or 5 in vitro produced donor embryos, were examined in order to define an optimal nuclear transfer protocol. The two activation protocols comprised calcium ionophore followed by either CHX or DMAP. Parthenogenetic blastocyst yields were greater (P

  15. Embryo cloning by nuclear transfer in the bovine species: first results

    OpenAIRE

    Ectors, Francis; Ectors, Fabien; Delval, Alain; Thonon, Fabienne; Beckers, Jean-François

    1993-01-01

    Le clonage par transfert de noyau fut réalisé pour la première fois chez les bovins par Prather et collaborateurs en 1987. Vu l'importance économique de ce mode de multiplication, de nombreuses équipes de recherche affinent la technique et étudient sa mise en application sur le terrain. Alors que l'énucléation de l'ovocyte receveur, l'injection et la fusion du blastomère donneur se réalisent avec succès, la maturation ovocytaire, la culture et la congélation de l'embryon reconstitué posent en...

  16. Evaluation of Bovine Embryo Biopsy Techniques according to Their Ability to Preserve Embryo Viability

    Directory of Open Access Journals (Sweden)

    M. Cenariu

    2012-01-01

    Full Text Available The purpose of this research was to evaluate three embryo biopsy techniques used for preimplantation genetic diagnosis (PGD in cattle and to recommend the least invasive one for current use, especially when PGD is followed by embryo cryopreservation. Three hundred bovine embryos were biopsied by either one of the needle, aspiration or microblade method, and then checked for viability by freezing/thawing and transplantation to recipient cows. The number of pregnancies obtained after the transfer of biopsied frozen/thawed embryos was assessed 30 days later using ultrasounds. The results were significantly different between the three biopsy methods: the pregnancy rate was of 57% in cows that received embryos biopsied by needle, 43% in cows that received embryos biopsied by aspiration, and 31% in cows that received embryos biopsied by microblade. Choosing an adequate biopsy method is therefore of great importance in embryos that will undergo subsequent cryopreservation, as it significantly influences their viability after thawing.

  17. In vitro production of bovine embryos

    DEFF Research Database (Denmark)

    Stroebech, L.; Mazzoni, Gianluca; Pedersen, Hanne Skovsgaard;

    2015-01-01

    In vitro production (IVP) of bovine embryos has become a widespread technology implemented in cattle breeding and production. The implementation of genomic selection and systems biology adds great dimensions to the impact of bovine IVP. The physical procedures included in the IVP process can still...

  18. [Bovine progressive degenerative myeloencephalopathy ("Weaver syndrome") in brown Swiss x Braunvieh cattle: reproductive occurrences, results of embryo transfer].

    Science.gov (United States)

    Tenhumberg, H; Trela, T; Matzke, P; Averdunk, G; Dirksen, G

    1994-12-01

    In the time between 1989 and 1991 seven Brown Swiss heifers, which had clinical signs of the Weaver syndrome were kept at the Bavarian Institute of Animal Breeding in Grub. This was in order to investigate this hereditary trait further. The number of animals carrying this genetic defect was increased by means of embryo transfer. Both cycle observations and ovary controls by means of rectal palpation resulted largely in physiological data and findings. All seven animals responded to superovulation treatment which was induced by sequential doses of p-FSH (32 mg) or of a single dose of 2,000 IU PMSG. The donors were flushed a total of 32 times without problem. On average 5.3 ova were recovered, 2.8 of which were viable and suitable for transfer. These are statistically only 50% of the normal value in a routine ET programme. Following the transfer of fresh and frozen embryos the pregnancy rate was 53%. There was only one abortion observed from 48 pregnancies.

  19. Expression Profile of Genes as Indicators of Developmental Competence and Quality of In Vitro Fertilization and Somatic Cell Nuclear Transfer Bovine Embryos

    Science.gov (United States)

    Monteleone, Melisa Carolina; Mucci, Nicolas; Kaiser, German Gustavo; Brocco, Marcela; Mutto, Adrián

    2014-01-01

    Reproductive biotechnologies such as in vitro fertilization (IVF) and somatic cell nuclear transfer (SCNT) enable improved reproductive efficiency of animals. However, the birth rate of in vitro-derived embryos still lags behind that of their in vivo counterparts. Thus, it is critical to develop an accurate evaluation and prediction system of embryo competence, both for commercial purposes and for scientific research. Previous works have demonstrated that in vitro culture systems induce alterations in the relative abundance (RA) of diverse transcripts and thus compromise embryo quality. The aim of this work was to analyze the RA of a set of genes involved in cellular stress (heat shock protein 70-kDa, HSP70), endoplasmic reticulum (ER) stress (immunoglobulin heavy chain binding protein, Bip; proteasome subunit β5, PSMB5) and apoptosis (BCL-2 associated X protein, Bax; cysteine aspartate protease-3, Caspase-3) in bovine blastocysts produced by IVF or SCNT and compare it with that of their in vivo counterparts. Poly (A) + mRNA was isolated from three pools of 10 blastocysts per treatment and analyzed by real-time RT-PCR. The RA of three of the stress indicators analyzed (Bax, PSMB5 and Bip) was significantly increased in SCNT embryos as compared with that of in vivo-derived blastocysts. No significant differences were found in the RA of HSP70 and Caspase-3 gene transcripts. This study could potentially complement morphological analyses in the development of an effective and accurate technique for the diagnosis of embryo quality, ultimately aiding to improve the efficiency of assisted reproductive techniques (ART). PMID:25269019

  20. Expression profile of genes as indicators of developmental competence and quality of in vitro fertilization and somatic cell nuclear transfer bovine embryos.

    Directory of Open Access Journals (Sweden)

    Maria Jesús Cánepa

    Full Text Available Reproductive biotechnologies such as in vitro fertilization (IVF and somatic cell nuclear transfer (SCNT enable improved reproductive efficiency of animals. However, the birth rate of in vitro-derived embryos still lags behind that of their in vivo counterparts. Thus, it is critical to develop an accurate evaluation and prediction system of embryo competence, both for commercial purposes and for scientific research. Previous works have demonstrated that in vitro culture systems induce alterations in the relative abundance (RA of diverse transcripts and thus compromise embryo quality. The aim of this work was to analyze the RA of a set of genes involved in cellular stress (heat shock protein 70-kDa, HSP70, endoplasmic reticulum (ER stress (immunoglobulin heavy chain binding protein, Bip; proteasome subunit β5, PSMB5 and apoptosis (BCL-2 associated X protein, Bax; cysteine aspartate protease-3, Caspase-3 in bovine blastocysts produced by IVF or SCNT and compare it with that of their in vivo counterparts. Poly (A + mRNA was isolated from three pools of 10 blastocysts per treatment and analyzed by real-time RT-PCR. The RA of three of the stress indicators analyzed (Bax, PSMB5 and Bip was significantly increased in SCNT embryos as compared with that of in vivo-derived blastocysts. No significant differences were found in the RA of HSP70 and Caspase-3 gene transcripts. This study could potentially complement morphological analyses in the development of an effective and accurate technique for the diagnosis of embryo quality, ultimately aiding to improve the efficiency of assisted reproductive techniques (ART.

  1. Bovine in vitro embryo production : An overview

    Directory of Open Access Journals (Sweden)

    V. S. Suthar

    Full Text Available Dairy industry perfected the application of the first reproductive biotechnology, i.e. artificial insemination (AI - a great success story and also remains the user of embryo transfer technology (ETT. In addition, recently the researchers taking interest to embraced the field of Transvaginal OocyteRecovery (TVOR and in vitro production (IVEP of embryos. IVF provides the starting point for the generation of reproductive material for a number of advanced reproduction techniques including sperm microinjection and nuclear transfer (cloning. In several countries commercial IVF facilities are already being employed by cattle ET operators. Various research groups have reported on modification of TVOR technique to give greater efficiency. Much research is still needed in domestic animal (Especially Indian species on mechanisms controlling embryo development and on development of totally in vitro system for embryo culture. [Vet World 2009; 2(12.000: 478-479`

  2. Aspects of energetic substrate metabolism of in vitro and in vivo bovine embryos

    International Nuclear Information System (INIS)

    Although the metabolism of early bovine embryos has not been fully elucidated, several publications have addressed this important issue to improve culture conditions for cattle reproductive biotechnologies, with the ultimate goal of producing in vitro embryos similar in quality to those developing in vivo. Here, we review general aspects of bovine embryo metabolism in vitro and in vivo, and discuss the use of metabolic analysis of embryos produced in vitro to assess viability and predict a viable pregnancy after transference to the female tract

  3. Aspects of energetic substrate metabolism of in vitro and in vivo bovine embryos

    Directory of Open Access Journals (Sweden)

    D.K. de Souza

    2015-03-01

    Full Text Available Although the metabolism of early bovine embryos has not been fully elucidated, several publications have addressed this important issue to improve culture conditions for cattle reproductive biotechnologies, with the ultimate goal of producing in vitro embryos similar in quality to those developing in vivo. Here, we review general aspects of bovine embryo metabolism in vitro and in vivo, and discuss the use of metabolic analysis of embryos produced in vitro to assess viability and predict a viable pregnancy after transference to the female tract.

  4. Aspects of energetic substrate metabolism of in vitro and in vivo bovine embryos

    Energy Technology Data Exchange (ETDEWEB)

    Souza, D.K. de [Laboratório de Biotecnologia da Saúde, Faculdade de Medicina, Universidade de Brasília, Brasília, DF (Brazil); Faculdade da Ceilândia, Universidade de Brasília, Brasília, DF (Brazil); Salles, L.P. [Laboratório de Biotecnologia da Saúde, Faculdade de Medicina, Universidade de Brasília, Brasília, DF (Brazil); Departamento de Biologia Molecular, Instituto de Biologia, Universidade de Brasília, Brasília, DF (Brazil); Rosa e Silva, A.A.M. [Laboratório de Biotecnologia da Saúde, Faculdade de Medicina, Universidade de Brasília, Brasília, DF (Brazil)

    2015-01-23

    Although the metabolism of early bovine embryos has not been fully elucidated, several publications have addressed this important issue to improve culture conditions for cattle reproductive biotechnologies, with the ultimate goal of producing in vitro embryos similar in quality to those developing in vivo. Here, we review general aspects of bovine embryo metabolism in vitro and in vivo, and discuss the use of metabolic analysis of embryos produced in vitro to assess viability and predict a viable pregnancy after transference to the female tract.

  5. The HIST1 Locus Escapes Reprogramming in Cloned Bovine Embryos

    Directory of Open Access Journals (Sweden)

    Byungkuk Min

    2016-05-01

    Full Text Available Epigenetic reprogramming is necessary in somatic cell nuclear transfer (SCNT embryos in order to erase the differentiation-associated epigenetic marks of donor cells. However, such epigenetic memories often persist throughout the course of clonal development, thus decreasing cloning efficiency. Here, we explored reprogramming-refractory regions in bovine SCNT blastocyst transcriptomes. We observed that histone genes residing in the 1.5 Mb spanning the cow HIST1 cluster were coordinately downregulated in SCNT blastocysts. In contrast, both the nonhistone genes of this cluster, and histone genes elsewhere remained unaffected. This indicated that the downregulation was specific to HIST1 histone genes. We found that, after trichostatin A treatment, HIST1 histone genes were derepressed, and DNA methylation at their promoters was decreased to the level of in vitro fertilization embryos. Therefore, our results indicate that the reduced expression of HIST1 histone genes is a consequence of poor epigenetic reprogramming in SCNT blastocysts.

  6. Efficient delivery of DNA into bovine preimplantation embryos by multiwall carbon nanotubes

    Science.gov (United States)

    Munk, Michele; Ladeira, Luiz O.; Carvalho, Bruno C.; Camargo, Luiz S. A.; Raposo, Nádia R. B.; Serapião, Raquel V.; Quintão, Carolina C. R.; Silva, Saulo R.; Soares, Jaqueline S.; Jorio, Ado; Brandão, Humberto M.

    2016-01-01

    The pellucid zone (PZ) is a protective embryonic cells barrier against chemical, physical or biological substances. This put, usual transfection methods are not efficient for mammal oocytes and embryos as they are exclusively for somatic cells. Carbon nanotubes have emerged as a new method for gene delivery, and they can be an alternative for embryos transfection, however its ability to cross the PZ and mediated gene transfer is unknown. Our data confirm that multiwall carbon nanotubes (MWNTs) can cross the PZ and delivery of pDNA into in vitro-fertilized bovine embryos. The degeneration rate and the expression of genes associated to cell viability were not affected in embryos exposed to MWNTs. Those embryos, however, had lower cell number and higher apoptotic cell index, but this did not impair the embryonic development. This study shows the potential utility of the MWNT for the development of new method for delivery of DNA into bovine embryos. PMID:27642034

  7. Single-embryo transfer versus multiple-embryo transfer.

    Science.gov (United States)

    Gerris, Jan

    2009-01-01

    Despite the progress made in assisted reproductive technology, live birth rates remain disappointingly low. Multiple-embryo transfer has been an accepted practice with which to increase the success rate. This has led to a higher incidence of multiple-order births compared with natural conception, which not only increase the risk of mortality and morbidity to both mother and children but are also associated with social and economic consequences. Elective single-embryo transfer (eSET) was developed in an effort to increase singleton pregnancies in assisted reproduction. Studies comparing eSET with multiple-embryo transfer highlight the benefit of this approach and suggest that, with careful patient selection and the transfer of good-quality embryos, the risk of a multiple-order pregnancy can be reduced without significantly decreasing live birth rates. Although the use of eSET has gradually increased in clinical practice, its acceptance has been limited by factors such as availability of funding and awareness of the procedure. An open discussion of eSET is warranted in an effort to enable a broader understanding by physicians and patients of the merits of this approach. Ultimately, eSET may provide a more cost-effective, potentially safer approach to patients undergoing assisted reproduction technology.

  8. Non-surgical embryo transfer in pigs

    NARCIS (Netherlands)

    Hazeleger, W.

    1999-01-01

    Embryo transfer in pigs has been performed surgically for a long time. However, a less invasive, non-surgical, procedure of embryo transfer could be a valuable tool for research (to study embryo survival and embryo-uterus interactions) and practical applications (export, prevention of disease transm

  9. Embryo-maternal interactions leading to embryonic development and survival in the bovine : role of progesterone and prostaglandins

    OpenAIRE

    Torres, Ana Catarina Belejo Mora

    2012-01-01

    Tese de Doutoramento em Ciências Veterinárias. Especialidade de Clínica The objectives of this thesis were to evaluate steroidogenic and prostanoid embryo-maternal interactions leading to embryonic development and survival in cattle, and to evaluate therapeutic strategies at embryo transfer (ET) designed to enhance embryo survival. In vitro experiments (three experimental chapters) - bovine early (Day 7) embryos i) had transcription of genes coding for enzymes progesterone (P4)...

  10. Oocyte-secreted factors in oocyte maturation media enhance subsequent development of bovine cloned embryos.

    Science.gov (United States)

    Su, Jianmin; Wang, Yongsheng; Zhang, Lei; Wang, Bo; Liu, Jun; Luo, Yan; Guo, Zekun; Quan, Fusheng; Zhang, Yong

    2014-04-01

    Successful in vitro maturation (IVM) and oocyte quality both affect the subsequent development of cloned embryos derived from somatic-cell nuclear transfer (SCNT). Developmental competence is usually lower in oocytes matured in vitro compared with those that matured in vivo, possibly due to insufficient levels of oocyte-secreted factors (OSFs) and disrupted oocyte-cumulus communication. This study investigated the effects of OSFs secreted by denuded oocytes (DOs) during IVM on the subsequent developmental competence of cloned bovine embryos. Cumulus-oocyte complexes (COCs) from antral follicles of slaughtered-cow ovaries collected from an abattoir were divided into four groups: COCs co-cultured with and without DOs in maturation media used for SCNT, as well as COCs co-cultured with and without DOs in maturation media used for in vitro fertilization (IVF). Based on the developmental competence and embryo quality of bovine embryos generated from these four groups, we found that co-culturing the COCs with DOs enhanced the in vitro development of IVF and cloned bovine embryos, and potentially generated more high-quality cloned blastocysts that possessed locus-specific histone modifications at levels similar to in vitro-fertilized embryos. These results strongly suggest that co-culturing COCs with DOs enhances subsequent developmental competence of cloned bovine embryo. PMID:24420374

  11. Demi-embryo production from hatching of zona-drilled bovine and rabbit blastocysts.

    Science.gov (United States)

    Skrzyszowska, M; Smorag, Z; Katska, L

    1997-09-01

    It is known that the pregnancy rate resulting after transfer of bisected embryos is lower than after transfer of whole embryos. The main reason is the reduced cell number in the demi-embryo which is less than 1 2 of that in the intact embryo, since a number of blastomeres is damaged as a result of the procedure used in conventional embryo splitting. The aim of our experiment was to develop a non-invasive procedure which would limit cell losses during microsurgery. The experiment was carried out on bovine IVM-IVF embryos at middle, late and expanded blastocyst stage and rabbit embryos at late blastocyst stage cultured in vitro from in vivo produced zygotes. The zona pellucida of these embryos was drilled on the line between the inner cell mass and the trophoblast using a glass microneedle (embryo configuration, connected by a very thin cell bridge (figure eight in shape). To separate the parts of the embryo, the cell bridge was cut using a glass microneedle. During the separation only a few cells were damaged. As a result of the procedure 4 20 (20.0%), 48 144 (33.3%) and 3 40 (7.5%) middle, late and expanded blastocysts hatched according to the pattern described. The developed procedure could be considered as a non-invasive alternative to conventional embryo splitting.

  12. Comparison of the efficacy of conventional slow freezing and rapid cryopreservation methods for bovine embryos

    NARCIS (Netherlands)

    Wagtendonk-de Leeuw, van A.M.; den Daas, J.H.; Kruip, T.A.; Rail, W.F.

    1995-01-01

    Day 7 bovine morulae and early blastocysts were randomly assigned to one of four cryopreservation methods: (i) a modified conventional controlled slow freezing and stepwise dilution after thawing; and three methods which enable direct transfer of the embryo into the recipient upon thawing: (ii) conv

  13. PreImplantation Factor (PIF correlates with early mammalian embryo development-bovine and murine models

    Directory of Open Access Journals (Sweden)

    Coulam Carolyn B

    2011-05-01

    Full Text Available Abstract Background PreImplantation Factor (PIF, a novel peptide secreted by viable embryos is essential for pregnancy: PIF modulates local immunity, promotes decidual pro-adhesion molecules and enhances trophoblast invasion. To determine the role of PIF in post-fertilization embryo development, we measured the peptide's concentration in the culture medium and tested endogenous PIF's potential trophic effects and direct interaction with the embryo. Methods Determine PIF levels in culture medium of multiple mouse and single bovine embryos cultured up to the blastocyst stage using PIF-ELISA. Examine the inhibitory effects of anti-PIF-monoclonal antibody (mAb added to medium on cultured mouse embryos development. Test FITC-PIF uptake by cultured bovine blastocysts using fluorescent microscopy. Results PIF levels in mouse embryo culture medium significantly increased from the morula to the blastocyst stage (ANOVA, P = 0.01. In contrast, atretic embryos medium was similar to the medium only control. Detectable - though low - PIF levels were secreted already by 2-cell stage mouse embryos. In single bovine IVF-derived embryos, PIF levels in medium at day 3 of culture were higher than non-cleaving embryos (control (P = 0.01 and at day 7 were higher than day 3 (P = 0.03. In non-cleaving embryos culture medium was similar to medium alone (control. Anti-PIF-mAb added to mouse embryo cultures lowered blastocyst formation rate 3-fold in a dose-dependent manner (2-way contingency table, multiple groups, X2; P = 0.01 as compared with non-specific mouse mAb, and medium alone, control. FITC-PIF was taken-up by cultured bovine blastocysts, but not by scrambled FITC-PIF (control. Conclusions PIF is an early embryo viability marker that has a direct supportive role on embryo development in culture. PIF-ELISA use to assess IVF embryo quality prior to transfer is warranted. Overall, our data supports PIF's endogenous self sustaining role in embryo development and the

  14. Can Chlamydia abortus be transmitted by embryo transfer in goats?

    Science.gov (United States)

    Oseikria, M; Pellerin, J L; Rodolakis, A; Vorimore, F; Laroucau, K; Bruyas, J F; Roux, C; Michaud, S; Larrat, M; Fieni, F

    2016-10-01

    The objectives of this study were to determine (i) whether Chlamydia abortus would adhere to or penetrate the intact zona pellucida (ZP-intact) of early in vivo-derived caprine embryos, after in vitro infection; and (ii) the efficacy of the International Embryo Transfer Society (IETS) washing protocol for bovine embryos. Fifty-two ZP-intact embryos (8-16 cells), obtained from 14 donors were used in this experiment. The embryos were randomly divided into 12 batches. Nine batches (ZP-intact) of five embryos were incubated in a medium containing 4 × 10(7)Chlamydia/mL of AB7 strain. After incubation for 18 hours at 37 °C in an atmosphere of 5% CO2, the embryos were washed in batches in 10 successive baths of a phosphate buffer saline and 5% fetal calf serum solution in accordance with IETS guidelines. In parallel, three batches of ZP-intact embryos were used as controls by being subjected to similar procedures but without exposure to C. abortus. The 10 wash baths were collected separately and centrifuged for 1 hour at 13,000 × g. The washed embryos and the pellets of the 10 centrifuged wash baths were frozen at -20 °C before examination for evidence of C. abortus using polymerase chain reaction. C. abortus DNA was found in all of the infected batches of ZP-intact embryos (9/9) after 10 successive washes. It was also detected in the 10th wash fluid for seven batches of embryos, whereas for the two other batches, the last positive wash bath was the eighth and the ninth, respectively. In contrast, none of the embryos or their washing fluids in the control batches were DNA positive. These results report that C. abortus adheres to and/or penetrates the ZP of in vivo caprine embryos after in vitro infection, and that the standard washing protocol recommended by the IETS for bovine embryos, failed to remove it. The persistence of these bacteria after washing makes the embryo a potential means of transmission of the bacterium during embryo transfer from

  15. Cloning in cattle: from embryo splitting to somatic nuclear transfer.

    Science.gov (United States)

    Heyman, Y; Vignon, X; Chesné, P; Le Bourhis, D; Marchal, J; Renard, J P

    1998-01-01

    The ability to obtain genetically identical offspring in cattle (clones) is useful for research and for potential applications to breeding schemes. Experimental possibilities for generating such animals have evolved considerably in the last two decades. Embryo splitting has become a relatively simple technique but is limited to twinning. Embryonic nuclear transfer has improved and is associated with sexing to generate sets of clones despite a great variability of results between parent embryos. The factors of progress are reviewed here. Recently, somatic cells used as a source of nuclei in bovine nuclear transfer has been demonstrated. Here we present the results of the developmental potential of nuclei from skin and muscle cells.

  16. Embryo aggregation does not improve the development of interspecies somatic cell nuclear transfer embryos in the horse.

    Science.gov (United States)

    Gambini, Andrés; De Stéfano, Adrián; Jarazo, Javier; Buemo, Carla; Karlanian, Florencia; Salamone, Daniel Felipe

    2016-09-01

    The low efficiency of interspecies somatic cell nuclear transfer (iSCNT) makes it necessary to investigate new strategies to improve embryonic developmental competence. Embryo aggregation has been successfully applied to improve cloning efficiency in mammals, but it remains unclear whether it could also be beneficial for iSCNT. In this study, we first compared the effect of embryo aggregation over in vitro development and blastocyst quality of porcine, bovine, and feline zona-free (ZF) parthenogenetic (PA) embryos to test the effects of embryo aggregation on species that were later used as enucleated oocytes donors in our iSCNT study. We then assessed whether embryo aggregation could improve the in vitro development of ZF equine iSCNT embryos after reconstruction with porcine, bovine, and feline ooplasm. Bovine- and porcine-aggregated PA blastocysts had significantly larger diameters compared with nonaggregated embryos. On the other hand, feline- and bovine-aggregated PA embryos had higher blastocyst cell number. Embryo aggregation of equine-equine SCNT was found to be beneficial for embryo development as we have previously reported, but the aggregation of three ZF reconstructed embryos did not improve embryo developmental rates on iSCNT. In vitro embryo development of nonaggregated iSCNT was predominantly arrested around the stage when transcriptional activation of the embryonic genome is reported to start on the embryo of the donor species. Nevertheless, independent of embryo aggregation, equine blastocyst-like structures could be obtained in our study using domestic feline-enucleated oocytes. Taken together, these results reported that embryo aggregation enhance in vitro PA embryo development and embryo quality but effects vary depending on the species. Embryo aggregation also improves, as expected, the in vitro embryo development of equine-equine SCNT embryos; however, we did not observe positive effects on equine iSCNT embryo development. Among oocytes

  17. Efeito do ibuprofeno administrado uma hora antes da inovulação de embriões bovinos Effect of ibuprofen administered one hour before the bovine embryo transfer

    Directory of Open Access Journals (Sweden)

    H.J. Narváez

    2010-06-01

    Full Text Available Avaliou-se o efeito do ibuprofeno administrado uma hora antes da inovulação de embriões bovinos, com o objetivo de melhorar a taxa de prenhez. Após a avaliação da resposta ao protocolo de sincronização do estro, 76 fêmeas selecionadas como receptoras de embriões foram distribuídas em três grupos (G experimentais: G1 (n=25 receptoras usadas como controle, G2 (n=30 receptoras que receberam ibuprofeno 5mg/kg, I.M, uma hora antes da inovulação dos embriões, e G3 (n=21 receptoras que receberam uma matriz polimérica de liberação controlada de ibuprofeno administrado por via subcutânea. As taxas de prenhez foram de 16% (4/25, 43,3% (13/30 e 14,2% (3/21, para G1, G2 e G3, respectivamente. Observou-se diferença (PThe effect of the administered ibuprofen was evaluated one hour before the embryo transfer of bovine embryos in order to improve pregnancy rates. After evaluating the response to protocol synchronization of estrus, 76 Females selected as the recipients of embryos were distributed into three experimental groups: G1 (n = 25 surrogate cows used as control, G2 (n = 30 surrogate cows that received 5mg/kg ibuprofen, IM, one hour before the embryo transfer, and G3 (n = 20 surrogate cows that received an array polymeric release of controlled ibuprofen subcutaneously administered. The pregnancy rates were 16% (4/25, 43.3% (13/30, and 14.2% (3/21 for G1, G2, and G3, respectively. There was statistical difference (P<0.024 on pregnancy rate of G2, in comparison with those of G1 and G3. The administration of ibuprofen intramuscularly one hour before the embryo transfer resulted in better pregnancy rate in Nellore surrogate cows.

  18. Developmental disparity between in vitro-produced and somatic cell nuclear transfer bovine days 14 and 21 embryos

    DEFF Research Database (Denmark)

    Alexopoulos, Natalie I.; Maddox-Hyttel, Poul; Tveden-Nyborg, Pernille Yde;

    2008-01-01

    , immunohistochemistry, and transmission electron microscopy to establish in vivo developmental milestones. Following morphological examination, samples were characterized for the presence of epiblast (POU5F1), mesoderm (VIM), and neuroectoderm (TUBB3). On D14, only 25, 15, and 7% of IVP, SUZI, and HMC embryos were...... of VIM and TUBB3 was less distinct in SCNT embryos when compared with IVP embryos, indicating slower or compromised development. In conclusion, SCNT and to some degree, IVP embryos displayed a high rate of embryonic mortality before D14 and surviving embryos displayed reduced quality with respect...

  19. Nucleolar remodeling in nuclear transfer embryos

    DEFF Research Database (Denmark)

    Laurincik, Jozef; Maddox-Hyttel, Poul

    2007-01-01

    to develop fully functional nucleoli. In bovine in vivo developed embryos, a range of important nucleolar proteins (e.g., topoisomerase I, upstream binding factor and RNA polymerase I, fibrillarin, nucleophosmin and nucleolin) become localized to the nucleolar anlage over several cell cycles. This relocation...

  20. Proteomic analysis of the early bovine yolk sac fluid and cells from the day 13 ovoid and elongated preimplantation embryos

    DEFF Research Database (Denmark)

    Jensen, Pernille L; Beck, Hans Christian; Petersen, Tonny S;

    2014-01-01

    The bovine blastocyst hatches 8 to 9 days after fertilization, and this is followed by several days of preimplantation development during which the embryo transforms from a spherical over an ovoid to an elongated shape. As the spherical embryo enlarges, the cells of the inner cell mass...... slightly later stage cell differentiation in the developing bovine embryo. In vitro-produced Day 6 embryos were transferred into a recipient heifer and after 7 days of further in vivo culture, ovoid and elongated Day 13 embryos were recovered by flushing both uterine horns after slaughter. The primitive YS...... differentiate into the hypoblast and epiblast, which remain surrounded by the trophectoderm. The formation of the hypoblast epithelium is also accompanied by a change in the fluid within the embryo, i.e., the blastocoel fluid gradually alters to become the primitive yolk sac (YS) fluid. Our previous research...

  1. Embryo Transfer (Techniques, Donors, and Recipients).

    Science.gov (United States)

    Phillips, Patrick E; Jahnke, Marianna M

    2016-07-01

    Commercial embryo transfer has evolved as an art and as a science since the early 1970s. Today's multiple ovulation embryo transfer is a widely used reproductive tool on many farms and is performed by veterinarians throughout the world. Propagation of the female genomes of select donors, through embryo transfer, has allowed a rapid progression of genetic gain in many breeds, much like what happened with artificial insemination since the 1940s. Advancement of this technology is migrating to in vitro fertilization technology today, allowing a higher volume of offspring to be produced with sex selection in the laboratory. PMID:27140299

  2. Research on Growth Behavior of Embryos for Bovine and Murine on Primary Murine Embryos Fibroblast Cell Feeder Layer

    Institute of Scientific and Technical Information of China (English)

    AN Li-long; XIAO Mei; FENG Xiu-Liang; DOU Zhong-ying; QIU Huai; YANG Qi; LEI An-min; YANG Chun-rong; GAO Zhi-min

    2002-01-01

    The difference in growth behavior between bovine embryos and murine embryos was studied on PMEF(primary murine embryos fibroblast)feeder layer. The results showed as follows: With embryos having attached, bovine embryonic trophoblast formed a transparent membranous structure covering on inner cell mass (ICM), however, murine embryonic trophoblast formed disc structure. Bovine embryos formed four kinds of ICM colonies with different morphology including the mass-like, the net-like, the stream-like and the mixture-like colonies. Compared with Murine ICM, the bovine ICM grew more fast. So, the bovine ICM was passaged at first after a culture of approximately 5 - 6 days in vitro, but murine ICM was passaged at first after an attachment of 3 - 4 days on PMEF feeder layer. The mixture colonies of bovine ICM differentiated very early, while the others differentiated very late. Most ICM-like mass of Bovine grew in a defined spot, but bovine ICMs like stream and ICMs like net proliferated fast and dispersed quickly. We found that the single blastomeres derived from late bovine morula and late murine morula formed sub-blastophere; moreover, the bovine ICM cell would differentiate rapidly if the trophoblast was removed.

  3. Cellular and molecular deviations in bovine in vitro-produced embryos are related to the large offspring syndrome

    NARCIS (Netherlands)

    Lazzari, G.; Wrenzycki, C.; Herrmann, D.; Duchi, R.; Kruip, T.; Niemann, H.; Galli, C.

    2002-01-01

    The large offspring syndrome (LOS) is observed in bovine and ovine offspring following transfer of in vitro-produced (IVP) or cloned embryos and is characterized by a multitude of pathologic changes, of which extended gestation length and increased birthweight are predominant features. In the presen

  4. Effect of Embryo Transfer on the Bovine Estrus Synchronization and Conception%胚胎移植受体牛同期发情及受胎效果的研究

    Institute of Scientific and Technical Information of China (English)

    李刚

    2012-01-01

    The method of CIDR+E2 and twice use of uterine PGF2α infusion method were used in this study, embryo transfer for cattle estrus synchronization and fresh embryo transfer pregnancy was observed. The results showed that receptor bovine estrous rate at 24--48h was 85.5% and 51.6%, there was significantly difference between them (P〈0.05). Cattle before transplantation luteal receptor inspection pass rate was 79.3%, 77.8%, there was no significant difference (P〉0.05). Conception rate was 45.8% and 42.4%, the difference was not significant (P〉0.05). Estrus observed after 24h, 6.5d for direct examination Cattle with luteinizing for A, B grade can be used for transplantation.%本试验分别采用CIDR+E2法和两次PGF2α子宫输注法,对胚胎移植受体牛做同期发情处理,鲜胚移植观察其妊娠情况。试验结果表明两种处理方法的受体牛在24-48h发情率分别为85.5%和51.6%,两者之间差异显著(P〈0.05)。移植前对受体牛进行黄体检查,黄体合格率分另11为79.3%和77.8%,差异不显著(P〉0.05);受胎率分别为45.8%和42.4%,差异不显著(P〉0.05)。24h后观察发情,6.5d直检,黄体A、B级者用于移植。

  5. To transfer fresh or thawed embryos?

    DEFF Research Database (Denmark)

    Pinborg, Anja

    2012-01-01

    Worldwide freezing and thawing of embryos has been increasingly used since the first infant was born as a result of this technique in 1984. The use of frozen embryo replacement (FER) currently even exceeds the number of fresh cycles performed in some countries. This article discusses the pros...... in the subsequent development of fetus and child. Because larger and more detailed data sets are available for early cleavage-stage embryo freezing and slow freezing, they are the main focus of this review....... and cons of FER versus fresh-embryo transfer with regard to both single-cycle and cumulative pregnancy and delivery rates. The review discusses the obvious advantages of FER: minimizing the proportion of pharmacological and surgical treatments, and lowering the risk of ovarian hyperstimulation syndrome...

  6. Economic optimization of the number of recipients in bovine embryo transfer programs Otimização econômica do número de receptoras em programas de transferência de embriões em bovinos

    Directory of Open Access Journals (Sweden)

    Renato Travassos Beltrame

    2007-06-01

    Full Text Available Purchase and maintenance of recipient females account for a large proportion of the costs and determine the number of calves that can be produced in an embryo transfer program. However, the large variability of embryo production by the donors and the need to purchase and synchronize the recipients before knowing the number of embryos collected make it difficult for the decision maker to identify the ideal number of recipient females to allocate. An ex-ante evaluation to determine the optimal number of recipient females was carried out through a sensitivity analysis for the ratio between the number of recipients and donors in a simulation model. The variability for the number of embryos collected was accounted for by applying the Monte Carlo simulation technique, assuming normal distribution and known values for mean and variance. The simulation considered monthly intervals between collections, during a 24 months program. The effect of embryo freezing on the number of pregnancies was considered by introducing a stock of frozen embryos into the mathematical model. Optimal recipient/donor ratio and the cost per pregnancy were compared for three recipient synchronization protocols (prostaglandin, progesterone - P4 and Ovsynch, based on the expected performance for synchronization, conception and transfer/treated rates for each protocol. Stochastic simulation associated with sensitivity analysis was effective in identifying the optimal donor to recipient ratio. Freezing embryos is effective to reduce the operational costs per pregnancy. The estimated optimal recipient/donor ratio was 20 for prostaglandin and 16.7 for the other protocols. The P4 protocol, although the most expensive, resulted in the lowest pregnancy cost estimation followed by prostaglandin and Ovsynch.A aquisição e manutenção de receptoras representam grande proporção dos custos e determinam o número de produtos gerados em um programa de transferência de embriões. Entretanto

  7. Nucleologenesis and embryonic genome activation are defective in interspecies cloned embryos between bovine ooplasm and rhesus monkey somatic cells

    Directory of Open Access Journals (Sweden)

    Han Yong-Mahn

    2009-07-01

    Full Text Available Abstract Background Interspecies somatic cell nuclear transfer (iSCNT has been proposed as a tool to address basic developmental questions and to improve the feasibility of cell therapy. However, the low efficiency of iSCNT embryonic development is a crucial problem when compared to in vitro fertilization (IVF and intraspecies SCNT. Thus, we examined the effect of donor cell species on the early development of SCNT embryos after reconstruction with bovine ooplasm. Results No apparent difference in cleavage rate was found among IVF, monkey-bovine (MB-iSCNT, and bovine-bovine (BB-SCNT embryos. However, MB-iSCNT embryos failed to develop beyond the 8- or 16-cell stages and lacked expression of the genes involved in embryonic genome activation (EGA at the 8-cell stage. From ultrastructural observations made during the peri-EGA period using transmission electron microscopy (TEM, we found that the nucleoli of MB-iSCNT embryos were morphologically abnormal or arrested at the primary stage of nucleologenesis. Consistent with the TEM analysis, nucleolar component proteins, such as upstream binding transcription factor, fibrillarin, nucleolin, and nucleophosmin, showed decreased expression and were structurally disorganized in MB-iSCNT embryos compared to IVF and BB-SCNT embryos, as revealed by real-time PCR and immunofluorescence confocal laser scanning microscopy, respectively. Conclusion The down-regulation of housekeeping and imprinting genes, abnormal nucleolar morphology, and aberrant patterns of nucleolar proteins during EGA resulted in developmental failure in MB-iSCNT embryos. These results provide insight into the unresolved problems of early embryonic development in iSCNT embryos.

  8. N, N-Dimethylglycine decreases oxidative stress and improves in vitro development of bovine embryos.

    Science.gov (United States)

    Takahashi, Toshikiyo; Sasaki, Kouya; Somfai, Tamas; Nagai, Takashi; Manabe, Noboru; Edashige, Keisuke

    2016-04-22

    The antioxidant effect of N, N-dimethylglycine (DMG) on in vitro-produced (IVP) bovine embryos was examined. After in vitro fertilization, presumptive zygotes were cultured with or without 0.1 μM DMG under different oxygen tensions. The percentage of embryos developing to the blastocyst stage was lowest under a 20% oxygen concentration without DMG, and it was significantly increased (P culture medium significantly (P invitro development of IVP bovine embryos by acting as an antioxidant. PMID:26875568

  9. Holistic differential analysis of embryo-induced alterations in the proteome of bovine endometrium in the preattachment period.

    Science.gov (United States)

    Berendt, Frank J; Fröhlich, Thomas; Schmidt, Susanne E M; Reichenbach, Horst-Dieter; Wolf, Eckhard; Arnold, Georg J

    2005-07-01

    During the peri-implantation period, molecular signaling between embryo and endometrium (layer of tissue lining the uterus lumen) is supposed to be crucial for the maintenance of pregnancy. To investigate embryo-induced alterations in the proteome of bovine endometrium in the preattachment period (day 18), we used monozygotic cattle twins (generated by embryo splitting) as a model eliminating genetic variability as a source for proteome differences. One of the twins was pregnant after the transfer of two in vitro produced blastocysts, while the corresponding twin received a sham-transfer and served as a nonpregnant control. The two-dimensional fluorescence difference gel electrophoresis (2-D DIGE) analysis of the endometrium samples of three twin pairs (pregnant/nonpregnant) revealed four proteins with significantly higher abundance (p embryo-maternal interactions.

  10. Effect of growth factors on oocyte maturation and allocations of inner cell mass and trophectoderm cells of cloned bovine embryos.

    Science.gov (United States)

    Arat, Sezen; Caputcu, Arzu Tas; Cevik, Mesut; Akkoc, Tolga; Cetinkaya, Gaye; Bagis, Haydar

    2016-08-01

    This study was conducted to determine the additive effects of exogenous growth factors during in vitro oocyte maturation (IVM) and the sequential culture of nuclear transfer (NT) embryos. Oocyte maturation and culture of reconstructed embryos derived from bovine granulosa cells were performed in culture medium supplemented with either epidermal growth factor (EGF) alone or a combination of EGF with insulin-like growth factor-I (IGF-I). The maturation rates of oocytes matured in the presence of EGF or the EGF + IGF-I combination were significantly higher than those of oocytes matured in the presence of only fetal calf serum (FCS) (P 0.05). IGF-I alone or in combination with EGF in sequential embryo culture medium significantly increased the ratio of inner cell mass (ICM) to total blastocyst cells (P media of cloned bovine embryos increased the ICM without changing the total cell number. These unknown and uncontrolled effects of growth factors can alter the allocation of ICM and trophectoderm cells (TE) in NT embryos. A decrease in TE cell numbers could be a reason for developmental abnormalities in embryos in the cloning system. PMID:26444069

  11. Single embryo transfer - state of the art.

    Science.gov (United States)

    De Neubourg, Diane; Gerris, Jan

    2003-12-01

    Every practitioner active in the field of assisted reproduction treatment is aware of the risks and complications related to twin and higher-order multiple pregnancies. Introduction of single embryo transfer (SET) into IVF/intracytoplasmic sperm injection (ICSI) is one of the possible ways of reducing the rate of twin pregnancy. Careful selection of patients, in combination with elective SET, has been shown to decrease the twin pregnancy rate while maintaining a stable ongoing pregnancy rate. The combination of a woman younger than 38 years of age, in her first or second IVF/ICSI cycle and with an embryo with a high implantation potential is the key to successful SET. This article will discuss embryo selection and patient selection and review the data published on SET. In the Centre for Reproductive Medicine at Middelheim Hospital, 39% of all transfers in 2002 were SET; the ongoing pregnancy rate remained stable at 30.6%. The twin (multiple) pregnancy rate declined to 11.7%. Particular attention should be drawn to the augmenting effect of the pregnancy rate of frozen-thawed cycles. Health economic data available so far subscribe the plea for SET.

  12. Post-hatching development of the porcine and bovine embryo - defining criteria for expected development in vivo and in vitro

    DEFF Research Database (Denmark)

    Vejlsted, Morten; Du, Yutao; Vajta, Gabor;

    2006-01-01

    without the need for transfer to recipient animals. Such a system would require (1) definition of milestones of expected post-hatching embryonic development in vivo; and (2) development of adequate culture systems. We propose a stereomicroscopical staging system for post-hatching embryos defining......) Somite stage(s) where paraxial mesoderm gradually condensates to form somites. Post-hatching development of bovine embryos in vitro is compromised and although hatching occurs and elongation can be physically provoked by culture in agarose tunnels, the embryonic disk characterizing the pre-streak stage 1...

  13. Research Progress on Technique of Frozen Embryo Transfer in Sheep

    Institute of Scientific and Technical Information of China (English)

    SHE Qiu-sheng; HU Jian-ye; LOU Peng-yan; TAO Jing; XIE Zhao-hui

    2011-01-01

    The paper introduced the research progress on the technique of frozen embryo transfer in sheep, illustrated selection of donors and receptors, superovulation, synchronization of estrus, embryo cryopreservation and embryo transplantation. Frozen embryo transfer in sheep is another breakthrough in the high-quality sheep raising, and this technique in China is in its infancy recommendation stage, but it will be comprehensively popularized in the future.

  14. Splitting of IVP bovine blastocyst affects morphology and gene expression of resulting demi-embryos during in vitro culture and in vivo elongation.

    Science.gov (United States)

    Velasquez, Alejandra E; Castro, Fidel O; Veraguas, Daniel; Cox, Jose F; Lara, Evelyn; Briones, Mario; Rodriguez-Alvarez, Lleretny

    2016-02-01

    Embryo splitting might be used to increase offspring yield and for molecular analysis of embryo competence. How splitting affects developmental potential of embryos is unknown. This research aimed to study the effect of bovine blastocyst splitting on morphological and gene expression homogeneity of demi-embryos and on embryo competence during elongation. Grade I bovine blastocyst produced in vitro were split into halves and distributed in nine groups (3 × 3 setting according to age and stage before splitting; age: days 7-9; stage: early, expanded and hatched blastocysts). Homogeneity and survival rate in vitro after splitting (12 h, days 10 and 13) and the effect of splitting on embryo development at elongation after embryo transfer (day 17) were assessed morphologically and by RT-qPCR. The genes analysed were OCT4, SOX2, NANOG, CDX2, TP1, TKDP1, EOMES, and BAX. Approximately 90% of split embryos had a well conserved defined inner cell mass (ICM), 70% of the halves had similar size with no differences in gene expression 12 h after splitting. Split embryos cultured further conserved normal and comparable morphology at day 10 of development; this situation changes at day 13 when embryo morphology and gene expression differed markedly among demi-embryos. Split and non-split blastocysts were transferred to recipient cows and were recovered at day 17. Fifty per cent of non-split embryos were larger than 100 mm (33% for split embryos). OCT4, SOX2, TP1 and EOMES levels were down-regulated in elongated embryos derived from split blastocysts. In conclusion, splitting day-8 blastocysts yields homogenous demi-embryos in terms of developmental capability and gene expression, but the initiation of the filamentous stage seems to be affected by the splitting.

  15. An investigation into the possibility of bluetongue virus transmission by transfer of infected ovine embryos

    Directory of Open Access Journals (Sweden)

    Estelle H. Venter

    2011-02-01

    Full Text Available Bluetongue (BT, a disease that affects mainly sheep, causes economic losses owing to not only its deleterious effects on animals but also its associated impact on the restriction of movement of livestock and livestock germplasm. The causative agent, bluetongue virus (BTV, can occur in the semen of rams and bulls at the time of peak viraemia and be transferred to a developing foetus. The risk of the transmission of BTV by bovine embryos is negligible if the embryos are washed according to the International Embryo Transfer Society (IETS protocol. Two experiments were undertaken to determine whether this holds for ovine embryos that had been exposed to BTV. Firstly, the oestrus cycles of 12 ewes were synchronised and the 59 embryos that were obtained were exposed in vitro to BTV-2 and BTV-4 at a dilution of 1 x 102.88 and 1 x 103.5 respectively. In the second experiment, embryos were recovered from sheep at the peak of viraemia. A total of 96 embryos were collected from BTV-infected sheep 21 days after infection. In both experiments half the embryos were washed and treated with trypsin according to the IETS protocol while the remaining embryos were neither washed nor treated. All were tested for the presence of BTV using cell culture techniques. The virus was detected after three passages in BHK-21 cells only in one wash bath in the first experiment and two unwashed embryos exposed to BTV-4 at a titre of 1 x 103.5. No embryos or uterine flush fluids obtained from viraemic donors used in the second experiment were positive for BTV after the standard washing procedure had been followed. The washing procedure of the IETS protocol can thus clear sheep embryos infected with BTV either in vitro or in vivo.

  16. Is embryo transfer a useful technique for small community farmers?

    International Nuclear Information System (INIS)

    Four main aspects of embryo technology are dealt with in this paper. The first analyses the reasons for the poor selection of recipients for embryo transfer, the second relates to inaccurate evaluation of embryos at least under tropical conditions, the third proposes alternative methods to evaluate embryos for selection and freezing, and the fourth analyses the feasibility of establishing this technique as a biotechnology approach for improving production in small community tropical farms. (author)

  17. Intrafallopian transfer of gametes and early stage embryos for in vivo culture in cattle.

    Science.gov (United States)

    Wetscher, F; Havlicek, V; Huber, T; Gilles, M; Tesfaye, D; Griese, J; Wimmers, K; Schellander, K; Müller, M; Brem, G; Besenfelder, U

    2005-07-01

    It may be possible to avoid inadequate in vitro culture conditions by incubating gametes or embryos in the oviducts for a short time. Ideally, an optimized procedure should be devised, combining in vitro and in vivo systems, in order to achieve synchronization in cattle. We transferred gametes as well as embryos in various stages of development and placed them into the oviducts. Embryos were recovered on Day 7 by flushing of oviducts and uterine horns. Blastocyst rates were determined on Day 7 and on Day 8. Experimental designs included transfer of in vitro matured cumulus oocyte complexes into previously inseminated heifers (COCs group), transfer of in vitro matured COCs simultaneously with capacitated spermatozoa (GIFTs group), transfer of four to eight cell stage embryos developed in vitro after IVM/IVF (Cleaved Stages group) and a group of solely in vitro produced embryos (IVP control group). Our results indicate that in vivo culture of IVM/IVF embryos in the homologous bovine oviduct has a positive influence on subsequent pre-implantation development. In addition, we have evidence that in vitro maturation and in vivo fertilization cannot be synchronized. PMID:15935840

  18. Cryopreservation of Equine Embryos and First Report of a Native Colombian Breed Born by Transfer of an Equine Vitrified Embryo

    Directory of Open Access Journals (Sweden)

    Nadya Nathalie Martínez

    2014-06-01

    Full Text Available The aim of this paper is to report on the success of a cryopreservation procedure of equine embryos to achieve a viable pregnancy. Equine embryos were collected on day 6-6.5 (<300 μm, n = 24 and subjected to two cryopreservation techniques: group 1 (n = 12, vitrified, exposing them to a VS1 (Gli [1.4 M] 5 min, VS2 (Gli [1.4 M] + EG [3.6 M] and VS3 (Gli [3.4M] + EG [4.6 M] 1 min solution. They were packed in 0.25 ml straws and immersed in liquid nitrogen; group 2 (n = 12, slow freezing: exposed to a freezing solution (1.8 M EG + 0.1 M sucrose for 10 minutes, packed into 0.25 ml straws, brought to the embryos freezer, exposed to a freezing curve and immersed in liquid nitrogen. Following defrosting, cryoprotectants were removed from the 24 embryos in one step; they were submerged in culture medium DMEM/F12 + 10% of fetal bovine serum (FBS and incubated under controlled atmosphere (5% CO2, 5% N2, 90% O2 for 48 h. Embryonic development was evaluated in 75% of the vitrified embryos (n = 4; 20% of the embryos were subjected to slow freezing (n = 1. No significant difference was observed in the groups regarding embryonic development, but a greater survival tendency on the vitrified embryos was noted. Also, one of these vitrified embryos was transferred to a receiver, achieving a viable pregnancy and the birth of a living foal.

  19. Molecular Characterization of the First Bovine Herpesvirus 4 (BoHV-4 Strains Isolated from In Vitro Bovine Embryos production in Argentina.

    Directory of Open Access Journals (Sweden)

    Erika González Altamiranda

    Full Text Available Bovine herpesvirus 4 (BoHV-4 is increasingly considered as responsible for various problems of the reproductive tract. The virus infects mainly blood mononuclear cells and displays specific tropism for vascular endothelia, reproductive and fetal tissues. Epidemiological studies suggest its impact on reproductive performance, and its presence in various sites in the reproductive tract highlights its potential transmission in transfer-stage embryos. This work describes the biological and genetic characterization of BoHV-4 strains isolated from an in vitro bovine embryo production system. BoHV-4 strains were isolated in 2011 and 2013 from granulosa cells and bovine oocytes from ovary batches collected at a local abattoir, used as "starting material" for in vitro production of bovine embryos. Compatible BoHV-4-CPE was observed in the co-culture of granulosa cells and oocytes with MDBK cells. The identity of the isolates was confirmed by PCR assays targeting three ORFs of the viral genome. The phylogenetic analyses of the strains suggest that they were evolutionary unlinked. Therefore it is possible that BoHV-4 ovary infections occurred regularly along the evolution of the virus, at least in Argentina, which can have implications in the systems of in vitro embryo production. Thus, although BoHV-4 does not appear to be a frequent risk factor for in vitro embryo production, data are still limited. This study reveals the potential of BoHV-4 transmission via embryo transfer. Moreover, the high variability among the BoHV-4 strains isolated from aborted cows in Argentina highlights the importance of further research on the role of this virus as an agent with the potential to cause reproductive disease in cattle. The genetic characterization of the isolated strains provides data to better understand the pathogenesis of BoHV-4 infections. Furthermore, it will lead to fundamental insights into the molecular aspects of the virus and the means by which these

  20. Evaluating recipient and embryo factors that affect pregnancy rates of embryo transfer in beef cattle.

    Science.gov (United States)

    Spell, A R; Beal, W E; Corah, L R; Lamb, G C

    2001-07-15

    The objectives of this experiment were to determine the effects of corpus luteum characteristics, progesterone concentration, donor-recipient synchrony, embryo quality, type, and developmental stage on pregnancy rates after embryo transfer. We synchronized 763 potential recipients for estrus using one of two synchronization protocols: two doses of PGF2alpha (25 mg i.m.) given 11 d apart (Location 1); and, a single norgestomet implant for 7 d with one dose of PGF2alpha (25 mg i.m.) 24 h before implant removal (Location 2). At embryo transfer, ovaries were examined by rectal palpation and ultrasonography. Of the 526 recipients presented for embryo transfer, 122 received a fresh embryo and 326 received a frozen embryo. Pregnancy rates were greater (P 0.1). There was a significant, positive simple correlation between CL diameter or luteal tissue volume and plasma progesterone concentration (r = 0.15, P quality. PMID:11480620

  1. The current status and future of commercial embryo transfer in cattle.

    Science.gov (United States)

    Hasler, John F

    2003-12-15

    A commercially viable cattle embryo transfer (ET) industry was established in North America during the early 1970s, approximately 80 years after the first successful embryo transfer was reported in a mammal. Initially, techniques for recovering and transferring cattle embryos were exclusively surgical. However, by the late 1970s, most embryos were recovered and transferred nonsurgically. Successful cryopreservation of embryos was widespread by the early 1980s, followed by the introduction of embryo splitting, in vitro procedures, direct transfer of frozen embryos and sexing of embryos. The wide spread adoption of ethylene glycol as a cryoprotectant has simplified the thaw-transfer procedures for frozen embryos. The number of embryos recovered annually has not grown appreciably over the last 10 years in North America and Europe; however, there has been significant growth of commercial ET in South America. Within North America, ET activity has been relatively constant in Holstein cattle, whereas there has been a large ET increase in the Angus breed and a concomitant ET decrease in some other beef breeds. Although a number of new technologies have been adopted within the ET industry in the last decade, the basic procedure of superovulation of donor cattle has undergone little improvement over the last 20 years. The export-import of frozen cattle embryos has become a well-established industry, governed by specific health regulations. The international movement of embryos is subject to sudden and dramatic disturbances, as exemplified by the 2001 outbreak of foot and mouth disease in Great Britain. It is probable that there will be an increased influence of animal rights issues on the ET industry in the future. Several companies in North America are currently commercially producing cloned cattle. The sexing of bovine semen with the use of flow cytometry is extremely accurate and moderate pregnancy rates in heifers have been achieved in field trials, but sexed semen

  2. Effects of donor cells on in vitro development of cloned bovine embryos

    Institute of Scientific and Technical Information of China (English)

    Jing Fu; Pengfei Guan; Leiwen Zhao; Hua Li; Shuzhen Huang; Fanyi Zeng; Yitao Zeng

    2008-01-01

    The donor cells from different individuals and with different foreign genes introduced were investigated to determine their effects on the efficiency of somatic cell nuclear transfer (SCNT). The bovine ear fibroblast from different individuals was isolated, cultured, and then transfected with foreign genes to establish the stable cell lines, which were used as donor cells for nuclear transfer. The ooeytes were obtained through ovum pick up operation. After in vitro maturation, the M II phase oocytes were selected as receptors for nuclear transfer.The reconstructed embryos were cultured in vitro and observed at 2 h, 48 h, and 7 days after transfer to assess the rate of fusion using cleaved and blastoeyst as the parameters of SCNT efficiency. The donor cells from different individuals (04036, 06081, 06088, and 06129)had no obvious effect on the fusion and cleaved rate, whereas there was significant difference in the blastocyst rate (P0.05). It was concluded that the genetic background of the donor cells could affect the effi-ciency of SCNT, while the introduction of foreign genes into the donor cells had no obvious effect on the efficiency. This study provides useful information for the SCNT and would benefit in promoting the efficiency.

  3. Viability of bovine demi embryo after splitting of fresh and frozen thawed embryo derived from in vitro embryo production

    Directory of Open Access Journals (Sweden)

    M Imron

    2007-06-01

    Full Text Available In vivo embryo production was limited by number of donor, wide variability respond due to superovulation program and also immunoactifity of superovulation hormone (FSH. Splitting technology could be an alternative to increase the number of transferrable embryos into recipien cows. Splitting is done with cutting embryo becoming two equal pieces (called demi embrio base on ICM orientation. The objective of this research was to determine the viability of demi embryo obtained from embryo splitting of fresh and frozen thawed embryo. The results showed that demi embryos which performed blastocoel reexpansion 3 hours after embryo splitting using fresh and frozen thawed embryos were 76.9 and 76.2% respectively. Base on existention of inner cell mass (ICM, the number of demi embryos developed with ICM from fresh and frozen thawed embryos were not significantly different (90.6 and 85.7% respectively. The cell number of demi embryo from fresh embryos splitting was not different compared with those from frozen thawed embryos (36.1 and 35.9 respectively. These finding indicated that embryo splitting can be applied to frozen thawed embryos with certain condition as well as fresh embryos.

  4. Genomic DNA methylation patterns in bovine preimplantation embryos derived from in vitro fertilization

    Institute of Scientific and Technical Information of China (English)

    HOU Jian; LIU Lei; LEI TingHua; CUI XiuHong; AN XiaoRong; CHEN YongFu

    2007-01-01

    By using the approach of immunofluorescence staining with an antibody against 5-methylcytosine (5MeC), the present study detected the DNA methylation patterns of bovine zygotes and preimplantation embryos derived from oocyte in vitro maturation (IVM), in vitro fertilization (IVF) and embryo in vitro culture (IVC). The results showed that: a) paternal-specific demethylation occurred in 61.5% of the examined zygotes, while 34.6% of them showed no demethylation; b) decreased methylation level was observed after the 8-cell stage and persisted through the morula stage, however methylation levels were different between blastomeres within the same embryos; c) at the blastocyst stage, the methylation level was very low in inner cell mass, but high in trophectoderm cells. The present study suggests, at least partly, that IVM/IVF/IVC may have effects on DNA methylation reprogramming of bovine zygotes and early embryos.

  5. Genomic DNA methylation patterns in bovine preim-plantation embryos derived from in vitro fertilization

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    By using the approach of immunofluorescence staining with an antibody against 5-methylcytosine (5MeC), the present study detected the DNA methylation patterns of bovine zygotes and preimplanta-tion embryos derived from oocyte in vitro maturation (IVM), in vitro fertilization (IVF) and embryo in vitro culture (IVC). The results showed that: a) paternal-specific demethylation occurred in 61.5% of the examined zygotes, while 34.6% of them showed no demethylation; b) decreased methylation level was observed after the 8-cell stage and persisted through the morula stage, however methylation levels were different between blastomeres within the same embryos; c) at the blastocyst stage, the methyla-tion level was very low in inner cell mass, but high in trophectoderm cells. The present study suggests, at least partly, that IVM/IVF/IVC may have effects on DNA methylation reprogramming of bovine zygotes and early embryos.

  6. Lysophosphatidic Acid Signaling in Late Cleavage and Blastocyst Stage Bovine Embryos

    OpenAIRE

    Ana Catarina Torres; Dorota Boruszewska; Mariana Batista; Ilona Kowalczyk-Zieba; Patricia Diniz; Emilia Sinderewicz; Jean Sebastian Saulnier-Blache; Izabela Woclawek-Potocka; Luis Lopes-da-Costa

    2014-01-01

    Lysophosphatidic acid (LPA) is a known cell signaling lipid mediator in reproductive tissues. In the cow, LPA is involved in luteal and early pregnancy maintenance. Here, we evaluated the presence and role of LPA in bovine early embryonic development. In relevant aspects, bovine embryos reflect more closely the scenario occurring in human embryos than the mouse model. Transcription of mRNA and protein expression of enzymes involved in LPA synthesis (ATX and cPLA2) and of LPA receptors (LPAR1–...

  7. Effect of oocyte quality and activation protocols on bovine embryo development following intracytoplasmic sperm injection

    OpenAIRE

    KORKMAZ, Ömer; KÜPLÜLÜ, Şükrü; AĞCA, Yüksel; POLAT, İbrahim Mert

    2013-01-01

    The purpose of this study was to investigate the effects of oocyte quality and activation protocols on the in vitro developmental competence of bovine embryos after intracytoplasmic sperm injection (ICSI). Bovine oocytes were grouped as being of excellent, good, and poor quality. All of the oocytes were activated using a calcium ionophore only, ethanol only, and 6-dimethylaminopurine (6-DMAP) following calcium ionophore. For the excellent quality oocytes, cleavage rates after ICSI were 70% in...

  8. 牛胚胎的体外生产技术%In vitro Bovine Embryo Production

    Institute of Scientific and Technical Information of China (English)

    马云; 窦忠英

    2001-01-01

    应用体外受精技术,可以有效地利用淘汰的高产奶牛和屠宰的青年母牛卵巢中的未成熟卵,大量、廉价地实验室生产胚胎。这对加速我国高产奶牛群和良种肉牛群的繁殖,是一项行之有效的胚胎工程技术。借鉴国外实验室生产牛胚胎的经验和根据我们实验室的实践,本文比较详细地叙述了体外受精牛胚胎(试管牛)的生产过程,包括卵母细胞的获得、培养、精子获能、体外受精以及受精后的胚胎培养、超低温冷冻保存与解冻后的胚胎移植等。%Using the immature ovum from eliminated high production dairy cow and slaughtered young cow, a number of embryos can be produced by in vitro insemination, it is an effective method to speed reproducing of high production dairy cow and improved beef breeds. According to abroad experience and our practices, the procedure of bovine embryo production in vitro, including oocyte collection, in vitro cultivation, sperm capacitation, in vitro insemination and embryo cultivation, freezing, thaw and transfer were reviewed in this paper.

  9. Effects of steroidal glycoalkaloids from potatoes (Solanum tuberosum) on in vitro bovine embryo development.

    Science.gov (United States)

    Wang, S; Panter, K E; Gaffield, W; Evans, R C; Bunch, T D

    2005-02-01

    alpha-Solanine and alpha-chaconine are two naturally occurring steroidal glycoalkaloids in potatoes (Solanum tuberosum), and solanidine-N-oxide is a corresponding steroidal aglycone. The objective of this research was to screen potential cyto-toxicity of these potato glycoalkaloids using bovine oocyte maturation, in vitro fertilization techniques and subsequent embryonic development as the in vitro model. A randomized complete block design with four in vitro oocyte maturation (IVM) treatments (Experiment 1) and four in vitro embryo culture (IVC) treatments (Experiment 2) was used. In Experiment 1, bovine oocytes (n=2506) were matured in vitro in medium supplemented with 6 microM of alpha-solanine, alpha-chaconine, solanidine-N-oxide or IVM medium only. The in vitro matured oocytes were then subject to routine IVF and IVC procedures. Results indicated that exposure of bovine oocytes to the steroidal glycoalkaloids during in vitro maturation inhibited subsequent pre-implantation embryo development. Potency of the embryo-toxicity varied between these steroidal glycoalkaloids. In Experiment 2, IVM/IVF derived bovine embryos (n=2370) were cultured in vitro in medium supplemented with 6 microM of alpha-solanine, alpha-chaconine, solanidine-N-oxide or IVC medium only. The results showed that the pre-implantation embryo development is inhibited by exposure to these glycoalkaloids. This effect is significant during the later pre-implantation embryo development period as indicated by fewer numbers of expanded and hatched blastocysts produced in the media containing these alkaloids. Therefore, we conclude that in vitro exposure of oocytes and fertilized ova to the steroidal glycoalkaloids from potatoes inhibits pre-implantation embryo development. Furthermore, we suggest that ingestion of Solanum species containing toxic amounts of glycoalkaloids may have negative effects on pre-implantation embryonic survival.

  10. Serial Nuclear Transfer of Goat-Rabbit Interspecies Reconstructed Embryos

    Institute of Scientific and Technical Information of China (English)

    ZHANG Zhi-guo; CHENG Li-zi; ZHANG Xiao-rong; LIU Ya; JING Ren-tao; WANG Cun-li; ZHAO Huan; LI Bin; CAO Chen-chong; LI Dong-wei

    2005-01-01

    The experiments of serial nuclear transfer were conducted between Boer goat and rabbit. The enucleated oocytes of rabbit were used as recipients while the blastomeres of goat morula was used as nuclear donor. The reconstructed embryos developing to morula were used as donor for serial cloning. As a result, two generations of reconstructed embryos were obtained, including 58 first generation reconstructed embryos and 14 second generation reconstructed embryos. The fusion rates were 79.5 and 70%, respectively. and there was no significant difference between them (P>0.05). The cleavage rates were 75.9 and 28.6% respectively with significant difference (P<0.01). No blastocyst was obtained from the second generation reconstructed embryos while 13.8% of first generation reconstructed embryos developed to blastocyst.

  11. Influence of antral follicle size on oocyte characteristics and embryo development in the bovine

    DEFF Research Database (Denmark)

    Lequarre, Anne Sophie; Vigneron, Céline; Ribaucour, Fabrice;

    2005-01-01

    The developmental competence of bovine oocytes isolated from antral follicles of different sizes was assessed in three European laboratories (Belgium, UCL; Denmark, DIAS; France, INRA). Using the same protocol for in vitro production of embryos, the oocytes isolated from follicles with a diameter...

  12. 体细胞核移植技术生产转入溶菌酶基因(hLYZ)牛胚胎的研究%Human Lysozyme Gene (hLYZ) Transgenic Bovine Embryos Produced by Somatic Cell Nuclear Transfer

    Institute of Scientific and Technical Information of China (English)

    熊显荣; 李文哲; 王丽君; 王勇胜; 苏建民; 华松; 张涌

    2011-01-01

    The aim of this study was to investigate the effect of different sizes and treatments of donor cells on the in vitro development of bovine embryos after genetic modification and selection, so as to establish an effective system for production of transgenic bovine embryos.Bovine fetal fibroblasts were transfeected with the recombinant plasmid of mammary gland specific expression vector pEBH which containing human lysozyme (hLYZ) gene.Transgenic positive cells which were obtained through G418 selection were used as donor cells for production of transgenic cloned embryos.The result showed that genetic modification and screening of donor cells were harmful for the development of transgenic cloned embryos, the blastocyst rate decreased significantly;referred to internal diameter of injective needle, cells with the diameter of 15~20 μm were selected and used as nuclear transfer donor cells, the fusion rate and blastocyst rate of transgenic cloned embryos were significant higher than that of the other groups (P < 0.05); Compared with serum starvation and contact restrain groups, the cleavage rate and blastocyst rate of transgenic cloned embryos which were constructed with plant cells were significantly higher (P < 0.05); The expression of GFP was observed in each stage of in vitro development in all transgenic cloned embryos, but the expression levels seemed to be vary among individuals and even among different developmental stages of the same individual.We chose I 0 embroys randomly for PCR detection and found that all of the embroys were positive.Above all, we obtained hLYZ transgenic cloned embryos by somatic cell nuclear transfer technique, the reconstructed embryos can develop to blastocysts successfully; and when plant cells with diameter of 15~20 μm were used as donor cell, the developmental competence of somatic cell cloned embryos is improved significantly.%本研究主要探讨经基因修饰和筛选后转基因供体细胞的挑选及处理对牛转

  13. Raman-based noninvasive metabolic profile evaluation of in vitro bovine embryos

    Science.gov (United States)

    dos Santos, Érika Cristina; Martinho, Herculano; Annes, Kelly; da Silva, Thais; Soares, Carlos Alexandre; Leite, Roberta Ferreira; Milazzotto, Marcella Pecora

    2016-07-01

    The timing of the first embryonic cell divisions may predict the ability of an embryo to establish pregnancy. Similarly, metabolic profiles may be markers of embryonic viability. However, in bovine, data about the metabolomics profile of these embryos are still not available. In the present work, we describe Raman-based metabolomic profiles of culture media of bovine embryos with different developmental kinetics (fast x slow) throughout the in vitro culture. The principal component analysis enabled us to classify embryos with different developmental kinetics since they presented specific spectroscopic profiles for each evaluated time point. We noticed that bands at 1076 cm-1 (lipids), 1300 cm-1 (Amide III), and 2719 cm-1 (DNA nitrogen bases) gave the most relevant spectral features, enabling the separation between fast and slow groups. Bands at 1001 cm-1 (phenylalanine) and 2892 cm-1 (methylene group of the polymethylene chain) presented specific patterns related to embryonic stage and can be considered as biomarkers of embryonic development by Raman spectroscopy. The culture media analysis by Raman spectroscopy proved to be a simple and sensitive technique that can be applied with high efficiency to characterize the profiles of in vitro produced bovine embryos with different development kinetics and different stages of development.

  14. Desarrollo de embriones de bovino obtenidos por fecundación in vitro cultivados con células oviductales o medio condicionado y transferidos a hembras receptoras Bovine embryo development produced by in vitro fertilization cultured with oviductal cell or conditioned medium and transfer to recipients

    Directory of Open Access Journals (Sweden)

    M RATTO

    1999-01-01

    (4- or 8-cell embryos was 62,7 % (64/102 for the zygotes co-cultured with oviductal cells and 66,7 % (100/150 for the zygotes cultured in conditioned medium. The development to the morula stage was 17,6 % (18/102 for the zygotes co-cultured with oviductal cells and 13,3 % (20/150 for the zygotes cultured in conditioned medium. A statiscally significant difference was not found in the development of 4, 8-cell embryos or morula. The development of embryos up to the blastocyst stage was 15,7 % (16/102 for the zygotes co-cultured with oviductal cells. Two blastocysts were transferred to the uterine horn ipsilateral to the CL in two recipients by non-surgical embryo transfer. Pregnancy was confirmed by ultrasonography at 42 and 57 days detecting the presence of one conceptus in each animal. This work has shown that in vitro inseminated of bovine oocytes with espermatozoa prepared with modified BO and co-cultured with oviductal cells, can develop to the blastocysts stage, unlike those that were cultured with conditioned medium. Finally, it is important to mention that this is the first communication in Chile of pregnancy after in vitro fertilization of bovine oocytes

  15. Screening of biotechnical parameters for production of bovine inter-subspecies embryonic chimeras by the aggregation of tetraploid Bos indicus and diploid crossbred Bos taurus embryos.

    Science.gov (United States)

    Razza, Eduardo M; Satrapa, Rafael A; Emanuelli, Isabele P; Barros, Ciro M; Nogueira, Marcelo F G

    2016-03-01

    The aggregation of a tetraploid zebu embryo (Bos indicus, a thermotolerant breed) with a diploid taurine embryo (Bos taurus, a thermosensitive breed) should create a complete taurine fetus, whose extra-embryonic components, e.g., the chorion, is derived mainly from the zebu embryo. These zebu-derived extra-embryonic components may interact positively with the taurine embryo/fetus during pregnancy in a tropical environment. We tested different parameters for the production of tetraploid Nelore (Bos indicus) embryos to be combined via aggregation with crossbred Bos taurus (diploid) embryos in order to produce viable chimeric blastocysts. Bovine (Bos indicus or crossbred Bos taurus) embryos were produced in vitro according to standard procedures. Two-cell Bos indicus embryos were submitted to electrofusion with varying numbers of pulses (1 or 2), voltages (0.4, 0.5, 0.75, 1.0, 1.4 and 5.0 kV/cm) and time (20, 25, 50 and 60 μs) to produce tetraploid embryos. Electrofused embryos were cultured with crossbred non-fused embryos to form chimeras that developed until the blastocyst stage. The best fusion parameter was 0.75 kV/cm for 60 μs. Four chimeric blastocysts (tetraploid Nelore with diploid crossbred Holstein) were formed after 31 attempts in 4 replicates (13%). We established an optimal procedure for the production of tetraploid Bos indicus (4n) embryos and embryonic chimeras by aggregation of crossbred Bos taurus (2n) with Bos indicus (4n) embryos. This technique would be valid in applied research, by producing exclusively taurine calves, but with placental elements from the Bos indicus breed, following transfer of these chimeras into recipient cows.

  16. Morphokinetic-related response to stress in individually cultured bovine embryos.

    Science.gov (United States)

    Silva, T; Santos, E C; Annes, K; Soares, C A; Leite, R F; Lima, C B; Milazzotto, M P

    2016-09-15

    The kinetics of in vitro-produced (IVP) bovine embryos is related to embryo viability, metabolism, and epigenetic patterns. Therefore, we believe that embryos with different speeds of development also respond differently to stress. In the present study, we performed global metabolic analysis (matrix-assisted laser desorption ionization time of flight mass spectrometry [MALDI-TOF]) of culture media, characterized apoptotic events (Terminal deoxynucleotidyl transferase dUTP nick end labeling [TUNEL] and caspase quantitation), and quantified transcript abundance of stress-related gene (real-time quantitative polymerase chain reaction [qRT-PCR]) in IVP bovine embryos with different developmental kinetics to investigate possible markers of stress response. For this purpose, embryos were considered "fast" if they presented four or more cells at 40 hours post insemination (hpi). Embryos presenting two cells at this time were classified as "slow". Evaluations were performed at 40 hpi, 112 hpi, and 186 hpi. Metabolome analysis revealed several metabolites differentially represented between groups at all time points related with energy, lipid and amino acids metabolism, and stress response. There was no difference in TUNEL positive cells between groups in any of the time points analyzed. Nevertheless, at 112 hpi, classified as a critical phase because of the genome activation, the amount of caspase 3 and 7 and total caspase were higher in slow when compared to fast group. Transcript abundance analysis of candidate genes (GRP78, HSP60, SOD1, and MORF4L2) was also different among groups. In conclusion, IVP bovine embryos of different development speeds respond differentially to the environmental stress leading to different metabolome patterns and apoptosis activation throughout the culture.

  17. Morphokinetic-related response to stress in individually cultured bovine embryos.

    Science.gov (United States)

    Silva, T; Santos, E C; Annes, K; Soares, C A; Leite, R F; Lima, C B; Milazzotto, M P

    2016-09-15

    The kinetics of in vitro-produced (IVP) bovine embryos is related to embryo viability, metabolism, and epigenetic patterns. Therefore, we believe that embryos with different speeds of development also respond differently to stress. In the present study, we performed global metabolic analysis (matrix-assisted laser desorption ionization time of flight mass spectrometry [MALDI-TOF]) of culture media, characterized apoptotic events (Terminal deoxynucleotidyl transferase dUTP nick end labeling [TUNEL] and caspase quantitation), and quantified transcript abundance of stress-related gene (real-time quantitative polymerase chain reaction [qRT-PCR]) in IVP bovine embryos with different developmental kinetics to investigate possible markers of stress response. For this purpose, embryos were considered "fast" if they presented four or more cells at 40 hours post insemination (hpi). Embryos presenting two cells at this time were classified as "slow". Evaluations were performed at 40 hpi, 112 hpi, and 186 hpi. Metabolome analysis revealed several metabolites differentially represented between groups at all time points related with energy, lipid and amino acids metabolism, and stress response. There was no difference in TUNEL positive cells between groups in any of the time points analyzed. Nevertheless, at 112 hpi, classified as a critical phase because of the genome activation, the amount of caspase 3 and 7 and total caspase were higher in slow when compared to fast group. Transcript abundance analysis of candidate genes (GRP78, HSP60, SOD1, and MORF4L2) was also different among groups. In conclusion, IVP bovine embryos of different development speeds respond differentially to the environmental stress leading to different metabolome patterns and apoptosis activation throughout the culture. PMID:27298151

  18. 1-cell embryo transfer into pseudopregnant recipient female mouse

    OpenAIRE

    sprotocols

    2014-01-01

    ### Abstract 1-cell embryo transfer is best performed after allowing injected embryos a little recovery time in culture. This allows better evaluation of the cells' survival - those that have been damaged during the injection process will undergo cytoplasmic condensation, causing the cellular material to become less glossy and darker in color as the cytoplasm shrinks away from the zona pellucida. This should be balanced against the increased survival rate with decreased in vitro exposure....

  19. Coagulansin-A has beneficial effects on the development of bovine embryos in vitro via HSP70 induction

    OpenAIRE

    Khan, Imran; Lee, Kyeong-Lim; Fakruzzaman, Md.; Song, Seok-Hwan; Ihsan-ul-Haq,; Mirza, Bushra; Yan, Chang Guo; Kong, Il-Keun

    2016-01-01

    Treatment with the steroidal lactone, coagulansin-A, improves bovine oocyte maturation and embryo development in vitro by inducing heat shock protein 70 (HSP70), which reduces the levels of reactive oxygen species (ROS), DNA damage and inflammation.

  20. Children conceived by in vitro fertilisation after fresh embryo transfer

    OpenAIRE

    D'Souza, S; Rivlin, E; Cadman, J; Richards, B; Buck, P; Lieberman, B.

    1997-01-01

    AIMS—To compare the outcome in in vitro fertilisation (IVF) children (after fresh embryo transfer) from multiple and singleton births with one another, and with normally conceived control children.
METHODS—A cohort of 278 children (150 singletons, 100 twins, 24 triplets and four quadruplets), conceived by IVF after three fresh embryos had been transferred, born between October 1984 and December 1991, and 278 normally conceived control children (all singletons), were followed up for four years...

  1. Expression of intracellular interferon-alpha confers antiviral properties in transfected bovine fetal fibroblasts and does not affect the full development of SCNT embryos.

    Directory of Open Access Journals (Sweden)

    Dawei Yu

    Full Text Available Foot-and-mouth disease, one of the most significant diseases of dairy herds, has substantial effects on farm economics, and currently, disease control measures are limited. In this study, we constructed a vector with a human interferon-α (hIFN-α (without secretory signal sequence gene cassette containing the immediate early promoter of human cytomegalovirus. Stably transfected bovine fetal fibroblasts were obtained by G418 selection, and hIFN-α transgenic embryos were produced by somatic cell nuclear transfer (SCNT. Forty-six transgenic embryos were transplanted into surrogate cows, and five cows (10.9% became pregnant. Two male cloned calves were born. Expression of hIFN-α was detected in transfected bovine fetal fibroblasts, transgenic SCNT embryos, and different tissues from a transgenic SCNT calf at two days old. In transfected bovine fetal fibroblasts, expression of intracellular IFN-α induced resistance to vesicular stomatitis virus infection, increased apoptosis, and induced the expression of double-stranded RNA-activated protein kinase gene (PKR and the 2'-5'-oligoadenylate synthetase gene (2'-5' OAS, which are IFN-inducible genes with antiviral activity. Analysis by qRT-PCR showed that the mRNA expression levels of PKR, 2'-5' OAS, and P53 were significantly increased in wild-type bovine fetal fibroblasts stimulated with extracellular recombinant human IFN-α-2b, showing that intracellular IFN-α induces biological functions similar to extracellular IFN-α. In conclusion, expression of intracellular hIFN-α conferred antiviral properties in transfected bovine fetal fibroblasts and did not significantly affect the full development of SCNT embryos. Thus, IFN-α transgenic technology may provide a revolutionary way to achieve elite breeding of livestock.

  2. Establishment of multiple ovulation and embryo transfer (MOET) technology for goats in Sri Lanka

    International Nuclear Information System (INIS)

    Multiple ovulation and embryo transfer (MOET) has been done successfully in goats in some countries (Chen et al., 2008). It can be used to multiply the genetically superior animals and to make elite herds with increased production potential. There have been no previous reports on successful MOET in goats in Sri Lanka. Therefore, this study was carried out to establish techniques for in vivo production and transfer of goat embryos in Sri Lanka. Genetically superior does (n = 7) were subjected to super ovulation for in vivo embryo production using a protocol modified from that of Batt et al (1993). Progesterone releasing intravaginal pessaries (45 mg, Cronolone) was inserted on Day 1 of the programme. The does in group 1 (n = 3) were stimulated on Day 8 with injections of pure porcine Follicular Stimulating Hormone (pFSH), while those in group 2 (n = 4) were stimulated with pure ovine Follicular Stimulating Hormone (oFSH). Equine Chorionic Gonadotrophin (eCG) was given to all does in the evening of Day 8. Subsequent injections of pFSH (group 1) or oFSH (group 2) were given in the morning and evening on Day 9 and Day 10. All does were injected with prostaglandin analogue (263 μg/ml cloprostenol sodium) in the morning of Day 9 and vaginal pessaries were removed in the evening of Day 10. On Day 11, pFSH or oFSH was injected in the morning and Gonadotropin Releasing Hormone (GnRH) was injected in the evening. Immediately after the GnRH injection does were exposed to breeding with a genetically superior Jamnapari buck for 48 hours. Embryos were collected surgically 7 d after oestrus, by flushing of the uterus with embryo flushing medium containing lactated Ringer's solution with 1% bovine serum albumin at 37 deg C through a mid ventral laparotomy. The quality of the embryos was assessed microscopically and those considered to be of good and excellent quality were transferred surgically to oestrus synchronized recipient goats (n = 6) 7 d post-oestrus. The ovarian

  3. Direct and Osmolarity-Dependent Effects of Glycine on Preimplantation Bovine Embryos.

    Science.gov (United States)

    Herrick, Jason R; Lyons, Sarah M; Greene, Alison F; Broeckling, Corey D; Schoolcraft, William B; Krisher, Rebecca L

    2016-01-01

    Concentrations of glycine (Gly) in embryo culture media are often lower (~0.1 mM) than those in oviductal or uterine fluids (≥1.2 mM). The objective of this study was to determine direct and osmolarity-dependent effects of physiological concentrations of Gly on blastocyst formation and hatching, cell allocation to the trophectoderm (TE) and inner cell mass (ICM), and metabolic activity of bovine embryos. In experiment 1, zygotes were cultured with 100 or 120 mM NaCl and 0 or 1 mM Gly for the first 72 h of culture. Blastocyst formation and hatching were improved (Pcultured with 100 compared to 120 mM NaCl. Inclusion of 1 mM Gly improved (Pcultured with 120 mM NaCl, suggesting bovine embryos can utilize Gly as an osmolyte. In experiment 2, embryos were cultured with 0.1, 1.1, 2.1, or 4.1 mM Gly (100 mM NaCl) for the final 96 h of culture. Blastocyst development was not affected (P>0.05) by Gly, but hatching (0.1 mM Gly, 18.2%) was improved (Pcultured with 1.1 (31.4%) or 2.1 (29.4%) mM Gly. Blastocyst, TE, and ICM cell numbers were not affected (P>0.05) by Gly in either experiment. Blastocysts produced alanine, glutamine, pyruvate, and urea and consumed aspartate, but this metabolic profile was not affected (P>0.05) by Gly. In conclusion, Gly (1.0 mM) improves the development of both early and late stage embryos, but beneficial effects are more pronounced for early embryos exposed to elevated osmolarity. PMID:27459477

  4. Opportunities for embryo transfer in the age of DNA testing

    Science.gov (United States)

    Embryo transfer (ET) has contributed to increasing selection intensity in cattle breeding for many years. Preimplantation DNA testing offers the opportunity to increase selection response further through increasing within-family selection intensity. Further increases in between-family selection inte...

  5. Efficacy of in vitro embryo transfer in lactating dairy cows using fresh or vitrified embryos produced in a novel embryo culture medium.

    Science.gov (United States)

    Block, J; Bonilla, L; Hansen, P J

    2010-11-01

    Objectives were to determine whether pregnancy success could be improved in lactating cows with timed embryo transfer when embryos were produced in vitro using a medium designed to enhance embryo development and survival after cryopreservation. In experiment 1, embryos (n=569 to 922) were cultured in either modified synthetic oviduct fluid or a serum-free medium, Block-Bonilla-Hansen-7 (BBH7). Development to the blastocyst stage was recorded at d 7, and selected blastocysts (n=79 to 114) were vitrified using open pulled straws. Culture of embryos in BBH7 increased development to the blastocyst stage (41.9±2.0 vs. 14.7±2.0%) and advanced blastocyst stages (expanded, hatching, hatched; 31.1±1.3 vs. 6.4±1.3%) at d 7 and resulted in higher hatching rates at 24h postwarming compared with embryos cultured in modified synthetic oviduct fluid (59.0±0.5 vs. 26.7±0.5%). In experiment 2, embryos were produced using X-sorted semen and cultured in BBH7. At d 7 after insemination, embryos were transferred fresh or following vitrification. Lactating Holstein cows were either subjected to timed artificial insemination (TAI) on the day of presumptive ovulation or used as embryo recipients 7 d later. Embryo recipients received an embryo if a corpus luteum was present. The percentage of cows pregnant at d 32, 46, and 76 of gestation was higher among cows that received fresh embryos compared with TAI cows or cows that received vitrified embryos. At d 76, for example, the proportion and percentage pregnant was 47/150 (31.3%) for cows subjected to TAI, 48/95 (50.5%) for cows receiving fresh embryos, and 39/141 (27.7%) for cows receiving a vitrified embryo. No difference was observed in the percentage of cows pregnant among TAI cows and those that received vitrified embryos. There was a service or transfer number × treatment interaction because differences in pregnancy rate between embryo transfer recipients and cows bred by TAI were greater for cows with more than 3 services or

  6. Comparison of transcriptomic landscapes of bovine embryos using RNA-Seq

    Directory of Open Access Journals (Sweden)

    Khatib Hasan

    2010-12-01

    Full Text Available Abstract Background Advances in sequencing technologies have opened a new era of high throughput investigations. Although RNA-seq has been demonstrated in many organisms, no study has provided a comprehensive investigation of the bovine transcriptome using RNA-seq. Results In this study, we provide a deep survey of the bovine embryonic transcriptomes, the first application of RNA-seq in cattle. Embryos cultured in vitro were used as models to study early embryonic development in cattle. RNA amplified from limited amounts of starting total RNA were sequenced and mapped to the reference genome to obtain digital gene expression at single base resolution. In particular, gene expression estimates from more than 1.6 million unannotated bases in 1785 novel transcribed units were obtained. We compared the transcriptomes of embryos showing distinct developmental statuses and found genes that showed differential overall expression as well as alternative splicing. Conclusion Our study demonstrates the power of RNA-seq and provides further understanding of bovine preimplantation embryonic development at a fine scale.

  7. Culture of bovine embryos in polyester mesh sections: the effect of pore size and oxygen tension on in vitro development.

    Science.gov (United States)

    Somfai, T; Inaba, Y; Aikawa, Y; Ohtake, M; Kobayashi, S; Akai, T; Hattori, H; Konishi, K; Imai, K

    2010-12-01

    The purpose of this study was to assess the feasibility of polyester mesh culture for the in vitro production of bovine embryos, as polyester mesh is an alternative way for tracking individual embryos throughout culture using time-lapse cinematography (TLC). Bovine embryos were isolated during in vitro culture using sections of three different polyethylene terephthalate (PET) mesh products. In vitro matured and fertilized bovine oocytes were cultured in the 217 × 217, 230 × 230 or 238 × 238-μm openings of PET mesh sections or in simple micro-drops (control) for 7 days under either 20% or 5% O(2) tensions. No difference in embryo developmental rates was found between the culture groups in terms of cleavage, blastocyst formation and blastocyst expansion irrespective of O(2) tension. In contrast, under 20% O(2) tension, blastocysts that developed in PET mesh with 217 × 217-μm opening had significantly higher numbers of total and trophectoderm (TE) cells than control embryos; however, the numbers and proportions of inner cell mass (ICM) cells did not differ. Under 5% O(2) tension, no difference was found among the culture groups in the numbers of total, ICM and TE cells in embryos. All three PET mesh products investigated in this study were proven to be effective to prevent embryo movement. The results demonstrate that bovine embryos can be cultured in PET mesh sections without negative side-effects and suggest that embryo distance determined by the mesh affects embryo quality at atmospheric oxygen tension. Polyethylene terephthalate mesh with 217 × 217-μm openings was found to be the most suitable for further application in TLC. PMID:19845884

  8. The dangers of disease transmission by artificial insemination and embryo transfer.

    Science.gov (United States)

    Philpott, M

    1993-01-01

    This review summarizes the major infectious diseases of the three major agricultural species (cattle, sheep and pigs) and horses, and presents the evidence for and against the possibility of infectious agents being transmitted between animals via the venereal route or by the use of semen or early embryos in commercial artificial insemination (AI) or embryo transfer (ET). Cattle feature most prominently in the widespread distribution of frozen semen, and national and international organizations have set out guidelines to work towards disease-free bull studs with semen free from potential pathogens. With the control of major epizootic diseases, attention has been focused on such diseases as IBR, BVD and blue tongue, where clinical signs are rarely evident but the detection of virus in semen is of great importance. New information on the relevance of bacterial disease such as Mycobacterium paratuberculosis, campylobacteriosis and leptospirosis is reviewed, along with details of the mycoplasma and ureaplasma species of the bull's genital tract. Bovine spongiform encephalopathy (BSE) has attracted much research and semen is not regarded as a source of infection. New work on the pathogenesis of a number of diseases and the use of new biotechnology in diagnosis is included. The International Embryo Transfer Society (IETS) has encouraged a great deal of experimental work--much originating in Canada--on the risk of transmission of disease from donors to recipients via a 7-day-old blastocyst. There has been much success in demonstrating that with an approved protocol of handling the embryos, to date there is very little danger in disease transmission with both viruses and bacteria. The mycoplasma group appear more intractable and the role of BSE is still being evaluated. In sheep, scrapie, Brucella ovis infection and blue tongue feature in current work. In the pig there is a surge in international movement of pig semen, and Aujeszky's disease and the new so-called Blue Ear

  9. Generation of bovine transgenics using somatic cell nuclear transfer

    Directory of Open Access Journals (Sweden)

    Stice Steven L

    2003-11-01

    Full Text Available Abstract The ability to produce transgenic animals through the introduction of exogenous DNA has existed for many years. However, past methods available to generate transgenic animals, such as pronuclear microinjection or the use of embryonic stem cells, have either been inefficient or not available in all animals, bovine included. More recently somatic cell nuclear transfer has provided a method to create transgenic animals that overcomes many deficiencies present in other methods. This review summarizes the benefits of using somatic cell nuclear transfer to create bovine transgenics as well as the possible opportunities this method creates for the future.

  10. Homologous recombination and non-homologous end-joining repair pathways in bovine embryos with different developmental competence

    International Nuclear Information System (INIS)

    This study investigated the expression of genes controlling homologous recombination (HR), and non-homologous end-joining (NHEJ) DNA-repair pathways in bovine embryos of different developmental potential. It also evaluated whether bovine embryos can respond to DNA double-strand breaks (DSBs) induced with ultraviolet irradiation by regulating expression of genes involved in HR and NHEJ repair pathways. Embryos with high, intermediate or low developmental competence were selected based on the cleavage time after in vitro insemination and were removed from in vitro culture before (36 h), during (72 h) and after (96 h) the expected period of embryonic genome activation. All studied genes were expressed before, during and after the genome activation period regardless the developmental competence of the embryos. Higher mRNA expression of 53BP1 and RAD52 was found before genome activation in embryos with low developmental competence. Expression of 53BP1, RAD51 and KU70 was downregulated at 72 h and upregulated at 168 h post-insemination in response to DSBs induced by ultraviolet irradiation. In conclusion, important genes controlling HR and NHEJ DNA-repair pathways are expressed in bovine embryos, however genes participating in these pathways are only regulated after the period of embryo genome activation in response to ultraviolet-induced DSBs.

  11. Homologous recombination and non-homologous end-joining repair pathways in bovine embryos with different developmental competence

    Energy Technology Data Exchange (ETDEWEB)

    Henrique Barreta, Marcos [Universidade Federal de Santa Catarina, Campus Universitario de Curitibanos, Curitibanos, SC (Brazil); Laboratorio de Biotecnologia e Reproducao Animal-BioRep, Universidade Federal de Santa Maria, Santa Maria, RS (Brazil); Garziera Gasperin, Bernardo; Braga Rissi, Vitor; Cesaro, Matheus Pedrotti de [Laboratorio de Biotecnologia e Reproducao Animal-BioRep, Universidade Federal de Santa Maria, Santa Maria, RS (Brazil); Ferreira, Rogerio [Centro de Educacao Superior do Oeste-Universidade do Estado de Santa Catarina, Chapeco, SC (Brazil); Oliveira, Joao Francisco de; Goncalves, Paulo Bayard Dias [Laboratorio de Biotecnologia e Reproducao Animal-BioRep, Universidade Federal de Santa Maria, Santa Maria, RS (Brazil); Bordignon, Vilceu, E-mail: vilceu.bordignon@mcgill.ca [Department of Animal Science, McGill University, Ste-Anne-De-Bellevue, QC (Canada)

    2012-10-01

    This study investigated the expression of genes controlling homologous recombination (HR), and non-homologous end-joining (NHEJ) DNA-repair pathways in bovine embryos of different developmental potential. It also evaluated whether bovine embryos can respond to DNA double-strand breaks (DSBs) induced with ultraviolet irradiation by regulating expression of genes involved in HR and NHEJ repair pathways. Embryos with high, intermediate or low developmental competence were selected based on the cleavage time after in vitro insemination and were removed from in vitro culture before (36 h), during (72 h) and after (96 h) the expected period of embryonic genome activation. All studied genes were expressed before, during and after the genome activation period regardless the developmental competence of the embryos. Higher mRNA expression of 53BP1 and RAD52 was found before genome activation in embryos with low developmental competence. Expression of 53BP1, RAD51 and KU70 was downregulated at 72 h and upregulated at 168 h post-insemination in response to DSBs induced by ultraviolet irradiation. In conclusion, important genes controlling HR and NHEJ DNA-repair pathways are expressed in bovine embryos, however genes participating in these pathways are only regulated after the period of embryo genome activation in response to ultraviolet-induced DSBs.

  12. Green tea polyphenols added to IVM and IVC media affect transcript abundance, apoptosis, and pregnancy rates in bovine embryos.

    Science.gov (United States)

    Wang, Zhengguang; Fu, Chunquan; Yu, Songdong

    2013-01-01

    Three experiments were conducted to examine the effects of green tea polyphenols (GTP) during IVM and IVC on apoptosis and relative transcript abundance (RA) of three genes controlling antioxidant enzymes, as well as subsequent pregnancy rates. In experiment 1, oocytes were matured in the presence of 0, 10, 15, or 25 μM GTP for 24 hours. The GTP dose applied to IVM medium was followed by the same dose supplemented to IVC medium, so oocytes and embryos of a given group were cultured in similar conditions. This resulted in a total of four groups (three experimental groups and the control). After IVF, presumptive zygotes were cultured in medium containing 0 to 25 μM GTP for 8 days. The addition of 15 μM GTP during IVM and IVC increased RA of SOD1, CAT, and GPX genes in blastocysts compared with the control (P < 0.05). Increase in GTP doses from 15 to 25 μM did not further increase the transcript level. In experiment 2, effects of GTP doses on apoptosis were investigated in bovine blastocysts. Two of the applied GTP doses (10 and 15 μM) decreased the apoptotic index (AI) in blastocysts (7.4% and 6.2% respectively) compared with the control (9.3%; P < 0.05). However, the highest GTP dose used (25 μM) caused an increase in AI compared with a dose of 15 μM (P < 0.05). Considering the results of experiment 1 and 2, the effects of 15 μM GTP treatment during IVM and IVC on pregnancy rate was evaluated after embryo transfer in experiment 3. Cows receiving embryos treated with 15 μM GTP had higher pregnancy rates on Day 30 (34.8% vs. 28.6%) and Day 60 (34.8% vs. 23.9%) than those receiving control embryos (P < 0.05). In conclusion, addition of 15 μM GTP during IVM and IVC improved pregnancy rates; this improvement seemed to be associated with the increase of RA of antioxidant enzyme genes and the decrease in AI in bovine blastocysts.

  13. Ovarian response, embryo recovery and results of embryo transfer in a Hungarian native pig breed.

    Science.gov (United States)

    Rátky, J; Brüssow, K P; Solti, L; Torner, H; Sarlós, P

    2001-09-15

    The objective of the study was to use embryo transfer (ET) for propagation of the Swallow Belly Mangalica population. Mangalica is a native Hungarian pig breed adapted to extreme climate and housing conditions and distinguished for excellent meat and fat quality. However, due to their weak reproductive characteristics and relatively high fat proportion, Mangalica pigs have been replaced by modern breeds. Now, there is an increased interest again to safeguard the properties of this breed. We conducted two experiments. First, we used a total of 18 puberal Mangalica gilts to determine an optimal superovulatory treatment. Following estrus synchronization with Regumate, we injected gilts with either 750, 1000 or 1250 IU PMSG, followed by 750 IU hCG 80 h later. We scanned ovaries endoscopically 3 days after hCG administration. The application of 1000 and 1250 IU PMSG resulted in a higher rate of ovulation compared to 750 IU (24.2 +/- 3.6 and 21.0 +/- 2.3 vs. 13.7 +/- 2.7 Pfollicular cysts increased after administration of 1250 IU PMSG compared to 750 and 1000 IU (2.0 +/- 1.3 vs. 0.3 +/- 0.7 and 0.2 +/- 0.3, P<0.05). Thus, we chose 1000 IU PMSG for further stimulation of Mangalica gilts. In the second experiment, we induced superovulation in 10 Mangalica donor gilts by 1000 IU PMSG and 750 IU hCG. Gilts were fixed-time inseminated, and then five days later embryo collection was carried out surgically (n=6) or endoscopically (n=4). Out of the 187 ova recovered, 92.5% were at the morula/blastocyst stage. The embryo recovery rate was higher following surgical flushing than following endoscopy (91.5 +/- 4.4% vs. 71.4 +/- 12.7%, P<0.05). Altogether 143 embryos were transferred surgically or endoscopically into 8 Landrace recipients. Surgical and endoscopic transfer of Mangalica embryos into Landrace gilts resulted in pregnancies in 3 and 2 gilts, respectively; thus the overall farrowing rate was 62.5%. The birth of 59 Mangalica piglets from 5 embryo recipients equals an

  14. Time-lapse cinematography-compatible polystyrene-based microwell culture system: a novel tool for tracking the development of individual bovine embryos.

    Science.gov (United States)

    Sugimura, Satoshi; Akai, Tomonori; Somfai, Tamás; Hirayama, Muneyuki; Aikawa, Yoshio; Ohtake, Masaki; Hattori, Hideshi; Kobayashi, Shuji; Hashiyada, Yutaka; Konishi, Kazuyuki; Imai, Kei

    2010-12-01

    We have developed a polystyrene-based well-of-the-well (WOW) system using injection molding to track individual embryos throughout culture using time-lapse cinematography (TLC). WOW culture of bovine embryos following in vitro fertilization was compared with conventional droplet culture (control). No differences between control- and WOW-cultured embryos were observed during development to the blastocyst stage. Morphological quality and inner cell mass (ICM) and trophectoderm (TE) cell numbers were not different between control- and WOW-derived blastocysts; however, apoptosis in both the ICM and TE cells was reduced in WOW culture (P < 0.01). Oxygen consumption in WOW-derived blastocysts was closer to physiological level than that of control-derived blastocysts. Moreover, WOW culture improved embryo viability, as indicated by increased pregnancy rates at Days 30 and 60 after embryo transfer (P < 0.05). TLC monitoring was performed to evaluate the cleavage pattern and the duration of the first cell cycle of embryos from oocytes collected by ovum pickup; correlations with success of pregnancy were determined. Logistic regression analysis indicated that the cleavage pattern correlated with success of pregnancy (P < 0.05), but cell cycle length did not. Higher pregnancy rates (66.7%) were observed for animals in which transferred blastocysts had undergone normal cleavage, identified by the presence of two blastomeres of the same size without fragmentation, than among those with abnormal cleavage (33.3%). These results suggest that our microwell culture system is a powerful tool for producing and selecting healthy embryos and for identifying viability biomarkers. PMID:20739661

  15. Use of versapoint to refashion the cervical canal to overcome unusually difficult embryo transfers and improve in-vitro fertilization-embryo transfer outcome: A case series

    OpenAIRE

    Nalini Mahajan; Ila Gupta

    2011-01-01

    Background : Smooth atraumatic embryo transfer is paramount for the success of in-vitro fertilization (IVF). In difficult cases, cervical canal manipulation may be required. Aim : To see if surgical correction of the cervical canal or cervical canal refashioning could improve ease of embryo transfer. Setting : Private infertility and IVF hospital. Design : Prospective study. Materials and Methods : Patients: 11 women with failed 1-3 IVF cycles with history of extremely difficult embryo transf...

  16. BIRTH OF TWIN CALVES THROUGH MULTIPLE OVULATION AND EMBRYO TRANSFER: A CASE REPORT

    Directory of Open Access Journals (Sweden)

    P.S. Sengupta

    2013-06-01

    Full Text Available A surrogate Gir cow delevered twin calves after receiving embryo from a donor Sahiwal cow at Multiple Ovulation and Embryo Transfer laboratory, Haringhata, West Bengal, India is reported.

  17. The effect of IVM and IVC media on in vitro development of bovine embryos

    Directory of Open Access Journals (Sweden)

    E.T Mergawati

    2000-12-01

    Full Text Available The purpose of this study was to examine the effect of medium combination of IVM and IVC on the in vitro development of bovine embryos. The study involved 4 groups in a 2 (IVM medium x 2 (IVC medium factorial in a randomized block design. Each group was replicated for 5 times. The treatments were as follows: TCM-199/CR1aa (T1; TCM-199/SOF (T2; B- 199/CR1aa (T3 and B-199/SOF (T4. Oocytes were aspirated from ovaries collected at local abattoirs using aspiration medium of PBS supplemented with 3% FCS and 0.1% Penicillin and Streptomycin. The oocytes were matured in medium of TCM-199 or B-199 supplemented with 10% FCS, hormones: 10μg/ml FSH+ 10μg/ml hCG+ 1μg/ml Estradiol. Maturation was maintained at 37oC for 22 hours in 5% CO2 incubator with high humidity. A method of BRACKETT & Oliphant (BO was used to fertilize the matured oocytes. The fertilization was incubated for 7 hours in the 5% CO2 incubator. Two culture media of CR1aa or SOF/AA/BSA were used to develop the fertilized oocytes undergo to morula and blastocyst embryos. The findings showed that the proportion of oocytes cleaved and formation of blastocysts were affected significantly by a combination of IVM and IVC media (P<0.05. A combination of B-199/SOF (T4 resulted in a higher blastocyst rate (32% than others (T3= 29%; T2=T1= 23%. This study suggests that either SOF/AA/BSA or CR1aa has similar competence in development of bovine embryos in vitro.

  18. Tumor necrosis factor alpha inhibits in vitro bovine embryo development through a prostaglandin mediated mechanism

    Directory of Open Access Journals (Sweden)

    Jackson Lauren R

    2012-03-01

    Full Text Available Abstract Mastitis or other infectious diseases have been related to reduced fertility in cattle. Inflammatory cytokines such as tumor necrosis factor α (TNFα are released in response to infection and may have negative effects on embryo development. In the current study the effect of exposure to TNFα on the development of in vitro fertilized bovine embryos was examined. Indomethacin, a prostaglandin synthesis inhibitor, was used to determine if blockade of prostaglandin synthesis would alter the effects of TNFα. Ovaries were obtained from a local abattoir and immature COC were isolated from 2-10 mm follicles, in vitro matured and fertilized. After fertilization, groups of presumptive zygotes were randomly placed into either control development medium, medium containing 25 ng/mL TNFα or medium containing 25 ng/mL TNFα plus 1 μg/mL indomethacin. The proportion of blastocysts formed was assessed at day 7 of culture. Fewer embryos exposed to TNFα alone reached the blastocyst stage (17.5 ± 2.4%, P

  19. In Vitro Developmental Potential of Cloned Embryos Derived from Bovine Somatic Cells and Rabbits Oocyte

    Institute of Scientific and Technical Information of China (English)

    LIU Ya; LI Bin; ZHAO Huan; CHENG Li-zi; ZHANG Xiao-rong; CHEN Da-yuan; ZHANG Yun-hai; ZHANG Zhi-guo; JING Ren-tao; WANG Cun-li; ZHANG Mei-lin; LI Dong-wei

    2003-01-01

    180 reconstituted embryos were produced by nuclear transplantation using bovine ear fibroblasts at G0 or non-G0 stage as donor nuclei and oocytes collected from superovulated multiparous or young rabbits as recipients. After cultivation in two kinds of medium M199+ 10%FBS or RD+ 10%FBS, 112 of them developed to 2-cell stage (62.2%) and 26 to morula stage (14.4%) and 20 of them eventually developed to blastocyst stage (11. 1% ). There is no significant difference for the cleavage rates in two groups of reconstituted embryos derived from G0-stage and non-G0 stage donor cells respectively. However, G0-stage donor cells could result in higher rate of 8-cell - 16-cell stage embryos significantly (P<0.05), as well as higher rate of blastocysts (P<0.01). It seems that using two different culture systems had no significant effects on the cleavage rate, morula rate or blastocyst rate (P>0.05).

  20. Monitoring in-vitro bovine embryo development during the first days after fertilization (Conference Presentation)

    Science.gov (United States)

    Kandel, Mikhail E.; Rubessa, Marcello; Fernandes, Daniel; Nguyen, Tan H.; Wheeler, Matthew B.; Popescu, Gabriel

    2016-03-01

    Conventional label-based contrast enhancement techniques (e.g., fluorescence) frequently modify the genetic makeup of tagged cells, making them poor candidates for use in in-vitro fertilization applications. Instead, we choose a label-free form of contrast, based on interferometric imaging, sensitive to optical path length differences. Compared to, single HeLa cells, typical mammalian ova and embryos are more than an order of magnitude thicker. As a result, regions of large phase variation lead to phase wrapping and an overall reduction in signal intensity occurs due to multiple scattering. These effects manifest themselves in low-spatial frequencies (blurs), with the desired details buried in the background. We present a phase shifting interferometer that yields the derivative of the phase, a quantity whose value is particularly sensitive to local variations and fine details. We demonstrate that our new real-time imaging platform is valuable in measuring the multiday development of bovine embryos. Reconstructing the derivative of the image phase and amplitude, we characterize the motion of previously low-contrast structures, which are relevant for embryo viability tests.

  1. Coagulansin-A has beneficial effects on the development of bovine embryos in vitro via HSP70 induction

    Science.gov (United States)

    Khan, Imran; Lee, Kyeong-Lim; Fakruzzaman, Md.; Song, Seok-Hwan; Ihsan-ul-Haq; Mirza, Bushra; Yan, Chang Guo; Kong, Il-Keun

    2016-01-01

    Coagulansin-A (withanolide) is the steroidal lactone obtained from Withania coagulans which belong to Solanaceae family. The present study investigated the effects of coagulansin-A on bovine oocyte maturation and embryo development in vitro. All these oocytes were aspirated from the ovaries obtained from Korean Hanwoo cows at a local abattoir. To determine whether coagulansin-A has beneficial effects on bovine oocyte maturation in vitro, 355 oocytes per group (control and treated) in seven replicates were subjected with different concentrations (1, 2.5, 5, 7.5 and 10 μM) of coagulansin-A. The coagulansin-A was added in the in vitro maturation (IVM) media followed by in vitro fertilization (IVF) and then in vitro culture (IVC). Only treatment with 5 μM coagulansin-A remarkably (P<0.05) improved embryos development (Day 8 blastocyst) having 27.30 and 40.01% for control and coagulansin-A treated groups respectively. Treatment with 5 μM coagulansin-A significantly induced activation of heat shock protein 70 (HSP70) (P<0.05). Immunofluorescence analysis revealed that 5 μM coagulansin-A treatment also significantly inhibited oxidative stress and inflammation during bovine embryo development in vitro by decreasing 8-oxoguanosine (8-OxoG) (P<0.05) and nuclear factor-κB (NF-κB) (P<0.05). The expressions of HSP70 and NF-κB were also conformed through real-time PCR (RT-PCR). Additionally, the terminal deoxynucleotidyl transferase dUTP nick-end labelling (TUNEL) assay confirmed that coagulansin-A treatment significantly improved the embryo quality and reduced bovine embryo DNA damage (P<0.05). The present study provides new information regarding the mechanisms by which coagulansin-A promotes bovine embryo development in vitro. PMID:26831738

  2. Comparison of pregnancy rate between fresh embryo transfers and frozen-thawed embryo transfers following ICSI treatment

    Directory of Open Access Journals (Sweden)

    Zahra Basirat

    2016-01-01

    Full Text Available Background: The use of assisted reproductive technology (ART is increasing in the world. The rate, efficacy and safety of ART are very different among countries. There is an increase in the use of intra cytoplasmic sperm injection (ICSI, single fresh embryo transfer (ET and frozen-thawed embryo transfer (FET. Objective: The objective of this study was to compare pregnancy rate in fresh ET and FET. Materials and Methods: In this retrospective cross-sectional study 1014 ICSI-ET cycles (426 fresh ET and 588 FET from 753 women undergoing ICSI treatment referred to Fatemezahra Infertility and Reproductive Health Research Center in Babol, Iran from 2008 to 2013 were reviewed. Results: There were no significant differences between biochemical pregnancy rate (23% versus 18.8%, OR 1.301; 95% CI .95-1.774, gestational sac (95.6% versus 100% in FET, OR 0.60; 95% CI 0.54-0.67, and fetal heart activity (87.2% versus 93.6% OR .46; 95% CI .16-1.32 in fresh ET and FET cycles, respectively. P< 0.05 was considered statistically significant for all measures. Conclusion: Although, the result showed no significantly difference between the fresh ET and the FET cycles, however the embryos are able to be stored for subsequent ART. Therefore, we recommend FET cycles as an option alongside the fresh ET.

  3. Regulation of pluripotency of inner cell mass and growth and differentiation of trophectoderm of the bovine embryo by colony stimulating factor 2.

    Science.gov (United States)

    Dobbs, Kyle B; Khan, Firdous A; Sakatani, Miki; Moss, James I; Ozawa, Manabu; Ealy, Alan D; Hansen, Peter J

    2013-12-01

    Colony-stimulating factor 2 (CSF2) enhances competence of the bovine embryo to establish and maintain pregnancy after the embryo is transferred into a recipient. Mechanisms involved could include regulation of lineage commitment, growth, or differentiation of the inner cell mass (ICM) and trophectoderm (TE). Experiments were conducted to evaluate regulation by CSF2 of pluripotency of the ICM and differentiation and growth of the TE. Embryos were cultured with 10 ng/ml recombinant bovine CSF2 or a vehicle control from Days 5 to 7 or 6 to 8 postinsemination. CSF2 increased the number of putative zygotes that developed to blastocysts when the percent of embryos becoming blastocysts in the control group was low but decreased blastocyst yield when blastocyst development in controls was high. ICM isolated from blastocysts by lysing the trophectoderm using antibody and complement via immunosurgery were more likely to survive passage when cultured on mitomycin C-treated fetal fibroblasts if derived from blastocysts treated with CSF2 than if from control blastocysts. There was little effect of CSF2 on characteristics of TE outgrowths from blastocysts. The exception was a decrease in outgrowth size for embryos treated with CSF2 from Days 5 to 7 and an increase in expression of CDX2 when treatment was from Days 6 to 8. Expression of the receptor subunit gene CSF2RA increased from the zygote stage to the 9-16 cell stage before decreasing to the blastocyst stage. In contrast, CSF2RB was undetectable at all stages. In conclusion, CSF2 improves competence of the ICM to survive in a pluripotent state and alters TE outgrowths. Actions of CSF2 occur through a signaling pathway that is likely to be independent of CSF2RB.

  4. IVF and embryo transfer: historical origin and development.

    Science.gov (United States)

    Biggers, John D

    2012-08-01

    IVF and embryo transfer for the treatment of human infertility has now resulted in the birth of over 4 million babies. The technique did not arise as a quantum event but was built on the efforts of many earlier workers in the fields of reproductive endocrinology and development. One should remember the famous saying of Isaac Newton: 'If I have seen further than most, it is because I have stood on the shoulder's of giants'. Ethical and moral issues have always arisen when investigators study early mammalian development, particularly human development. This paper documents these earlier studies and also draws attention to the ethical and moral arguments that inevitably arose. PMID:22695311

  5. Risk assessment of transmission of bovine viral diarrhea virus (BVDV) in abattoir-derived in vitro produced embryos.

    Science.gov (United States)

    Perry, G H

    2007-07-01

    Bovine virus diarrhea virus (BVDV) is a pathogen of the bovine reproductive system causing reduced conception rates, abortions and persistently infected calves. Most if not all strains of BVDV are transmissible by natural mating and AI. For international trade, it is recommended that in vitro fertilized embryos be washed according to the IETS Manual. However, BVDV may not be entirely washed out, resulting in possible transmission risks to recipients. Donor cows, donor bulls and biological agents are all possible sources of contamination. The process for producing in vitro produced (IVP) embryos is complex and non-standard, and some procedures can contribute to spread of BVDV to uninfected embryos. The structure of the zone pellucida (ZP) of IVP embryos permits adherence of BVDV to the ZP. To estimate the risk of producing infected recipients and persistently infected calves from abattoir-derived IVP embryos, a quantitative risk assessment model using Microsoft Excel and Palisade @Risk was developed. Assumptions simplified some of the complexities of the IVP process. Uncertainties due to incomplete or variable data were addressed by incorporating probability distributions in the model. Model variables included: disease prevalence; the number of donor cows slaughtered for ovaries; the number of oocytes collected, selected and cultured; the BVDV status of ovaries, semen, biological compounds and its behavior in the IVP embryo process. The model used the Monte Carlo method to simulate the IVP process. When co-culture cells derived from donor cows of unknown health status were used for in vitro culture (IVC), the probability of a recipient cow at risk of infection to BVDV per oocyte selected for IVP processing averaged 0.0006. However, when co-culture free from BVDV was used, the probability was 1.2 x 10(-5). Thus, for safe international trade in bovine IVP embryos (i.e. negligible risks of transmission of BVDV), co-culture cells, if used during IVC for producing IVP

  6. Expression of the gap junction gene connexin43 (Cx43) in preimplantation bovine embryos derived in vitro or in vivo.

    Science.gov (United States)

    Wrenzycki, C; Herrmann, D; Carnwath, J W; Niemann, H

    1996-09-01

    In this study we have examined the presence of mRNA encoding connexin 43 (Cx43) in bovine embryos derived in vivo and in vitro. Cumulus-oocyte complexes, immature and matured oocytes liberated from cumulus cells, zygotes, 2-4-cell and 8-16-cell embryos, morulae, blastocysts and hatched blastocysts were produced in vitro from ovaries obtained from an abattoir using TCM 199 supplemented with hormones and 10% oestrous cow serum for maturation. Cumulus-oocyte complexes matured for 24 h were exposed to bull spermatozoa for 19 h and then cultured in TCM 199 supplemented with 10% oestrous cow serum to the desired developmental stage. Morulae and blastocysts derived in vivo were collected from superovulated donor cows. Total RNA was extracted from pools of 60-200 bovine oocytes or embryos using a modified phenol-chloroform extraction method and analysed by reverse transcriptase polymerase chain reaction. Before reverse transcription, aliquots of DNase-digested embryonic RNA were tested by polymerase chain reaction using bovine-specific primers to control for residual genomic DNA contamination. DNA-free, total RNA was reverse transcribed after preincubation with the Cx43 specific 3'primer. The resultant cDNA was amplified by polymerase chain reaction using Cx43 specific primers that define a 516 bp fragment of Cx43. The reverse transcriptase polymerase chain reaction product was verified by restriction enzyme analysis with Alu I and sequencing. Assays were repeated at least twice for each developmental stage and provided identical results between replicates. Cx43 transcripts were detected in bovine morulae and blastocysts grown in vivo. In contrast, whereas the early in vitro stages from cumulus-oocyte complexes to morulae expressed Cx43, blastocysts and hatched blastocysts did not have detectable concentrations of mRNA from this gene. Restriction enzyme cutting revealed three fragments of the predicted size (139, 177, 200 bp). The amplified product showed 100% identity

  7. Effects of Seasons on Embryo Transfer of Cattle in Different Climatic Zones

    Institute of Scientific and Technical Information of China (English)

    Wang; Feng; Wang; Shenyuan; Zhang; Dong; Han; Lidong; Li; Lu; Huang; Chunhua; Zhong; Gang; Han; Jinlong; Wang; Bingping; Liu; Yiyi; Liu; Caiyun; Pan; Jing; Zhao; Zhichao; Zhou; Huanmin; Zhang; Li

    2014-01-01

    The paper was to evaluate the efficient seasons for embryo transfer of cattle in different climatic zones in China. Three climatic zones(mid-temperate zone,warm temperate zone,subtropical zone) were designed,and embryo transfers had been carried out in spring,summer,autumn and winter from 2009 to 2011,respectively. The total number of transplant recipient cattle was 22 208. The results showed that the best seasons for embryo transfers varied with different climatic zones. In mid-temperate zone,summer and autumn were better while summer was the best,and the rate of pregnant was 50. 67%(the number of pregnant cattle was 8 005). In warm temperate zone,spring and autumn were better while autumn was the best,and the rate of pregnant was 54. 99 %(the number of pregnant cattle was 551). In subtropical zone,spring and winter were better while winter was the best,and the rate of pregnant was 56. 94 %(the number of pregnant cattle was 328). The seasonal average temperatures and relative humidity of the best seasons in different climatic zones were concentrated on the same region. In mid-temperate zone,the mean temperature ranged between 22. 4 ℃ and 24. 2 ℃,and the mean relative humidity ranged from 44% to 55. 3%. In warm zone,the mean temperature ranges were between 14. 2 ℃ and 16. 2 ℃,and the mean relative humidity ranges were from 59. 3% to 71. 6%. In sub-tropical zone,the mean temperature ranges were between 3. 26 ℃ and 7. 73 ℃,and the mean relative humidity ranges were from 72% to 80. 6%. Therefore,the optimized conditions of temperature and humidity of season in different zones could be simulated. It is possible that we apply the program to bovine production in the similar agroecological zones,which is of great significance for improving the embryo transfer efficiency of livestock in production.

  8. Invited review: Genetic contributions underlying the development of preimplantation bovine embryos.

    Science.gov (United States)

    Kropp, J; Peñagaricano, F; Salih, S M; Khatib, H

    2014-03-01

    In recent years, it has become evident that genetic selection to improve milk production has resulted in a decline in dairy cattle fertility. Growing evidence suggests that the greatest loss occurs early in pregnancy around the time of embryo implantation. As a means to make genetic improvements and to assist in reproductive performance, use of artificial reproductive technologies such as artificial insemination and in vitro production of embryos have been widely used. Both of these technologies rely on the competence and quality of gametes for successful development of embryos. Often, selection of animals is based on the genetic merit of the animal, although specific fertility markers are relatively underdeveloped compared with markers for production traits. Similarly, current in vitro fertilization systems could benefit from a uniform method for selection of the best quality embryos to transfer into recipients for successful implantation and delivery of healthy offspring. As genetics underlie biological processes such as fertility, the need exists to further identify and characterize genes that affect fertility and development within both the parental gametes and the embryo. Furthermore, the magnitude of the contribution of each parental genome to the success of embryo development and pregnancy is not clear. As such, the objective of this review is to provide an overview of studies relating to genetic markers at the DNA level, parental and embryonic gene expression, and the effects of epigenetics on embryonic development. Future studies should exploit advances in molecular technologies to identify and classify genes underlying fertility and development to establish biomarkers and predictors for improved genetic selection. PMID:24377798

  9. Superovulation, collection and transfer of embryos and demi-embryos from Boran(Bos indicus ) cows and heifers.

    Science.gov (United States)

    Jordt, T; Lorenzini, E

    1988-08-01

    Twenty-three Boran(Bos indicus ) cows and heifers were superovulated with pregnant mare serum gonadotropin (PMSG); a total of four embryos and 4.1 +/- 0.3 (mean +/- SEM) ova per ova-producing donor resulted. Follicle stimulating hormone (FSH-P) was then used to superovulate 49 Boran cows for a total of 106 superovulations, of which 63 (59.4%) produced an average of 3.7 +/- 0.4 (mean +/- SEM) embryos. The embryo production was not influenced by either the season or the number of times(one to five) the cows were superovulated. A higher pregnancy rate was obtained when the selection of Boran recipients was based on their plasma-progesterone values (overall 52.5%, single embryos 63.3%, twin demi-embryos 45.8%) than when they were selected by palpation per rectum only (overall 43.8%, single embryos 50%, twin demi-embryos 36.4%). The twinning rate of twin demiembryos was 62.5%, whereas only single calves were born after transfer of two embryos per recipient. No pregnancies were produced following transfer of twin demi-embryos without zonae pellucidae. Transferring single demi-embryos gave a low pregnancy rate (13.3%). Twelve donor Boran cows (21 superovulations) bred with their fathers resulted in a high rate of early embryonic death; additionally, only 20.9% (overall) of the recipients became pregnant. Estrus synchronization of Boran cows with a progesterone releasing intravaginal device (PRID) for a short period (7 d) combined with one injection of prostaglandin (Day 6) produced a larger number of good quality recipients (70.5%) than using double prostaglandin injections (60%). PMID:16726476

  10. Co-expression network analysis to identify pluripotency biomarkers in bovine and porcine embryos

    DEFF Research Database (Denmark)

    Mazzoni, Gianluca; Freude, Karla Kristine; Hall, Vanessa Jane;

    stable culture of pluripotent cells in pig and cattle. Methods After quality control reads were pre-processed and mapped with STAR aligner. Post-mapping quality was checked with Qualimap. Finally the expression levels were estimated using HTSeq. Gene co-expression will be analyzed using a weighted...... network based method to identify highly co-expressed genes (module) and hub genes. Modules with a potential role in pluripotency will be identified with enrichment procedure and regulator genes identified with LemonTree algorithm. Differential wiring of modules among species will be evaluated. Expected...... results We expect to find out candidate pluripotency factors in porcine and bovine embryo. Acknowledgements We thank for the financial support from the EU project PluriSys, HEALTH-2007-B-223485....

  11. Multiple ovulation and embryo transfer (MOET in camels: An overview

    Directory of Open Access Journals (Sweden)

    Binoy S. Vettical

    2016-04-01

    Full Text Available Unlike in other domestic animal species like cattle, reproductive biotechnologies like Artificial Insemination (AI and Embryo Transfer (ET are not well developed and thus are not being used as routine breeding procedures in camels. One of the important objectives of this manuscript is to focus on analyzing the present status of Multiple Ovulation and Embryo Transfer (MOET in camels and its future perspectives. Camels are induced ovulators, thus require hormonal treatment to induce ovulation and control the follicular cycles, which is the main reason why protocols used in other domestic animal species cannot be directly used in this species. The review suggests that the best method for super stimulation of ovaries in camels is use of a combination of Equine Chorionic Gonadotropin (eCG and Follicle Stimulating Hormone (FSH at any stage after elimination of dominant follicle if any or at the early stage of the follicular wave and ovulation of the developed multiple follicles can be achieved by mating donors. The review highlights that a better pregnancy rate is achieved with recipients who ovulate 24 h after the donor.

  12. EFFECT OF FETAL SERUM AND FOLLICULAR LIQUOR SUPPLEMENTATION ON THE IN VITRO PRODUCTION OF BOVINE EMBRYOS

    Directory of Open Access Journals (Sweden)

    Clara Larocca

    2012-01-01

    Full Text Available To optimize In Vitro Fertilization (IVF results in bovines by lowering costs, comparing the efficiency of Fetal Calf Serum (FCS (high cost with respect to bovine Follicular Fluid (bFF (low cost and by quantifying embryo production. Cumulus-Oocyte-Complexes (COC obtained from a slaughter house, transported at 37°C in saline solution, were classified according to their quality in A, B, C and D. A and B oocytes were washed with modified Phosphate-Buffered Saline (m-PBS and three groups were randomly formed (GC; G1; G2 cultured in drops of the respective media (10 COC/drop of 100 μL, covered with mineral oil and placed in an incubator (38,5°C, 5% CO2 y 95% humidity. Maturation was done in Tissue Culture Medium (TCM-199 and antibiotics, for 22h and developed in CR1aa medium and antibiotics. The Control Group (CG was supplemented during both maturation and development stages with 5% FCS and 10% bFF; G1 with 5% FCS during maturation and development and G2 with 10% bFF during maturation and development. We obtained bFF from follicles larger than 15 mm, it was centrifuged (800G inactivating it. At fertilization Bracket and Oliphant (BO medium was used. Zygotes were evaluated every 48 h for 8 days since insemination, counting the Division Rate (DR and the Embryo Development (ED of compacted morulae. Χ2 Test was applied. For RD, G2 differs from G1 and CG (p0.05. During development no significant differences were found between groups (p>0.05. Results show that when using FCS, bFF or a combination of both, development results are similar. It is assumed that it is possible to substitute FCS with bFF."

  13. Live embryo imaging to follow cell cycle and chromosomes stability after nuclear transfer.

    Science.gov (United States)

    Balbach, Sebastian T; Boiani, Michele

    2015-01-01

    Nuclear transfer (NT) into mouse oocytes yields a transcriptionally and functionally heterogeneous population of cloned embryos. Most studies of NT embryos consider only embryos at predefined key stages (e.g., morula or blastocyst), that is, after the bulk of reprogramming has taken place. These retrospective approaches are of limited use to elucidate mechanisms of reprogramming and to predict developmental success. Observing cloned embryo development using live embryo cinematography has the potential to reveal otherwise undetectable embryo features. However, light exposure necessary for live cell cinematography is highly toxic to cloned embryos. Here we describe a protocol for combined bright-field and fluorescence live-cell imaging of histone H2b-GFP expressing mouse embryos, to record cell divisions up to the blastocyst stage. This protocol, which can be adapted to observe other reporters such as Oct4-GFP or Nanog-GFP, allowed us to quantitatively analyze cleavage kinetics of cloned embryos. PMID:25287344

  14. Effects of different nuclear recipients on developmental potential of mouse somatic nuclear transfer embryos

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    In order to investigate the effects of different kinds of nuclear recipients from Kunming (KM) mouse on developmental potential of somatic nuclear transfer em- bryos, the enucleated MⅡ oocytes, enucleated zygotes and 2-cell blastomere were used to produce cloned mouse embryos. Using fibroblast deriving from C57/BL6 ear tissue as nuclear donor, we produced cloned embryos by transferring the fibroblast nuclei into enucleated KM mouse oocytes (single nuclear transfer, SNT), transferring pronuclei from the SNT embryos into enucleated KM zygotes (nuclear into zygote, NZ), and 2-cell blastomere nuclei from SNT embryos into enucleated KM mouse oocytes (nuclear into oocytes, NO); tetraploid embryos (tetraploid embryos, TE) were obtained by fusing two blastomeres, one is from the SNT cloned embryos, and the other from normal 2-cell KM mouse embryos. In group SNT, the cloned embryos could not develop beyond 8-cell stage and the rate of 8-cell stage is only 0.3%; in group NO, the reconstructed embryos could develop to morula stage, the rate of 8-cell stage was significantly greater than that of SNT group (P < 0.05); in group NZ, the development rate was further improved, and the reconstructed embryos could develop into blastocyst stage, the rate of blastocyst was 1.9%; in group TE, as high as 62.3% of the reconstructed embryos could develop into blastocyst. Results suggested that different nuclear recipients could significantly affect the developmental potential of cloned mouse embryos; KM MⅡ oocyte cytoplasm was not so effective as zygotes to reprogram the mouse somatic cell nuclei; serial nuclear transfer could improve the developmental potential of cloned mouse embryos.

  15. Genetic analysis of superovulation and embryo transfer traits in Holstein cattle

    Science.gov (United States)

    The objectives of this study were to estimate variance components and investigate genomic regions of interest associated with superovulation and embryo transfer in dairy cattle. Superovulation and embryo transfer are methods commonly used by dairy producers to increase the rate of genetic gain achie...

  16. Current status and potential of embryo transfer and reproductive technology in dairy cattle.

    Science.gov (United States)

    Hasler, J F

    1992-10-01

    Significant use of embryo transfer in dairy cattle commenced with the introduction of nonsurgical embryo recovery in the mid-1970s and developed with the use of nonsurgical transfers in the late 1970s. Numbers of registered Holstein calves from embryo transfer doubled yearly through 1980, after which the rate of increase slowed; the total reached nearly 19,000 calves in 1990. However, the efficacy of superovulation procedures and commercial success rates of transferred fresh embryos have not improved the past 10 to 15 yr. Fertilization rates in superovulated donors remain low. Although embryo-splitting techniques were perfected in the early 1980s, they are not used widely. A practical, commercial embryo-sexing procedure remains unavailable. Recent significant improvement is apparent in the technology of ultrasound-guided oocyte collection and in vitro oocyte maturation, fertilization, and embryo culture. In the future, this technology may be used in conjunction with sperm separated by sex with a flow cytometer. Modest numbers of embryo clones have been produced in several commercial programs via nuclear transfer techniques. However, the efficiency of gene transfer experiments involving ova of cattle and other domestic species has been low. Recently, DNA probe technology has begun to provide genotype information for cattle and will ultimately be applied to embryos.

  17. Paternal breed effects on expression of IGF-II, BAK1 and BCL2-L1 in bovine preimplantation embryos

    DEFF Research Database (Denmark)

    Valleh, Mehdi Vafaye; Tahmoorespur, Mojtaba; Joupari, Morteza Daliri;

    2015-01-01

    Summary The effects of the paternal breed on early embryo and later pre- and postnatal development are well documented. Several recent studies have suggested that such paternal effects may be mediated by the paternally induced epigenetic modifications during early embryogenesis. The objective...... of this study was to investigate the effects of the paternal breed on the early embryonic development and relative expression of the maternally imprinted gene, IGF-II, and the apoptosis-related genes BAK1 and BCL2-L1 in in vitro produced (IVP) bovine embryos derived from two unrelated paternal breeds (Holstein...

  18. In situ DNA transfer to chicken embryos by biolistics

    Directory of Open Access Journals (Sweden)

    Luciana A. Ribeiro

    1999-12-01

    Full Text Available Fertilized chicken eggs were bombarded with a biolistic device. Transient expression of the lacZ gene under the control of a human cytomegalovirus (CMV promoter was assessed after in situ gene transfer using this approach. The influence of different pressures, vacuum levels and particles was tested. Survival rate improved as particle velocity decreased, but resulted in lower levels of expression. The best survival and expression were obtained with gold particles, a helium gas pressure of 600 psi and a vacuum of 600 mmHg. Under these conditions, all bombarded embryos showed b-galactosidase activity, indicating that this was an effective method for transformation of chicken embryos.Ovos fertilizados de galinha foram bombardeados através da técnica de biobalística. A expressão transiente do gene lacZ, sob o controle do promotor humano citomegalovírus, foi verificada após a transferência in situ. Diferentes níveis de pressão de gás hélio, vácuo e tipos de partículas foram testados. A taxa de sobrevivência aumentou à medida que a velocidade das partículas diminuíram, entretanto, o nível de expressão foi menor. Os melhores resultados, combinando taxa de sobrevivência e expressão, foram obtidos com partículas de ouro, 600 libras por polegada ao quadrado de hélio e 600 mmHg de vácuo. Nestas condições, todos os embriões bombardeados apresentaram atividade da b-galactosidase, indicando que esta técnica é eficiente para a transformação de embriões de galinhas.

  19. Transfer of human frozen-thawed embryos with further cleavage during culture increases pregnancy rates

    Directory of Open Access Journals (Sweden)

    Bharat V Joshi

    2010-01-01

    Full Text Available Aim: To compare the pregnancy rate following transfer of frozen-thawed embryos with or without overnight culture after thawing. Settings and Design: This is a retrospective analysis of frozen-thawed embryo transfer (FET cycles performed between January 2006 and December 2008. Materials and Methods: Out of 518 thaw cycles, 504 resulted in embryo transfers (ETs. Of the total FET cycles, 415 were performed after an overnight culture of embryos (group A; and in 89 cycles, ET was performed within 2 hours of embryo thawing (group B. Statistical Analysis: The data were statistically analyzed using chi-square test. Results: We observed that with FET, women ≤30 years of age had a significantly higher (P=0.003 pregnancy rate (PR=28.9% as compared to women >30 years of age (17.5%. A significantly higher (P<0.001FNx08 pregnancy rate was also observed in women receiving 3 frozen-thawed embryos (29% as compared to those who received less than 3 embryos (10.7%. The difference in PR between group A (PR=24.3% and group B (PR=20.3% was not statistically significant. However, within group A, ET with cleaved embryos showed significantly ( P≤0.01 higher pregnancy rate compared to the uncleaved embryos, depending on the number of cleaved embryos transferred. Conclusion: No significant difference was noticed between FETs made with transfer of embryos with overnight culture and those without culture. However, within the cultured group, transfer of embryos cleaved during overnight culture gave significantly higher PR than transfers without any cleavage.

  20. Methods for endometrial preparation in frozen-thawed embryo transfer cycles.

    Science.gov (United States)

    Kalem, Ziya; Kalem, Müberra Namlı; Gürgan, Timur

    2016-01-01

    Frozen-thawed (FT) embryo transfer is a procedure used for the storage and transfer of excess embryos obtained during in vitro fertilization- intracytoplasmic sperm injection cycles. In recent years, improvements in laboratory conditions and limitations on the number of embryos to be transferred have led to a progressive increase in FT embryo transfer cycles. However, the best solution for endometrial preparation in these cycles is still a matter of debate. In this study, we aimed to review the current methods of endometrial preparation in FT embryo transfer cycles. In light of the current literature, it is hard to determine which method is the best for endometrial preparation. It is therefore necessary to conduct randomized controlled studies in a prospective design, which will also evaluate the above-mentioned factors. PMID:27651727

  1. Analysis of Factors Influencing Pregnancy Rate in Frozen-thawed Embryo Transfer

    Institute of Scientific and Technical Information of China (English)

    Lu LI; Xiao-xi SUN; Jun-ling CHEN; Xiao-hong GAO; Yong-wei WANG; Jie-wei TAO; Li-nan CHENG

    2004-01-01

    Objective To analyse factors influencing the outcome of frozen-thawed embryo transfer (FET)Method A retrospective analysis was performed in our center on 129 thawing cycles from March 2001 to April 2003. The related parameters were compared between conceived and non-conceived cycles.Results There were totally 129 clinical pregnancies in these transfers (pregnancy rate: 27.1%). Frozen-thawed embryos were transferred to natural cycles and CC cycling and hormone replacement treatment had equal success. Groups of IVF and ICSI did not differ significantly in pregnancy rates (P>0. 05). The pregnancy rates for one, two, three and four pre-embryos transfer were 0, 20.0%,44.1% and 75.0%,respectively (P<0. 05). There were statistical differences between pregnancy group or non- pregnancy group in the endometrial thickness, CES, CES/No. Of embryo. A higher pregnancy rate was observed in embryo transfers which had at least one 4-cell grade I embryo (d 2)(P<0.01). Conclusions The most important factors influencing the implantation rate and pregnancy rate of frozen-thawed embryo transfer are age, endometrium thickness, and the number, morphology and growth rate of transferred frozen embryos of women participants.

  2. Establishment of pregnancies in large white sow recipients after transfer of cloned embryos of different pig breeds

    DEFF Research Database (Denmark)

    Schmidt, Mette; Kragh, Peter M.; Li, J.;

    2010-01-01

    Establishment of pregnancies in large white sow recipients after transfer of cloned embryos of different pig breeds......Establishment of pregnancies in large white sow recipients after transfer of cloned embryos of different pig breeds...

  3. Embryo transfer: a comparative biosecurity advantage in international movements of germplasm.

    Science.gov (United States)

    Thibier, M

    2011-04-01

    This paper uses cattle as a model to provide an overview of the hazards involved in the transfer of in vivo-derived and in vitro-produced embryos. While scientific studies in recent decades have led to the identification of pathogens that may be associated with both in vivo- and in vitro-derived embryos, those studies have also been the basis of appropriate disease control measures to reduce the risks to a negligible level. These disease control measures have been identified and assessed by the International Embryo Transfer Society's (lETS) Health and Safety Advisory Committee, the expert body that advises the World Organisation for Animal Health (OIE) on matters related to the safety of embryo transfer. Through the OIE's processes for developing and adopting international standards, the disease control measures identified by the IETS have been incorporated into the Terrestrial Animal Health Code. The basic principles rely on the crucial ethical roles of the embryo collection team and embryo transfer team, under the leadership of approved veterinarians. Decades of experience, with nearly 10 million embryos transferred, have demonstrated the very significant biosecurity advantage that embryo transfer technology has when moving germplasm internationally, provided that the international standards developed by the IETS and adopted by the OIE are strictly followed. PMID:21809763

  4. Bisphenol A affects early bovine embryo development and metabolism that is negated by an oestrogen receptor inhibitor

    Science.gov (United States)

    Choi, Bom-Ie; Harvey, Alexandra J.; Green, Mark P.

    2016-01-01

    Increasing evidence supports an association between exposure to endocrine disruptors, such as the xenoestrogen bisphenol A (BPA), a commonly used plasticiser, and the developmental programming of offspring health. To date however animal studies to investigate a direct causal have mainly focussed on supra-environmental BPA concentrations, without investigating the effect on the early embryo. In this study we investigated the effect of acute BPA exposure (days 3.5 to 7.5 post-fertilisation) at environmentally relevant concentrations (1 and 10 ng/mL) on in vitro bovine embryo development, quality and metabolism. We then examined whether culturing embryos in the presence of the oestrogen receptor inhibitor fulvestrant could negate effects of BPA and 17β-oestradiol (E2). Exposure to BPA or E2 (10 ng/mL) decreased blastocyst rate and the percentage of transferrable quality embryos, without affecting cell number, lineage allocation or metabolic gene expression compared to untreated embryos. Notably, blastocysts exposed to BPA and E2 (10 ng/mL) displayed an increase in glucose consumption. The presence of fulvestrant however negated the adverse developmental and metabolic effects, suggesting BPA elicits its effects via oestrogen-mediated pathways. This study demonstrates that even acute exposure to an environmentally relevant BPA concentration can affect early embryo development and metabolism. These may have long-term health consequences on an individual. PMID:27384909

  5. Bisphenol A affects early bovine embryo development and metabolism that is negated by an oestrogen receptor inhibitor.

    Science.gov (United States)

    Choi, Bom-Ie; Harvey, Alexandra J; Green, Mark P

    2016-01-01

    Increasing evidence supports an association between exposure to endocrine disruptors, such as the xenoestrogen bisphenol A (BPA), a commonly used plasticiser, and the developmental programming of offspring health. To date however animal studies to investigate a direct causal have mainly focussed on supra-environmental BPA concentrations, without investigating the effect on the early embryo. In this study we investigated the effect of acute BPA exposure (days 3.5 to 7.5 post-fertilisation) at environmentally relevant concentrations (1 and 10 ng/mL) on in vitro bovine embryo development, quality and metabolism. We then examined whether culturing embryos in the presence of the oestrogen receptor inhibitor fulvestrant could negate effects of BPA and 17β-oestradiol (E2). Exposure to BPA or E2 (10 ng/mL) decreased blastocyst rate and the percentage of transferrable quality embryos, without affecting cell number, lineage allocation or metabolic gene expression compared to untreated embryos. Notably, blastocysts exposed to BPA and E2 (10 ng/mL) displayed an increase in glucose consumption. The presence of fulvestrant however negated the adverse developmental and metabolic effects, suggesting BPA elicits its effects via oestrogen-mediated pathways. This study demonstrates that even acute exposure to an environmentally relevant BPA concentration can affect early embryo development and metabolism. These may have long-term health consequences on an individual. PMID:27384909

  6. Experimental cloning of embryos through human-rabbit inter-species nuclear transfer

    Institute of Scientific and Technical Information of China (English)

    JI Jingjuan; GUO Tonghang; TONG Xianhong; LUO Lihua; ZHOU Guixiang; FU Yingyun; LIU Yusheng

    2007-01-01

    Therapeutic cloning,which is based on human somatic cell nuclear transfer,is one of our major research objectives.Though inter-species nuclear transfer has been introduced to construct human somatic cell cloned embryos,the effects of type,passage,and preparation method of donor cells on embryo development remain unclear.In our experiment,cloned embryos were reconstructed with different passage and preparation methods of ossocartilaginous cell,skin fibroblast,and cumulus cells.The cumulus cell embryos showed significantly higher development rates than the other two (P<0.05).The development rate of embryos reconstructed with skin fibroblasts of different passage number and somatic cells of different chilling durations showed no significant difference.Also,fluorescence in situ hybridization (FISH)was conducted to detect nuclear derivation of the embryos.The result showed that the nuclei of the inter-species cloned embryo cells came from human.We conclude that (1)cloned embryos can be constructed through human-rabbit interspecies nuclear transfer;(2)different kinds of somatic cells result in different efficiency of nuclear transfer,while in vitro passage of the donor does not influence embryo development;(3)refrigeration is a convenient and efficient donor cell preparation method.Finally,it is feasible to detect DNA gcnotype through FISH.

  7. Clinical effectiveness of elective single versus double embryo transfer: meta-analysis of individual patient data from randomised trials

    NARCIS (Netherlands)

    D.J. McLernon; K. Harrild; C. Bergh; M.J. Davies; D. de Neubourg; J.C.M. Dumoulin; J. Gerris; J.A.M. Kremer; H. Martikainen; B.W. Mol; R.J. Norman; A. Thurin-Kjellberg; A. Tiitinen; A.P.A. van Montfoort; A.M. van Peperstraten; E. van Royen; S. Bhattacharya

    2010-01-01

    Objective To compare the effectiveness of elective single embryo transfer versus double embryo transfer on the outcomes of live birth, multiple live birth, miscarriage, preterm birth, term singleton birth, and low birth weight after fresh embryo transfer, and on the outcomes of cumulative live birth

  8. Evolution studies from embryo transfer and in vitro fertilization in Nelore cattle at Brazil

    Directory of Open Access Journals (Sweden)

    Renato Travassos Beltrame

    2010-01-01

    Full Text Available Data from Brazilian Association of Zebu Breeders were used to show the evolution of conventional embryo transfer (ET and in vitro fertilization techniques between 1997 and 2007 for the Nelore cattle in Brazil. Data from 109.775 conventional embryo transfer and 73.121 ovum pickup (OPU were used. The embryo transfer supremacy was observed until 2004 when a considerable increase of OPU number over ET occurred. More ET and OPU were carried out in Southeast and Mid-West than in others Brazilian regions. The average number of viable embryo along the analysis period is close to that reported by literature (between 5 and 7 embryos per flushing or OPU session.

  9. Producción in vitro de embriones bovinos: suplementación de los medios de cultivo con suero In vitro production of bovine embryos: serum supplementation to the culture media

    Directory of Open Access Journals (Sweden)

    N Mucci

    2006-01-01

    Full Text Available Las técnicas para producir embriones bovinos, en estadios de preimplantación mediante la maduración de ovocitos y su posterior fertilización in vitro, ofrece la posibilidad de obtener embriones a bajo costo para ser utilizados con fines de estudio (desarrollo embrionario temprano, transgénesis, clonación o con propósitos comerciales. Las condiciones de cultivo in vitro pueden influenciar significativamente el desarrollo embrionario, determinando cambios responsables de su menor calidad, comparados con los embriones producidos in vivo . En particular, la adición de suero a los medios de cultivo altera tanto la morfología embrionaria como su calidad, y su eliminación posibilitaría producir embriones de buena calidad para ser criopreservados y transferidos. Los objetivos de esta revisión son describir los aspectos generales del cultivo in vitro de embriones y resumir algunas hipótesis referidas a los mecanismos por los cuales el suero podría alterar el desarrollo y la calidad embrionaria.Techniques for producing bovine preimplantation embryos by in vitro maturation and fertilization, offers the potential for a large number of embryos at low cost to be used for basic scientific research (embryo development, transgenesis, cloning or for commercial purposes. Embryo culture conditions can influence in vitro embryo development significantly, provoking deviations responsible for low quality compared with in vivo counterparts. In particular, serum supplementation alters both embryo morphology and quality and eliminating serum from the media could be beneficial to produce good quality embryos for cryopreservation and embryo transfer. The objectives of this review are to describe the general aspects of in vitro embryo culture and summerize some hypotheses about the way serum could alter embryo development and quality.

  10. Post-hatching development of in vitro bovine embryos from day 7 to 14 in vivo versus in vitro.

    Science.gov (United States)

    Machado, G M; Ferreira, A R; Pivato, I; Fidelis, A; Spricigo, J F; Paulini, F; Lucci, C M; Franco, M M; Dode, M A

    2013-11-01

    This study evaluates the post-hatching development of in vitro-produced (IVP) embryos until Day 14. On Day 7, IVP embryos were either transferred to recipient uteruses or placed in a post-hatching development (PHD) system. As a control group, in vivo-produced (IVV), Day-7 embryos were also transferred to recipient uteruses. All groups were collected on Day 14 and were morphologically evaluated. Day-7 and Day-14 IVV and IVP embryos were used for quantification of eight genes (PLAC8, CD9, SLC2A1, SLC2A3, KRT8, SOD2, HSP1A1, and IFNT2) by reverse transcriptase qPCR. Day-14 embryos from the PHD system were smaller (2.92 ± 0.45 mm) and had a lower embryonic disk diameter (0.14 ± 0.00 mm) than those produced by IVV (24.18 ± 3.71; 0.29 ± 0.03 mm, respectively) or IVP (19.06 ± 2.43; 0.28 ± 0.01 mm) culture and transferred to the uterus (P > 0.05). Day-7 IVP embryos had a higher expression of the HSP1A1, SCL2A1, and SCL2A3 genes than IVV embryos. When these embryos were cultured in the uterus, no differences in gene expression were observed on Day 14. Conversely, Day-14 IVP embryos cultured in the PHD system showed a higher expression of PLAC8, SOD2, and SLC2A3 genes. It is concluded that Day-7 IVP embryos are different from IVV embryos in regards to gene expression, although exposure to the uterine environment during the elongation period allowed the IVP embryos to overcome this difference. In contrast, IVP embryos cultured in the PHD system were morphologically and molecularly different, being of poorer quality than those cultured in the uterus.

  11. Use of versapoint to refashion the cervical canal to overcome unusually difficult embryo transfers and improve in-vitro fertilization-embryo transfer outcome: A case series

    Directory of Open Access Journals (Sweden)

    Nalini Mahajan

    2011-01-01

    Full Text Available Background : Smooth atraumatic embryo transfer is paramount for the success of in-vitro fertilization (IVF. In difficult cases, cervical canal manipulation may be required. Aim : To see if surgical correction of the cervical canal or cervical canal refashioning could improve ease of embryo transfer. Setting : Private infertility and IVF hospital. Design : Prospective study. Materials and Methods : Patients: 11 women with failed 1-3 IVF cycles with history of extremely difficult embryo transfers (ETs despite undergoing cervical dilatation in the cycle prior to IVF. Interventions : Operative hysteroscopy using Versapoint for refashioning of the cervical canal. Main Outcome Measures : Ease of ET in the subsequent IVF cycle. Secondary outcome measure was to assess reproductive outcome. Results : Easy and atraumatic ET in the IVF cycle after procedure in 100% patients. PR was 46.5%. Conclusions : Use of Versapoint for refashioning the cervical canal can improve the quality of ET and PR.

  12. Effect of temporary meiosis block during prematuration of bovine cumulus-oocyte complexes on pregnancy rates in a commercial setting for in vitro embryo production.

    Science.gov (United States)

    Guemra, Samuel; da Silva Santo, Eriko; Zanin, Renato; Monzani, Paulo Sergio; Sovernigo, Tobias Canan; Ohashi, Otávio Mitio; Verde Leal, Cláudia Lima; Adona, Paulo Roberto

    2014-04-15

    Ovum pick up (OPU) associated with in vitro production (IVP) of embryos has been shown as an important tool in cattle breeding to increase the number of descendants from animals of high genetic value. In herds maintained distant from the laboratory, collecting cumulus-oocyte complexes (COCs) and transporting them to the laboratory may take several hours and decrease COCs viability, representing a challenge for commercial settings. In this study, a prematuration culture to induce temporary meiosis block was evaluated in a commercial scale IVP setting as a strategy to transport bovine OPU-derived COCs from Nelore and Brangus donors. Effects on embryo yield and pregnancy rates were assessed. Viable COCs from each donor were destined to one of the experimental groups (control, blocks 1 and 2). Control group COCs were placed in cryotubes with 1 mL TCM199-HEPES. In block groups (1 and 2), COCs were placed in cryotubes with 300 μL TCM 199 + 12 μM butyrolactone I (block medium). All groups were gassed and kept in a thermos bottle for 4 hours at 36 °C. Next, COCs in the control group were transferred to IVM medium and block 1 group to block medium, and cultured for 22 hours and 15 hours, respectively, at 38.5 °C and 5% CO2 in air. Block 2 COCs were kept in the cryotubes and in the thermos bottle for another 15 hours at 36 °C to simulate long-term transport conditions. After meiosis block in prematuration culture, blocks 1 and 2 COCs were matured in vitro for 22 hours as for the control group. After IVM, COCs in all groups were submitted to IVF and IVC, and blastocyst rates were evaluated on day 7. Embryos were transferred and pregnancy rates evaluated at 60 days of gestation. The mean total number of COCs retrieved by OPU did not differ between Nelore and Brangus donors (16.8 and 17.2, respectively, P > 0.05), but Nelore donors produced more viable COCs than Brangus (10.1 and 7.6, respectively, P Brangus cattle, respectively (P > 0.05). Pregnancy rates did

  13. Non-invasive assessment of in-vitro embryo quality to improve transfer success

    DEFF Research Database (Denmark)

    Højbøge, Tina Rødgaard; Heegaard, Peter M. H.; Callesen, Henrik

    2015-01-01

    embryos before the transfer to a recipient still remains challenging. Presently, the predominant non-invasive technique for selecting viable embryos is based on morphology, where parameters such as rates of cleavage and blastocyst formation as well as developmental kinetics are evaluated mostly...... subjectively. The simple morphological approach is, however, inadequate for the prediction of embryo quality, and several studies have focused on developing new non-invasive methods using molecular approaches based particularly on proteomics, metabolomics and most recently small non-coding RNA, including micro......RNA. This review outlines the potential of several non-invasive in-vitro methods based on analysis of spent embryo culture medium....

  14. Reducing twin pregnancy rates after IVF--elective single embryo transfer (eSET).

    LENUS (Irish Health Repository)

    Milne, P

    2010-01-01

    Multiple pregnancy is a major complication of IVF and is associated with increased maternal, fetal and neonatal morbidity. Elective single embryo transfer (eSET) during IVF, rather than the more standard transfer of two embryos (double embryo transfer or DET), has been shown to significantly reduce the multiple pregnancy rate associated with IVF, while maintaining acceptable pregnancy rates. Couples undergoing IVF in 2008 who met good prognostic criteria had eSET performed. Pregnancy and twinning rates were compared with those for similar couples in 2007 who had DET. Couples unsuccessful with a fresh cycle of treatment had subsequent frozen embryo transfer cycles with DET. The cumulative pregnancy rate was similar for each group. However there were no multiple pregnancies in the eSET group, compared to 4 twins of 5 pregnancies in the DET group. 96% of eligible couples agreed to eSET. ESET is successful in and acceptable to good prognosis Irish couples undergoing IVF.

  15. Non-synchronized endometrium and its correction in non-ovulatory cryopreserved embryo transfer cycles.

    Science.gov (United States)

    Gomaa, Hala; Casper, Robert F; Esfandiari, Navid; Bentov, Yaakov

    2015-04-01

    The aim of this case series study was to investigate the effect of adjusting the length of progesterone exposure on clinical pregnancy rates in cryopreserved embryo transfer cycles of patients with out-of-phase classic endometrial dating. Eighty infertile women with previous implantation failure and good-quality embryos underwent endometrial biopsy before cryopreserved embryo transfer and were included in this study. The main outcome measures were clinical pregnancy rate and histologic endometrial dating. After adjusting the length of progesterone exposure according to endometrial dating, a significantly higher implantation rate was observed in blastocyst transfers (P = 0.02) and the clinical pregnancy rate for all cycles was 36.4%, similar to that in patients with in-phase endometrium (22.5%). In conclusion, the use of classic histologic endometrial dating to estimate the timing of the window of implantation and to adjust progesterone exposure accordingly may increase the implantation rate in frozen embryo transfer cycles. PMID:25676169

  16. Nonsurgical embryo recovery and transfer in sheep and goats.

    Science.gov (United States)

    Fonseca, Jeferson F; Souza-Fabjan, Joanna Maria G; Oliveira, Maria Emília F; Leite, Ceci R; Nascimento-Penido, Paula Maria P; Brandão, Felipe Z; Lehloenya, Khoboso C

    2016-07-01

    The embryo transfer techniques used in small ruminants worldwide are based in surgical procedures. These actions are performed under general anesthesia which needs a combination of animal fasting and drugs for secure animal handling and surgery manipulations. Therefore, it involves risks to animal health and life. The major limiting sequels are adhesions formed by the abdominal surgery, in the ovaries, uterus, or between them. These occurrences can both compromise uterus accessing and oocyte capture and are responsible for decreasing success and limiting successive embryo collections. In contrast, nonsurgical embryo procedures can be performed in a relatively simplified way. Nonsurgical embryo recovery does not need animal prolonged starvation, drug retention is minimized, and donors can stay in a standing position. After the end of embryo recovery, donors are promptly restored to their routine housing and feeding. Furthermore, this technique does not need incisions and, therefore, can be used repetitively in superovulated or nonsuperovulated goats and sheep for embryo recovery-a similar procedure done in cattle. In Brazil, promising results are reported using nonsurgical embryo transfer in recipient goats, and studies are currently evaluating similar procedures in sheep. Therefore, this review aimed to present the current panorama of nonsurgical embryo transfer in sheep and goats.

  17. Non-invasive assessment of in-vitro embryo quality to improve transfer success

    OpenAIRE

    Højbøge, Tina Rødgaard; Peter M H Heegaard; Callesen, Henrik

    2015-01-01

    Although IVF has been performed routinely for many years to help couples with fertility problems and in relation to modern breeding of farm animals, pregnancy rates after transfer to a recipient have not improved during the last decade. Early prediction of the viability of in-vitro developed embryos before the transfer to a recipient still remains challenging. Presently, the predominant non-invasive technique for selecting viable embryos is based on morphology, where parameters such as rates ...

  18. Large baby syndrome in singletons born after frozen embryo transfer (FET)

    DEFF Research Database (Denmark)

    Pinborg, Anja; Henningsen, AA; Loft, A;

    2013-01-01

    Are singletons born after frozen embryo transfer (FET) at increased risk of being born large for gestational age (LGA) and if so, is this caused by intrinsic maternal factors or related to the freezing/thawing procedures?......Are singletons born after frozen embryo transfer (FET) at increased risk of being born large for gestational age (LGA) and if so, is this caused by intrinsic maternal factors or related to the freezing/thawing procedures?...

  19. Developmental kinetics of the first cell cycles of bovine in vitro PRODUCED EMBRYOS IN RELATION TO THEIR IN VITRO VIABILITY AND SEX

    DEFF Research Database (Denmark)

    Holm, P; Shukri, N.N; Vajta, Gabor;

    1998-01-01

    The development of bovine IVP-embryos was observed in a time-lapse culture system to determine cell cycle lengths of 1) embryos that developed into compact morulae (CM) or blastocysts (BL) within 174 h after insemination (viable), 2) embryos that arrested during earlier stages (nonviable) and 3...... were included (n=392), and the times of cleavage events noted. After culture, 100 CM or BL were randomly selected for sexing by PCR. BL developed equally well in the time-lapse and control culture systems (36 vs 38. The respective lengths of the first 4 cell cycles of viable embryos were 32.0 + 3.9, g...

  20. Monozygotic Triplets and Dizygotic Twins following Transfer of Three Poor-Quality Cleavage Stage Embryos

    Directory of Open Access Journals (Sweden)

    Reshef Tal

    2012-01-01

    Full Text Available Background. Assisted reproductive technology has been linked to the increased incidence of monozygotic twinning. It is of clinical importance due to the increased risk of complications in multiple pregnancies in general and in monozygotic twins in particular. Case. A 29-year-old female, nulligravida underwent her first IVF cycle. Three poor-quality cleavage stage embryos were transferred resulting in monochorionic triamniotic triplets and dichorionic diamniotic twins. Selective embryo reduction was performed at 12 weeks leaving dichorionic twins. The patient underwent emergency cesarean section due to preterm labor and nonreassuring fetal heart tracing at 30 weeks of gestation. Conclusion. Our case emphasizes that even embryos with significant morphological abnormalities should be considered viable and the possibility of simultaneous spontaneous embryo splitting must be factored into determining number of embryos to transfer.

  1. Numerical Chromosome Errors in Day 7 Somatic Nuclear Transfer Bovine Blastocysts

    DEFF Research Database (Denmark)

    Booth, Paul J.; VIUFF, Dorte; Tan, Shijian;

    2002-01-01

    amino acids, myoinositol, sodium citrate, and 5% cattle serum in microwells for 7 days, at which time nuclei from all blastocysts were extracted and chromosome aberrations were evaluated using dual-color fluorescent in situ hybridization with bovine chromosome 6- and 7-specific probes. Five embryo clone...

  2. Effect of exogenous glutathione in medium fertilization to improve blastocyst rates of bovine embryos

    Directory of Open Access Journals (Sweden)

    E Triwulaninngsih

    2002-06-01

    Full Text Available Glutathione (C10H17N3O6S is a tripeptide (γ-Glu-Cys-Gly widespread in living organism. Glutathione (GSH at the 5 mM concentration increased the motility and fertility of frozen-thawed sperm. Intracellulair glutathione improved the cleavage rate and embryo development to the blastocyst rate. Research on in vitro embryos production through the implementation of GSH during IVF (in vitro fertilization on embryo development has been conducted at the Laboratorium Reproductive of Physiology, Research Institute for Animal Production. Ovaries of beef cattle as source of oocytes were collected from the slaughterhouse in a thermo flask with 350C PBS as medium and transported to the laboratory. The oocytes were fertilized in vitro with selected motile sperm using Percoll gradient (90 and 45%. Ten COCs (cumulus oocytes complexes were transfered to 44 μl of fertilization medium (mTALP was performed with either 0; 0.25; 0.50; 0.75 and 1.00 mM of glutathione as treatment A, B, C, D and E respectively, and inseminated with 2 μl of capacitated sperm and added 2 μl of heparin and 2 μl of PHE (consisting of 20 μM penicillamine, 10 μM hypotaurine, 1 μM epinephrine. Incubation between sperm and oocytes in the 5% CO2 incubator and RH 90% in fertilization media (TALP for 20 hours. All zygotes were cultured in modification of CR1aa medium up to blastocyst and were fed serum 5 μl/50μl medium on day 6. Results of this experiment showed that the effect of concentration of glutathione (0, 0.25; 0.50; 0.75 and 1.00 mM on fertilization medium to the percentage of cleavage rates were 69.35 + 29.40; 79.07 + 13.17; 67.88 + 10.65; 98.10 + 3.30 and 82.62 + 24.19% not significant different (P>0.05 among treatments A, B, C, D dan E respectively.The precentages of morula and blastocyst for treatment D were 50.45 + 42.64% and 31.28 + 24.28%; while 36.55 + 24.13% and 17.74 + 17.86% for treatment E respectively.

  3. Effect of insulin-like growth factor-1 during in vitro oocyte maturation and in vitro culture of bovine embryos

    OpenAIRE

    Quetglas M.D.; Coelho L.A.; Garcia J.M.; Oliveira Filho E.B.; Esper C.R.

    2001-01-01

    The effects of insulin-like growth factor-I (IGF-I) on in vitro maturation (IVM) (experiment I) and on in vitro embryo development (experiment II) of bovine oocytes fertilized in vitro, were evaluated in terms of cleavage (CR), blastocyst (BR) and hatching (HR) rates. For IVM, immature cumulus-oocyte complexes were cultured in TCM-199 medium supplemented with Hepes, sodium bicarbonate, sodium pyruvate, additives, fetal calf serum (B-199 medium) and gonadotropins (14 U/ml PMSG and 7 U/ml hCG)....

  4. Kinetics of fertilization and development, and sex ratio of bovine embryos produced using the semen of different bulls.

    Science.gov (United States)

    Alomar, M; Tasiaux, H; Remacle, S; George, F; Paul, D; Donnay, I

    2008-08-01

    The between bulls variation in in vitro fertility and the shift of sex ratio towards male embryos are two problems affecting the in vitro production (IVP) of bovine embryos. Our objective was to evaluate the kinetics of fertilization, embryo development and the sex ratio of the resulting embryos using the frozen/thawed semen of four different bulls. In a first experiment, the kinetics of pronucleus (PN) formation was evaluated at 8, 12 and 18 h post-insemination (hpi). Based upon the pronuclei sizes and the distance between the two pronuclei, inseminated oocytes were classified in three PN stages. Differences between bulls were observed at each time point, but were more important at 12 hpi. At 8 and 12 hpi bull III showed a significantly faster PN evolution by comparison with the three other bulls (Pcinematography. The analysis of embryos reaching the blastocyst stage revealed significant differences in the mean time of first cleavage (range of 22.7-25.6h, P<0.05), while the lengths of the subsequent three cell cycles did not differ between bulls. The early mean time of first cleavage with bull III was associated with an early blastulation and a high blastocyst rate at Day 7, in opposition to what was observed with bull II showing a later timing of first cleavage (first cleavage 22.1 hpi versus 25.5 hpi; blastulation 140.4 hpi versus 152.5 hpi; D7 blastocyst rates: 31.3% versus 21.9%; P<0.05). In a third experiment, 65-76 Day 8 blastocysts per bull were sexed by PCR. Only blastocysts obtained with bull III showed a shift in sex ratio towards male embryos (76% male embryos; P<0.05). Such shift was already observed at the 2-cell and morula stages. In conclusion, the bull influences the kinetics of PN formation, of embryo development and the sex ratio of the embryos. Moreover, those parameters might be related. PMID:17629423

  5. Ploidy of Bovine Nuclear Transfer Blastocysts Blastomere Donors

    DEFF Research Database (Denmark)

    Booth, P J; VIUFF, D; THOMSEN, P D;

    2000-01-01

    reconstructed from in vitro produced embryo donors. In vitro matured oocytes were enucleated and then activated using calcium ionophore A23187 followed by 6- dimethylaminopurine (6-DMAP). Subsequently, embryos were reconstructed using blas- tomeres from day 4–5 in vitro produced donors. The embryos were...

  6. Effect of washing mineral oil on development of mouse embryos in vitro and in vivo after embryo transfer

    Institute of Scientific and Technical Information of China (English)

    Li Hui; Zhang Li-xuan; Zhong Yu; Zhu Kai; Zhang Tian; Wang Min-kang

    2008-01-01

    Objective:To establish a simple and effective washing procedure for both used and purchased mineral oil,that can be used for embryo culture.Methods:A complete test system has been used for this purpose.There are 3 steps in our new washing proto-col.First,the oil was mixed with 95% ethanol at 1:1,the bottle being shaken by hand for 10 minutes,then sepa-rated.Second,the oil was heated to boiling point with 0.31 mol/L NaCl for 30 minutes.Third,anhydrous Na:SO4 was put into the oil for further treatment.1-cell stage embryos of a KM strain mouse have been collected surgically and cultured.Cleavage and blastocyst stage development were recorded and some embryos were transferred into re-cipients.Results:The results show that recycled oil can promote the development from 2-cell to blastocyst stage(23.3%)when compared with that of control(16.9%).Offspring have been obtained at 44 %(7/16),16 %(3/19)from washed recycled oil and control oil respectively.Conclusion:This washing procedure is safe and effective for the used treatment and for other sources of mineral oil used for embryo culture.

  7. In Vitro Embryo Production and Transfer of Bubaline Embryos using Oocytes Derived from Transvaginal Ultrasound-Guided Follicular Aspiration (TUFA

    Directory of Open Access Journals (Sweden)

    FP Aquino

    2014-06-01

    Full Text Available Transvaginal ultrasound-guided follicular aspiration (TUFA has become a popular tool for embryo production in vitro due to its high degree of repeatability in terms of recovering oocytes from live animals. In Study 1, the quantity and quality of oocytes from Bulgarian Murrah buffalo cows (n=10 of varying ages (Group 1, 8-12; and Group 2, 13-17 years were assessed. Group 1 buffalo donor cows yielded significantly higher (P<0.05 number of oocytes vs Group 2 buffalo donor cows (71 vs 29 oocytes, respectively, though in terms of oocyte quality, no difference was observed. In Study 2, oocytes collected (n=100 in Study 1 were matured, fertilized in vitro and the resulting zygotes were cultured which developed to blastocyst stage embryos. The maturation, fertilization and blastocyst development rates obtained were 53.0%, 40.0% and 32.5%, respectively. In Study 3, the viability of resulting blastocyst stage embryos was determined by transferring to recipient cows. Of 10 recipients 1 got pregnant and delivered a 35 kg male calf after 310 days gestation period. Overall, the results of the studies conducted demonstrated the potential of TUFA technology in the in vitro production of embryos which eventually could be used in the production of live offspring.

  8. Effect of Equilibration Temperature on In vitro Viability and Subsequent Embryo Development of Vitrified-Warmed Immature Bovine Oocytes

    Directory of Open Access Journals (Sweden)

    Hajarian Hadi

    2010-01-01

    Full Text Available Problem statement: Vitrification is replacing conventional slow freezing to cryopreserve gametes and embryos especially for in vitro production of embryo in domestic animal species. However, the results are still not satisfactory. The aim of this experiment was to study the effect of different equilibration temperatures on in vitro viability of immature bovine oocytes after vitrification. Approach: Oocytes were obtained from slaughterhouse ovaries. Only grade one oocytes were used. Oocytes were equilibrated in three different temperatures: 32, 37, or 41°C. Immature oocytes were equilibrated in VS1 (7.5 Ethylene Glycol (EG + 7.5% DMSO for 10-12 min and then exposed to VS2 (15% EG + 15%DMSO + 0.5M sucrose for one min. Thereafter oocytes were loaded on hand-made Cryotop and directly plunged into liquid nitrogen. After warming, oocytes were examined for viability, maturation, cleavage and blastocyst production. Results: Oocytes that were equilibrated at 37°C had significantly higher (pConclusion: In conclusion, these results indicated that immature bovine oocytes can be equilibrated successfully at 37°C while higher or lower temperature can significantly decrease their subsequent viability and development.

  9. Superovulatory response and embryonic progressive in Iranian Qezel ewes treated with two different concentrations of bovine somatotropin

    Directory of Open Access Journals (Sweden)

    Amir Hossein Asgari Safdar

    2016-05-01

    Conclusions: The treatment 50 mg bovine somatotropins enhance the ratio and growth of the transferable embryos. Embryos of bST-50 treatment indicated an improved embryonic development but bST did not affect the pregnancy rates of transferred embryos.

  10. Effects of Multimodal Analgesia on the Success of Mouse Embryo Transfer Surgery

    OpenAIRE

    Parker, John M.; Austin, Jamie; Wilkerson, James; Carbone, Larry

    2011-01-01

    Multimodal analgesia is promoted as the best practice pain management for invasive animal research procedures. Universal acceptance and incorporation of multimodal analgesia requires assessing potential effects on study outcome. The focus of this study was to assess effects on embryo survival after multimodal analgesia comprising an opioid and nonsteroidal antiinflammatory drug (NSAID) compared with opioid-only analgesia during embryo transfer procedures in transgenic mouse production. Mice w...

  11. Functional full-term placentas formed from parthenogenetic embryos using serial nuclear transfer.

    Science.gov (United States)

    Hikichi, Takafusa; Ohta, Hiroshi; Wakayama, Sayaka; Wakayama, Teruhiko

    2010-09-01

    Mammalian parthenogenetic embryos invariably die in mid-gestation from imprinted gene defects and placental hypoplasia. Based on chimera experiments, trophoblastic proliferation is supposed to be inhibited in the absence of a male genome. Here, we show that parthenogenetic mouse embryonic cell nuclei can be reprogrammed by serial rounds of nuclear transfer without using any genetic modification. The durations of survival in uteri of cloned foetuses derived from green fluorescent protein (GFP)-labelled parthenogenetic cell nuclei were extended with repeated nuclear transfers. After five repeats, live cloned foetuses were obtained up to day 14.5 of gestation; however, they did not survive longer even when we repeated nuclear transfer up to nine times. All foetuses showed intestinal herniation and possessed well-expanded large placentas. When embryonic stem (ES) cells derived from fertilised embryos were aggregated with the cloned embryos, full-term offspring with large placentas were obtained from the chimeric embryos. Those placentas were derived from parthenogenetic cell nuclei, judging from GFP expression. The patterns of imprinted gene expression and methylation status were similar to their parthenogenetic origin, except for Peg10, which showed the same level as in the normal placenta. These results suggest that there is a limitation for foetal development in the ability to reprogramme imprinted genes by repeated rounds of nuclear transfer. However, the placentas of parthenogenetic embryos can escape epigenetic regulation when developed using nuclear transfer techniques and can support foetal development to full gestation.

  12. Finding biomarkers in non-model species: literature mining of transcription factors involved in bovine embryo development

    Directory of Open Access Journals (Sweden)

    Turenne Nicolas

    2012-08-01

    Full Text Available Abstract Background Since processes in well-known model organisms have specific features different from those in Bos taurus, the organism under study, a good way to describe gene regulation in ruminant embryos would be a species-specific consideration of closely related species to cattle, sheep and pig. However, as highlighted by a recent report, gene dictionaries in pig are smaller than in cattle, bringing a risk to reduce the gene resources to be mined (and so for sheep dictionaries. Bioinformatics approaches that allow an integration of available information on gene function in model organisms, taking into account their specificity, are thus needed. Besides these closely related and biologically relevant species, there is indeed much more knowledge of (i trophoblast proliferation and differentiation or (ii embryogenesis in human and mouse species, which provides opportunities for reconstructing proliferation and/or differentiation processes in other mammalian embryos, including ruminants. The necessary knowledge can be obtained partly from (i stem cell or cancer research to supply useful information on molecular agents or molecular interactions at work in cell proliferation and (ii mouse embryogenesis to supply useful information on embryo differentiation. However, the total number of publications for all these topics and species is great and their manual processing would be tedious and time consuming. This is why we used text mining for automated text analysis and automated knowledge extraction. To evaluate the quality of this “mining”, we took advantage of studies that reported gene expression profiles during the elongation of bovine embryos and defined a list of transcription factors (or TF, n = 64 that we used as biological “gold standard”. When successful, the “mining” approach would identify them all, as well as novel ones. Methods To gain knowledge on molecular-genetic regulations in a non model organism, we offer an

  13. Effects of oocyte quality, incubation time and maturation environment on the number of chromosomal abnormalities in IVF-derived early bovine embryos.

    Science.gov (United States)

    Demyda-Peyrás, Sebastian; Dorado, Jesus; Hidalgo, Manuel; Anter, Jaouad; De Luca, Leonardo; Genero, Enrique; Moreno-Millán, Miguel

    2013-01-01

    Chromosomal aberrations are one of the major causes of embryo developmental failures in mammals. The occurrence of these types of abnormalities is higher in in vitro-produced (IVP) embryos. The aim of the present study was to investigate the effect of oocyte morphology and maturation conditions on the rate of chromosomal abnormalities in bovine preimplantational embryos. To this end, 790 early cattle embryos derived from oocytes with different morphologies and matured under different conditions, including maturation period (24 v. 36h) and maturation media (five different serum supplements in TCM-199), were evaluated cytogenetically in three sequential experiments. The rates of normal diploidy and abnormal haploidy, polyploidy and aneuploidy were determined in each embryo. Throughout all the experiments, the rate of chromosomal abnormalities was significantly (P<0.05) affected by oocyte morphology and maturation conditions (maturation time and culture medium). Lower morphological quality was associated with a high rate of chromosome abnormalities (P<0.05). Moreover, polyploidy was associated with increased maturation time (P<0.01), whereas the maturation medium significantly (P<0.05) affected the rates of haploidy and polyploidy. In general, supplementing the maturation medium with oestrous cow serum or fetal calf serum resulted in higher rates of chromosomal aberrations (P<0.05) compared with the other serum supplements tested (bovine steer serum, anoestroues cow serum, bovine amniotic fluid and bovine serum albumin). On the basis of the results of the present study, we conclude that the morphological quality of oocytes and the maturation conditions affect the rate of chromosomal abnormalities in IVP bovine embryos.

  14. Cryopreservation and delayed embryo transfer-assisted reproductive technology registry and reporting implications.

    Science.gov (United States)

    Doody, Kevin J

    2014-07-01

    Clinics performing assisted reproductive technology (ART) procedures have collected data via registry and publicly reported pregnancy outcomes for more than 25 years. During this time, the practice of ART has changed considerably with frozen embryo transfer (FET) procedures contributing an increasing proportion of live births. Cycles initiated with the intent of embryo banking for the purpose of fertility preservation have been excluded from these public reports, because pregnancy outcomes are not immediately available. An unintended consequence of the common sense handling of fertility preservation has been that cycles performed with intentional short-term cryopreservation of all embryos for other indications have also been excluded from the report. Over the last few years, cryopreservation with short-term delayed transfer increasingly has been performed for reasons other than fertility preservation. The pregnancy outcomes of these cycles are expected within a reasonable time frame and should be transparently reported. The Society for Assisted Reproductive Technology has collaborated with the Centers for Disease Control and Prevention to "recapture" these cycles for the public reports. This recapture is done by linking the FET cycles to the stimulation cycles from which the embryos were derived and by changing the labels of the outcome success metrics. Stimulations using ART, initiated for the purpose of transferring embryos within 1 year will be included in the report despite any prospective intent to freeze all eggs or embryos. A positive outcome will be reported when a live birth results from the first embryo transfer following stimulation ("primary transfer"). Linkage of ovarian stimulation and egg-retrieval procedures to FET will also allow development of other success metrics to further benefit fertility patients.

  15. Avoiding multiple pregnancies in ART: a plea for single embryo transfer.

    Science.gov (United States)

    Gerris, J; Van Royen, E

    2000-09-01

    It has been generally accepted that triplets after IVF/intracytoplasmic sperm injection (ICSI) can and should be avoided by adopting a standard strategy of replacing no more than two embryos. However, there is an increasing awareness of the risks and costs and of the epidemic size of twin pregnancies after IVF/ICSI. This has resulted in efforts to replace no more than one embryo. However, this approach has been hampered by our relative inability to identify embryos with a very high implantation potential. To identify such embryos, a number of strategies are being considered, both at the two pronuclear (2PN), early cleavage and the blastocyst stages. At the 2PN stage, the polarity characteristics of the nucleoli have been shown to be correlated with a high implantation rate. Similarly, the morphological characteristics at day 2 and 3 have been used to describe top quality embryos in approximately 75% of all IVF/ICSI cycles. Blastocyst culture has resulted in very high implantation rates in the hands of some authors. No approach has shown its superiority at present, but initial experience with single embryo transfer (SET) at the early cleavage stage by Scandinavian and Belgian groups shows that an ongoing pregnancy rate of 35% and more can be achieved. Proper identification of patients at risk of a twin pregnancy after double embryo transfer is equally important. It is clear that mainly young patients (aged 80% of all twins) and are the main target group for twin prevention by SET of a top quality embryo at whatever stage. Therefore, in our opinion, although a further fine-tuning of both embryo and patient characteristics relating to a high risk for (twin) pregnancy is desirable, SET should be introduced carefully and progressively in each IVF/ICSI programme from now on. Correct counselling is very important and both public and private insurers will have to join in the discussion.

  16. Accidental Thawing of Embryos, Cryopreserved for Transfer. Two Italian cases, Milan and Rome.

    Science.gov (United States)

    Busardò, Francesco P; Vergallo, Gianluca Montanari; Turillazzi, Emanuela; Bolino, Giorgio; Vullo, Annamaria; Frati, Paola

    2016-01-01

    The bioethical and juridical debate on the status of frozen embryos sometimes adds new issues arising from new scientific evidence or by accidental occurrences that bring to the attention of the scientific community the need for new practical solutions. Within this scenario, there have been, in recent years, episodes concerning the accidental thawing of embryos, which have been cryopreserved for transfer. Two Italian cases (the Milan and the Rome cases) are here reported: the Milan case involves a couple undergoing artificial insemination. Three eggs were collected for insemination and two of them had been fertilized. During the night of 8/9 May 2007 a short circuit occurred, resulting in an electricity blackout, which caused the loss of the embryos in culture, which should have been transferred to the woman's uterus on 9 May. The couple applied for damage compensation from the hospital following the loss of the embryos. The case went to Court and the result was a judgment issued by the Milan civil court, which recognized that the centre was to blame for irreparable damage to the embryos. The Rome case, involves two couples (A and B) affected by sterility who applied to an authorized public centre to undergo an ART program. Following the medical procedures, two of the embryos produced were transferred to the woman in couple A and five were frozen, whereas three embryos produced by couple B were transferred to the uterus of the woman and six eggs were cryopreserved in the centre. Two years after the procedure there was an electricity blackout, and the backup electricity generator failed to function, causing the loss of the gametes and the embryos cryopreserved in the centre. Legal proceedings begun by the couples to obtain compensation for damages are still underway. The above reported cases have significantly intensified the bioethical debate on the lawfulness of such practices and on the fate of the cryopreserved embryos, at the same time opening new frontiers in

  17. Effect of cryopreservation and in vitro culture of bovine fibroblasts on histone acetylation levels and in vitro development of hand-made cloned embryos

    Science.gov (United States)

    Chacon, L.; Gomez, M.C.; Jenkins, J.A.; Leibo, S.P.; Wirtu, G.; Dresser, B.L.; Pope, C.E.

    2011-01-01

    In this study, the relative acetylation levels of histone 3 in lysine 9 (H3K9ac) in cultured and cryopreserved bovine fibroblasts was measured and we determined the influence of the epigenetic status of three cultured (C1, C2 and C3) donor cell lines on the in vitro development of reconstructed bovine embryos. Results showed that cryopreservation did not alter the overall acetylation levels of H3K9 in bovine fibroblasts analysed immediately after thawing (frozen/thawed) compared with fibroblasts cultured for a period of time after thawing. However, reduced cleavage rates were noted in embryos reconstructed with fibroblasts used immediately after thawing. Cell passage affects the levels of H3K9ac in bovine fibroblasts, decreasing after P1 and donor cells with lower H3K9ac produced a greater frequency of embryo development to the blastocyst stage. Cryopreservation did not influence the total cell and ICM numbers, or the ICM/TPD ratios of reconstructed embryos. However, the genetic source of donor cells did influence the total number of cells and the trophectoderm cell numbers, and the cell passage influenced the total ICM cell numbers. ?? Copyright Cambridge University Press 2010.

  18. cDNA microarray analysis of bovine embryo gene expression profiles during the pre-implantation period

    Directory of Open Access Journals (Sweden)

    Tokunaga Tomoyuki

    2004-11-01

    Full Text Available Abstract Background After fertilization, embryo development involves differentiation, as well as development of the fetal body and extra-embryonic tissues until the moment of implantation. During this period various cellular and molecular changes take place with a genetic origin, e.g. the elongation of embryonic tissues, cell-cell contact between the mother and the embryo and placentation. To identify genetic profiles and search for new candidate molecules involved during this period, embryonic gene expression was analyzed with a custom designed utero-placental complementary DNA (cDNA microarray. Methods Bovine embryos on days 7, 14 and 21, extra-embryonic membranes on day 28 and fetuses on days 28 were collected to represent early embryo, elongating embryo, pre-implantation embryo, post-implantation extra-embryonic membrane and fetus, respectively. Gene expression at these different time points was analyzed using our cDNA microarray. Two clustering algorithms such as k-means and hierarchical clustering methods identified the expression patterns of differentially expressed genes across pre-implantation period. Novel candidate genes were confirmed by real-time RT-PCR. Results In total, 1,773 individual genes were analyzed by complete k-means clustering. Comparison of day 7 and day 14 revealed most genes increased during this period, and a small number of genes exhibiting altered expression decreased as gestation progressed. Clustering analysis demonstrated that trophoblast-cell-specific molecules such as placental lactogens (PLs, prolactin-related proteins (PRPs, interferon-tau, and adhesion molecules apparently all play pivotal roles in the preparation needed for implantation, since their expression was remarkably enhanced during the pre-implantation period. The hierarchical clustering analysis and RT-PCR data revealed new functional roles for certain known genes (dickkopf-1, NPM, etc as well as novel candidate genes (AW464053, AW465434, AW

  19. Serial nuclear transfer improves the development of interspecies reconstructed giant panda (Aluropoda melanoleuca) embryos

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Interspecies somatic nuclear transfer (NT) may provide a new approach for preservation of the endangered rare species. Previous interspecies cloning studies have shown that a nucleus from a quiescent somatic cell supports early development of reconstructed embryos in the ooplasm from another species. In this study, we transferred nonquiescent somatic cells from a giant panda into the perivitelline space of the enucleated rabbit oocytes. After electrofusion (at the rate of 71.6%) and electrical activation, 4.2% of the panda-rabbit reconstructed embryos developed to blastocyst in vitro. For improving the development rate of reconstructed embryos, we used serial NT in this study, I.e. Blastomeres from reconstructed morulae were transferred into the perivitelline space of the enucleated rabbit oocytes. The fusion rates in the groups of serial Ⅰ, serial Ⅱ and serial Ⅲ were 79.5%, 84.1% and 78.0%, respectively, having no difference with that of somatic group. And the blastocyst rates in serial NT groups were 19.4%, 13.5% and 10.3%, respectively, which are significantly higher than that in somatic NT group. These results indicate that the nuclei from nonquiescent somatic cells can support early development of reconstructed embryos and serial NT can improve the development rate of interspecies reconstructed embryos.

  20. Effects of Histone Deacetylase Inhibitor Oxamflatin on In Vitro Porcine Somatic Cell Nuclear Transfer Embryos

    Science.gov (United States)

    Hou, Liming; Ma, Fanhua; Yang, Jinzeng; Riaz, Hasan; Wang, Yongliang; Wu, Wangjun; Xia, Xiaoliang; Ma, Zhiyuan; Zhou, Ying; Zhang, Lin; Ying, Wenqin; Xu, Dequan; Zuo, Bo; Ren, Zhuqing

    2014-01-01

    Abstract Low cloning efficiency is considered to be caused by the incomplete or aberrant epigenetic reprogramming of differentiated donor cells in somatic cell nuclear transfer (SCNT) embryos. Oxamflatin, a novel class of histone deacetylase inhibitor (HDACi), has been found to improve the in vitro and full-term developmental potential of SCNT embryos. In the present study, we studied the effects of oxamflatin treatment on in vitro porcine SCNT embryos. Our results indicated that the rate of in vitro blastocyst formation of SCNT embryos treated with 1 μM oxamflatin for 15 h postactivation was significantly higher than all other treatments. Treatment of oxamflatin decreased the relative histone deacetylase (HDAC) activity in cloned embryos and resulted in hyperacetylation levels of histone H3 at lysine 9 (AcH3K9) and histone H4 at lysine 5 (AcH4K5) at pronuclear, two-cell, and four-cell stages partly through downregulating HDAC1. The suppression of HDAC6 through oxamflatin increased the nonhistone acetylation level of α-tubulin during the mitotic cell cycle of early SCNT embryos. In addition, we demonstrated that oxamflatin downregulated DNA methyltransferase 1 (DNMT1) expression and global DNA methylation level (5-methylcytosine) in two-cell-stage porcine SCNT embryos. The pluripotency-related gene POU5F1 was found to be upregulated in the oxamflatin-treated group with a decreased DNA methylation tendency in its promoter regions. Treatment of oxamflatin did not change the locus-specific DNA methylation levels of Sus scrofa heterochromatic satellite DNA sequences at the blastocyst stage. Meanwhile, our findings suggest that treatment with HDACi may contribute to maintaining the stable status of cytoskeleton-associated elements, such as acetylated α-tubulin, which may be the crucial determinants of donor nuclear reprogramming in early SCNT embryos. In summary, oxamflatin treatment improves the developmental potential of porcine SCNT embryos in vitro. PMID

  1. What is the optimal means of preparing the endometrium in frozenthawed embryo transfer cycles? A systematic review and meta-analysis

    NARCIS (Netherlands)

    Groenewoud, Eva R.; Cantineau, Astrid E. P.; Kollen, Boudewijn J.; Macklon, Nick S.; Cohlen, Ben J.

    2013-01-01

    Frozenthawed embryo transfer (FET) enables surplus embryos derived from IVF or IVF-ICSI treatment to be stored and transferred at a later date. In recent years the number of FET cycles performed has increased due to transferring fewer embryos per transfer and improved laboratory techniques. Currentl

  2. A randomized controlled, non-inferiority trial of modified natural versus artificial cycle for cryo-thawed embryo transfer

    NARCIS (Netherlands)

    Groenewoud, E. R.; Cohlen, B. J.; Al-Oraiby, A.; Brinkhuis, E. A.; Broekmans, F. J M; De Bruin, J. P.; Van Den Dool, G.; Fleisher, K.; Friederich, J.; Goddijn, M.; Hoek, A.; Hoozemans, D. A.; Kaaijk, E. M.; Koks, C. A M; Laven, J. S E; Van Der Linden, P. J Q; Manger, A. P.; Slappendel, E.; Spinder, T.; Kollen, B. J.; Macklon, N. S.

    2016-01-01

    studyquestion: Are live birth rates (LBRs) after artificial cycle frozen-thawed embryo transfer (AC-FET) non-inferior to LBRs after modified natural cycle frozen-thawed embryo transfer (mNC-FET)? summaryanswer: AC-FET is non-inferior to mNC-FET with regard to LBRs, clinical and ongoing pregnancy rat

  3. The onset of foreign gene transcription in nuclear-transferred embryos of fish

    Institute of Scientific and Technical Information of China (English)

    孙永华; 陈尚萍; 汪亚平; 朱作言

    2000-01-01

    The transcriptional onset ot hGH-transgene in fish was studied in the following three cases: the first is in MThGH-transgenic F4 common carp (Cyprinus carpio) embryos, the second is in nuclear-transferred embryos supported by the transgenic F4 embryonic nuclei, and the third is in nuclear-transferred embryos supported by the transgenic F4 tail-fin nuclei. RT-PCR results show that the hGH-transgene initiates its transcriptional activity from early-gastrula stage, the early blastula stage and even 16-cell stage in the first, second and third cases, respectively. It looks like that fish egg cytoplasm could just offer a very restricted reprogramming on transcriptional activity of specific gene in differentiated cell nuclei by nuclear transplantation.

  4. The onset of foreign gene transcription in nuclear-transferred embryos of fish

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The transcriptional onset of hGH-transgene in fish was studied in the following three cases: the first is in MThGH-transgenic F4 common carp (Cyprinus carpio) embryos, the second is in nuclear-transferred embryos supported by the transgenic F4 embryonic nuclei, and the third is in nuclear-transferred embryos supported by the transgenic F4 tail-fin nuclei. RT-PCR results show that the hGH-transgene initiates its transcriptional activity from early-gastrula stage, the early blas-tula stage and even 16-cell stage in the first, second and third cases, respectively. It looks like that fish egg cytoplasm could just offer a very restricted reprogramming on transcriptional activity of specific gene in differentiated cell nuclei by nuclear transplantation.

  5. The impact of laser-assisted hatching on the outcome of frozen human embryo transfer cycles.

    Science.gov (United States)

    Kanyo, Katalin; Zeke, Jozsef; Kriston, Rita; Szücs, Zoltan; Cseh, Sandor; Somoskoi, Bence; Konc, Janos

    2016-10-01

    Biochemical modifications of zona pellucida (ZP) result in zona hardening. Zona hardening (ZH) is induced by several factors such as advancing maternal age, in vitro culture conditions and cryopreservation and adversely effects implantation. The objective of the clinical study was to determine whether or not laser-assisted hatching (LAH) applied on day 3 frozen embryos improves the outcome of frozen embryo transfer (FET) cycles in patients with recurrent implantation failure and/or advanced female age. In total, 413 patients of different ages with recurrent implantation failure (maximum three cycles) were involved into the study. Patients were allocated randomly into LAH and control groups. On the day of FET, after thawing and just before FET, the ZP was thinned using a laser system. In the control group no treatment was applied on frozen embryo before transfer. The main outcome measures were clinical pregnancy rate. Overall, the results indicate a tendency that LAH increased (P = 0.08) clinical pregnancy. However, for patients older than 37 years, LAH increased pregnancy rates significantly (P = 0.03). In the LAH and control groups, the age of patients and the number of transferred embryos influenced pregnancy rates (P = 0.01). For patients older than 37 years, no effect of number of transferred embryos was detected (P = 0.14). The incidence of multiple pregnancies also increased in the LAH group (P = 0.01). In conclusion, in older woman, to overcome the negative effect of zona hardening, LAH could be performed on frozen embryos as a routine strategy before FET in frozen cycles in order to increase the possibility of pregnancy formation.

  6. Successful pregnancy following the transfer of vitrified blastocyst which developed from poor quality embryos on day 3

    Directory of Open Access Journals (Sweden)

    Xiao-jian Zhang

    2011-01-01

    Full Text Available Background: The selection of pre-embryos for transferred is based on morphological appearance. But some poor quality cleaved embryos also can be cultured to the blastocyst stage and implanted.Objective: To assess the clinical pregnancy outcomes of blastocyst transfer which developed from poor quality embryos. Materials and Methods: A total of 109 cleaved embryos with poor quality were cultured to day 5/day 6 and 27 (24.8% blastocysts were collected from the 15 cycles/patients undergoing conventional IVF. All the blastocysts were cooling with fast-freezing. Then the blastocysts were warmed for transfer. Results: All of 25 vitrified blastocysts (92.6% survived after warming and were transferred to 15 patients. Five of the women became pregnant. Conclusion: Our results suggest that vitrified human day 5/day 6 blastocyst transfer which develop from poor quality embryo at day 3 can contribute to increasing cumulative pregnancy rates in assisted reproduction

  7. Term delivery following pyometra after in vitro fertilization and embryo transfer

    OpenAIRE

    Sathya Balasubramanyam; Thankam R Varma

    2014-01-01

    A 30 year old woman presented 12 days after embryo transfer with lower abdominal pain and orange vaginal discharge. She was diagnosed to have pyometra. A conservative management with drainage of the pyometra was followed by an uneventful pregnancy and term delivery. Conservative management in a case of pregnancy with pyometra needs close supervision to ensure maternal and fetal well being.

  8. Term delivery following pyometra after in vitro fertilization and embryo transfer.

    Science.gov (United States)

    Balasubramanyam, Sathya; Varma, Thankam R

    2014-04-01

    A 30 year old woman presented 12 days after embryo transfer with lower abdominal pain and orange vaginal discharge. She was diagnosed to have pyometra. A conservative management with drainage of the pyometra was followed by an uneventful pregnancy and term delivery. Conservative management in a case of pregnancy with pyometra needs close supervision to ensure maternal and fetal well being. PMID:25191031

  9. Term delivery following pyometra after in vitro fertilization and embryo transfer

    Directory of Open Access Journals (Sweden)

    Sathya Balasubramanyam

    2014-01-01

    Full Text Available A 30 year old woman presented 12 days after embryo transfer with lower abdominal pain and orange vaginal discharge. She was diagnosed to have pyometra. A conservative management with drainage of the pyometra was followed by an uneventful pregnancy and term delivery. Conservative management in a case of pregnancy with pyometra needs close supervision to ensure maternal and fetal well being.

  10. Cryo-thawed embryo transfer : natural versus artificial cycle. A non-inferiority trial. (ANTARCTICA trial)

    NARCIS (Netherlands)

    Groenewoud, Eva R.; Macklon, Nick S.; Cohlen, Ben J.

    2012-01-01

    Background: Frozen thawed embryo transfer (FET) is a cost-effective adjunct to IVF or IVF-ICSI treatment. In order to optimize treatment outcome, FET should be carried out during a period of optimal endometrial receptivity. To optimize implantation several methods for endometrium preparation have be

  11. Acupuncture on the day of embryo transfer: a randomized controlled trial of 635 patients

    DEFF Research Database (Denmark)

    Andersen, Dorota; Løssl, Kristine; Nyboe Andersen, Anders;

    2010-01-01

    sperm injection (ICSI) were included. In 314 patients, embryo transfer was accompanied by acupuncture according to the principles of traditional Chinese medicine. In the control group, 321 patients received placebo acupuncture using a validated placebo needle. In the acupuncture group and the placebo...

  12. Production of bovine cloned embryos with donor cells frozen at a slow cooling rate in a conventional freezer (20 C)

    Science.gov (United States)

    Chacon, L.; Gomez, M.C.; Jenkins, J.A.; Leibo, S.P.; Wirtu, G.; Dresser, B.L.; Pope, C.E.

    2009-01-01

    Summary Usually, fibroblasts are frozen in dimethyl sulphoxide (DMSO, 10% v/v) at a cooling rate of 1 C/min in a low-temperature (80 C) freezer (LTF) before storage in liquid nitrogen (LN2); however, a LTF is not always available. The purpose of the present study was to evaluate apoptosis and viability of bovine fibroblasts frozen in a LTF or conventional freezer (CF; 20 C) and their subsequent ability for development to blastocyst stage after fusion with enucleated bovine oocytes. Percentages of live cells frozen in LTF (49.5%) and CF (50.6%) were similar, but significantly less than non-frozen control (88%). In both CF and LTF, percentages of live apoptotic cells exposed to LN2 after freezing were lower (4% and 5%, respectively) as compared with unexposed cells (10% and 18%, respectively). Cells frozen in a CF had fewer cell doublings/24 h (0.45) and required more days (9.1) to reach 100% confluence at the first passage (P) after thawing and plating as compared with cells frozen in a LTF (0.96 and 4.0 days, respectively). Hypoploidy at P12 was higher than at P4 in cells frozen in either a CF (37.5% vs. 19.2%) or in a LTF (30.0% vs. 15.4%). A second-generation cryo-solution reduced the incidence of necrosis (29.4%) at 0 h after thawing as compared with that of a first generation cryo-solution (DMEM + DMSO, 60.2%). The percentage of apoptosis in live cells was affected by cooling rate (CF = 1.9% vs. LFT = 0.7%). Development of bovine cloned embryos to the blastocyst stage was not affected by cooling rate or freezer type. ?? 2009 Cambridge University Press.

  13. Conjugative transfer of resistance determinants among human and bovine Streptococcus agalactiae

    OpenAIRE

    Tatiana Castro Abreu Pinto; Natália Silva Costa; Ana Beatriz de Almeida Corrêa; Ivi Cristina Menezes Oliveira; Marcos Correa de Mattos; Alexandre Soares Rosado; Leslie Claude Benchetrit

    2014-01-01

    Streptococcus agalactiae (GBS) is a major source of human perinatal diseases and bovine mastitis. Erythromycin (Ery) and tetracycline (Tet) are usually employed for preventing human and bovine infections although resistance to such agents has become common among GBS strains. Ery and Tet resistance genes are usually carried by conjugative transposons (CTns) belonging to the Tn916 family, but their presence and transferability among GBS strains have not been totally explored. Here we evaluated ...

  14. Can Characteristics of Reciprocal Translocations Predict the Chance of Transferable Embryos in PGD Cycles?

    Directory of Open Access Journals (Sweden)

    Elsbeth Dul

    2014-04-01

    Full Text Available Translocation carriers have an increased risk of miscarriage or the birth of a child with congenital anomalies. Preimplantation genetic diagnosis (PGD is performed in translocation carriers to select for balanced embryos and, thus, increase the chance of an ongoing pregnancy. However, a common experience is that reciprocal translocation carriers produce a high percentage of unbalanced embryos, which cannot be transferred. Therefore, the pregnancy rates in PGD in this patient group are low. In a cohort of 85 reciprocal translocation carriers undergoing PGD we have searched for cytogenetic characteristics of the translocations that can predict the percentage of balanced embryos. Using shape algorithms, the most likely segregation mode per translocation was determined. Shape algorithm, breakpoint location, and relative chromosome segment sizes proved not to be independent predictors of the percentage of balanced embryos. The ratio of the relative sizes of the translocated segments of both translocation chromosomes can give some insight into the chance of transferable embryos: Very asymmetrical translocations have a higher risk of unbalanced products (p = 0.048. Counseling of the couples on the pros and cons of all their reproductive options remains very important.

  15. Role of ooplasm in nuclear and nucleolar remodeling of intergeneric somatic cell nuclear transfer embryos during the first cell cycle

    DEFF Research Database (Denmark)

    Østrup, Olga; Strejcek, Frantisek; Petrovicova, Ida;

    2011-01-01

    -cell stage embryos were processed at different points in time post activation (2 hpa, 4 hpa, 8 hpa, and 12 hpa) for detailed nuclear and nucleolar analysis by TEM, and immunofluorescence for visualization of nucleolar proteins related to transcription (UBF) and processing (fibrillarin). Bovine and porcine...

  16. Simulated Microgravity Influences Bovine Oocyte In Vitro Fertilization and Preimplantation Embryo Development

    Science.gov (United States)

    The aim of this study was to investigate whether in vitro fertilization and preimplantation embryos exposed to a simulated microgravity environment in vitro would improve, or be deleterious to, their fertilization and embryonic development. A Rotating Cell Culture System™ (RCCS) bioreactor with a Hi...

  17. Perinatal outcomes of children born after frozen-thawed embryo transfer: a Nordic cohort study from the CoNARTaS group

    DEFF Research Database (Denmark)

    Wennerholm, UB; Henningsen, Anna-Karina Aaris; Romundstad, LB;

    2013-01-01

    What are the risks of adverse outcomes in singletons born after frozen-thawed embryo transfer (FET)?......What are the risks of adverse outcomes in singletons born after frozen-thawed embryo transfer (FET)?...

  18. Increasing The Number of Embryos Transferred from Two to Three, Does not Increase Pregnancy Rates in Good Prognosis Patients

    Directory of Open Access Journals (Sweden)

    Mahnaz Ashrafi

    2015-10-01

    Full Text Available Background: To compare the pregnancy outcomes after two embryos versus three embryos transfers (ETs in women undergoing in vitro fertilization (IVF/intracytoplasmic sperm injection (ICSI cycles. Materials and Methods: This retrospective study was performed on three hundred eighty seven women with primary infertility and with at least one fresh embryo in good quality in order to transfer at each IVF/ICSI cycle, from September 2006 to June 2010. Patients were categorized into two groups according to the number of ET as follows: ET2 and ET3 groups, indicating two and three embryos were respectively transferred. Pregnancy outcomes were compared between ET2 and ET3 groups. Chi square and student t tests were used for data analysis. Results: Clinical pregnancy and live birth rates were similar between two groups. The rates of multiple pregnancies were 27 and 45.2% in ET2 and ET3 groups, respectively. The rate of multiple pregnancies in young women was significantly increased when triple instead of double embryos were transferred. Logistic regression analysis indicated two significant prognostic variables for live birth that included number and quality of transferred embryos; it means that the chance of live birth following ICSI treatment increased 3.2-fold when the embryo with top quality (grade A was transferred, but the number of ET had an inverse relationship with live birth rate; it means that probability of live birth in women with transfer of two embryos was three times greater than those who had three ET. Conclusion: Due to the difficulty of implementation of the elective single-ET technique in some infertility centers in the world, we suggest transfer of double instead of triple embryos when at least one good quality embryo is available for transfer in women aged 39 years or younger. However, to reduce the rate of multiple pregnancies, it is recommended to consider the elective single ET strategy.

  19. Live birth in a woman with recurrent implantation failure and adenomyosis following transfer of refrozen-warmed embryos

    Science.gov (United States)

    Safari, Somayyeh; Faramarzi, Azita; Khalili, Mohammad Ali

    2016-01-01

    The aim was to report a healthy live birth using re-vitrified-warmed cleavage-stage embryos derived from supernumerary warmed embryos after frozen embryo transfer (ET) in a patient with recurrent implantation failure (RIF). The case was a 39-year-old female with a history of polycystic ovarian syndrome and adenomyosis, along with RIF. After ovarian hyperstimulation, 33 cumulus-oocyte complexes were retrieved and fertilized with conventional in vitro fertilization and intracytoplasmic sperm injection. Because of the risk of ovarian hyperstimulation syndrome, 16 grade B and C embryos were vitrified. After 3 and 6 months, 3 and 4 B–C warmed embryos were transferred to the uterus, respectively. However, implantation did not take place. Ten months later, four embryos were warmed, two grade B 8-cell embryos were transferred, and two embryos were re-vitrified. One year later, the two re-vitrified cleavage-stage embryos were warmed, which resulted in a successful live birth. This finding showed that following first warming, it is feasible to refreeze supernumerary warmed embryos for subsequent ET in patients with a history of RIF. PMID:27689042

  20. Live birth in a woman with recurrent implantation failure and adenomyosis following transfer of refrozen-warmed embryos.

    Science.gov (United States)

    Safari, Somayyeh; Faramarzi, Azita; Agha-Rahimi, Azam; Khalili, Mohammad Ali

    2016-09-01

    The aim was to report a healthy live birth using re-vitrified-warmed cleavage-stage embryos derived from supernumerary warmed embryos after frozen embryo transfer (ET) in a patient with recurrent implantation failure (RIF). The case was a 39-year-old female with a history of polycystic ovarian syndrome and adenomyosis, along with RIF. After ovarian hyperstimulation, 33 cumulus-oocyte complexes were retrieved and fertilized with conventional in vitro fertilization and intracytoplasmic sperm injection. Because of the risk of ovarian hyperstimulation syndrome, 16 grade B and C embryos were vitrified. After 3 and 6 months, 3 and 4 B-C warmed embryos were transferred to the uterus, respectively. However, implantation did not take place. Ten months later, four embryos were warmed, two grade B 8-cell embryos were transferred, and two embryos were re-vitrified. One year later, the two re-vitrified cleavage-stage embryos were warmed, which resulted in a successful live birth. This finding showed that following first warming, it is feasible to refreeze supernumerary warmed embryos for subsequent ET in patients with a history of RIF. PMID:27689042

  1. What about the remaining twins since single-embryo transfer? How far can (should) we go?

    Science.gov (United States)

    De Neubourg, D; Gerris, J

    2006-04-01

    Single-embryo transfer (SET) and more specifically elective SET (eSET) have taken their place in good clinical IVF/ICSI practice. After the initial cautious search for the characteristics of the twin-prone patient and of the selection of the embryo with the highest implantation potential many centres have embarked on the (progressive) implementation of SET, either by conviction or forced by legislation or both. It was only because the ongoing pregnancy rates remained largely unaffected that SET was accepted. Generally speaking, it can be said that the twinning rate after IVF/ICSI has dropped by at least 50% simply by transferring only one good-quality embryo in the first and second fresh IVF/ICSI cycles in young women, without decrease in the overall pregnancy rate. Preventing 'the second half' of IVF/ICSI twins constitutes another and probably tougher challenge because the target group is a heterogeneous mix consisting of patients in very different clinical situations. Can we expand our experience for further twin prevention to women of older age and to cycles of higher rank without a significant drop in pregnancy rates? Can we extend it to more cryopreservation cycles? To have an idea of future target groups for increased application of SET, we analysed the remaining twins after double-embryo transfer (DET), and from these data we suggest expanding the eSET policy to women <38 years of age until the third cycle and to cryopreservation cycles.

  2. Effect of Adding Human Chorionic Gonadotropin to The Endometrial Preparation Protocol in Frozen Embryo Transfer Cycles

    Directory of Open Access Journals (Sweden)

    Maryam Eftekhar

    2012-01-01

    Full Text Available Background: Human chorionic gonadotropin (HCG, one of the initial embryonic signals, isprobably a major regulator of the embryo-endometrial relationship. This study aims to assess theadvantage of HCG supplementation during the secretory phase of hormonally prepared cycles forthe transfer of cryopreserved-thawed embryos.Materials and Methods: This study was a randomized clinical trial. Infertile women who werecandidates for frozen-thawed embryo transfers entered the study and were divided into two groups,HCG and control. The endometrial preparation method was similar in both groups: all women receivedestradiol valerate (6 mg po per day from the second day of the menstrual cycle and progesteronein oil (100 mg intramuscular (I.M. when the endometrial thickness reached 8 mm. Estradiol andprogesterone were continued until the tenth week of gestation. In the HCG group, patients received anHCG 5000 IU injection on the first day of progesterone administration and the day of embryo transfer.Results: In this study, 130 couples participated: 65 in the HCG group and 65 in the control group.There was no statistically significant difference between groups regarding basic characteristics.Implantation rate, chemical pregnancy, clinical pregnancy, ongoing pregnancy, and abortion rateswere similar in both groups.Conclusion: Although HCG has some advantages in assisted reproductive technology (ARTcycles, our study did not show any benefit of HCG supplementation during the secretory phase offrozen cycles (Registration Number: IRCT201107266420N4.

  3. Effects of Antioxidants on Development of In Vitro Fertilized Bovine Embryos

    OpenAIRE

    Anderson, Bret L.

    1995-01-01

    Free radicals are short-lived molecules that can cause decreased embryonic development in vitro. Antioxidants are molecules that block free radical formation or guard against their harmful effects. Many studies have linked exposure of media to light and culturing of embryos in high (20%) oxygen concentrations to free radical production. Some of the antioxidants used in culture media are superoxide dismutase (SOD), catalase, zinc (II), ethylenedinitrilo tetraacetic acid (EDTA), mannitol, vitam...

  4. Selection of reference genes for quantitative real-time PCR in bovine preimplantation embryos

    Directory of Open Access Journals (Sweden)

    Van Zeveren Alex

    2005-12-01

    Full Text Available Abstract Background Real-time quantitative PCR is a sensitive and very efficient technique to examine gene transcription patterns in preimplantation embryos, in order to gain information about embryo development and to optimize assisted reproductive technologies. Critical to the succesful application of real-time PCR is careful assay design, reaction optimization and validation to maximize sensitivity and accuracy. In most of the studies published GAPD, ACTB or 18S rRNA have been used as a single reference gene without prior verification of their expression stability. Normalization of the data using unstable controls can result in erroneous conclusions, especially when only one reference gene is used. Results In this study the transcription levels of 8 commonly used reference genes (ACTB, GAPD, Histone H2A, TBP, HPRT1, SDHA, YWHAZ and 18S rRNA were determined at different preimplantation stages (2-cell, 8-cell, blastocyst and hatched blastocyst in order to select the most stable genes to normalize quantitative data within different preimplantation embryo stages. Conclusion Using the geNorm application YWHAZ, GAPD and SDHA were found to be the most stable genes across the examined embryonic stages, while the commonly used ACTB was shown to be highly regulated. We recommend the use of the geometric mean of those 3 reference genes as an accurate normalization factor, which allows small expression differences to be reliably measured.

  5. Effects of bone morphogenic protein 4 (BMP4 and its inhibitor, Noggin, on in vitro maturation and culture of bovine preimplantation embryos

    Directory of Open Access Journals (Sweden)

    Fernandez-Martin Rafael

    2011-02-01

    Full Text Available Abstract Background BMP4 is a member of the transforming growth factor beta (TGFbeta superfamily and Noggin is a potent BMP inhibitor that exerts its function by binding to BMPs preventing interactions with its receptors. The aim of this work was to investigate the role of BMP4 and Noggin, on oocytes in vitro maturation (m experiments and embryos in vitro development (c experiments of bovine. Methods For m experiments, COCs were collected from slaughterhouse ovaries and in vitro matured in TCM with 100 ng/ml of either BMP4 or Noggin. After 24 h, the nuclear stage of the oocytes was determined by staining with Hoechst 33342. In addition, RT-qPCR was performed on MII oocytes to study the relative concentration of ZAR1, GDF9, BAX, MATER and HSP70 transcripts. Treated oocytes were submitted to parthenogenic activation (PA or in vitro fertilization (IVF and cultured in CR2. For c experiments, non-treated matured oocytes were submitted to PA or IVF to generate embryos that were exposed to 100 ng/ml of BMP4 or Noggin in CR2 until day nine of culture. Cleavage, blastocyst and hatching rates, expression pattern of the transcription factor Oct-4 in blastocysts and embryo cell number at day two and nine post-activation or fertilization were evaluated. Results We found that Noggin, as BMP4, did not affect oocyte nuclear maturation. Noggin supplementation up-regulated the expression of HSP70 and MATER genes in matured oocytes. Moreover, BMP4 during maturation increased the proportion of Oct-4 positive cells in parthenogenic embryos. On the other hand, when Noggin was added to embryo culture medium, developmental rates of parthenogenic and in vitro fertilized embryos were reduced. However, BMP4 addition decreases the development only for in vitro fertilized embryos. BMP4 and Noggin during culture reduced the proportion of Oct-4-expressing cells. Conclusions Our results show that BMP4 is implicated in bovine oocytes maturation and embryo development. Moreover

  6. Effects of multimodal analgesia on the success of mouse embryo transfer surgery.

    Science.gov (United States)

    Parker, John M; Austin, Jamie; Wilkerson, James; Carbone, Larry

    2011-07-01

    Multimodal analgesia is promoted as the best practice pain management for invasive animal research procedures. Universal acceptance and incorporation of multimodal analgesia requires assessing potential effects on study outcome. The focus of this study was to assess effects on embryo survival after multimodal analgesia comprising an opioid and nonsteroidal antiinflammatory drug (NSAID) compared with opioid-only analgesia during embryo transfer procedures in transgenic mouse production. Mice were assigned to receive either carprofen (5 mg/kg) with buprenorphine (0.1 mg/kg; CB) or vehicle with buprenorphine (0.1 mg/kg; VB) in a prospective, double-blinded placebo controlled clinical trial. Data were analyzed in surgical sets of 1 to 3 female mice receiving embryos chimeric for a shared targeted embryonic stem-cell clone and host blastocyst cells. A total of 99 surgical sets were analyzed, comprising 199 Crl:CD1 female mice and their 996 offspring. Neither yield (pups weaned per embryo implanted in the surgical set) nor birth rate (average number of pups weaned per dam in the set) differed significantly between the CB and VB conditions. Multimodal opioid-NSAID analgesia appears to have no significant positive or negative effect on the success of producing novel lines of transgenic mice by blastocyst transfer. PMID:21838973

  7. Cloned pigs derived from somatic cell nuclear transfer embryos cultured in vitro at low oxygen tension

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Pig cloning has great potential to human xenotransplantation. The present study was designed to establish a more efficient system for producing cloned pigs by somatic cell nuclear transfer (SCNT). Our approach was as follows: SCNT embryos were reconstructed by using fetal fibroblasts of Chinese miniature pig as donors and in vitro matured oocytes of prepubertal gilts as recipients. Reconstructed embryos were induced by electrical fusion/activation and cultured in BSA-containing North Carolina State University 23 medium (NCSU-23) or Porcine Zygote Medium (PZM-3) at the gas condition of 5% CO2, 7% O2, 88% N2. A total of 230 cloned embryos were transferred to three surrogate sows, producing three piglets. One of them is apparently healthy. The clonal provenance of the piglet was indicated by its coat color and confirmed by DNA microsatellite analysis. These results indicate that the use of in vitro matured oocytes from prepubertal gilts as recipient, combined with cloned embryos cultured at low oxygen tension is an effective way to produce cloned pigs.

  8. Refuting a misguided campaign against the goal of single-embryo transfer and singleton birth in assisted reproduction.

    Science.gov (United States)

    Stillman, Robert J; Richter, Kevin S; Jones, Howard W

    2013-10-01

    Much recent progress has been made by assisted reproductive technology (ART) professionals toward minimizing the incidence of multiple pregnancy following ART treatment. While a healthy singleton birth is widely considered to be the ideal outcome of such treatment, a vocal minority continues a campaign to advocate the benefits of multiple embryo transfer as treatment and twin pregnancy as outcome for most ART patients. Proponents of twinning argue four points: that patients prefer twins, that multiple embryo transfer maximizes success rates, that the costs per infant are lower with twins and that one twin pregnancy and birth is associated with no higher risk than two consecutive singleton pregnancies and births. We find fault with the reasoning and data behind each of these tenets. First, we respect the principle of patient autonomy to choose the number of embryos for transfer but counter that it has been shown that better patient education reduces their desire for twins. In addition, reasonable and evidentially supported limits may be placed on autonomy in exchange for public or private insurance coverage for ART treatment, and counterbalancing ethical principles to autonomy exist, especially beneficence (doing good) and non-maleficence (doing no harm). Second, comparisons between success rates following single-embryo transfer (SET) and double-embryo transfers favor double-embryo transfers only when embryo utilization is not comparable; cumulative pregnancy and birth rates that take into account utilization of cryopreserved embryos (and the additional cryopreserved embryo available with single fresh embryo transfer) consistently demonstrate no advantage to double-embryo transfer. Third, while comparisons of costs are system dependent and not easy to assess, several independent studies all suggest that short-term costs per child (through the neonatal period alone) are lower with transfers of one rather than two embryos. And, finally, abundant evidence conclusively

  9. Genetic variation in resistance of the preimplantation bovine embryo to heat shock.

    Science.gov (United States)

    Hansen, Peter J

    2014-12-01

    Reproduction is among the physiological functions in mammals most susceptible to disruption by hyperthermia. Many of the effects of heat stress on function of the oocyte and embryo involve direct effects of elevated temperature (i.e. heat shock) on cellular function. Mammals limit the effects of heat shock by tightly regulating body temperature. This ability is genetically controlled: lines of domestic animals have been developed with superior ability to regulate body temperature during heat stress. Through experimentation in cattle, it is also evident that there is genetic variation in the resistance of cells to the deleterious effects of elevated temperature. Several breeds that were developed in hot climates, including Bos indicus (Brahman, Gir, Nelore and Sahiwal) and Bos taurus (Romosinuano and Senepol) are more resistant to the effects of elevated temperature on cellular function than breeds that evolved in cooler climates (Angus, Holstein and Jersey). Genetic differences are expressed in the preimplantation embryo by Day 4-5 of development (after embryonic genome activation). It is not clear whether genetic differences are expressed in cells in which transcription is repressed (oocytes >100 µm in diameter or embryos at stages before embryonic genome activation). The molecular basis for cellular thermotolerance has also not been established, although there is some suggestion for involvement of heat shock protein 90 and the insulin-like growth factor 1 system. Given the availability of genomic tools for genetic selection, identification of genes controlling cellular resistance to elevated temperature could be followed by progress in selection for those genes within the populations in which they exist. It could also be possible to introduce genes from thermotolerant breeds into thermally sensitive breeds. The ability to edit the genome makes it possible to design new genes that confer protection of cells from stresses like heat shock. PMID:25472041

  10. Activation of ribosomal RNA genes in porcine embryos produced in vitro or by somatic cell nuclear transfer

    DEFF Research Database (Denmark)

    Bjerregaard, Bolette; Pedersen, Hanne Gervi; Jakobsen, Anne Sørig;

    2007-01-01

    nuclear transfer (SCNT) using fluorescence in situ hybridization (FISH) with an rDNA probe and subsequent visualization of the nucleolar proteins by silver staining. In the 205 IVP embryos investigated, all two-cell embryos (n = 34) were categorized as transcriptionally inactive. At the late four...

  11. Morphological characterization of pre- and peri-implantation in vitro cultured, somatic cell nuclear transfer and in vivo derived ovine embryos

    DEFF Research Database (Denmark)

    Tveden-Nyborg, Pernille Yde; Peura, T.T.; Hartwich, K.M.;

    2005-01-01

    The processes of cellular differentiation were studied in somatic cell nuvlear transfer (SCNT), in vitro cultured (IVC) and in vivo developed (in vivo) ovine embryos on days 7, 9, 11, 13, 17 and 19. SCNT embryos were constructed from in vitro matured oocytes and granulosa cells, and IVC embryos...... embryos had impaired hypoblast development, some lacking identifiable inner cell masses. On day 11, only in vivo and IVC embryos had developed an embryonic disc, and gastrulation was evident in half of in vivo embryos and one IVC embryo. By day 13, all in vivo embryos had completed gastrulation whereas...... IVC and SCNT embryos remained retarted. On days 17 and 19, in vivo embryos had significantly more somited and a more developed allantois than IVC and SCNT embryos. We conclude that IVC and particularly SCNT procedures cause a retardation of embryo development and cell differentiation at days 7...

  12. Increasing vaginal progesterone gel supplementation after frozen-thawed embryo transfer significantly increases the delivery rate

    DEFF Research Database (Denmark)

    Alsbjerg, Birgit; Polyzos, Nikolaos P; Elbaek, Helle Olesen;

    2013-01-01

    with oligoamenorrhoea (cycle length >34 days) or amenorrhoea undergoing oestradiol and progesterone priming prior to frozen-thawed embryo transfer. Patients treated with vaginal progesterone gel (Crinone 90 mg) twice daily had a lower risk of pregnancy loss (43.7%) compared with women treated once a day (67.4....... A total of 346 infertility patients with oligoamenorrhoea undergoing frozen-thawed embryo transfer after priming with oestradiol and vaginal progesterone gel were included. The vaginal progesterone dose was changed from 90 mg (Crinone) once a day to twice a day and the reproductive outcome during the two...... periods was compared. The pregnancy rate increased significantly after doubling of the progesterone dose (26.7% (90 mg) versus 38.4% (180 mg); P=0.021). Moreover, the early pregnancy loss rate decreased significantly (67.4% versus 43.7%, respectively; P=0.014), which significantly increased the delivery...

  13. Acupuncture on the day of embryo transfer: a randomized controlled trial of 635 patients

    DEFF Research Database (Denmark)

    Andersen, Dorthe; Løssl, Kristine; Nyboe Andersen, Anders;

    2010-01-01

    This prospective, randomized, controlled and double-blinded trial studied whether acupuncture in relation to embryo transfer could increase the ongoing pregnancy rates and live birth rates in women undergoing assisted reproductive therapy. A total of 635 patients undergoing IVF or intracytoplasmic...... sperm injection (ICSI) were included. In 314 patients, embryo transfer was accompanied by acupuncture according to the principles of traditional Chinese medicine. In the control group, 321 patients received placebo acupuncture using a validated placebo needle. In the acupuncture group and the placebo...... group, the ongoing pregnancy rates were 27% (95% CI 22-32) and 32% (95% CI 27-37), respectively. Live birth rates were 25% (95% CI 20-30) in the acupuncture group and 30% (95% CI 25-30) in the placebo group. The differences were not statistically significant. These results suggest that acupuncture...

  14. Serum Beta-hCG of 11 Days after Embryo Transfer to Predict Pregnancy Outcome

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Objective To assess the clinic value of a single maternal serum beta-human chorionic gonadotropin (β-hCG) assay 11 d after embryo transfer in ART pregnancies and to predict pregnancy outcome.Methods A total of 384 pregnancies after embryo transfer were included.Inviable pregnancies were defined as biochemical pregnancies,ectopic pregnancies and first trimester abortions.Ongoing pregnancies were defined as singleton pregnancies and multiple pregnancies whose gestation were achieved more than 12 weeks.Serum β-hCG concentrations were compared among different groups.Results On the post embryo transfer d 11,the mean β-hCG concentration of the ongoing pregnancy group (323.7±285.2 mIU/ml) was significantly higher than that of the inviable pregnancy group(81.4±68.1 mmIU/ml)(P<0.001).In multiple gestations,the levels of β-hCG were significantly higher compared with singleton pregnancies.If the β-hCG level was between 10 mIU/ml and 50 mIU/ml,the positive predictive value of biochemical pregnancies and ectopic pregnancies was 81.8%,the negative predictive value was 94.4%.If the level was less than 100 mIU/ml,the positive predictive value of first trimester abortions was 80.8% the negative predictive value was 77.8%.If the level was greater than 250 mIU/ml,the positive predictive value of multiple pregnancies was 83.3%.the negative predictive value was 74.4%.Conclusions A single serum β-hCG level on d 11 after embryo transfer has good predictive valuefor clinical pregnancy outcome in controlled ovarian stimulation cycles and helps to plan the subsequent follow-up.

  15. Relationship between the length of cell cycles, cleavage pattern and developmental competence in bovine embryos generated by in vitro fertilization or parthenogenesis.

    Science.gov (United States)

    Somfai, Tamás; Inaba, Yasushi; Aikawa, Yoshio; Ohtake, Masaki; Kobayashi, Shuji; Konishi, Kazuyuki; Imai, Kei

    2010-04-01

    This study was conducted to study the kinetics of initial cell divisions in relation with the cleavage patterns in viable (with the ability to develop to the blastocyst stage) and non-viable bovine embryos and parthenotes. The kinetics of in vitro development and cleavage patterns were observed by time lapse cinematography. The length of the first and second but not third cell cycle differed significantly between the viable and non-viable embryos after IVF or parthenogenesis. Viable embryos had significantly shorter first and second cell cycles than non-viable ones. The presence of fragments, protrusions and unequally-sized blastomeres was associated with an extended one-cell stage and reduced ability to develop to the blastocyst stage; however, the lengths of the second and third cell cycles were not altered. Oocytes showing direct division from one cell to 3 or 4 blastomeres showed similar developmental ability and embryonic cell numbers to those showing normal division, although, with a high frequency of chromosomal abnormalities. Our results suggest that the differences in the first cell cycles between viable and non-viable embryos were not sperm-related, whereas direct cleavage of 1-cell embryos to 3 or more blastomeres and protrusion formation are related to sperm-driven factors. The length of the first and second cell cycles and the cleavage pattern should be examined simultaneously to predict developmental competence of embryos at early cleavage stages. PMID:20035110

  16. Daily supplementation with ghrelin improves in vitro bovine blastocysts formation rate and alters gene expression related to embryo quality.

    Science.gov (United States)

    Dovolou, Eleni; Periquesta, Eva; Messinis, Ioannis E; Tsiligianni, Theodora; Dafopoulos, Konstantinos; Gutierrez-Adan, Alfonso; Amiridis, Georgios S

    2014-03-01

    Ghrelin is a gastric peptide having regulatory role in the reproductive system functionality, acting mainly at central level. Because the expression of ghrelin system (ghrelin and its receptor) has been detected in the bovine ovary, the objectives of the present study were to investigate whether ghrelin can affect the developmental potential of in vitro-produced embryos, and to test their quality in terms of relative abundance of various genes related to metabolism, apoptosis and oxidation. In the first experiment, in vitro-produced zygotes were cultured in the absence (control [C]) and in the presence of three concentrations of acylated ghrelin (200 pg/mL [Ghr200], 800 pg/mL [Ghr800]; and 2000 pg/mL [Ghr2000]); blastocyst formation rates were examined on Days 7, 8, and 9. In the second experiment, only the 800 pg/mL dose of ghrelin was used. Zygotes were produced as in experiment 1 and 24 hours post insemination they were divided into 4 groups; in two groups (C; without ghrelin; Ghr800 with ghrelin), embryos were cultured without medium replacement; in the remaining two groups (Control N and GhrN), the culture medium was daily renewed. A pool of Day-7 blastocysts were snap frozen for relative mRNA abundance of various genes related to metabolism, oxidation, implantation, and apoptosis. In experiment 3, embryos were produced as in experiment 2, but in the absence of serum (semi-defined culture medium). In experiment 1, no differences were detected between C, Ghr200, and Ghr2000, although fewer blastocysts were produced in group Ghr800 compared with C. In experiment 2, the lowest blastocysts yield was found in Ghr800, whereas daily renewal of ghrelin (Ghr800N) resulted to increased blastocysts formation rate, which on Day 7 was the highest among groups (P quality than controls. Our results imply a specific role of ghrelin in early embryonic development; however, the specific mode of its action needs further investigation. PMID:24332928

  17. Vitamin C enhances in vitro and in vivo development of porcine somatic cell nuclear transfer embryos

    International Nuclear Information System (INIS)

    Highlights: → Report for the first time that vitamin C has a beneficial effect on the development of porcine SCNT embryos. → The level of acH4K5 and Oct4 expression at blastocyst-stage was up-regulated after treatment. → A higher rate of gestation and increased number of piglets born were harvested in the treated group. -- Abstract: The reprogramming of differentiated cells into a totipotent embryonic state through somatic cell nuclear transfer (SCNT) is still an inefficient process. Previous studies revealed that the generation of induced pluripotent stem (iPS) cells from mouse and human fibroblasts could be significantly enhanced with vitamin C treatment. Here, we investigated the effects of vitamin C, to our knowledge for the first time, on the in vitro and in vivo development of porcine SCNT embryos. The rate of blastocyst development in SCNT embryos treated with 50 μg/mL vitamin C 15 h after activation (36.0%) was significantly higher than that of untreated SCNT embryos (11.5%). The enhanced in vitro development rate of vitamin C-treated embryos was associated with an increased acetylation level of histone H4 lysine 5 and higher Oct4, Sox2 and Klf4 expression levels in blastocysts, as determined by real-time PCR. In addition, treatment with vitamin C resulted in an increased pregnancy rate in pigs. These findings suggest that treatment with vitamin C is beneficial for enhancement of the in vitro and in vivo development of porcine SCNT embryos.

  18. Single embryo transfer and IVF/ICSI outcome: a balanced appraisal.

    Science.gov (United States)

    Gerris, Jan M R

    2005-01-01

    This review considers the value of single embryo transfer (SET) to prevent multiple pregnancies (MP) after IVF/ICSI. The incidence of MP (twins and higher order pregnancies) after IVF/ICSI is much higher (approximately 30%) than after natural conception (approximately 1%). Approximately half of all the neonates are multiples. The obstetric, neonatal and long-term consequences for the health of these children are enormous and costs incurred extremely high. Judicious SET is the only method to decrease this epidemic of iatrogenic multiple gestations. Clinical trials have shown that programmes with >50% of SET maintain high overall ongoing pregnancy rates ( approximately 30% per started cycle) while reducing the MP rate to <10%. Experience with SET remains largely European although the need to reduce MP is accepted worldwide. An important issue is how to select patients suitable for SET and embryos with a high putative implantation potential. The typical patient suitable for SET is young (aged <36 years) and in her first or second IVF/ICSI trial. Embryo selection is performed using one or a combination of embryo characteristics. Available evidence suggests that, for the overall population, day 3 and day 5 selection yield similar results but better than zygote selection results. Prospective studies correlating embryo characteristics with documented implantation potential, utilizing databases of individual embryos, are needed. The application of SET should be supported by other measures: reimbursement of IVF/ICSI (earned back by reducing costs), optimized cryopreservation to augment cumulative pregnancy rates per oocyte harvest and a standardized format for reporting results. To make SET the standard of care in the appropriate target group, there is a need for more clinical studies, for intensive counselling of patients, and for an increased sense of responsibility in patients, health care providers and health insurers.

  19. Vitamin C enhances in vitro and in vivo development of porcine somatic cell nuclear transfer embryos

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Yongye; Tang, Xiaochun; Xie, Wanhua; Zhou, Yan; Li, Dong; Zhou, Yang; Zhu, Jianguo; Yuan, Ting; Lai, Liangxue [Jilin Province Key Laboratory of Animal Embryo Engineering, College of Animal Science and Veterinary Medicine, Jilin University, 5333 Xi An DaLu, Changchun 130062 (China); Pang, Daxin, E-mail: pdx@jlu.edu.cn [Jilin Province Key Laboratory of Animal Embryo Engineering, College of Animal Science and Veterinary Medicine, Jilin University, 5333 Xi An DaLu, Changchun 130062 (China); Ouyang, Hongsheng, E-mail: ouyh@jlu.edu.cn [Jilin Province Key Laboratory of Animal Embryo Engineering, College of Animal Science and Veterinary Medicine, Jilin University, 5333 Xi An DaLu, Changchun 130062 (China)

    2011-07-29

    Highlights: {yields} Report for the first time that vitamin C has a beneficial effect on the development of porcine SCNT embryos. {yields} The level of acH4K5 and Oct4 expression at blastocyst-stage was up-regulated after treatment. {yields} A higher rate of gestation and increased number of piglets born were harvested in the treated group. -- Abstract: The reprogramming of differentiated cells into a totipotent embryonic state through somatic cell nuclear transfer (SCNT) is still an inefficient process. Previous studies revealed that the generation of induced pluripotent stem (iPS) cells from mouse and human fibroblasts could be significantly enhanced with vitamin C treatment. Here, we investigated the effects of vitamin C, to our knowledge for the first time, on the in vitro and in vivo development of porcine SCNT embryos. The rate of blastocyst development in SCNT embryos treated with 50 {mu}g/mL vitamin C 15 h after activation (36.0%) was significantly higher than that of untreated SCNT embryos (11.5%). The enhanced in vitro development rate of vitamin C-treated embryos was associated with an increased acetylation level of histone H4 lysine 5 and higher Oct4, Sox2 and Klf4 expression levels in blastocysts, as determined by real-time PCR. In addition, treatment with vitamin C resulted in an increased pregnancy rate in pigs. These findings suggest that treatment with vitamin C is beneficial for enhancement of the in vitro and in vivo development of porcine SCNT embryos.

  20. Effect of zona pellucida removal on early development of in vitro produced bovine embryos Efecto de la remoción de la zona pelúcida sobre el desarrollo temprano de embriones bovinos producidos in vitro

    OpenAIRE

    AE Velásquez; JR Manríquez; FO Castro; Ll Rodríguez-Alvarez

    2013-01-01

    During early embryo development the zona pellucida acts as a barrier against polyspermia and guarantees communication between blastomeres before and during compaction. However, the development of new technologies of embryo production such as "Handmade Cloning" demands removal of this membrane. The aim of this study was to determine the effect of zona pellucida removal on in vitro bovine embryo development. First, the consequences of zona pellucida removal was assessed by comparing the percent...

  1. Impact on outcome of frozen-thawed embryo transfer by reducing numbers of transferred embryos%减少移植胚胎数量对冻融胚胎移植结局的影响

    Institute of Scientific and Technical Information of China (English)

    江楠; 王丽萍; 常永富; 付伟平

    2011-01-01

    Objective To study the impact on pregnant outcome of reducing the number of embryos transferred from three to two in women at age less than 35 who received frozen-thawed embryo transfer (FET). Methods The analysis was performed on 90 FET cycles (77 infertile couples,less than 35 years old) with slow-freezing/rapid-thawing method, including 48 cycles with two embryos transferred and 42 cycles with three embryos transferred. The embryo survival rate, high quality embryo rate, clinical pregnancy rate, implantation rate and multiple pregnancies rate were analyzed. Results No significant differences in embryo survival rate (88.9% versus 88.1%), high quality embryo rate (89.6% versus 81.0%), clinical pregnancy rate (37.5% versus 42.9%), implantation rate (26.0% versus 18.3%) and multiple pregnancy rate (38.9% versus 16.7%) were observed between two and three embryos transferred group (all P > 0.05). However, there were 2 triple pregnancies in three embryos transferred group while none in two embryos transferred group. Conclusion Reducing the number of high quality embryos transferred from three to two in women at age of less than 35 years old who received FET,could decrease the incidence of triple pregnancy and keep the similar clinical pregnancy rate.%目的 探讨年龄0.05).但移植2个胚胎者中无三胎妊娠发生,而移植3个胚胎者中发生2次三胎妊娠.结论 冻融胚胎移植周期中,如果胚胎质量良好,年龄<35岁妇女移植胚胎数量从3个减少至2个,并不会明显降低临床妊娠率,同时可减少三胎妊娠的发生.

  2. Establishment of Multiple Ovulation and Embryo Transfer (MOET) technology for goats in Sri Lanka

    International Nuclear Information System (INIS)

    This study was conducted to determine a suitable follicular stimulating hormone (FSH) preparation for superovulation in goats, establish techniques for embryo production and transfer in goats, and to examine the feasibility of applying such techniques in Sri Lanka. Two groups of genetically superior does were inserted with progesterone releasing intravaginal pessaries (45 mg Cronolone) on d 1 of the programme. On d 8, the does in Group 1 (n = 3) and Group 2 (n = 4) were given 2.5 mL injections of pure porcine FSH (pFSH, 20 mg/mL) or pure ovine FSH (oFSH, 0.88 mg/mL), respectively. On the same day, all animals were injected with 300 IU pregnant mare serum gonadotropin (PMSG, 500 μg/mL). Subsequent injections of 1.25 mL pFSH or oFSH were given in the morning and evening on d 9 and 10. Does were injected with 197 μg prostaglandin F2α (PGF2α, 263 μg/mL) in the morning of d 9 and vaginal pessaries were removed on the even- ing of d 10. On d 11, 1.25 mL of pFSH or oFSH and 1 mL of luteinising hormone releasing hormone (LHRH, 50 μg/mL) injections were given in the morning and evening, respectively. On the same day, does in oestrus were bred to two Jamnapari bucks. Seven d post- oestrus, embryos were collected surgically, using embryo flushing medium. The quality of the embryos was assessed and the recovered embryos were transplanted surgically to oestrus synchronised goat recipients (n = 4/group) at 7 d post-oestrus. Following embryo transplantation, four does (Group 1, n = 1, Group 2, n = 3) were found to be pregnant by ultrasound scanning at 35 d into pregnancy. One healthy female offspring (Peradeniya Kumari) was born to Group 1. Another four goat kids were born to Group 2, while one kid died. In the same group, one abortion was reported. The results suggest that oFSH is better than pFSH for the superovulation of goats and that embryo transfer technology can be used in goats in Sri Lanka. (author)

  3. Use of purified FSH and LH for embryo production, cryopreservation by conventional freezing or vitrification and transfer of embryos in dairy ewes

    Directory of Open Access Journals (Sweden)

    Giovanni Martemucci

    2010-01-01

    Full Text Available Three experiments were carried out with the aim of evaluating the efficiency of techniques of in vivo production, storageand transfer of embryos in dairy sheep. Experiment I - For embryo production, thirty-one ewes were synchronized withFGA (vaginal sponges, 40 mg, 9 d and PGF2α (ICI; 50 μg, 7th d, and subdivided into three groups corresponding to thefollowing superovulatory treatments over 3 days with purified gonadotrophic preparations: A control, FSH/LH ratio = 1(250 IU p-FSH : 250 UI p-LH; B FSH/LH ratio = 2 (250 IU p-FSH : 125 IU p-LH and daily FSH/LH ratio of 3.4 – 1.7 –0.8 in the 3 days of treatment, respectively; C FSH/LH ratio = 2 (250 IU p-FSH : 125 IU p-LH and daily FSH/LH ratioof 5.0 – 1.0 – 0.3. On the 7th day after oestrus and mating, ovarian response and embryo production were evaluated.Experiment II – Three freezing methods were evaluated based upon post-thaw embryo quality: CF conventional slowfreezing by 1.5 M ethylene glycol (EG; V-1 one-step vitrification based on exposure of the embryos to one solution (EG7.15 M + ficoll 2.5 mM; V-3 vitrification in three steps, corresponding to three solutions at increasing concentration ofglycerol (GLY and EG (GLY 1.4 M; GLY 3.4 M + EG 1.4 M; GLY 4.6 M + EG 3.4 M. V-1 and V-3 frozen embryos weredirectly plunged in liquid nitrogen. At thawing, embryo viability was evaluated on the basis of morphological features.Experiment III – For embryo transfer, a total of 26 recipient ewes were synchronized with donors. On the 7th d fromoestrus, 11 recipient ewes received fresh embryos (Group FE – control and 15 recipients received vitrified-thawedembryos (Group VTE. Each recipient received 2 embryos. Superovulatory treatment B significantly advanced the onsetof oestrus compared to the control (27.3 vs 34.7 h; P10.8. Transferable embryos in Group B (7.2 resulted similar to Group A (5.3 and significantly (Pcompared to Group C (3.2. V3-method resulted in the highest (PCF- and V1-methods

  4. Elective single-embryo transfer: persuasive communication strategies can affect choice in a young British population.

    Science.gov (United States)

    van den Akker, O B A; Purewal, S

    2011-12-01

    This study tested the effectiveness of the framing effect and fear appeals to inform young people about the risks of multiple births and the option of selecting elective single-embryo transfer (eSET). A non-patient student sample (age (mean±SD) 23±5.5 years; n=321) were randomly allocated to one of seven groups: (1) framing effect: (1a) gain and (1b) loss frame; (2) fear appeal: (2a) high, (2b) medium and (2c) low fear; or (3) a control group: (3a) education and (3b) non-education. The primary outcome measure was the Attitudes towards Single Embryo Transfer questionnaire, before exposure to the messages (time 1) and immediately afterwards (time 2). Results revealed participants in the high fear, medium fear and gain condition demonstrated the most positive and significant differences (Ppromote immediate and hypothetical eSET preferences is more successful at promoting eSET than merely reporting educational content. Future research should investigate its application in a clinical population. A multiple pregnancy is a health risk to both infant and mother following IVF treatment. The aims of this study were to test the effectiveness of two persuasive communication techniques (the framing effect and fear appeals) to inform young people about the risks of multiple births and the hypothetical option of selecting elective single-embryo transfer (eSET) (i.e., only one embryo is transferred to the uterus using IVF treatment). A total of 321 non-patient student sample (mean age 23) were randomly allocated to read a message from one of seven groups: (1) framing effect: (1a) gain and (1b) loss frame; (2) fear appeal: (2a) high, (2b) medium and (2c) low fear; or (3) a control group: education (3a) and (3b) non-education. Participants completed the Attitudes towards Single Embryo Transfer questionnaire, before exposure to the messages (time 1) and immediately afterwards (time 2). Results revealed that participants in the high fear, medium fear and gain condition demonstrated

  5. The fate of frozen human embryos when transferred either on the day of thawing or after overnight culture

    Institute of Scientific and Technical Information of China (English)

    Yanhe Liu; Kelli Peirce; Kailin Yap; Kate McKenzie; Jay Natalwala; Vince Chapple; Margo Norman; Phillip Matson

    2012-01-01

    Objective:To study the performance of thawed zygotes and cleavage stage embryos transferred either on the day of thaw or after overnight culture.Methods:A retrospective study of864 frozen embryo transfer cycles.Cryosurvival rates per thawed embryo and implantation rates were analysed for embryos frozen onDay1,Day2 orDay3 relative to oocyte collection(Day0) and transferred on the day of thaw or after overnight culture, together with clinical pregnancy rates and prevalence of multiple gestations.Results:Survival ofDay3 embryos was significantly lower than those frozen onDay1(P=0.017) orDay2(P=0.015).Following overnight culture, resumption of mitosis of zygotes was more frequent thanDay2(P=0.000) which are in turn higher thanDay3(P=0.000) embryos.The implantation rate forDay2 embryos dividing overnight was significantly higher than those that did not divide for women <35 yrs(P=0.001) but not those women≥35 yrs(P=0.055).There were no differences in the implantation rates for those dividing or not after culture, for embryos frozen onDay3 for women <35 yrs(P=0.254) or≥35 yrs(P=0.403). Conclusions:Later cleavage stage post-thaw embryos survive and resume mitosis less frequently compared to earlier stages.Embryos not resuming mitosis after culture overnight can implant, particularlyDay3 embryos, suggesting that they can further increase the cumulative pregnancy rate per oocyte collection and that discarding them is wasteful.Overnight culture is best used for logistical reasons rather than a strategy to improve pregnancy rates.

  6. Production of rhesus monkey cloned embryos expressing monomeric red fluorescent protein by interspecies somatic cell nuclear transfer

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Hai-Ying; Kang, Jin-Dan; Li, Suo; Jin, Jun-Xue; Hong, Yu; Jin, Long; Guo, Qing; Gao, Qing-Shan; Yan, Chang-Guo; Yin, Xi-Jun, E-mail: yinxj33@msn.com

    2014-02-21

    Highlights: • Rhesus monkey cells were electroporated with a plasmid containing mRFP1, and an mRFP1-expressing cell line was generated. • For the first time, mRFP1-expressing rhesus monkey cells were used as donor cells for iSCNT. • The effect of VPA on the development of embryos cloned using iSCNT was determined. - Abstract: Interspecies somatic cell nuclear transfer (iSCNT) is a promising method to clone endangered animals from which oocytes are difficult to obtain. Monomeric red fluorescent protein 1 (mRFP1) is an excellent selection marker for transgenically modified cloned embryos during somatic cell nuclear transfer (SCNT). In this study, mRFP-expressing rhesus monkey cells or porcine cells were transferred into enucleated porcine oocytes to generate iSCNT and SCNT embryos, respectively. The development of these embryos was studied in vitro. The percentage of embryos that underwent cleavage did not significantly differ between iSCNT and SCNT embryos (P > 0.05; 71.53% vs. 80.30%). However, significantly fewer iSCNT embryos than SCNT embryos reached the blastocyst stage (2.04% vs. 10.19%, P < 0.05). Valproic acid was used in an attempt to increase the percentage of iSCNT embryos that developed to the blastocyst stage. However, the percentages of embryos that underwent cleavage and reached the blastocyst stage were similar between untreated iSCNT embryos and iSCNT embryos treated with 2 mM valproic acid for 24 h (72.12% vs. 70.83% and 2.67% vs. 2.35%, respectively). These data suggest that porcine-rhesus monkey interspecies embryos can be generated that efficiently express mRFP1. However, a significantly lower proportion of iSCNT embryos than SCNT embryos reach the blastocyst stage. Valproic acid does not increase the percentage of porcine-rhesus monkey iSCNT embryos that reach the blastocyst stage. The mechanisms underling nuclear reprogramming and epigenetic modifications in iSCNT need to be investigated further.

  7. Storage of bovine isolated follicles: a new alternative way to improve the recovery rate of viable embryos from ovarian follicles of slaughtered cows.

    Science.gov (United States)

    Pavlok, A; Cech, S; Kubelka, M; Lopatárová, M; Holý, L; Jindra, M

    2006-11-01

    The vitality of bovine oocytes stored in isolated follicles was examined. The aim of this work was to prolong the time of in vitro manipulation of oocytes before their maturation and develop a new alternative of oocyte "capacitation" to improve the quality of in vitro produced embryos. Follicles were dissected from the ovaries of slaughtered cows; subsequently, follicles were divided according to their diameter into three categories (2-3, 3-4 and 4-6 mm), and stored at 17-18 degrees C for 24 or 48 h in a modified tissue culture medium-199 (TCM-199) with reduced pH. After that time, the cumulus-oocyte complexes (COCs) were isolated, matured, fertilized, and embryos cultured in vitro for a total of 9 days. The percentage of total blastocysts, and hatched blastocysts developed from oocytes, initially kept ("capacitated") for 24h at 17-18 degrees C, within follicles of 3-6mm size categories, were significantly higher than that oocytes of the control [of control oocytes] (44.9 and 30.3% versus 36.2 and 20.4%, respectively). The oocytes of follicles stored for 48 h at 17-18 degrees C already had decreased developmental capacity. Interesting data were obtained when COCs of the 3-4 and 4-6 categories were additionally divided into two subgroups according to their presumed developmental history (originating from the supposed growing "fit" in contrast to the supposed regressing "unfit" follicles). The higher improvement in the rate of hatched blastocysts from 24h stored oocytes was observed only in the subgroup originated from "fit" COCs (15.3 versus 25.0%, and 20.0 versus 34.4%, in the 3-4 and 4-6mm categories, respectively). The transfer of 26 blastocysts (developed of follicles kept for 24h at 17-18 degrees C) to 26 recipient heifers resulted in 18 pregnancies. Storage of follicles at 17-18 degrees C in vitro resulted not only in recovery of higher numbers of blastocysts of better quality but also facilitated the safe transport of follicles for a long distance. The

  8. Study on Sex Determination of Bovine Pre-implantation Embryos By Bovine Y Chromosome Repeated Sequence%利用牛Y染色体重复序列进行早期胚胎性别鉴定的研究

    Institute of Scientific and Technical Information of China (English)

    王世银; 张伟; 张兆旺; 赵兴绪

    2011-01-01

    本试验利用Y染色体重复序列作为雄性特异性引物,以肿瘤坏死因子(TNF-α) 内标引物建立多重PCR体系,进行牛早期胚胎性别鉴定.共设计四对引物一Y染色体重复序列外引物和内引物,其大小分别为534bp和480bp;肿瘤坏死因子外引物和内引物大小分别为357bp和272bp.试验结果表明,优化后的多重PCR体系的灵敏度分别达到3个胚胎细胞,准确率100%,可以满足早期胚胎性别鉴定的需要.%In this study, we designed four pairs of primers which the amplifiment products length were 534bp, 480bp, 357bp and 272bp respectively according to Y chromosome repeated sequence and tumor necrosis factor alpha(TNF-α) for sex determination of bovine embryo.The result shows that these four pairs of primers all have highly specificity and stability.The Multi-PCR need only 3 cells DNA to determine the sex of embryo, so it is more suitable for sex determination of bovine embryo.

  9. In vivo survival of domestic cat oocytes after vitrification, intracytoplasmic sperm injection and embryo transfer.

    Science.gov (United States)

    Pope, C E; Gómez, M C; Kagawa, N; Kuwayama, M; Leibo, S P; Dresser, B L

    2012-02-01

    We evaluated: (1) cleavage rate after IVF or intracytoplasmic sperm injection (ICSI) of in vivo- and in vitro-matured oocytes after vitrification (experiment 1); and (2) fetal development after transfer of resultant ICSI-derived embryos into recipients (experiment 2). In vivo-matured cumulus-oocyte complexes (COCs) were recovered from gonadotropin-treated donors at 24 h after LH treatment. In vitro-matured oocytes were obtained by mincing ovaries (from local veterinary clinics) and placing COCs into maturation medium for 24 h. Mature oocytes were denuded and cryopreserved in a vitrification solution of 15% DMSO, 15% ethylene glycol, and 18% sucrose. In experiment 1, for both in vivo- and in vitro-matured oocytes, cleavage frequencies after IVF of control and vitrified oocytes and after ICSI of vitrified oocytes were not different (P > 0.05). After vitrification, blastocyst development occurred only in IVF-derived, in vitro-matured oocytes. In experiment 2, 18 presumptive zygotes and four two-cell embryos derived by ICSI of vitrified in vitro-matured oocytes and 19 presumptive zygotes produced from seven in vivo- and 12 in vitro-matured oocytes were transferred by laparoscopy into the oviducts of two recipients, respectively. On Day 21, there were three fetuses in one recipient and one fetus in the other. On Days 63 and 66 of gestation, four live kittens were born. In vivo viability of zygotes and/or embryos produced via ICSI of vitrified oocytes was established by birth of live kittens after transfer to recipients. PMID:22015162

  10. Study on relationship between perifollicular blood flow and in vitro fertilization-embryo transfer

    Institute of Scientific and Technical Information of China (English)

    Yan Zhang; Jing Yang; Wangming Xu

    2008-01-01

    Objective: To study the relationship between perifoUicular blood flow and follicule development, oocyte maturing rate, fertilizing rate, cleaving rate, embryo quality and the outcomes of embryo transfer. Methods: The samples were selected from 66 suffers who underwent in vitro fertilization(IVF)or intracytoplasmic sperm injection(ICSI). Eeach patients' perifollicular blood flow(diameter≥12mm )was estimated on the day of human chorionic gonadotropin(HCG)administration. Results:Among 66 cycles, 26(39.4%) cycles resulted in pregnancy, perifollicular blood flow resistance index(Rl), peak systolic velocity/end diastasis velocity(S/D) of non-preg-nant group was significantly higher than that of the pregnant group (P < 0.004). When RI<0.49, the pregnancy rates, fecundation rates, fertilization rates, metaphase numbers for the of second meiosis oocytes increased evidently(P<0.05), but there were no statistical difference in gonadotropin dosage, cycle frequency, infertility years, ages, estradiol(E2)on the day of HCG administration,numbers of oocyet retrieved and high-quality embryo rates (P > 0.05 ). There were no statistical difference between non-pregnant group and pregnant group in S and D (P>0.05). There was no correlation between periFollicular blood flow RI and follicular diameter by linear regression analysis. Conclusion:Our study shows that perifollicular blood flow RI and S/D are effective indices of predicting the pregnancy outcome of IVF-ET.

  11. Successful transfer of frozen N'Dama embryos from the Gambia to Kenya.

    Science.gov (United States)

    Jordt, T; Mahon, G D; Touray, B N; Ngulo, W K; Morrison, W I; Rawle, J; Murray, M

    1986-05-01

    Frozen embryos from N'Dama cattle were successfully transferred from The Gambia to Kenya. Of the 26 N'Dama cows used 12 were successfully programmed to superovulate and of these seven produced 30 embryos that were collected seven days after oestrus/service. Five N'Dama bulls were used for natural service. In Kenya 29 embryos were implanted into 29 Boran heifers seven days (+/- 1) after the induction of synchronised oestrus. Eleven pregnancies were established and after one abortion of unknown aetiology at seven and a half months five female and five male calves were born and subsequently reared. During programming the N'Dama cows showed prolonged anoestrus leading to the necessity of oestrus induction using intravaginal progesterone releasing coils; pregnant mare serum gonadotrophin gave better superovulation than follicle stimulating hormone. One N'Dama bull proved to be subfertile. The success of the project has demonstrated the potential of this technique to make disease-free N'Dama available for research purposes and for the promotion of livestock development programmes in tsetse-infested areas using trypanotolerant cattle. PMID:3738996

  12. Effects of different nuclear transfer and activation methods on the development of mouse somatic cell cloned embryos

    Institute of Scientific and Technical Information of China (English)

    Wang ErYao; YU Yang; Li XueMei; JIAO LiHong; Wang Liu

    2007-01-01

    A group of adult somatic cell cloned mice were obtained by using cumulus cells as nuclei donor cells. To study the effect of different nuclear transfer (NT) and activation methods on the development of mouse cloned embryos, embryos were reconstructed using two traditional NT methods (electrofusion and direct injection) and four activation treatments (electric pulse, ethanol, SrCl2 and electric pulse combined with SrCl2). The data showed that the efficiency of reconstruction using the direct injection method is significantly higher (90.7%) than that of the electrofusion method (49.7%). Parthenogenetic embryos can develop to blastocyst stage with three activation conditions, including ethanol, electric pulse and SrCl2; however, the rates of development to blastocyst after ethanol and electric pulse activation (52.4%, 54.2%) are significantly lower than after SrCl2 activation (76.9%). Treatment of embryos for 6 h with 10 mmol/L SrCl2 was found to be the best condition for activation of parthenogenetic as well as reconstructed embryos. By contrast, reconstructed embryos failed to develop to blastocyst stage after being activated by ethanol. The use of either injection or electrofusion for embryo reconstruction affected the pre-implantation development. However, after transfer in pseudopregnant mice, cloned mice were obtained from both methods.

  13. Active caspase-3 and ultrastructural evidence of apoptosis in spontaneous and induced cell death in bovine in vitro produced pre-implantation embryos

    DEFF Research Database (Denmark)

    Gjørret, Jakob O.; Fabian, Dusan; Avery, Birthe;

    2007-01-01

    In this study we investigated chronological onset and involvement of active caspase-3, apoptotic nuclear morphology, and TUNEL-labeling, as well as ultrastructural evidence of apoptosis, in both spontaneous and induced cell death during pre-implantation development of bovine in vitro produced...... staining for detection of apoptotic nuclear morphology, and subjected to fluorescence microscopy. Additionally, treated and untreated blastocysts were fixed and processed for ultrastructural identification of apoptosis. Untreated embryos revealed no apoptotic features at 2- and 4-cell stages. However......, active caspase-3 and apoptotic nuclear morphology were observed in an untreated 8-cell stage, and TUNEL-labeling was observed from the 16-cell stage. Blastomeres concurrently displaying all apoptotic features were present in a few embryos at 16-cell and morula stages and in all blastocysts. All three...

  14. Improved preimplantation development of bovine ICSI embryos generated with spermatozoa pretreated with membrane-destabilizing agents lysolecithin and Triton X-100.

    Science.gov (United States)

    Zambrano, Fabiola; Aguila, Luis; Arias, María E; Sánchez, Raúl; Felmer, Ricardo

    2016-10-01

    In cattle, intracytoplasmic sperm injection (ICSI) has a low efficiency. The acrosome content may be responsible for this effect because of the large amount of hydrolytic enzymes that are released within the oocyte. With the aim of removing the acrosome and destabilize the membranes, cryopreserved bovine spermatozoa were treated with lysolecithin (LL) and Triton X-100 (TX) at different concentrations. We evaluated the membrane integrity, the acrosome integrity, DNA integrity, and the variation of phospholipase C zeta. The rates of development (cleavage and blastocysts) were also evaluated along with pronuclear formation and the embryo quality. Spermatozoa incubated with LL and TX (0.01%, 0.02%, 0.03%, and 0.04%) decreased (P embryonic development, without affecting the quality of the embryos produced by this technique. PMID:27325573

  15. Trichostatin A specifically improves the aberrant expression of transcription factor genes in embryos produced by somatic cell nuclear transfer

    OpenAIRE

    Kimiko Inoue; Mami Oikawa; Satoshi Kamimura; Narumi Ogonuki; Toshinobu Nakamura; Toru Nakano; Kuniya Abe; Atsuo Ogura

    2015-01-01

    Although mammalian cloning by somatic cell nuclear transfer (SCNT) has been established in various species, the low developmental efficiency has hampered its practical applications. Treatment of SCNT-derived embryos with histone deacetylase (HDAC) inhibitors can improve their development, but the underlying mechanism is still unclear. To address this question, we analysed gene expression profiles of SCNT-derived 2-cell mouse embryos treated with trichostatin A (TSA), a potent HDAC inhibitor t...

  16. Heterotopic triplet pregnancy: Report of a patient with remnant tubal ectopic and intrauterine twin pregnancy after frozen-thawed embryo transfer

    OpenAIRE

    Okamura, Yoshinori; Arakane, Futoshi; Nagayoshi-Taura, Yumiko; Honda, Ritsuo; Ohba, Takashi; Katabuchi, Hidetaka; オカムラ, ヨシノリ; アラカネ, フトシ; ナガヨシ-タウラ, ユミコ; ホンダ, リツオ; オオバ, タカシ; カタブチ, ヒデタカ; 岡村, 佳則; 荒金, 太; 永吉(田浦), 裕三子

    2011-01-01

    A case of heterotopic triplet pregnancy after frozen-thawed embryo transfer is presented. The patient conceived after transfer of three frozen-thawed embryos at a fertility clinic where she had previously undergone laparoscopic left salpingectomy due to pyosalpinx. Approximately 4 weeks after the embryo transfer, she presented with a complaint of abnormal genital bleeding and was diagnosed as having a dichorionic twin pregnancy by ultrasound. One week later, she was referred to our hospital b...

  17. IVF outcomes in average- and poor-prognosis infertile women according to the number of embryos transferred.

    Science.gov (United States)

    Vega, Mario G; Gleicher, Norbert; Darmon, Sarah K; Weghofer, Andrea; Wu, Yan-Guang; Wang, Qi; Zhang, Lin; Albertini, David F; Barad, David H; Kushnir, Vitaly A

    2016-09-01

    Outcome measures of IVF success, which account for effectiveness of IVF and perinatal outcome risks, have recently been described. The association between number of embryos transferred in average and poor-prognosis IVF patients, and the chances of having good or poor IVF and perinatal outcomes, was investigated. Good IVF and perinatal outcome was defined as the birth of a live, term, normal-weight infant (≥2500 g). Poor IVF and perinatal outcome was defined as no live birth or birth of a very low weight neonate (<1500 g) or severe prematurity (birth at <32 weeks gestation). Each neonate was analysed as a separate outcome. A total of 713 IVF cycles in 504 average and poor-prognosis patients from January 2010 to December 2013 were identified. The odds of having good IVF and perinatal outcomes increased by 28% for each additional embryo transferred. The odds of poor IVF and perinatal outcome decreased by 32% with an additional embryo transferred. The likelihood of live birth with good perinatal outcome in average- and poor-prognosis patients after IVF increases with additional embryos being transferred. These data add to recently reported evidence in favour of multiple embryo transfer in older women and those with average or poor IVF prognosis.

  18. Health and Safety Advisory Committee (HASAC) of the International Embryo Transfer Society (IETS) has managed critical challenges for two decades.

    Science.gov (United States)

    Thibier, M; Stringfellow, D A

    2003-02-01

    The International Embryo Transfer Society (IETS) was founded in 1974. Early members used the society as a forum for the exchange of scientific and technical information relevant to a newly emerging embryo transfer industry. The impact that embryo transfer could have on the international trade of livestock genetics was clear by 1982, so the IETS commissioned the Import/Export Committee. The initial challenge for this Committee was to deal with concerns about disease transmission via embryo transfer. Many of the early concerns have been dispelled, but at the time they threatened the continued development of a fledgling industry. Over the past two decades, many new critical challenges have been met and managed by this Committee, which was recently renamed the Health and Safety Advisory Committee (HASAC). Assessing risks of animal disease transmission via reproductive technologies and establishing protocols for managing these risks are still major issues for HASAC. However, additional concerns have developed as views of the society changed and as novel applications of biotechnology in farm animals were identified. This paper is intended to chronicle some of the major changes and challenges that were managed by members of the HASAC and its Subcommittees from the early years of embryo transfer to the current millennium with technological advances in molecular biology.

  19. Aberrant Expression of Xist in Aborted Porcine Fetuses Derived from Somatic Cell Nuclear Transfer Embryos

    Directory of Open Access Journals (Sweden)

    Lin Yuan

    2014-11-01

    Full Text Available Cloned pigs generated by somatic cell nuclear transfer (SCNT show a greater ratio of early abortion during mid-gestation than normal controls. X-linked genes have been demonstrated to be important for the development of cloned embryos. To determine the relationship between the expression of X-linked genes and abortion of cloned porcine fetuses, the expression of X-linked genes were investigated by quantitative real-time polymerase chain reaction (q-PCR and the methylation status of Xist DMR was performed by bisulfate-specific PCR (BSP. q-PCR analysis indicated that there was aberrant expression of X-linked genes, especially the upregulated expression of Xist in both female and male aborted fetuses compared to control fetuses. Results of BSP suggested that hypomethylation of Xist occurred in aborted fetuses, whether male or female. These results suggest that the abnormal expression of Xist may be associated with the abortion of fetuses derived from somatic cell nuclear transfer embryos.

  20. Success of frozen embryo transfer: Does the type of gonadotropin influence the outcome?

    Directory of Open Access Journals (Sweden)

    Hesham Al-Inany

    2010-05-01

    Full Text Available Hesham Al-Inany1, Pieter van Gelder21Egyptian IVF-ET Center, Maadi, Egypt; 2PSCT BV, Den Haag, The NetherlandsObjectives: To determine whether there is a difference in outcome between different ovulationinduced cycles after frozen-thawed embryo transfer (FET.Methods: We searched the Cochrane Menstrual Disorders and Subfertility Group’s trials register in May 2009, the Cochrane Central Register of Controlled Trials (Cochrane Library, Issue 1, 2008, ISI Web of Knowledge (1985 to August 2009, and reference lists of articles. Relevant conference proceedings were hand-searched and researchers in the field were contacted. Randomized controlled trials and retrospective studies were included, comparing the various cycle regimens and different methods during FET in assisted reproductive technology, ie, in vitro fertilization and intracytoplasmic sperm injection.Results: Using the agonist long protocol for downregulation, five trials provided extractable data for live-birth rates, ongoing pregnancy, and clinical pregnancy rates following FET. One trial provided extractable data for clinical pregnancy rate. There was no evidence of a significant difference in any outcome between the users of urinary gonadotropins versus recombinant follicle-stimulating hormone. Data on implantation and miscarriage rates following FET were not available for analysis.Conclusions: It seems that clinical pregnancy rate after FET is not influenced by the type of gonadotropins used. Research should be directed towards improving freezing and thawing techniques.Keywords: infertility, assisted reproductive technology, frozen embryo transfer, gonadotropins

  1. Generating Transgenic Mice by Lentiviral Transduction of Spermatozoa Followed by In Vitro Fertilization and Embryo Transfer.

    Science.gov (United States)

    Chandrashekran, Anil; Casimir, Colin; Dibb, Nick; Readhead, Carol; Winston, Robert

    2016-01-01

    Most transgenic technologies rely on the oocyte as a substrate for genetic modification. Transgenics animals are usually generated by the injection of the gene constructs (including lentiviruses encoding gene constructs or modified embryonic stem cells) into the pronucleus of a fertilized egg followed by the transfer of the injected embryos into the uterus of a foster mother. Male germ cells also have potential as templates for transgenic development. We have previously shown that mature sperm can be utilized as template for lentiviral transduction and as such used to generate transgenic mice efficiently with germ line capabilities. We provide here a detailed protocol that is relatively simple, to establish transgenic mice using lentivirally transduced spermatozoa. This protocol employs a well-established lentiviral gene delivery system (usual for somatic cells) delivering a variety of transgenes to be directly used with sperm, and the subsequent use of these modified sperm in in vitro fertilization studies and embryo transfer into foster female mice, for the establishment of transgenic mice. PMID:27317176

  2. Endometrial thickness, Caucasian ethnicity, and age predict clinical pregnancy following fresh blastocyst embryo transfer: a retrospective cohort

    Directory of Open Access Journals (Sweden)

    Santoro Nanette

    2009-04-01

    Full Text Available Abstract Background In-vitro fertilization (IVF with blastocyst as opposed to cleavage stage embryos has been advocated to improve success rates. Limited information exists on which to predict which patients undergoing blastocyst embryo transfer (BET will achieve pregnancy. This study's objective was to evaluate the predictive value of patient and cycle characteristics for clinical pregnancy following fresh BET. Methods This was a retrospective cohort study from 2003–2007 at an academic assisted reproductive program. 114 women with infertility underwent fresh IVF with embryo transfer. We studied patients undergoing transfer of embryos at the blastocyst stage of development. Our main outcome of interest was clinical pregnancy. Clinical pregnancy and its associations with patient characteristics (age, body mass index, FSH, ethnicity and cycle parameters (thickness of endometrial stripe, number eggs, available cleaving embryos, number blastocysts available, transferred, and cryopreserved, and embryo quality were examined using Student's T test and Mann-Whitney-U tests as appropriate. Multivariable logistic regression models were created to determine independent predictors of CP following BET. Receiver Operating Characteristic analyses were used to determine the optimal thickness of endometrial stripe for predicting clinical pregnancy. Results Patients achieving clinical pregnancy demonstrated a thicker endometrial stripe and were younger preceding embryo transfer. On multivariable logistic regression analyses, Caucasian ethnicity (OR 2.641, 95% CI 1.054–6.617, thickness of endometrial stripe, (OR 1.185, 95% CI 1.006–1.396 and age (OR 0.879, 95% CI 0.789–0.980 predicted clinical pregnancy. By receiver operating characteristic analysis, endometrial stripe ≥ 9.4 mm demonstrated a sensitivity of 83% for predicting clinical pregnancy following BET. Conclusion In a cohort of patients undergoing fresh BET, thicker endometrial stripe, Caucasian

  3. Pregnancy, Delivery, and Neonatal Outcomes of In Vitro Fertilization-Embryo Transfer in Patient with Previous Cesarean Scar.

    Science.gov (United States)

    Zhang, Ningyuan; Chen, Hua; Xu, Zhipeng; Wang, Bin; Sun, Haixiang; Hu, Yali

    2016-01-01

    BACKGROUND What role should previous cesarean section play in affecting clinical pregnancy outcomes and avoiding the complications of in vitro fertilization? In this article, we focus on elective single-embryo transfer (eSET) versus double-embryo transfer (DET) and assess the clinical efficacy and safety of eSET in patients who have a previous cesarean scar. MATERIAL AND METHODS The pregnancy, delivery, and neonatal outcomes of 130 patients who had a previous cesarean scar and received in vitro fertilization-embryo transfer (IVF-ET) were retrospectively analyzed. The number of transferred embryos was chosen depending on patients' desire after acknowledging all benefits and risks, including eSET (eSET group, n=56) and DET (DET group, n=74). A total of 101 patients with previous vaginal delivery receiving IVF-ET in the same period were included as a control group. RESULTS The pregnancy rates, multiple birth rates, abortion rates, ectopic pregnancy rates, gestational age at delivery, preterm birth rates, neonatal birth weight, and take-home baby rates were similar between the previous cesarean section group and the previous vaginal delivery group. A previous cesarean section scar did not affect embryo implantation and pregnancy outcomes in IVF. In the eSET and DET groups of previous cesarean section patients, the embryo implantation rates, pregnancy rates, abortion rates, and take-home baby rates were similar. However, the rate of multiple pregnancies reached 50% in the DET group, which led to more preterm births and lower birth weight. CONCLUSIONS Elective single-embryo transfer is a well-accepted strategy to avoid multiple pregnancies and improve the obstetric and neonatal outcomes of singleton pregnancy in IVF patients with a previous cesarean section. PMID:27636504

  4. Horizontal gene transfers link a human MRSA pathogen to contagious bovine mastitis bacteria.

    Directory of Open Access Journals (Sweden)

    Thomas Brody

    Full Text Available BACKGROUND: Acquisition of virulence factors and antibiotic resistance by many clinically important bacteria can be traced to horizontal gene transfer (HGT between related or evolutionarily distant microflora. Comparative genomic analysis has become an important tool for identifying HGT DNA in emerging pathogens. We have adapted the multi-genome alignment tool EvoPrinter to facilitate discovery of HGT DNA sequences within bacterial genomes and within their mobile genetic elements. PRINCIPAL FINDINGS: EvoPrinter analysis of 13 different Staphylococcus aureus genomes revealed that one of the human isolates, the hospital epidemic methicillin-resistant MRSA252 strain, uniquely shares multiple putative HGT DNA sequences with different causative agents of bovine mastitis that are not found in the other human S. aureus isolates. MRSA252 shares over 14 different DNA sequence blocks with the bovine mastitis ET3 S. aureus strain RF122, and many of the HGT DNAs encode virulence factors. EvoPrinter analysis of the MRSA252 chromosome also uncovered virulence-factor encoding HGT events with the genome of Listeria monocytogenes and a Staphylococcus saprophyticus associated plasmid. Both bacteria are also causal agents of contagious bovine mastitis. CONCLUSIONS: EvoPrinter analysis reveals that the human MRSA252 strain uniquely shares multiple DNA sequence blocks with different causative agents of bovine mastitis, suggesting that HGT events may be occurring between these pathogens. These findings have important implications with regard to animal husbandry practices that inadvertently enhance the contact of human and livestock bacterial pathogens.

  5. Uterine artery blood flow in the periimplantation period in embryo transfer cycles

    Institute of Scientific and Technical Information of China (English)

    Ursula Zollner; Marie-Theres Specketer; Klaus-Peter Zollner; Johannse Dietl

    2012-01-01

    Objective:To assess the role of the uterine artery blood flow in the prediction of implantation in women undergoing embryo transfer during the periimplantation period.Methods:A total of 233 couples were included in this prospective study.All patients had embryo transfer,125 were performed inin-vitro fertilization/intracytoplasmic sperm injection(IVF/ICSI) and108 in cryo cycles.Ultrasound measurements were performed immediately before transfer.The pulsatility index(PI),Resistance index(RI) and the peak systolic velocity(PSV) were measured in both uterine arteries using endovaginal ultrasound.Results:InIVF/ICSI cycles the doppler parameters PI(2.48vs.2.15),RI(0.78vs.1.30) andPSV(60 vs.63) did not differ significantly between the pregnant and non-pregnant group.The pregnancy rate per transfer was similar in women showing an unilateral(24%), bilateral(33%) or no(27%) notch in the uterine blood flow.In cryo cycles the uterine artery blood flow parametersPI(3.2vs.3.0),RI(0.9vs.0.9) andPSV(53.2vs.51.2) did not differ either between pregnant and not pregnant patients.Conclusions:Previous studies were aiming at the measurement of arterial doppler parameters during the follicular phase which may not be adequate for the prediction of implantation.However, our results show that doppler studies during the early luteal phase of assisted reproductive technology cycles are not indicative for the likelihood of pregnancy, too.

  6. A comparison of implantation, miscarriage and pregnancy rates of single and double day 3 embryo transfer between fresh and frozen thawed transfer cycles: a retrospective study

    Institute of Scientific and Technical Information of China (English)

    Liu Liu; Tong Xiaomei; Jiang Lingying; Tinchiu Li; Zhou Feng; Zhang Songying

    2014-01-01

    Background Reduced endometrial receptivity in hyperstimulated cycles may lead to a lower implantation rate and a lower clinical pregnancy rate,but it is unclear if it is also associated with an increase in pregnancy loss rate.The aim of this study was to compare the implantation,miscarriage,and pregnancy rates between fresh and frozen thawed transfer of one or two day-3 embryos,with a view to understanding whether or not reduced endometrial receptivity encountered in hyperstimulated cycles is associated with an increase in miscarriage rate.Methods This study involved a consecutive series of 1 551 single day-3 embryo transfer cycles and consecutive 5 919 double day-3 embryo transfer cycles in the Assisted Reproductive Unit of the Sir Run Run Shaw Hospital,Hangzhou,China,between January 2010 and December 2012.Results The implantation and clinical pregnancy rates (single embryo 30.7% and double embryos 33.4% and 51.4%)using fresh cycle were both significantly lower than that of frozen-thawed cycles (single embryo 35.8% and double embryos 38.1% and 57.8%).There was no difference in biochemical loss or clinical miscarriage rates between the two groups.Conclusions Impairment of endometrial receptivity associated with ovarian hyperstimulation leads to implantation failure at a very early stage,resulting in an increased number of non-pregnancy.It does not lead to increase in biochemical or clinical losses.The significantly reduced ongoing pregnancy rates in both fresh single and double embryo transfer are therefore due to failure to achieve a pregnancy,rather than pregnancy loss after conception.

  7. Difficult embryo transfers or blood on catheter and assisted reproductive outcomes: a systematic review and meta-analysis.

    Science.gov (United States)

    Phillips, James A S; Martins, Wellington P; Nastri, Carolina O; Raine-Fenning, Nicholas J

    2013-06-01

    We performed a systematic review and meta-analysis to examine whether a difficult embryo transfer or the presence of blood on the transfer catheter affects assisted reproduction outcomes. We searched the following databases: Cochrane Central Register of Controlled Trials (CENTRAL), Database of Abstracts of Reviews of Effects (DARE), MEDLINE, Embase, PsycINFO, Cumulative Index to Nursing and Allied Health Literature (CINAHL), and Literatura Latino-Americana e do Caribe em Ciências da Saúde (LILACS). We aimed to determine the risk ratio (RR) associated with difficult embryo transfer or the presence of blood on the transfer catheter for the following outcomes: live birth, clinical pregnancy, and miscarriage. We identified 3066 papers, of which 194 were reviewed and nine were included. The outcome of live birth was not reported in any of the included studies and the effect on miscarriage was too imprecise for any conclusions. Pooled analysis of five studies demonstrated lower clinical pregnancy rates following a non-easy embryo transfer (RR=0.75; 95% CI=0.66-0.86). This included three studies showing subjectively difficult transfers reducing clinical pregnancies (RR=0.67; 95% CI=0.51-0.87) and two studies in which the need for additional manoeuvers reduced clinical pregnancies (RR=0.78; 95% CI=0.67-0.91). The presence of blood on the transfer catheter did not affect clinical pregnancy rates (RR=0.96; 95% CI=0.82-1.14) in five studies. We concluded that low quality evidence suggests that a difficult embryo transfer but not a bloody catheter reduces the chance of achieving a clinical pregnancy. More good quality studies are needed to evaluate the effect of difficult embryo transfer and the presence of blood on the catheter on the main outcomes of assisted reproduction.

  8. Use of sexed sorted semen for fixed-time artificial insemination or fixed-time embryo transfer of in vitro-produced embryos in cattle.

    Science.gov (United States)

    Pellegrino, C A G; Morotti, F; Untura, R M; Pontes, J H F; Pellegrino, M F O; Campolina, J P; Seneda, M M; Barbosa, F A; Henry, M

    2016-08-01

    Artificial insemination and in vitro embryo production are powerful tools for disseminating superior genetic qualities and improving the reproductive performance of dairy and beef cattle. In conjunction with these biotechnologies, sexed-sorted semen has been used to obtain offspring of a predetermined sex. This study compared the pregnancy rates obtained using in vitro fertilization/timed embryo transfer (IVF/TET) and timed artificial insemination (TAI), both performed using sexed-sorted (Y-chromosome-bearing) semen obtained from the same bull. For the in vitro embryo production, the ovaries of 250 Nelore cows with known histories were collected in the slaughterhouse and used for IVF. After evaluation of the recipients (IVF/TET group; n = 974), the resultant embryos were transferred to the females with corpus luteum (n = 822). The pregnancy-related data for this group were compared with those for the TAI group (n = 974). Ultrasonography was performed at 60 days to determine the pregnancy status and confirm the sex of the fetus. A total of 2008 oocytes produced 1050 embryos, with 52% of them reaching the blastocyst stage. The pregnancy rate and the accuracy in determining the fetal sex were 35.4% (345/974) and 95.07% (328/345), respectively, for the IVF/TET group and 30% (293/974; P < 0.05) and 94.88% (278/293), respectively, for the TAI group. In the present study, we concluded that male calves could be better obtained using IVF/TET rather than TAI; therefore, this strategy can be considered to increase the pregnancy rate of beef cattle. PMID:27068357

  9. Effects of Two Types of Melatonin-Loaded Nanocapsules with Distinct Supramolecular Structures: Polymeric (NC) and Lipid-Core Nanocapsules (LNC) on Bovine Embryo Culture Model

    Science.gov (United States)

    Komninou, Eliza Rossi; Remião, Mariana Härter; Lucas, Caroline Gomes; Domingues, William Borges; Basso, Andrea Cristina; Jornada, Denise Soledade; Deschamps, João Carlos; Beck, Ruy Carlos Ruver; Pohlmann, Adriana Raffin; Bordignon, Vilceu; Seixas, Fabiana Kömmling; Campos, Vinicius Farias; Guterres, Silvia Stanisçuaski; Collares, Tiago

    2016-01-01

    Melatonin has been used as a supplement in culture medium to improve the efficiency of in vitro produced mammalian embryos. Through its ability to scavenge toxic oxygen derivatives and regulate cellular mRNA levels for antioxidant enzymes, this molecule has been shown to play a protective role against damage by free radicals, to which in vitro cultured embryos are exposed during early development. In vivo and in vitro studies have been performed showing that the use of nanocapsules as active substances carriers increases stability, bioavailability and biodistribution of drugs, such as melatonin, to the cells and tissues, improving their antioxidant properties. These properties can be modulated through the manipulation of formula composition, especially in relation to the supramolecular structures of the nanocapsule core and the surface area that greatly influences drug release mechanisms in biological environments. This study aimed to evaluate the effects of two types of melatonin-loaded nanocapsules with distinct supramolecular structures, polymeric (NC) and lipid-core (LNC) nanocapsules, on in vitro cultured bovine embryos. Embryonic development, apoptosis, reactive oxygen species (ROS) production, and mRNA levels of genes involved in cell apoptosis, ROS and cell pluripotency were evaluated after supplementation of culture medium with non-encapsulated melatonin (Mel), melatonin-loaded polymeric nanocapsules (Mel-NC) and melatonin-loaded lipid-core nanocapsules (Mel-LNC) at 10−6, 10−9, and 10−12 M drug concentrations. The highest hatching rate was observed in embryos treated with 10−9 M Mel-LNC. When compared to Mel and Mel-NC treatments at the same concentration (10−9 M), Mel-LNC increased embryo cell number, decreased cell apoptosis and ROS levels, down-regulated mRNA levels of BAX, CASP3, and SHC1 genes, and up-regulated mRNA levels of CAT and SOD2 genes. These findings indicate that nanoencapsulation with LNC increases the protective effects of

  10. In vitro manipulation techniques of porcine embryos: a meta-analysis related to transfers, pregnancies and piglets.

    Science.gov (United States)

    Liu, Ying; Li, Juan; Løvendahl, Peter; Schmidt, Mette; Larsen, Knud; Callesen, Henrik

    2015-03-01

    During the last 17 years, considerable advancements have been achieved in the production of pigs, transgenic and non-transgenic, by methods of somatic cell nuclear transfer, in vitro fertilisation, intracytoplasmic sperm injection, microinjection and sperm-mediated gene transfer by artificial insemination. Therefore, a review of the overall efficiency for the developmental competence of embryos produced by these in vitro methods would be useful in order to obtain a more thorough overview of this growing area with respect to its development and present status. In this review a meta-analysis was used to analyse data collected from all published articles with a focus on zygotes and embryos for transfer, pregnancy, full-term development and piglets born. It was generally concluded that an increasing level of in vitro manipulation of porcine embryos decreased the overall efficiency for production of piglets. The techniques of nuclear transfer have been developed markedly through the increasing number of studies performed, and the results have become more stable. Prolonged in vitro culture period did not lead to any negative effect on nuclear transfer embryos after their transfer and it resulted in a similar or even higher litter size. More complete information is needed in future scientific articles about these in vitro manipulation techniques to establish a more solid basis for the evaluation of their status and to reveal and further investigate any eventual problems. PMID:25482653

  11. In vitro manipulation techniques of porcine embryos: a meta-analysis related to transfers, pregnancies and piglets.

    Science.gov (United States)

    Liu, Ying; Li, Juan; Løvendahl, Peter; Schmidt, Mette; Larsen, Knud; Callesen, Henrik

    2015-03-01

    During the last 17 years, considerable advancements have been achieved in the production of pigs, transgenic and non-transgenic, by methods of somatic cell nuclear transfer, in vitro fertilisation, intracytoplasmic sperm injection, microinjection and sperm-mediated gene transfer by artificial insemination. Therefore, a review of the overall efficiency for the developmental competence of embryos produced by these in vitro methods would be useful in order to obtain a more thorough overview of this growing area with respect to its development and present status. In this review a meta-analysis was used to analyse data collected from all published articles with a focus on zygotes and embryos for transfer, pregnancy, full-term development and piglets born. It was generally concluded that an increasing level of in vitro manipulation of porcine embryos decreased the overall efficiency for production of piglets. The techniques of nuclear transfer have been developed markedly through the increasing number of studies performed, and the results have become more stable. Prolonged in vitro culture period did not lead to any negative effect on nuclear transfer embryos after their transfer and it resulted in a similar or even higher litter size. More complete information is needed in future scientific articles about these in vitro manipulation techniques to establish a more solid basis for the evaluation of their status and to reveal and further investigate any eventual problems.

  12. Current Status of Comprehensive Chromosome Screening for Elective Single-Embryo Transfer

    Directory of Open Access Journals (Sweden)

    Ming-Yih Wu

    2014-01-01

    Full Text Available Most in vitro fertilization (IVF experts and infertility patients agree that the most ideal assisted reproductive technology (ART outcome is to have a healthy, full-term singleton born. To this end, the most reliable policy is the single-embryo transfer (SET. However, unsatisfactory results in IVF may result from plenty of factors, in which aneuploidy associated with advanced maternal age is a major hurdle. Throughout the past few years, we have got a big leap in advancement of the genetic screening of embryos on aneuploidy, translocation, or mutations. This facilitates a higher success rate in IVF accompanied by the policy of elective SET (eSET. As the cost is lowering while the scale of genome characterization continues to be up over the recent years, the contemporary technologies on trophectoderm biopsy and freezing-thaw, comprehensive chromosome screening (CCS with eSET appear to be getting more and more popular for modern IVF centers. Furthermore, evidence has showen that, by these avant-garde techniques (trophectoderm biopsy, vitrification, and CCS, older infertile women with the help of eSET may have an opportunity to increase the success of their live birth rates approaching those reported in younger infertility patients.

  13. Single-embryo transfer reduces clinical pregnancy rates and live births in fresh IVF and Intracytoplasmic Sperm Injection (ICSI cycles: a meta-analysis

    Directory of Open Access Journals (Sweden)

    Oliveira João

    2009-04-01

    Full Text Available Abstract Background It has become an accepted procedure to transfer more than one embryo to the patient to achieve acceptable ongoing pregnancy rates. However, transfers of more than a single embryo increase the probability of establishing a multiple gestation. Single-embryo transfer can minimize twin pregnancies but may also lower live birth rates. This meta-analysis aimed to compare current data on single-embryo versus double-embryo transfer in fresh IVF/ICSI cycles with respect to implantation, ongoing pregnancy and live birth rates. Methods Search strategies included on-line surveys of databases from 1995 to 2008. Data management and analysis were conducted using the Stats Direct statistical software. The fixed-effect model was used for odds ratio (OR. Fixed-effect effectiveness was evaluated by the Mantel Haenszel method. Seven trials fulfilled the inclusion criteria. Results When pooling results under the fixed-effect model, the implantation rate was not significantly different between double-embryo transfer (34.5% and single-embryo transfer group (34.7% (P = 0.96; OR = 0.99, 95% CI 0.78, 1.25. On the other hand, double-embryo transfer produced a statistically significantly higher ongoing clinical pregnancy rate (44.5% than single-embryo transfer (28.3% (P Conclusion Meta-analysis with 95% confidence showed that, despite similar implantation rates, fresh double-embryo transfer had a 1.64 to 2.60 times greater ongoing pregnancy rate and 1.44 to 2.42 times greater live birth rate than single-embryo transfer in a population suitable for ART treatment.

  14. Pregnancy rates, prenatal and postnatal survival of offspring, and litter sizes after reciprocal embryo transfer in DBA/2JHd, C3H/HeNCrl and NMRI mice.

    Science.gov (United States)

    Rose, C; Schwegler, H; Hanke, J; Yilmazer-Hanke, D M

    2012-06-01

    Success of embryo transfer is often a limiting factor in transgenic procedures and rederivation efforts, and depends on the genetic background of the donor and recipient strains used. Here we show that embryo transfer to DBA/2J females is possible, and present data on pre- and postnatal success rates after reciprocal embryo transfer using the inbred DBA/2J and C3H/HeN, and outbred NMRI strains. The highest embryo yield was achieved in outbred NMRI females, but embryo yields were similar in DBA/2J and C3H/HeN mice following superovulation despite poor estrus cycle synchronization in DBA/2J females. In-strain transfer of DBA/2J blastocysts (transfer of embryos to recipients from the same strain) resulted in pregnancy rates (57.1%) similar to those obtained following in-strain transfer of C3H/HeN (60.0%) and NMRI mice (83.3%), although the prenatal survival rate of blastocysts was low. Moreover, from the pups born only half survived the postnatal period after transfer of DBA/2J and C3H/HeN blastocysts to DBA/2J recipients. These problems were not observed when transferring NMRI-blastocysts to C3H/HeN and DBA/2J mothers. The number of blastocysts transferred also had a positive effect on the success of embryo transfer. In conclusion, C3H/HeN and DBA/2J females can be used as recipients for embryo transfer procedures for certain donor strains like NMRI, as one major determinant seems to be the genetic background of the embryos transferred. We also recommend to increase the number of DBA/2J blastocysts transferred, and to foster the DBA/2J pups to other DBA/2J mothers postnatally for in-strain transfer of DBA/2J mice. PMID:22401828

  15. Preferences of subfertile women regarding elective single embryo transfer : additional in vitro fertilization cycles are acceptable, lower pregnancy rates are not

    NARCIS (Netherlands)

    Twisk, Moniek; van der Veen, Fulco; Repping, Sjoerd; Heineman, Maas-Jan; Korevaar, Johanna C.; Bossuyt, Patrick M. M.

    2007-01-01

    With identical pregnancy rates after elective single embryo transfer (ET) and double ET strategies consisting of three cycles of IVF or intracytoplasmic sperm injection (ICSI) plus transfers of thawed/frozen embryos if available, 46% of the women undergoing IVF/ICSI favor elective single ET. If elec

  16. Evaluation of an effective multifaceted implementation strategy for elective single-embryo transfer after in vitro fertilization

    NARCIS (Netherlands)

    Kreuwel, I.A.M.; Peperstraten, A.M. van; Hulscher, M.E.J.L.; Kremer, J.A.M.; Grol, R.P.T.M.; Nelen, W.L.D.M.; Hermens, R.P.M.G.

    2013-01-01

    STUDY QUESTION: What is the relationship between the rate of elective single-embryo transfer (eSET) and couples' exposure to different elements of a multifaceted implementation strategy? SUMMARY ANSWER: Additional elements in a multifaceted implementation strategy do not result in an increased eSET

  17. An ideal oocyte activation protocol and embryo culture conditions for somatic cell nuclear transfer using sheep oocytes.

    Science.gov (United States)

    Patel, Hiren; Chougule, Shruti; Chohan, Parul; Shah, Naval; Bhartiya, Deepa

    2014-10-01

    Pluripotent stem cells are possibly the best candidates for regenerative medicine, and somatic cell nuclear transfer (SCNT) is one of the viable options to make patient-specific embryonic stem cells. Till date efficacy of SCNT embryos is very low and requires further improvement like ideal oocyte activation and in vitro culture system. The aim of the present study was to evaluate ideal oocyte activation using different stimulation protocols and to study the effect of cumulus co-culture conditions on embryo development. Results demonstrate that between electric stimulation and chemical stimulation using calcium ionomycin and ionophore, best oocyte activation was obtained using calcium ionomycin (5 microM for 5 min) which resulted in 83% cleavage followed by 7% of early blastocyst which further increased to 15% when a cumulus bed was also introduced during embryo culture. Sequential modified Charles Rosenkrans 2 (mCR2) medium was used for embryo culture in which glucose levels were increased from 1 mM to 5 mM from Day 3 onwards. SCNT using cumulus cells as donor somatic cell, calcium ionomycin to activate the reconstructed oocyte and embryo culture on a cumulus bed in sequential mCR2 medium, resulted in the development of 6% embryos to early blastocyst stage. Such technological advances will make SCNT a viable option to make patient-specific pluripotent stem cell lines in near future.

  18. EFFECT OF BODY CONDITION AND SEASON ON THE YIELD AND QUALITY OF CATTLE EMBRYOS

    Directory of Open Access Journals (Sweden)

    Elena Kubovičová

    2013-02-01

    Full Text Available Unsatisfactory reproductive performance in dairy cows has been associated with environmental influences, such as season, chronic and acute changes in dietary intake and body composition. These factors can affect fertility especially ovarian function and yield and quality of oocytes and embryos. In our study the cow ´s body condition affected the overall embryo recovery rate (proportion of collected embryos to palpated corpora lutea. The significantly higher number of embryos was collected from cows with BCS 2.5- 2.75 (68.32 % embryo recovery rate and 3.0- 3.25 (63.30 % compared to the cows with BCS 2.0-2.25 (53.33% and 3.5-4.0 (47.87%; P0.05. On the contrary, the yield of transferable embryos was higher (P<0.05 during the autumn months (78.94% compared to spring (58.38% or summer (60.00% months. In conclusion, body condition and season may affect the yield and quality of bovine embryos. Higher embryo yield was recorded in average BCS (2.5-3.25 cows, whilst most transferable embryos were obtained in the higher BCS (3.5-4.0. Our results indicate that the best season for collection of transferable bovine embryos is autumn.

  19. Spindle formation and microtubule organization during first division in reconstructed rat embryos produced by somatic cell nuclear transfer.

    Science.gov (United States)

    Tomioka, Ikuo; Mizutani, Eiji; Yoshida, Tomoyuki; Sugawara, Atsushi; Inai, Kentaro; Sasada, Hiroshi; Sato, Eimei

    2007-08-01

    The present study was conducted to demonstrate the spindle formation and behavior of chromosomes and microtubules during first division in reconstructed rat embryos produced by somatic cell nuclear transfer (SCNT) with cumulus cell nuclei. To demonstrate the effect of oocyte aging after ovulation on the cleavage of SCNT embryos, micromanipulation was carried out 11, 15 and 18 h after injection of hCG. SCNT oocytes were activated by incubation in culture medium supplemented with 5 microM ionomycin for 5 min followed by treatment with 2 mM 6-dimethylaminopurine (6-DMAP) in mR1ECM for 2-3 h. For immunocytochemical observation, the SCNT embryos were incubated with monoclonal anti-alpha-tubulin antibody and then fluorescein isothiocyanate (FITC)-conjugated goat anti-mouse IgG. Cleavage rates were significantly higher for oocytes collected after 15 and 18 h rather than for those collected 11 h after injection of hCG (56 and 53%, respectively vs. 28%; P<0.05). Premature chromosome condensation occurred before activation of the SCNT oocytes, but adequate spindle formation was only rarely observed. The distribution of microtubules in SCNT embryos after activation was different from those of fertilized and parthenogenic oocytes, i.e., a dense microtubule organization shaped like a ring was observed. Eighteen to 20 h post-activation, most SCNT embryos were in the 2-cell stage, but no nucleoli were clearly visible, which was quite different from the fertilized oocytes. In addition, first division with and without small cellular bodies containing DNA was observed in the rat SCNT embryos in some cases. The present study suggests that reorganization of transferred nuclei in rat SCNT embryos may be inadequate in terms of formation of the mitotic assembly and nucleolar reorganization. PMID:17446658

  20. Role of Embryo Glue as a transfer medium in the outcome of fresh non-donor in-vitro fertilization cycles

    Directory of Open Access Journals (Sweden)

    Neeta Singh

    2015-01-01

    Full Text Available Background: EmbryoGlue is a hyaluronan-enriched embryo transfer (ET medium which aids in implantation of embryos, hence, improves pregnancy rates in in-vitro fertilization-ET cycles (IVF-ET. Aim: To evaluate the role of EmbryoGlue in improving implantation and pregnancy rates. Design and Setting: A prospective case-control study conducted at assisted reproductive center of a tertiary care hospital. Method: In 42 women undergoing IVF, embryos were transferred into 50 μL of EmbryoGlue for 10 min prior to transfer inside uterine cavity. In the control group (n = 42, embryos were transferred to conventional blastocyst culture medium. Statistical analysis was performed using SPSS IBM version 19.0. Results: Clinical pregnancy rate in the study group was 7% higher than the control group. The difference, however, was not statistically significant. In addition, no improvement in implantation rates was observed in the study group. However, significant difference (P = 0.04 in clinical pregnancy rate was observed with the EmbryoGlue in patients with previous IVF failure. In the study group, 50% patients (6/12 with previous IVF failure had successful implantation, but in the control group none of the patients (0/11 with previous implantation failure could achieve pregnancy. Conclusion: It is difficult to conclude a favourable role of EmbryoGlue in IVF-ET cycles with a good prognosis. However, in patients with recurrent implantation failure, it may be considered as a useful transfer medium.

  1. PENICILLIN-STREPTOMYCIN IN THE CULTURE MEDIUM DURING IN VITRO MATURATION (IVM OF BOVINE OOCYTES AFFECTS NUCLEAR MATURATION AND SUBSEQUENT EMBRYO DEVELOPMENT

    Directory of Open Access Journals (Sweden)

    SHIRAZI A

    2001-01-01

    Full Text Available Introduction: Standard concentrations of antibiotics in culture media are thought to have no detectable toxic effects on the cultured cells. However, since antibiotics are biologically active substances, the possibility that they interfere to some extent with cellular processes occurring in the cultured cells can not always be totally excluded. This study, therefore, was conducted to assess whether the presence of penicllin-streptomycin (pen-strep during in vitro maturation (IVM of bovine cumulus oocyte complexes (COCs affect nuclear and cytoplasmic maturation and subsequent embryo development. Materials and Methods: Bovine COCs were matured at 39oC in a humidified atmosphere with 5 % CO2 in air for 24 h in: 1- culture medium M 199 supplemented with 10 % FCS (Fetal calf serum, 0.05 IU/ml rhFSH (recombinant human FSH and 100 units penicillin and 100 ?g streptomycin/ ml. 2- culture medium M 199 without FCS and rhFSH in the presence of pen-strep. Cultures without antibiotics served as control. Six series of experiments, each consisted of at least 3 replicates, were performed. Results: In vitro maturation in the presence of pen-strep in culture medium supplemented with FCS and rhFSH significantly (P<0.05 increased the percentage of MII oocytes, however, when the COCs were divided, on the basis of appearance of the cumulus investment, into bright and dark groups, this effect was less obvious in both types of COCs, 76% vs 72% in bright COCs (P= 0.149 or 83% vs 80% in dark COCs (P=0.296 in treated and control groups respectively. The percentage of oocytes with type III of cortical granules (CGs distribution was not affected in the presence of pen-strep. The COCs expansion after IVM was not affected by the presence of antibiotics in culture medium. The subsequent embryo development of IVM/IVF produced ova, which were exposed to pen-strep during IVM, was significantly (P<0.05 decreased with respect to blastocyst formation by day 9. In vitro maturation in

  2. Cryopreservation and single embryo transfer%玻璃化冷冻技术与单胚胎移植

    Institute of Scientific and Technical Information of China (English)

    徐蓓; 朱桂金

    2013-01-01

    The pregnancy rates increased gradually with the development of assisted reproduction technology (ART). Nevertheless, multiple pregnancy has become the major complication beside ovarian hy-perstimulation syndrome(OHSS). Now more and more centers begin to develop single embryo transfer to decrease multiple pregnancy rate. The method for increasing the successful rates of single embryo transfer is single blastocyst transfer. To realize single blastocyst transfer,it should firstly improve the culture condition to increase blastocyst rate. Secondly, preservation of surplus blastocysts, which could ensure the cumulative pregnancy rate,needs safe and simple freezing method. However, programmed cryopreservation is liable to produce ice crystals due to the large volume, numerous and varied cells of blastocyst and blasto-coele existed. Therefore vitrification cryopreservation emerged as required in recent decades, which could significantly improve the freezing effects of blastocyst. The pregnancy rate of frozen embryo transfer (FET)by vitrification cryopreservation is similar to that in fresh cycles,which is essential for single blastocyst transfer in future.

  3. Nuclear and nuclear reprogramming during the first cell cycle in bovine nuclear transfer embryos

    DEFF Research Database (Denmark)

    Østrup, Olga; Petrovicova, Ida; Strejcek, Frantisek;

    2009-01-01

    , somatic cell nuclei introduced into enucleated oocytes displayed chromatin condensation, partial nuclear envelope breakdown, nucleolar desegregation and transcriptional quiescence already at 0.5 hpa. Somatic cell cytoplasm remained temporally attached to introduced nucleus and nucleolus was partially...

  4. Effect of Traditional Chinese Herbs Combined with Low Dose Human Menopausal Gonadotropin Applied in Frozen-thawed Embryo Transfer

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Objective: To assess embryo implantation rate (IR) and pregnancy rate (PR) in women who received Bushen Wengong Decoction (补肾温宫汤, BSWGD), a Chinese herbal formula, combined with low dose of human menopausal gonadotropin (hMG) prior to frozen-thawed embryo transfer (FET). Methods: A total of 262 subjects (674 transferred embryos) who received FET were analyzed retrospectively. In them,122 women were under 30 years old, 106 between 30-35 years and 32 over 35 years. The 85 subjects with normal ovulation were assigned to Group A, the natural menstruation cycling group, on whom no pre-transfer treatment was applied. The other 177 subjects with abnormal ovulation were assigned to Group B, and subdivided, according to the pre-transfer treatment they received, into three groups, Group B1 (50 cases) received BSWGD, Group B2 (58 cases) received hMG and Group B3 (69 cases) received both BSWGD and low dose hMG. The IR and PR of FET in the four groups were compared, and the effect of the embryo cryotime on PR of FET were compared also. Besides, the influencing factors to FET were analyzed. Results: IR and PR were significantly higher in all age sects of Group B3 than those in Group A, showing significant difference (P< 0.05). IR and PR in subjects in age sects of <30 years and > 35 years in group B3 were signifi cantly higher than those in Group B1 ( P<0.05), but no significant difference was shown in the two parameters between Group B 2 and Group B3 ( P>0.05). PR in the subjects who received embryos with cryo-time of > 200 days was significantly lower than that in those with cryo-time of < 100 days (P<0.05). Embryo cryo-time, endometrial thickness, use of BSWGD and use of hMG were of significance in FET ( P< 0.05).Conclusion: A programmed cycle of BSWGD combined with low dose of hMG could improve the embryo IR and PR of FET. Embryo cryo-time, endometrial thickness, and the use of BSWGD and hMG are of significance for FET.

  5. Efficient edition of the bovine PRNP prion gene in somatic cells and IVF embryos using the CRISPR/Cas9 system.

    Science.gov (United States)

    Bevacqua, R J; Fernandez-Martín, R; Savy, V; Canel, N G; Gismondi, M I; Kues, W A; Carlson, D F; Fahrenkrug, S C; Niemann, H; Taboga, O A; Ferraris, S; Salamone, D F

    2016-11-01

    The recently developed engineered nucleases, such as zinc-finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated nuclease (Cas) 9, provide new opportunities for gene editing in a straightforward manner. However, few reports are available regarding CRISPR application and efficiency in cattle. Here, the CRISPR/Cas9 system was used with the aim of inducing knockout and knock-in alleles of the bovine PRNP gene, responsible for mad cow disease, both in bovine fetal fibroblasts and in IVF embryos. Five single-guide RNAs were designed to target 875 bp of PRNP exon 3, and all five were codelivered with Cas9. The feasibility of inducing homologous recombination (HR) was evaluated with a reporter vector carrying EGFP flanked by 1 kbp PRNP regions (pHRegfp). For somatic cells, plasmids coding for Cas9 and for each of the five single-guide RNAs (pCMVCas9 and pSPgRNAs) were transfected under two different conditions (1X and 2X). For IVF zygotes, cytoplasmic injection was conducted with either plasmids or mRNA. For plasmid injection groups, 1 pg pCMVCas9 + 0.1 pg of each pSPgRNA (DNA2X) was used per zygote. In the case of RNA, two amounts (RNA1X and RNA2X) were compared. To assess the occurrence of HR, a group additionally cotransfected or coinjected with pHRegfp plasmid was included. Somatic cell lysates were analyzed by polymerase chain reaction and surveyor assay. In the case of embryos, the in vitro development and the genotype of blastocysts were evaluated by polymerase chain reaction and sequencing. In somatic cells, 2X transfection resulted in indels and large deletions of the targeted PRNP region. Regarding embryo injection, higher blastocyst rates were obtained for RNA injected groups (46/103 [44.6%] and 55/116 [47.4%] for RNA1X and RNA2X) than for the DNA2X group (26/140 [18.6%], P genetic modifications (29) was higher than the total number of gene-edited embryos, as

  6. Efficient edition of the bovine PRNP prion gene in somatic cells and IVF embryos using the CRISPR/Cas9 system.

    Science.gov (United States)

    Bevacqua, R J; Fernandez-Martín, R; Savy, V; Canel, N G; Gismondi, M I; Kues, W A; Carlson, D F; Fahrenkrug, S C; Niemann, H; Taboga, O A; Ferraris, S; Salamone, D F

    2016-11-01

    The recently developed engineered nucleases, such as zinc-finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated nuclease (Cas) 9, provide new opportunities for gene editing in a straightforward manner. However, few reports are available regarding CRISPR application and efficiency in cattle. Here, the CRISPR/Cas9 system was used with the aim of inducing knockout and knock-in alleles of the bovine PRNP gene, responsible for mad cow disease, both in bovine fetal fibroblasts and in IVF embryos. Five single-guide RNAs were designed to target 875 bp of PRNP exon 3, and all five were codelivered with Cas9. The feasibility of inducing homologous recombination (HR) was evaluated with a reporter vector carrying EGFP flanked by 1 kbp PRNP regions (pHRegfp). For somatic cells, plasmids coding for Cas9 and for each of the five single-guide RNAs (pCMVCas9 and pSPgRNAs) were transfected under two different conditions (1X and 2X). For IVF zygotes, cytoplasmic injection was conducted with either plasmids or mRNA. For plasmid injection groups, 1 pg pCMVCas9 + 0.1 pg of each pSPgRNA (DNA2X) was used per zygote. In the case of RNA, two amounts (RNA1X and RNA2X) were compared. To assess the occurrence of HR, a group additionally cotransfected or coinjected with pHRegfp plasmid was included. Somatic cell lysates were analyzed by polymerase chain reaction and surveyor assay. In the case of embryos, the in vitro development and the genotype of blastocysts were evaluated by polymerase chain reaction and sequencing. In somatic cells, 2X transfection resulted in indels and large deletions of the targeted PRNP region. Regarding embryo injection, higher blastocyst rates were obtained for RNA injected groups (46/103 [44.6%] and 55/116 [47.4%] for RNA1X and RNA2X) than for the DNA2X group (26/140 [18.6%], P < 0.05). In 46% (26/56) of the total sequenced blastocysts, specific gene editing was

  7. Absence of nucleolus formation in raccoon dog-porcine interspecies somatic cell nuclear transfer embryos results in embryonic developmental failure.

    Science.gov (United States)

    Jeon, Yubyeol; Nam, Yeong-Hee; Cheong, Seung-A; Kwak, Seong-Sung; Lee, Eunsong; Hyun, Sang-Hwan

    2016-08-25

    Interspecies somatic cell nuclear transfer (iSCNT) can be a solution for preservation of endangered species that have limited oocytes. It has been reported that blastocyst production by iSCNT is successful even if the genetic distances between donors and recipients are large. In particular, domestic pig oocytes can support the development of canine to porcine iSCNT embryos. Therefore, we examined whether porcine oocytes may be suitable recipient oocytes for Korean raccoon dog iSCNT. We investigated the effects of trichostatin A (TSA) treatment on iSCNT embryo developmental patterns and nucleolus formation. Enucleated porcine oocytes were fused with raccoon dog fibroblasts by electrofusion and cleavage, and blastocyst development and nucleolus formation were evaluated. To our knowledge, this study is the first in which raccoon dog iSCNT was performed using porcine oocytes; we found that 68.5% of 158 iSCNT embryos had the ability to cleave. However, these iSCNT embryos did not develop past the 4-cell stage. Treatment with TSA did not affect iSCNT embryonic development; moreover, the nuclei failed to form nucleoli at 48 and 72 h post-activation (hpa). In contrast, pig SCNT embryos of the control group showed 18.8% and 87.9% nucleolus formation at 48 and 72 hpa, respectively. Our results demonstrated that porcine cytoplasts efficiently supported the development of raccoon dog iSCNT embryos to the 4-cell stage, the stage of porcine embryonic genome activation (EGA); however, these embryos failed to reach the blastocyst stage and showed defects in nucleolus formation.

  8. Hepes na produção de embriões bovinos in vitro Hepes on in vitro production of bovine embryos

    Directory of Open Access Journals (Sweden)

    Marcelo Marcos Montagner

    2000-06-01

    of pH changes in maturation and embryo development media, buffered with different HEPES concentrations. Initially, the effect of different concentrations of HEPES (0, 12.5 and 25.0mM on the variation of pH in the maturation (modified TCM-199 and embryonic development (modified KSOM media was evaluated at room temperature (25ºC and in an atmosphere of 5% CO2 in air at 39ºC. In another experiment, the effect of HEPES on in vitro oocyte maturation was determined. Oocytes were maturated in TCM-199 modified either with 25.0mM of HEPES (HEPES group; n = 137 or without HEPES (control group; n = 142, performing 7 replicates and evaluating the rate of blastocyst. In this study, the medium used for fertilization was Fert-TALP while for embryo development was KSOM with 10% of fetal bovine serum with monolayer of oviduct epithelial cells. A third experiment was designed to determine the effect of HEPES on embryo development. The zygotes were divided in two groups and co-incubated with oviduct epithelial cells in modified KSOM with 10% of fetal bovine serum without HEPES (n = 95 or with 25.0mM of HEPES (n = 92. For this experiment, it was used embryos with two or more cells and the embryo development was considered from cleavage to expanded blastocyst (Bx, 7 and 9 days after insemination. The oocytes and embryos were incubated at temperature of 39ºC, an atmosphere containing 5% CO2 in air and saturated humidity. The media with 25.0mM of HEPES were more efficient in minimizing the range of pH than those with 12.5mM or without HEPES. To determine the effect of HEPES during in vitro oocyte maturation, the percentage of Bl considered either the total number of oocytes or the total number of cleavages was higher in the HEPES group (21.9% or 42.9%, respectively than those obtained in the control group (10.56% or 16.67%, respectively. When HEPES was added to embryo culture medium, the percentage of Bx (45.65% was higher than that obtained in medium without HEPES (11.58%; p<0.01. The

  9. Inheritance of resistance of bovine preimplantation embryos to heat shock: relative importance of the maternal versus paternal contribution.

    Science.gov (United States)

    Block, J; Chase, C C; Hansen, P J

    2002-09-01

    Brahman preimplantation embryos are less affected by exposure to heat shock than Holstein embryos. Two experiments were conducted to test whether the ability of Brahman embryos to resist the deleterious effects of heat shock was a result of the genetic and cellular contributions from the oocyte, spermatozoa, or a combination of both. In the first experiment, Brahman and Holstein oocytes were collected from slaughterhouse ovaries and fertilized with spermatozoa from an Angus bull. A different bull was used for each replicate to eliminate bull effects. On day 4 after fertilization, embryos >or= 9 cells were collected and randomly assigned to control (38.5 degrees C) or heat shock (41 degrees C for 6 hr) treatments. The proportion of embryos developing to the blastocyst (BL) and advanced blastocyst (ABL; expanded and hatched) stages was recorded on day 8. Heat shock reduced the number of embryos produced from Holstein oocytes that developed to BL (P Brahman oocytes (BL = 42.1 +/- 4.8% vs. 55.6 +/- 4.8% for 38.5 and 41 degrees C, respectively; ABL = 17.6 +/- 4.2% vs. 32.4 +/- 4.2%). In the second experiment, oocytes from Holstein cows were fertilized with semen from bulls of either Brahman or Angus breeds. Heat shock of embryos >or= 9 cells reduced development to BL (P Brahman (BL = 54.3 +/- 7.7% vs. 23.4 +/- 7.7%; ABL = 43. +/- 7.4% vs. 7.9 +/- 7.4%, for 38.5 and 41 degrees C, respectively) and Angus bulls (BL = 57.9 +/- 7.7% vs. 31.0 +/- 7.7%; ABL = 33.6 +/- 7.4% vs. 18.4 +/- 7.4%, for 38.5 and 41 degrees C, respectively). There were no breed x temperature interactions. Results suggest that the oocyte plays a more significant role in the resistance of Brahman embryos to the deleterious effects of heat shock than the spermatozoa. PMID:12211058

  10. The role of RNA polymerase I transcription and embryonic genome activation in nucleolar development in bovine preimplantation embryos

    DEFF Research Database (Denmark)

    Østrup, Olga; Strejcek, F.; Petrovicova, I.;

    2008-01-01

    The aim of the present study was to investigate the role of RNA polymerase I (RPI) transcription in nucleolar development during major transcriptional activation (MTA) in cattle. Late eight-cell embryos were cultured in the absence (control group) or presence of actinomycin D (AD) (RPI inhibition...

  11. Factors affecting survival rates of in vitro produced bovine embryos after vitrification and direct in-straw rehydration

    DEFF Research Database (Denmark)

    Vajta, Gábor; Holm, Peter; Greve, Torben;

    1996-01-01

    The aim of this work was to investigate the possibilities of simplification, and to outline the limits of application, of a vitrification method for cow embryos. Morulae and blastocysts were produced by in vitro fertilization of slaughterhouse-derived, in vitro matured oocytes with frozen-thawed ...

  12. Development of a generally applicable morphokinetic algorithm capable of predicting the implantation potential of embryos transferred on Day 3

    Science.gov (United States)

    Petersen, Bjørn Molt; Boel, Mikkel; Montag, Markus; Gardner, David K.

    2016-01-01

    STUDY QUESTION Can a generally applicable morphokinetic algorithm suitable for Day 3 transfers of time-lapse monitored embryos originating from different culture conditions and fertilization methods be developed for the purpose of supporting the embryologist's decision on which embryo to transfer back to the patient in assisted reproduction? SUMMARY ANSWER The algorithm presented here can be used independently of culture conditions and fertilization method and provides predictive power not surpassed by other published algorithms for ranking embryos according to their blastocyst formation potential. WHAT IS KNOWN ALREADY Generally applicable algorithms have so far been developed only for predicting blastocyst formation. A number of clinics have reported validated implantation prediction algorithms, which have been developed based on clinic-specific culture conditions and clinical environment. However, a generally applicable embryo evaluation algorithm based on actual implantation outcome has not yet been reported. STUDY DESIGN, SIZE, DURATION Retrospective evaluation of data extracted from a database of known implantation data (KID) originating from 3275 embryos transferred on Day 3 conducted in 24 clinics between 2009 and 2014. The data represented different culture conditions (reduced and ambient oxygen with various culture medium strategies) and fertilization methods (IVF, ICSI). The capability to predict blastocyst formation was evaluated on an independent set of morphokinetic data from 11 218 embryos which had been cultured to Day 5. PARTICIPANTS/MATERIALS, SETTING, METHODS The algorithm was developed by applying automated recursive partitioning to a large number of annotation types and derived equations, progressing to a five-fold cross-validation test of the complete data set and a validation test of different incubation conditions and fertilization methods. The results were expressed as receiver operating characteristics curves using the area under the

  13. Effects of Embryo Transfer on Emotional Behaviors in C57BL/6 Mice.

    Science.gov (United States)

    Lerch, Sandra; Tolksdorf, Gabriele; Schütz, Patrizia; Brandwein, Christiane; Dormann, Christof; Gass, Peter; Chourbaji, Sabine

    2016-01-01

    Microbiologic standardization plays a key role in the management of animal facilities because contamination of stock could affect the health status and wellbeing of animals and thereby induce artifacts in biomedical research. One common method to avoid the dissemination of pathogens is embryo transfer (ET). Although disturbances in the perinatal environment may cause long-lasting effects on the behavior and physiology of mouse offspring, the influences of ET during this sensitive phase have not yet been addressed. Our study investigated the effects of various components of ET (anesthesia, surgery, recipient strain) on the behavior of dams (exploration, nest-building) and offspring (nest-building, exploration, anxiety, and social and depressive-like behaviors). For ET, the donor strain C57BL/6N and a standard protocol were used. Whereas treatment with anesthesia-analgesia did not affect maternal behavior, female offspring demonstrated overall effects on weight gain and corticosterone levels. Compared with naturally delivered female offspring, dams obtained through ET demonstrated decreased exploration and nest-building. In addition, female ET-derived offspring had enhanced levels of anxiety and increased social interest. Furthermore, ET-derived dams obtained by using NMRI as the recipient strain showed increased exploratory behavior compared with that of dams obtained by using C57 mice as recipients. Compared with using C57 as recipients, both sexes of offspring transferred into NMRI recipients weighed more, and female mice showed a depressive-like phenotype. Our findings suggest that ET, now considered to be a routine procedure in animal husbandry, bears the risk of introducing artifacts. PMID:27657704

  14. Controlled ultraviolet resonance energy transfer between bovine serum albumin donors and cadmium sulfide quantum dots acceptors

    Science.gov (United States)

    Ghali, Mohsen; El-Kemary, Maged; Ramadan, Mahmoud

    2015-08-01

    We report on Förester resonance nergy transfer (FRET) within a bioconjugated system composed of cadmium sulfide (CdS) quantum dots (QDs) and transport protein bovine serum albumin (BSA). The optical properties of these two elements of the bioconjugate were exploited to produce FRET in the ultraviolet (UV) region with a maximum efficiency of 22% from BSA donors to QD acceptors. In contrast to previous studies, which were limited to FRET in the visible light, we used 2.6 nm CdS QDs because they emit light with a shorter wavelength (∼370 nm) that facilitates the UV-FRET process. UV-FRET was controlled by tuning the spectral overlap between BSA and CdS QDs.

  15. Transfer of auxinic herbicide resistance from Brassica kaber to Brassica juncea and Brassica rapa through embryo rescue

    OpenAIRE

    Mithila, J.; Hall, J Christopher

    2013-01-01

    Auxinic herbicides are widely used in agriculture to selectively control broadleaf weeds. Prolonged use of auxinic herbicides has resulted in the evolution of resistance to these herbicides in some biotypes of Brassica kaber (wild mustard), a common weed in agricultural crops. In this study, auxinic herbicide resistance from B. kaber was transferred to Brassica juncea and Brassica rapa, two commercially important Brassica crops, by traditional breeding coupled with in vitro embryo rescue. A h...

  16. Study of the morphology of ovary and cytology of oocyte as basis for establishing methods IVM, IVF and embryo transfer

    OpenAIRE

    Smiljaković T.; Poleksić V.; Petrović M.M.; Pejcić S.; Trenkovski S.; Stojanović Lj.; Alm H.

    2007-01-01

    In three types of domestic animals: cattle, pigs and sheep, morphology of ovary was studied. Results such as differences in shape, size within and between species, number of follicles in maturation, changes in sex cycle, connection of follicles in maturation are presented in figures. By method of aspiration, oocytes were isolated from ovaries and their cytology analyzed as the first step in methods of in vitro maturation (IVM), followed by in vitro fertilization (IVF) and embryo transfer. The...

  17. The integrated HIV-1 provirus in patient sperm chromosome and its transfer into the early embryo by fertilization.

    Directory of Open Access Journals (Sweden)

    Dian Wang

    Full Text Available Complete understanding of the route of HIV-1 transmission is an important prerequisite for curbing the HIV/AIDS pandemic. So far, the known routes of HIV-1 transmission include sexual contact, needle sharing, puncture, transfusion and mother-to-child transmission. Whether HIV can be vertically transmitted from human sperm to embryo by fertilization is largely undetermined. Direct research on embryo derived from infected human sperm and healthy human ova have been difficult because of ethical issues and problems in the collection of ova. However, the use of inter-specific in vitro fertilization (IVF between human sperm and hamster ova can avoid both of these problems. Combined with molecular, cytogenetical and immunological techniques such as the preparation of human sperm chromosomes, fluorescent in situ hybridization (FISH, and immunofluorescence assay (IFA, this study mainly explored whether any integrated HIV provirus were present in the chromosomes of infected patients' sperm, and whether that provirus could be transferred into early embryos by fertilization and maintain its function of replication and expression. Evidence showed that HIV-1 nucleic acid was present in the spermatozoa of HIV/AIDS patients, that HIV-1 provirus is present on the patient sperm chromosome, that the integrated provirus could be transferred into early embryo chromosomally integrated by fertilization, and that it could replicate alongside the embryonic genome and subsequently express its protein in the embryo. These findings indicate the possibility of vertical transmission of HIV-1 from the sperm genome to the embryonic genome by fertilization. This study also offers a platform for the research into this new mode of transmission for other viruses, especially sexually transmitted viruses.

  18. Evaluation of bovine (Bos indicus) ovarian potential for in vitro embryo production in the Adamawa plateau (Cameroon)

    OpenAIRE

    Kouamo, J.; Dawaye, S.M.; Zoli, A. P.; Bah, G. S.

    2014-01-01

    An abattoir study was conducted to evaluate the ovarian potential of 201 local zebu cattle from Ngaoundere, Adamawa region (Cameroon) for in vitro embryo production (IVEP). The ovaries were excised, submerged in normal saline solution (0.9%) and transported to the laboratory for a detailed evaluation. Follicles on each ovary were counted, their diameters (Φ) measured and were grouped into 3 categories: small (Φ 8 mm). Each ovary was then sliced in...

  19. Successful laparoscopic management of a rare complication after embryo transfer: ovarian pregnancy. A case report and up-to-date literature review.

    Science.gov (United States)

    Aydin, Turgut; Yucel, Burak; Aksoy, Huseyin; Ekemen, Suheyla

    2016-01-01

    Ovarian pregnancy (OP) after embryo transfer is very rare. Due to the rarity and the asymptomatic nature, there are still difficulties in diagnosis and treatment. The traditional operative treatment for OP has been oophorectomy. However, the desire to maintain reproductive capability and improvements in laparoscopy have more recently led to conservative laparoscopic techniques. This rare complication could be diagnosed early and managed by a conservative laparoscopic approach. Here we present a survey of the literature and a case of successful laparoscopic management of ovarian pregnancy after intracytoplasmic sperm injection and embryo transfer. The current case is the first case in the literature in which ovarian pregnancy occurred after a single embryo transfer. We also summarize the literature about management of ovarian pregnancy after embryo transfer.

  20. Superovulation of the Cloned Cattle Derived from Somatic Cells and the Transfer of the Vitrified-Thawed Embryos of the Cloning Cattle

    Institute of Scientific and Technical Information of China (English)

    DONG Ya-juan; BAI Xue-jin; LI Jian-dong; CHENG Ming

    2004-01-01

    In this experiment, it was designed to carry out superovulation on the two cloned cattles, vitrification and transfer of the embryos recovered from them. First of all, it was carried out vitrification on embryos obtained by IVF. Results showed that there were no significant differences between the blastocysts (obtained by IVF) vitrified in EPS10 and these in EPS20 on the resuscitative rate and the developmental rate. The hatched rate of the blastocysts vitrified in EPS10 (31.3%, 35/112) was significantly higher than that in EPS20 (12.2%, 13/107) (P<0.01), so EPS20 was selected as the vitrification solution to freeze the embryos recovered from the cloned cattle. After superovulation, six (four usable embryos) and ten (nine usable embryos) embryos were respectively recovered from Kangkang and Shuanghuang. Two embryos were selected from the recovered embryos of each cloned cattle to freeze in EPS20, subsequently thawed and transferred into luteal ipsilateral uterine horns of 4 Holstein recipient cows after synchronization of estrus, respectively. At last, one recipient cow (No. 9908) became pregnant and delivered one healthy calf (descendant of the cloned cattle-Shuangshuang). The results of this experiment show that the cloned cattle as well as common cattle had better response to the exotic FSH and better ability to multiovulation, the embryos recovered from the cloned cattle can be vitrificated.

  1. A biolistic process for in vitro gene transfer into chicken embryos

    Directory of Open Access Journals (Sweden)

    L.A. Ribeiro

    2001-09-01

    Full Text Available Chicken embryos kept in culture medium were bombarded using a high helium gas pressure biolistic device. To optimize the factors that affect transformation efficiency, the lacZ gene under control of the human cytomegalovirus immediate early enhancer/promoter was used as a reporter gene. There was an inverse relationship between survival rate and transformation efficiency. The best conditions obtained for high embryo survival and high transformation efficiency were achieved with 800 psi helium gas pressure, 500 mmHg vacuum, gold particles, an 8 cm DNA-coated microparticle flying distance to the embryo and embryo placement 0.5 cm from the center of the particle dispersion cone. Under these conditions, transformation efficiency was 100%, survival rate 25% and the number of expression units in the embryo body cells ranged from 100 to 1,000. Expression of green fluorescent protein was also detected in embryos bombarded under optimal conditions. Based on the results obtained, the biolistic process can be considered an efficient method for the transformation of chicken embryos and therefore can be used as a model system to study transient gene expression and tissue-specific promoters.

  2. Transfer of Dicamba Tolerance from Sinapis arvensis to Brassica napus via Embryo Rescue and Recurrent Backcross Breeding.

    Directory of Open Access Journals (Sweden)

    M Jugulam

    Full Text Available Auxinic herbicides (e.g. dicamba are extensively used in agriculture to selectively control broadleaf weeds. Although cultivated species of Brassicaceae (e.g. Canola are susceptible to auxinic herbicides, some biotypes of Sinapis arvensis (wild mustard were found dicamba resistant in Canada. In this research, dicamba tolerance from wild mustard was introgressed into canola through embryo rescue followed by conventional breeding. Intergeneric hybrids between S. arvensis (2n = 18 and B. napus (2n = 38 were produced through embryo rescue. Embryo formation and hybrid plant regeneration was achieved. Transfer of dicamba tolerance from S. arvensis into the hybrid plants was determined by molecular analysis and at the whole plant level. Dicamba tolerance was introgressed into B. napus by backcrossing for seven generations. Homozygous dicamba-tolerant B. napus lines were identified. The ploidy of the hybrid progeny was assessed by flow cytometry. Finally, introgression of the piece of DNA possibly containing the dicamba tolerance gene into B. napus was confirmed using florescence in situ hybridization (FISH. This research demonstrates for the first time stable introgression of dicamba tolerance from S. arvensis into B. napus via in vitro embryo rescue followed by repeated backcross breeding. Creation of dicamba-tolerant B. napus varieties by this approach may have potential to provide options to growers to choose a desirable herbicide-tolerant technology. Furthermore, adoption of such technology facilitates effective weed control, less tillage, and possibly minimize evolution of herbicide resistant weeds.

  3. Transfer of Dicamba Tolerance from Sinapis arvensis to Brassica napus via Embryo Rescue and Recurrent Backcross Breeding.

    Science.gov (United States)

    Jugulam, M; Ziauddin, Asma; So, Kenny K Y; Chen, Shu; Hall, J Christopher

    2015-01-01

    Auxinic herbicides (e.g. dicamba) are extensively used in agriculture to selectively control broadleaf weeds. Although cultivated species of Brassicaceae (e.g. Canola) are susceptible to auxinic herbicides, some biotypes of Sinapis arvensis (wild mustard) were found dicamba resistant in Canada. In this research, dicamba tolerance from wild mustard was introgressed into canola through embryo rescue followed by conventional breeding. Intergeneric hybrids between S. arvensis (2n = 18) and B. napus (2n = 38) were produced through embryo rescue. Embryo formation and hybrid plant regeneration was achieved. Transfer of dicamba tolerance from S. arvensis into the hybrid plants was determined by molecular analysis and at the whole plant level. Dicamba tolerance was introgressed into B. napus by backcrossing for seven generations. Homozygous dicamba-tolerant B. napus lines were identified. The ploidy of the hybrid progeny was assessed by flow cytometry. Finally, introgression of the piece of DNA possibly containing the dicamba tolerance gene into B. napus was confirmed using florescence in situ hybridization (FISH). This research demonstrates for the first time stable introgression of dicamba tolerance from S. arvensis into B. napus via in vitro embryo rescue followed by repeated backcross breeding. Creation of dicamba-tolerant B. napus varieties by this approach may have potential to provide options to growers to choose a desirable herbicide-tolerant technology. Furthermore, adoption of such technology facilitates effective weed control, less tillage, and possibly minimize evolution of herbicide resistant weeds. PMID:26536372

  4. Normal epigenetic inheritance in mice conceived by in vitro fertilization and embryo transfer

    Institute of Scientific and Technical Information of China (English)

    Lei LI; Fang LE; Li-ya WANG; Xiang-rong XU; Hang-ying LOU; Ying-ming ZHENG; Jiang-zhong SHENG; He-feng HUANG; Fan JIN

    2011-01-01

    An association between assisted reproductive technology (ART) and neurobehavioral imprinting disorders has been reported in many studies,and it seems that ART may interfere with imprint reprogramming.However,it has never been explored whether epigenetic errors or imprinting disease susceptibility induced by ART can be inherited transgenerationally.Hence,the aim of this study was to determine the effect of in vitro fertilization and embryo transfer (IVF-ET) on transgenerational inheritance in an inbred mouse model.Mice derived from IVF-ET were outcrossed to wild-type C57BL/6J to obtain their female and male line F2 and F3 generations.Their behavior,morphology,histology,and DNA methylation status at several important differentially methylated regions (DMRs) were analyzed by Morris water maze,hematoxylin and eosin (H&E) staining,and bisulfite genomic sequencing.No significant differences in spatial learning or phenotypic abnormality were found in adults derived from IVF (F1) and female and male line F2 and F3 generations.A borderline trend of hypomethylation was found in H19 DMR CpG island 3 in the female line-derived F3 generation (0.40±0.118,P=0.086).Methylation status in H19/Igf2 DMR island 1,Igf2 DMR,KvDMR,and Snrpn DMR displayed normal patterns.Methylation percentage did not differ significantly from that of adults conceived naturally,and the expression of the genes they regulated was not disturbed.Transgenerational integrity,such as behavior,morphology,histology,and DNA methylation status,was maintained in these generations,which indicates that exposure of female germ cells to hormonal stimulation and gamete manipulation might not affect the individuals and their descendents.

  5. Efecto del ácido linoleico conjugado sobre la proporción de sexos y calidad de embriones bovinos producidos in vitro Effect of conjugated linoleic acid on sex ratio and quality of in vitro produced bovine embryos

    Directory of Open Access Journals (Sweden)

    NA Gómez

    2013-01-01

    Full Text Available El objetivo de este estudio fue evaluar el efecto de la suplementación del medio de cultivo con ácido linoleico conjugado (CLA sobre el clivaje, producción, proporción de sexos y calidad embrionaria en embriones bovinos producidos in vitro al día 7 de cultivo. Se fertilizaron 308 CCO suplementados en cultivo con 100 µM del isómero de CLA Cis-9 Trans-11 y Cis-10-Trans-12 y 257 CCO en el grupo control; la producción de embriones fue 25,32% vs 35,40% respectivamente con diferencia significativa (P The aim of this study was to evaluate the effect of culture medium supplementation with conjugated linoleic acid (CLA on embryo cleavage, embryo production, sex ratio and embry o quality in in vitro produced bovine embryos at day 7 of culture. 308 COCs were used for the group supplemented with 100 µM of the CLA isomer Cis-9 trans-11 and Cis-10-Trans-12 and 257 COCs for the untreated control group; the embryo production was 25.32% vs 35.40%, respectively, with significant difference between them (P < 0.05. The embryos were classified according to the IETS in Mo, Bt, Bl and Bx stages for morphological and molecular analysis. PCR was used for sex determination; embryo quality was assessed as grade 1 (excellent or good and Grade 2 (regular. The results showed no significant difference in the proportion of embryos male:female for any of the stages in the CLA supplemented group achieving the expected natural ratio (50:50, while the control maintained a greater number of males. The CLA improved quality in Bl and Bt stages for both females and males (P < 0.05 having a greater number of grade 1 embryos in supplemented group, while control embryos were more in grade 2. In conclusion, CLA adversely affects the production of bovine embryos in vitro, but the sex ratio equals the natural one in all stages and improves embryo quality in some stages of early development.

  6. Vitrificación como técnica de crioconservación de embriones bovinos Vitrification as a technique of bovine embryo cryopreservation

    Directory of Open Access Journals (Sweden)

    M CELESTINOS

    2002-01-01

    less expensive freezing procedures. One of these is vitrification, a process of solidification that uses a highly concentrated solution which does not crystallize during freezing; its viscosity increases with the descent of the temperature until the formation of an amorphous solid state similar to glass. For this reason, exposure and freezing rates should be quick enough to avoid toxicity and the formation of intracellular ice, which can cause embryonic damage. In order for the embryos to support the osmotic shock, they should be equilibriated with a less concentrated crioprotectant solution before being exposed to the vitrificant solution for their later freezing. Several vitrification procedures have been published about embryo preservation, using different crioprotectants, concentration, volume, addition method, temperatures, exposition time, freezing rate, thawing procedure and dilution, to maintain the function, normal structure and viability of the embryo. These techniques have also been experimented with in vitro and in vivo produced embryos, at different developmental stages. This paper aims to review the present bovine embryo cryopreservation methods, particulary vitrification, as well as to mention the latest procedures and progress so that new researchers may have an updated literature review to start their works.

  7. Tiger, Bengal and Domestic Cat Embryos Produced by Homospecific and Interspecific Zona-Free Nuclear Transfer.

    Science.gov (United States)

    Moro, L N; Jarazo, J; Buemo, C; Hiriart, M I; Sestelo, A; Salamone, D F

    2015-10-01

    The aim of this study was to evaluate three different cloning strategies in the domestic cat (Felis silvestris) and to use the most efficient to generate wild felid embryos by interspecific cloning (iSCNT) using Bengal (a hybrid formed by the cross of Felis silvestris and Prionailurus bengalensis) and tiger (Panthera tigris) donor cells. In experiment 1, zona-free (ZP-free) cloning resulted in higher fusion and expanded blastocyst rates with respect to zona included cloning techniques that involved fusion or injection of the donor cell. In experiment 2, ZP-free iSCNT and embryo aggregation (2X) were assessed. Division velocity and blastocyst rates were increased by embryo aggregation in the three species. Despite fewer tiger embryos than Bengal and cat embryos reached the blastocyst stage, Tiger 2X group increased the percentage of blastocysts with respect to Tiger 1X group (3.2% vs 12.1%, respectively). Moreover, blastocyst cell number was almost duplicated in aggregated embryos with respect to non-aggregated ones within Bengal and tiger groups (278.3 ± 61.9 vs 516.8 ± 103.6 for Bengal 1X and Bengal 2X groups, respectively; 41 vs 220 ± 60 for Tiger 1X and Tiger 2X groups, respectively). OCT4 analysis also revealed that tiger blastocysts had higher proportion of OCT4-positive cells with respect to Bengal blastocysts and cat intracytoplasmic sperm injection blastocysts. In conclusion, ZP-free cloning has improved the quality of cat embryos with respect to the other cloning techniques evaluated and was successfully applied in iSCNT complemented with embryo aggregation. PMID:26332056

  8. Expression of LIF, VEGF,CD57 and CD68 after the transfer of rat embryos to mouse uteri

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    The high failure rate of interspecific pregnancy is a major obstacle to the successful interspecific cloning of mammals. To investigate the reasons for the failure of interspecfic pregnancy between rats and mice, we transferred rat blastocysts into mouse uteri on the third day of pseudopregnancy (D3). Our previous study showed that intact rat embryos could still be observed in mouse uteri on D9. In the present study, we found that expression of CD57 and CD68 increased significantly at the maternal-fetal interface following the transfer of rat embryos. Similarly, Leukaemia inhibitory factor (LIF) expression increased, but vascular endothelial growth factor (VEGF) expession decreased. In a co-culture system, the percentage of rat ectoplacental cones (EPCs) with adhesion and outgrowth and outgrowth area on mouse uterine decidual cells were less than that of mouse EPCs. These results indicate that an increase in the immunological rejection response and a decrease in the invasiveness of rat embryos may be important reasons for the failure of interspecific pregnancy between rat and mouse.

  9. Effective Oocyte Vitrification and Survival Techniques for Bovine Somatic Cell Nuclear Transfer.

    Science.gov (United States)

    Park, Min Jee; Lee, Seung Eun; Kim, Eun Young; Lee, Jun Beom; Jeong, Chang Jin; Park, Se Pill

    2015-06-01

    Bovine somatic cell nuclear transfer (SCNT) using vitrified-thawed (VT) oocytes has been studied; however, the cloning efficiency of these oocytes is not comparable with that of nonvitrified (non-V) fresh oocytes. This study sought to optimize the survival and cryopreservation of VT oocytes for SCNT. Co-culture with feeder cells that had been preincubated for 15 h significantly improved the survival of VT oocytes and their in vitro developmental potential following SCNT in comparison to co-culture with feeder cells that had been preincubated for 2, 5, or 24 h (pEVT) group, 13.7%; VT group, 15.0%; p<0.05] and was comparable with that of the non-V group (25.9%). The reactive oxygen species level was significantly lower in the EAVT group than in the other vitrification groups (p<0.05). mRNA levels of maternal genes (ZAR1, BMP15, and NLRP5) and a stress gene (HSF1) were lower in the vitrification groups than in the non-V group (p<0.05), whereas the level of phospho-p44/42 mitogen-activated protein kinase did not differ among the groups. Among the vitrification groups, blastocysts in the EAVT group had the best developmental potential, as judged by their high mRNA expression of developmental potential-related genes (POU5f1, Interferon-tau, and SLC2A5) and their low expression of proapoptotic (CASP3) and stress (Hsp70) genes. This study demonstrates that SCNT using bovine frozen-thawed oocytes can be successfully achieved using optimized vitrification and co-culture techniques. PMID:25984830

  10. SGO1 maintains bovine meiotic and mitotic centromeric cohesions of sister chromatids and directly affects embryo development.

    Directory of Open Access Journals (Sweden)

    Feng-Xia Yin

    Full Text Available Shugoshin (SGO is a critical factor that enforces cohesion from segregation of paired sister chromatids during mitosis and meiosis. It has been studied mainly in invertebrates. Knowledge of SGO(s in a mammalian system has only been reported in the mouse and Hela cells. In this study, the functions of SGO1 in bovine oocytes during meiotic maturation, early embryonic development and somatic cell mitosis were investigated. The results showed that SGO1 was expressed from germinal vesicle (GV to the metaphase II stage. SGO1 accumulated on condensed and scattered chromosomes from pre-metaphase I to metaphase II. The over-expression of SGO1 did not interfere with the process of homologous chromosome separation, although once separated they were unable to move to the opposing spindle poles. This often resulted in the formation of oocytes with 60 replicated chromosomes. Depletion of SGO1 in GV oocytes affected chromosomal separation resulting in abnormal chromosome alignment at a significantly higher proportion than in control oocytes. Knockdown of SGO1 expression significantly decreased the embryonic developmental rate and quality. To further confirm the function(s of SGO1 during mitosis, bovine embryonic fibroblast cells were transfected with SGO1 siRNAs. SGO1 depletion induced the premature dissociation of chromosomal cohesion at the centromere and along the chromosome arm giving rise to abnormal appearing mitotic patterns. The results of this study infer that SGO1 is involved in the centromeric cohesion of sister chromatids and chromosomal movement towards the spindle poles. Depletion of SGO1 causes arrestment of cell division in meiosis and mitosis.

  11. The prediction of ovarian response in in vitro fertilization-embryo transfer cycles

    Institute of Scientific and Technical Information of China (English)

    LIN Wen-qin; Yu Rong; YE Bi-lu; LIN Jin-ju; YANG Hai-yan

    2008-01-01

    Objective:To indentify the predictive parameters of ovarian response in in vitro fertilization-embryo transfer(IVF-ET)cycles,Methods:one hundred and six patients were administered GnRH-a/ rFSH/ human chorionic gonadotrophin(HCG)for ovarian stimulation.The following predictive parameters were examined,age,the basal level of FSH,mean ovarian volume,total of antral follicle count(AFC),the ratio of FSH/LH and the concentration of inhibin B after down-regulation.Ovarian response was assessed by the number of oocytes retrieved and the dosage of gonado-trophins(Can)used.Results:Correlations between predictive parameters and assessment indices of ovarian response were as follows:The basal FSH level and the ratio of FSH/LH showed a negative correlation with the number of oocyte obtained(r =-0.192,r=-0.227,respectively),while AFC and the concentration of inhibin B after down-regulation posi-tively correlated with the number of oocyte obtained(r=0.316,r=0.523,respectively),AFC,the mean ovarian volume and the concentration of inhibin B after down-regulation showed a negative correlation with the dosage of Gn used(r=-0.245,r=-0.294,r=-0.241,respectively),the ratio of FSH/ LH showed a positive correlation with the dosage of Gn used(r=0.255).Multiple regression analysis showed that inhibin B was the best predictor for the number of retrieved oocyte,followed by AFC,while the ovarian volume was the most important predictive parameter for the dosage of Gn used,then the ratio of FSH/LH.Conclusions:The level of inhibin B after down-regulation is the best predictor of ovarian response in IVF-ET.However,it's not yet a widely available test due to its high cost.AFC,by simple,non-invasive trsnsvaginal ultra-sound scan,could be the first applicable option in predicting ovarian response before IVF treatment.

  12. No peri- and postnatal effects on calves born after transfer of in vitro produced embryos vitrified by the open pulled straw (OPS) method

    DEFF Research Database (Denmark)

    Jacobsen, H.; Holm, P.; Schmidt, M.;

    2003-01-01

    The general objective of this study was to perform follow-up studies including selected peri- and postnatal characteristics on calves born after transfer of in vitro produced (IVP) embryos vitrified by the 'Open Pulled Straw' (OPS) method. An overall pregnancy rate of 16% after transfer of the OP...

  13. Reproductive efficiency of asymptomatic Theileria equi carriers mares submitted to an embryo transfer program

    Directory of Open Access Journals (Sweden)

    Luciana L. Bezerra

    2015-03-01

    Full Text Available This study aimed to assess and evaluate the effects of Theileria equi infection on embryonic recovery, gestation and early embryonic loss. Thirteen Mangalarga Marchador Theileria equi positive donors (diagnosed through nested-PCR and 40 embryos receptors were used. Donors were submitted to two embryo collections in two consecutive estrous cycles (GId; after, the same mares were treated with imidocarb dipropionate (1.2mg/kg IM. in order to collect more embryos in two more estrous cycles (GIId. Receptors were divided into two groups (control and with treated with 20 animals each, where one group was the control (GIr and the other one (GIIr treated with 1.2mg/kg IM of imidocarb dipropionate assessing the gestation rate at 15, 30, 45 and 60 days. After 52 embryo collections, the embryonic recovery rates were 53.84% (14/26 and 65.38% (17/26 (p> 0.05 for GId and GIId, respectively. The gestation rate was 70% (14/20 (p>0.05 at 15, 30, 45 and 60 days in group GIr and for GIIr was 85% (17/20 (p>0.05 at 15 days, 80% (16/20 (p>0.05 at 30, 45 and 60 days. The treatment with imidocarb dipropionate did not cause significant improvement in the reproductive efficiency at an ET program.

  14. GnRH agonist versus GnRH antagonist in in vitro fertilization and embryo transfer (IVF/ET).

    Science.gov (United States)

    Depalo, Raffaella; Jayakrishan, K; Garruti, Gabriella; Totaro, Ilaria; Panzarino, Mariantonietta; Giorgino, Francesco; Selvaggi, Luigi E

    2012-04-13

    Several protocols are actually available for in Vitro Fertilization and Embryo Transfer. The review summarizes the main differences and the clinic characteristics of the protocols in use with GnRH agonists and GnRH antagonists by emphasizing the major outcomes and hormonal changes associated with each protocol. The majority of randomized clinical trials clearly shows that in "in Vitro" Fertilization and Embryo Transfer, the combination of exogenous Gonadotropin plus a Gonadotropin Releasing Hormone (GnRH) agonist, which is able to suppress pituitary FSH and LH secretion, is associated with increased pregnancy rate as compared with the use of gonadotropins without a GnRH agonist. Protocols with GnRH antagonists are effective in preventing a premature rise of LH and induce a shorter and more cost-effective ovarian stimulation compared to the long agonist protocol. However, a different synchronization of follicular recruitment and growth occurs with GnRH agonists than with GnRH antagonists. Future developments have to be focused on timing of the administration of GnRH antagonists, by giving a great attention to new strategies of stimulation in patients in which radio-chemotherapy cycles are needed.

  15. GnRH agonist versus GnRH antagonist in in vitro fertilization and embryo transfer (IVF/ET

    Directory of Open Access Journals (Sweden)

    Depalo Raffaella

    2012-04-01

    Full Text Available Abstract Several protocols are actually available for in Vitro Fertilization and Embryo Transfer. The review summarizes the main differences and the clinic characteristics of the protocols in use with GnRH agonists and GnRH antagonists by emphasizing the major outcomes and hormonal changes associated with each protocol. The majority of randomized clinical trials clearly shows that in “in Vitro” Fertilization and Embryo Transfer, the combination of exogenous Gonadotropin plus a Gonadotropin Releasing Hormone (GnRH agonist, which is able to suppress pituitary FSH and LH secretion, is associated with increased pregnancy rate as compared with the use of gonadotropins without a GnRH agonist. Protocols with GnRH antagonists are effective in preventing a premature rise of LH and induce a shorter and more cost-effective ovarian stimulation compared to the long agonist protocol. However, a different synchronization of follicular recruitment and growth occurs with GnRH agonists than with GnRH antagonists. Future developments have to be focused on timing of the administration of GnRH antagonists, by giving a great attention to new strategies of stimulation in patients in which radio-chemotherapy cycles are needed.

  16. Transgenerational inheritance of the insulin-resistant phenotype in embryo-transferred intrauterine growth-restricted adult female rat offspring.

    Science.gov (United States)

    Thamotharan, Manikkavasagar; Garg, Meena; Oak, Shilpa; Rogers, Lisa M; Pan, Gerald; Sangiorgi, Frank; Lee, Paul W N; Devaskar, Sherin U

    2007-05-01

    To determine mechanisms underlying the transgenerational presence of metabolic perturbations in the intrauterine growth-restricted second-generation adult females (F2 IUGR) despite normalizing the in utero metabolic environment, we examined in vivo glucose kinetics and in vitro skeletal muscle postinsulin receptor signaling after embryo transfer of first generation (F1 IUGR) to control maternal environment. Female F2 rats, procreated by F1 pre- and postnatally nutrient- and growth-restricted (IUGR) mothers but embryo transferred to gestate in control mothers, were compared with similarly gestating age- and sex-matched control (CON) F2 progeny. Although there were no differences in birth weight or postnatal growth patterns, the F2 IUGR had increased hepatic weight, fasting hyperglycemia, hyperinsulinemia, and unsuppressed hepatic glucose production, with no change in glucose futile cycling or clearance, compared with F2 CON. These hormonal and metabolic aberrations were associated with increased skeletal muscle total GLUT4 and pAkt concentrations but decreased plasma membrane-associated GLUT4, total pPKCzeta, and PKCzeta enzyme activity, with no change in total SHP2 and PTP1B concentrations in IUGR F2 compared with F2 CON. We conclude that transgenerational presence of aberrant glucose/insulin metabolism and skeletal muscle insulin signaling of the adult F2 IUGR female offspring is independent of the immediate intrauterine environment, supporting nutritionally induced heritable mechanisms contributing to the epidemic of type 2 diabetes mellitus.

  17. IVF policy and global/local politics: the making of multiple-embryo transfer regulation in Taiwan.

    Science.gov (United States)

    Wu, Chia-Ling

    2012-08-01

    This paper analyzes the regulatory trajectory of multiple-embryo transfer in in-vitro fertilization (IVF) in Taiwan. Taking a latecomer to policy-making as the case, it argues the importance of conceptualizing the global/local dynamics in policy-making for assisted reproductive technology (ART). The conceptual framework is built upon recent literature on standardization, science policy, and global assemblage. I propose three interrelated features that reveal the "global in the local": (1) the power relationships among stakeholders, (2) the selected global form that involved actors drew upon, and (3) the re-contextualized assemblage made of local networks. Data included archives, interviews, and participant observation. In different historical periods the specific stakeholders selected different preferred global forms for Taiwan, such as Britain's code of ethics in the 1990s, the American guideline in the early 2000s, and the European trend in the mid-2000s. The global is heterogeneous. The failure to transfer the British regulation, the revision of the American guideline by adding one more embryo than it specified, and the gap between the cited European trend and the "no more than four" in Taiwan's 2007 Human Reproduction Law all show that the local network further transforms the selected global form, confining it to rhetoric only or tailoring it to local needs. Overall, Taiwanese practitioners successfully maintained their medical autonomy to build a 'flexible standardization'. Multiple pregnancy remains the most common health risk of IVF in Taiwan. PMID:22607747

  18. Effect of Gutai Decoction (固胎汤) on the Abortion Rate of in vitro Fertilization and Embryo Transfer

    Institute of Scientific and Technical Information of China (English)

    LIU Ying; WU Jing-zhi

    2006-01-01

    Objective: To study the effect of Chinese herbal medicine Gutai Decoction (固胎汤, GTD) on the abortion rate of in vitro fertilization and embryo transfer (IVF-ET). Methods: Observed were two hundred and forty-seven women having received IVF-ET and with β-human chorionic gonadotropin (β-HCG) > 25 IU/L on the 14th day after transferring. All were treated conventionally with progesterone 20-80 mg per day after transferring and if necessary the treatment was supplemented with Progynova 2 - 4 mg per day, with the medication withdrawn gradually from the 9th week of pregnancy till stopped completely. Among them 131 cases received GTD medication additionally, for 109 cases of whom the medication started from the 2nd day of transferring (taken as Group A) and for the other 22 cases from the 14th day after transferring (taken as Group B), the other 116 cases with no additional GTD treatment given were taken as the control group, with the medication lasting to the 12th week. The abortion rate in them was observed. Results: The abortion rate in Group A, Group B and the control group was 12.84%, 13.64% and 23.28%, respectively, the difference between the GTD treated groups and the control group was significant (P<0.05). Conclusion: Chinese medicine GTD could reduce abortion rate in women receiving IVF-ET.

  19. Evaluation of bovine (Bos indicus) ovarian potential for in vitro embryo production in the Adamawa plateau (Cameroon).

    Science.gov (United States)

    Kouamo, J; Dawaye, S M; Zoli, A P; Bah, G S

    2014-01-01

    An abattoir study was conducted to evaluate the ovarian potential of 201 local zebu cattle from Ngaoundere, Adamawa region (Cameroon) for in vitro embryo production (IVEP). The ovaries were excised, submerged in normal saline solution (0.9%) and transported to the laboratory for a detailed evaluation. Follicles on each ovary were counted, their diameters (Φ) measured and were grouped into 3 categories: small (Φ 8 mm). Each ovary was then sliced into a petri dish; the oocytes were recovered in Dulbecco's phosphate buffered saline, examined under a stereoscope (x10) and graded into four groups based on the morphology of cumulus oophorus cells and cytoplasmic changes of the oocytes. Grade I (GI): oocytes with more than 4 layers of bunch of compact cumulus cells mass with evenly granulated cytoplasm; grade II (GII): oocyte with at least 2-4 layers of compact cumulus cell mass with evenly granulated cytoplasm; grade III (GIII): oocyte with at least one layer of compact cumulus cell mass with evenly granulated cytoplasm; grade IV (GIV): denuded oocyte with no cumulus cells or incomplete layer of cumulus cell or expanded cells and having dark or unevenly granulated cytoplasm. The effects of both ovarian (ovarian localization, corpus luteum, size and weight of ovary) and non-ovarian factors (breed, age, body condition score (BCS) and pregnancy status of cow) on the follicular population and oocyte recovery rate were determined. There were an average of 16.75±0.83 follicles per ovary. The small, medium and large follicles were 8.39±0.60, 8.14±0.43 and 0.21±0.02 respectively. Oocyte recovery was 10.97±0.43 per ovary (65%). Oocytes graded I, II, III and IV were 3.53±0.19 (32.21%), 2.72±0.15 (24.82%), 2.24±0.15 (20.43%) and 2.47±0.20 (22.54%) respectively. The oocyte quality index was 2.26. Younger non pregnant cows having BCS of 3 and large ovaries presented higher number of follicles and oocyte quality (P quality (grade I and II) acceptable for IVEP constituted 57

  20. Evaluation of bovine (Bos indicus) ovarian potential for in vitro embryo production in the Adamawa plateau (Cameroon)

    Science.gov (United States)

    Kouamo, J.; Dawaye, S.M.; Zoli, A.P.; Bah, G.S.

    2014-01-01

    An abattoir study was conducted to evaluate the ovarian potential of 201 local zebu cattle from Ngaoundere, Adamawa region (Cameroon) for in vitro embryo production (IVEP). The ovaries were excised, submerged in normal saline solution (0.9%) and transported to the laboratory for a detailed evaluation. Follicles on each ovary were counted, their diameters (Φ) measured and were grouped into 3 categories: small (Φ 8 mm). Each ovary was then sliced into a petri dish; the oocytes were recovered in Dulbecco’s phosphate buffered saline, examined under a stereoscope (x10) and graded into four groups based on the morphology of cumulus oophorus cells and cytoplasmic changes of the oocytes. Grade I (GI): oocytes with more than 4 layers of bunch of compact cumulus cells mass with evenly granulated cytoplasm; grade II (GII): oocyte with at least 2-4 layers of compact cumulus cell mass with evenly granulated cytoplasm; grade III (GIII): oocyte with at least one layer of compact cumulus cell mass with evenly granulated cytoplasm; grade IV (GIV): denuded oocyte with no cumulus cells or incomplete layer of cumulus cell or expanded cells and having dark or unevenly granulated cytoplasm. The effects of both ovarian (ovarian localization, corpus luteum, size and weight of ovary) and non-ovarian factors (breed, age, body condition score (BCS) and pregnancy status of cow) on the follicular population and oocyte recovery rate were determined. There were an average of 16.75±0.83 follicles per ovary. The small, medium and large follicles were 8.39±0.60, 8.14±0.43 and 0.21±0.02 respectively. Oocyte recovery was 10.97±0.43 per ovary (65%). Oocytes graded I, II, III and IV were 3.53±0.19 (32.21%), 2.72±0.15 (24.82%), 2.24±0.15 (20.43%) and 2.47±0.20 (22.54%) respectively. The oocyte quality index was 2.26. Younger non pregnant cows having BCS of 3 and large ovaries presented higher number of follicles and oocyte quality (P BCS and ovarian size must be taken into account to

  1. Evaluation of bovine (Bos indicus ovarian potential for in vitro embryo production in the Adamawa plateau (Cameroon

    Directory of Open Access Journals (Sweden)

    J. Kouamo

    2014-12-01

    Full Text Available An abattoir study was conducted to evaluate the ovarian potential of 201 local zebu cattle from Ngaoundere, Adamawa region (Cameroon for in vitro embryo production (IVEP. The ovaries were excised, submerged in normal saline solution (0.9% and transported to the laboratory for a detailed evaluation. Follicles on each ovary were counted, their diameters (Φ measured and were grouped into 3 categories: small (Φ 8 mm. Each ovary was then sliced into a petri dish; the oocytes were recovered in Dulbecco’s phosphate buffered saline, examined under a stereoscope (x10 and graded into four groups based on the morphology of cumulus oophorus cells and cytoplasmic changes of the oocytes. Grade I (GI: oocytes with more than 4 layers of bunch of compact cumulus cells mass with evenly granulated cytoplasm; grade II (GII: oocyte with at least 2-4 layers of compact cumulus cell mass with evenly granulated cytoplasm; grade III (GIII: oocyte with at least one layer of compact cumulus cell mass with evenly granulated cytoplasm; grade IV (GIV: denuded oocyte with no cumulus cells or incomplete layer of cumulus cell or expanded cells and having dark or unevenly granulated cytoplasm. The effects of both ovarian (ovarian localization, corpus luteum, size and weight of ovary and non-ovarian factors (breed, age, body condition score (BCS and pregnancy status of cow on the follicular population and oocyte recovery rate were determined. There were an average of 16.75±0.83 follicles per ovary. The small, medium and large follicles were 8.39±0.60, 8.14±0.43 and 0.21±0.02 respectively. Oocyte recovery was 10.97±0.43 per ovary (65%. Oocytes graded I, II, III and IV were 3.53±0.19 (32.21%, 2.72±0.15 (24.82%, 2.24±0.15 (20.43% and 2.47±0.20 (22.54% respectively. The oocyte quality index was 2.26. Younger non pregnant cows having BCS of 3 and large ovaries presented higher number of follicles and oocyte quality (P < 0.05 compared with other animals. Oocytes with

  2. No Peri- and Postnatal Effects on Calves Born After Transfer of in Vitro Produced Embryos Vitrified by the Open Pulled Straw (OPS Method

    Directory of Open Access Journals (Sweden)

    Callesen H

    2003-06-01

    Full Text Available The general objective of this study was to perform follow-up studies including selected peri- and postnatal characteristics on calves born after transfer of in vitro produced (IVP embryos vitrified by the 'Open Pulled Straw' (OPS method. An overall pregnancy rate of 16% after transfer of the OPS-vitrified IVP embryos was achieved and resulted in birth of 9 calves, with 11 AI calves serving as controls. There were no immediate or long-term effects on these calves with respect to birth weight, gestation length, perinatal mortality, growth rate, disease susceptibility and reproductive performance.

  3. DNA methylation in porcine preimplantation embryos developed in vivo or produced by in vitro fertilization, parthenogenetic activation and somatic cell nuclear transfer

    DEFF Research Database (Denmark)

    Deshmukh, Rahul Shahaji; Østrup, Olga; Østrup, Esben;

    2011-01-01

    ), in vitro fertilized (IVF), somatic cell nuclear transfer (SCNT) and parthenogenetically activated (PA) embryos were evaluated for DNA methylation quantification at different developmental stages. Fertilized (IV and IVF) one-cell stages lacked a substantial active demethylation of the paternal genome....... Embryos produced under in vitro conditions had higher levels of DNA methylation than IV. A lineage-specific DNA methylation (hypermethylation of the inner cell mass and hypomethylation of the trophectoderm) was observed in porcine IV late blastocysts, but was absent in PA- and SCNT-derived blastocysts...... despite the occurrence of de novo methylation in early blastocysts. Comparable levels of DNA methylation were found in IV embryos and in 50% and 14% of SCNT early and late blastocysts, respectively. In conclusion, DNA methylation patterns were adversely affected by in vitro embryo production...

  4. Transgenic expression of green fluorescent protein in caprine embryos produced through electroporation-aided sperm-mediated gene transfer.

    Science.gov (United States)

    Kumar Pramod, R; Kumar, Rakesh; Mitra, Abhijit

    2016-01-15

    Current methods of transgenic animal production are afflicted by low efficiency and high cost. Recently, the electroporation aided sperm-mediated gene transfer (SMGT) emerges as a promising alternative with variable success rate. Among the domestic animal species, the electroporation-aided SMGT is less investigated in goats, except a few reports in which attempts have been made using the auto-uptake method of SMGT. In this study, we report an optimized electroporation condition for SMGT of caprine sperm cells. Results of this study demonstrated that electroporation of caprine sperm cells at 300 V for 200 mS in TALP medium allowed the maximum uptake of foreign DNA with minimum adverse effects on the vital semen parameters viz., progressive motility, viability, and membrane and acrosome integrity. Further, DNA binding assay revealed DNA uptake by 81.3% sperm cells when 1.0 μg of DNA was used under optimum electroporation conditions as compared to 16.5% on simple incubation. The qPCR analysis showed four-fold more (Pelectroporation than incubation. A similar cleavage rate was observed after IVF using either electroporated (23.20 ± 1.20) or non-electroporated (25.20 ± 2.41) sperm cells suggesting the absence of adverse effect of electroporation on the fertilizing ability. Out of the 116 embryos produced by electroporated sperm, five (4.31%) embryos showed the expression of the foreign gene. In conclusion, our results confirm that using optimized electroporation conditions, the caprine sperm cells can uptake foreign DNA effectively with minimum negative effect on the semen parameters and could produce transgenic embryos.

  5. Does transferring three or more embryos make sense for a well-defined population of infertility patients undergoing IVF/ICSI?

    Science.gov (United States)

    Masschaele, T.; Gerris, J.; Vandekerckhove, F.; De Sutter, P.

    2012-01-01

    Recently, there has been a marked increase in the use of Single Embryo Transfer (SET) subsequent to In Vitro Fertilization (IVF) and Intracytoplasmic Sperm Injection (ICSI), with the aim of reducing the risk of multiple gestations. However, critics have stated that by reducing the number of embryos transferred, a group of women with an a priori reduced chance of pregnancy are at particular greater risk of undertreatment. This group includes women who are of a certain age (≥ 40 years) or have already received a number of – failed – IVF attempts. We wanted to study whether the practice of three or more embryos being transferred would be of added value to these patients and whether the strategy of Heavy Load Transfer (HLT) is likely to boost the pregnancy rates to an acceptable level. We performed both a literature study and a retrospective cohort analysis of 7,850 IVF/ICSI cycles of early cleavage stage embryo transfer. Notwithstanding the limitations associated with this approach, we contend that HLT in the group of patients with poor prognosis should be recommended. This article outlines a suggested protocol within the legal framework relevant to Belgium. PMID:24753889

  6. The analysis of clinical outcome of frozen-thawing embryo transfer after whole embryo cryopreservation%全胚冷冻后冻融周期移植的临床结局分析

    Institute of Scientific and Technical Information of China (English)

    李攀; 姜宏; 陈京京; 范静

    2014-01-01

    Objective To analyze the outcomes of frozen-thawed blastocyst and cleavage embryo transfer after whole embryos cryopreservation. Methods The data of 489 IVF-ET cycles in reproductive medicine center of our hospital from September 2012 to August 2013 were analyzed retrospectively. Whole embryos cryopreservation in 214 patients were carried out with vitrification method and served as group A , 275 cycles performed fresh embryo transfer were served as group B. Then group A and group B were subdivided into group A1 (83 cycles),group A2 (131 cycles), group B1 (120 cycles)and group B2 (155 cycles)according to blastocyst transfer or cleavage-stage embryo transfer. The clinical outcomes of all groups were compared each other. Results The pregnancy rate and embryo implantation rate in group A1 were significantly higher(71.1% ,53.0%)than those in group A2, B1 and B2 (A2 group: 57.3%, 34.0%, B1group: 55.0%,42.1%, B2 group: 52.9%, 32.7%,respectively)(P<0.05). The embryo implantation rate in group B1 were higher than those in group B2 (P < 0.05). Conclusion F-ET after whole embryos freezing could significantly improve the embryo utilization rate and clinical pregnancy rate. Frozen-thawed blastocyst transfer could get better clinical outcomes than frozen-thawed cleavage-stage embryo transfer.%目的:分析全胚冷冻对体外受精/卵泡浆内单精子注射-胚胎移植(IVF/ICSI-ET)临床结局的影响。方法:回顾性分析2012年9月至2013年8月我院生殖医学中心489个移植周期的临床资料,其中214个全胚冷冻后首次冻融移植周期(F-ET)为A组,275个新鲜移植周期为B 组,并根据移植胚胎发育阶段将各组进一步分为囊胚移植组(A1组,B1组)和卵裂胚移植组(A2组,B2组),比较各组的临床结局。结果:A1组临床妊娠率和胚胎种植率(71.1%、53.0%)显著高于 A2、B1和 B2组(分别为:57.3%、34.0%;55.0%、42.1%;52.9%、32.7

  7. Production of human lysozyme-transgenic cloned porcine embryos by somatic nuclear transfer

    Institute of Scientific and Technical Information of China (English)

    Qiuyan Li; Hengxi Wei; Ying Guo; Yan Li; Rui Zhao; Yufang Ma; Zhengquan Yu; Bo Tang; Lei Zhang; Yunping Dai; Ning Li

    2009-01-01

    Due to their physiology and organ size, pigs have significant potential as human disease models and as organ transplantation donors. Genetic modification of pigs could provide benefits for both agriculture and human medicine. In this study, five fetal pig fibroblast cell lines from two species (Wuzhishan and Landrace pigs) were transfected using double-marked human lysozyme (HLY) plasmids (pBC1-HLY-GFP-NEO) by a liposome-mediated method. The ratio of green fluorescent protein (GFP)-expressing cells was >95% in sw7, sw8, s1w3 and s1w6 cell lines, but only 49.3% in slw9 cells. Cells from the four highly transgenic lines were used as nuclear donors to construct embryos, which were then cultured after fusion and activation by electric stimulation. The rate of cleavage was 76.7%, 48 h after acti-vation. After 7 days, 18.5% of cleaved eggs had developed to the blastocyst stage and 93.3% of blastocysts were GFP-positive. These results indicate that transgenic fetal pig fibroblast cell lines could be obtained by a liposome-mediated method, though the transfection efficiency varied between cell lines. Reconstructed embryos derived from transgenic cells could successfully develop into blastocysts, most of which were GFP-positive.

  8. DNA methylation status of H19 and Xist genos in lungs of somatic cell nuclear transfer bovines

    Institute of Scientific and Technical Information of China (English)

    CHEN Jie; LI DongJie; LIU YanQin; ZHANG Cui; DAI YunPing; LI ShiJie; LINing

    2008-01-01

    In somatic cell nuclear transfer (SCNT) technologies, the donor cell's nuclei need to be epigenetically reprogrammed for embryonic development. The incomplete reprogramming of donor cell nuclei has been implicated as a primary reason for the low efficiency of SCNT. DNA methylation is a major epige- netic modification of the genome that regulates crucial aspects of genome function, including estab-lishment of genomic imprinting. In order to make sure whether the DNA methylation reprogramming is efficient in SCNT animals, we analyzed the DNA methylation status of two imprinting genes, H19 and Xist, in lungs of deceased SCNT bovines that died within 48 h of birth using bisulfite sequencing analysis. Our findings demonstrated that cloned bovines showed significantly lower DNA methylation of H19 than controls (P<0.05), and three tested CpGs sites (1, 2, 3) exhibited unmethylation in one cloned bovine (9C3); however, Xist showed similar DNA methylation levels between clones and con- trols, and both showed hypermethylation (96.11% and 86.67%).

  9. 冷冻前和复苏后胚胎质量对临床结局的影响%Impact of embryo factors on clinical outcome in frozen - thawed embryo transfer cycles

    Institute of Scientific and Technical Information of China (English)

    陆小溦; 孙贻娟; 张爱军; 谷瑞环; 冯云

    2012-01-01

    Objective: To investigate the influence of post -thaw survival rate or pre -freeze embryo qulity and clinical pregnancy rate in frozen -thawed embryo transfer program (FET). Methods: A total of 683 FET of the patients aged <35 years and who were only transferred 2 embryos in a cycle were analyzed retrospectively from Jan 2008 to Dec 2010. Cases were divided into 4 groups according to the pre - freeze embryo qulity and post - thaw survival rate ( Group A: fully intact embryos and ≥1 pre - freeze high - qulity embryo, Group B: fully intact embryos and no pre - freeze high - qulity embryo, Group C: post - thaw damaged embryos and ≥1 pre - freeze high - qulity embryo, Group D: post - thaw damaged embryos and no pre - freeze high - qulity embryo). And clinical pregnancy rate and multiple pregnancy rate were compared between groups. Results: There was no significant difference in the clinical pregnancy rate and multiple pregnancy rate between Group A and Group C, or Group B and Group D, or Group A + B and Group C + D. But there was a significantly higher pregnancy rate in Group A and Group C (A vs. B,A vs. D,A + C vs. B + D, P < 0. 01; C vs. B, P < 0. 05). Conclusions: If there was any pre - freeze high - qulity embryo in the post — thaw embyos, they will have a higher pregnancy rate.%目的 将复苏后胚胎按冷冻前是否含有优胚和/或是否同冻存时分组,比较他们的妊娠结局,探讨是冷冻前的胚胎质量还是复苏后胚胎是否同冻存时,对妊娠结局的影响更大.方法 本中心2008.01 - 2010.12期间促排获得胚胎并冻存,患者年龄<35岁,复苏并移植2枚胚胎的683周期,共分四组:A组同冻存时,≥1枚优胚;B组同冻存时,0枚优胚;C组≥1枚有球溶解可用胚,≥1枚冻前优胚;D组≥1枚有球溶解可用胚,0枚冻前优胚.分析各组妊娠率、多胎率的差异.结果 总复苏率94.08%,冻前优胚复苏率98.01% (395/403),高于冻前非优胚复苏率92.56% (971/1049),P<0

  10. EFSA Panel on Biological Hazards (BIOHAZ); Scientific Opinion on the risk of transmission of classical scrapie via in vivo derived embryo transfer in ovine animals

    DEFF Research Database (Denmark)

    Hald, Tine; Baggesen, Dorte Lau

    . Under natural exposure conditions, animals that are heterozygous or homozygous A136R154R171 display respectively a low or negligible risk of being infected. The genetic control of the susceptibility to classical scrapie is also likely to impact on the risk of transmitting the disease via embryo transfer......The risk of transmission of classical scrapie via the transfer of in vivo derived embryo in ovines was assessed, taking into account the scientific information made available since the last EFSA opinion on this topic (2010) (see http://www.efsa.europa.eu/en/efsajournal/pub/1429.htm). The potential...... impact of PrP genotype of the embryo and/or of the ram and donor ewe on this risk was also assessed. The new data made available over the last three years further reinforce the view that classical scrapie could be vertically transmitted in sheep. Since the possibility of such vertical transmission...

  11. Cryo-thawed embryo transfer: natural versus artificial cycle. A non-inferiority trial.(ANTARCTICA trial

    Directory of Open Access Journals (Sweden)

    Groenewoud Eva R

    2012-09-01

    Full Text Available Abstract Background Frozen thawed embryo transfer (FET is a cost- effective adjunct to IVF or IVF-ICSI treatment. In order to optimize treatment outcome, FET should be carried out during a period of optimal endometrial receptivity. To optimize implantation several methods for endometrium preparation have been proposed. In natural cycle FET (NC-FET, the endometrium develops under endogenous hormonal stimulation. The development of the dominant follicle and endometrium is monitored by ultrasound and FET is timed after triggering ovulation induction or determination of the spontaneous LH surge. In an artificial cycle FET (AC-FET estrogens and progesterone are administered to prepare the endometrium for implantation. While the currently available data show no significant difference in pregnancy rates between these methods, well designed randomized controlled trials are lacking. Moreover there is little literature on difference in cancellation rates, cost-efficiency and adverse events. Methods and design In this randomized, multi-centre, non-inferiority trial we aim to test the hypothesis that there is no significant difference in live birth rates between patients undergoing NC-FET versus AC-FET. The primary outcome will be live birth rate per embryo transfer procedure. Secondary outcomes will be ongoing and clinical pregnancy rate, cancellation rate, (serious adverse events and cost-efficiency. Based on a live birth rate of 20% and a minimal clinical important difference of 7,5% (one-sided alpha 2,5%, beta 20% a total of 1150 patients will be needed. Analyzes will be performed using both per protocol as well as intention to treat analyses. Discussion This prospective, randomized, non –inferiority trial aims to address the hypothesis that there is no significant difference in live birth rates between patients undergoing NC-FET versus patients undergoing AC-FET. Moreover it addresses cost-efficiency as well as the perceived burden of both treatments

  12. 人胚胎移植后剩余胚胎继续体外培养潜能的研究%Potential development to blastocyst of the surplus embryos from human embryo transfer

    Institute of Scientific and Technical Information of China (English)

    薛侠; 赵皖秋; 张四林; 秦臻; 师娟子

    2011-01-01

    Objective To explore the developmental potential of the surplus embryos from human embryo transfer during IVF - ET cycles. Methods All embryos with non - pronucleus (0PN), a single pronudeus (1PN), a number of pronucleus (≥3PN) and 2 pronudeus delaying development in cleavage stage (2PN) were cultured into blastula by the sequential method. Results ① 314 Surplus embryos were collected and formed 152 blastulas(48.41% ) after the sequential culture, among which 53 (34.87%) were high-quality blastula. ② The embryo grade on Day 3 was related to blastocyst rate. The higher embryo grade, the higher blastula formation (54.39%, 52.39%, 49.61% and 21.62% ); ③ Blastocyst formation rates of embryos in 1PN embryos,0PN embryos and D3 from blastocyst embryos classified Ⅲ had higher rates of blastula formation than D3 from blastocyst embryos classified beyond Ⅲ ( P < 0.05). Conclusion Embryos of level Ⅲ and above in D3 are still opportunities for blastocyst formation. The 0PN, 1PN embryo cleavage embryos developed from the D3 can continue to develop in high - quality, until after the formation of blastocysts, and then a pre- implantation genetic diagnosis. If the karyotype is aneuploid karyotype, then it should be frozen or transplanted first. The measures above can improve the oocyte retrieval in patients with accumulation of a single pregnancy, which can also provide resources for embryonic stem cell research.%目的 探讨IVF新鲜周期中D3可用胚胎移植和冷冻后剩余胚胎继续培养的价值.方法 通过囊胚序贯培养法将无原核(0PN)、单个原核(1PN)、多个原核(≥3PN)和卵裂期发育延缓的2原核(2PN)废弃胚胎培养至囊胚期.结果 ① 314枚剩余胚胎于D5~D7形成152枚囊胚(48.41%),其中53枚为优质囊胚(34.87%); ② 胚胎级别越高,囊胚形成率越高(54.39%、52.39%、49.61%和21.62%); ③ 0PN和1PN卵裂发育而来的D3优质胚胎、2PN卵裂发育来的D3评分为Ⅲ级

  13. Evaluation of a commercial bovine colostrum replacer for achieving passive transfer of immunity in springbok calves (Antidorcas marsupialis).

    Science.gov (United States)

    Thompson, Kimberly A; Lamberski, Nadine; Kass, Philip H; Coons, David; Chigerwe, Munashe

    2013-09-01

    Failure of passive transfer (FPT) occurs in ruminant species when there is inadequate absorption of immunoglobulins from the colostrum. In zoologic establishments, FPT can be a common occurrence in hand-raised ruminant neonates fed insufficient amounts of colostrum replacer (CR) and/or poor-quality CR. The goals of this study were to investigate the efficacy of a commercial bovine CR at achieving adequate passive transfer of immunity and evaluate tests to assess FPT in nondomestic ruminant species. In the past several years, research in dairy calves has shown that passive transfer rates can be dramatically improved if the CR dose is doubled. The treatment group (n = 10) consisted of springbok (Antidorcas marsupialis) calves fed Land O'Lakes bovine commercial CR at a dose of > or = 4.68 g of immunoglobulin G (22 g of CR powder) per kilogram of animal's body weight divided into five feedings over 24 hr. The control group (n=7) consisted of calves that nursed from their dams. This study reported no significant difference between the proportion of calves with adequate passive transfer in the treatment (80%) and control (71%) groups (P= 1.00). Morbidity and mortality rates until weaning were 0% in both groups. The study also determined the sensitivity and specificity values (n = 37) for five serum tests (y-glutamyl-transferase [GGT], globulin, glutaraldehyde coagulation [GC], sodium sulfite turbidity test, and total protein) used to determine passive transfer status in springbok calves. This study recommends the following serum tests and cutoff ranges for determining FPT in springbok calves: globulin or = 28 min. PMID:24063080

  14. Parabolic trend in endometrial thickness at embryo transfer in in vitro fertilization/intracytoplasmic sperm injection cases with clinical pregnancy evidence.

    Science.gov (United States)

    Lamanna, Giuseppina; Scioscia, Marco; Lorusso, Filomenamila; Serrati, Giuseppe; Selvaggi, Luigi E; Depalo, Raffaella

    2008-10-01

    Sonographic measurement of endometrial thickness at embryo transfer is thought to be a good predictor of the success of in vitro fertilization/intracytoplasmic sperm injection cycles because the clinical pregnancy rate increases as the endometrium thickens. Nevertheless, a retrospective analysis of a study population of 606 patients showed a decrease of clinical pregnancy rates in the setting of extreme endometrial thicknesses.

  15. Long term costs and effects of reducing the number of twin pregnancies in IVF by single embryo transfer: the TwinSing study

    Directory of Open Access Journals (Sweden)

    van Goudoever Johannes B

    2010-10-01

    Full Text Available Abstract Background Pregnancies induced by in vitro fertilisation (IVF often result in twin gestations, which are associated with both maternal and perinatal complications. An effective way to reduce the number of IVF twin pregnancies is to decrease the number of embryos transferred from two to one. The interpretation of current studies is limited because they used live birth as outcome measure and because they applied limited time horizons. So far, research on long-term outcomes of IVF twins and singletons is scarce and inconclusive. The objective of this study is to investigate the short (1-year and long-term (5 and 18-year costs and health outcomes of IVF singleton and twin children and to consider these in estimating the cost-effectiveness of single embryo transfer compared with double embryo transfer, from a societal and a healthcare perspective. Methods/Design A multi-centre cohort study will be performed, in which IVF singletons and IVF twin children born between 2003 and 2005 of whom parents received IVF treatment in one of the five participating Dutch IVF centres, will be compared. Data collection will focus on children at risk of health problems and children in whom health problems actually occurred. First year of life data will be collected in approximately 1,278 children (619 singletons and 659 twin children. Data up to the fifth year of life will be collected in approximately 488 children (200 singletons and 288 twin children. Outcome measures are health status, health-related quality of life and costs. Data will be obtained from hospital information systems, a parent questionnaire and existing registries. Furthermore, a prognostic model will be developed that reflects the short and long-term costs and health outcomes of IVF singleton and twin children. This model will be linked to a Markov model of the short-term cost-effectiveness of single embryo transfer strategies versus double embryo transfer strategies to enable the

  16. Effects of Scriptaid on Cell Cycle and Histone Acetylation of Ovine Nuclear Donor Cumulus Cells and their Ability to Support the Development of Somatic Cell Nuclear Transfer Embryos

    Directory of Open Access Journals (Sweden)

    Hui Cao

    2015-10-01

    Full Text Available Compelling evidence suggests that histone deacetylase inhibitor (HDACi influences the development of somatic cell nuclear transfer (SCNT embryos. The current study was conducted to determine the effect of pretreatment of donor cumulus cells with Scriptaid (a novel HDACi on cell cycle, histone acetylation and cloning embryos development in ovine. First, we optimized the efficiency of Scriptaid in a dose (0, 0.1, 0.2, 0.4 and 0.8 μmol/L and time-dependent (0, 12, 24, 36, and 48 h manner on the developmental capacity of these embryos. Then, we quantitatively assessed the alterations of acetylation levels in histone H3 lysine 9 (acH3K9 and histone H4 lysine 12 (acH4K12 of cumulus cells and SCNT embryos by immunofluorescence staining. Furthermore, we detected the proportion of G0/G1 phase cells in cumulus cells. We found a significantly improved blastocyst development rates of cloning embryos derived from donor cumulus cells pretreated with a mild dose (0.2 μmol/L of Scriptaid for 24 hours (21/86 [24.39%] vs. 11/85 [12.91%]; P<0.05. Meanwhile, the levels of acH3K9 and acH4K12 were also improved significantly in cumulus cells and SCNT embryos (P<0.05. Moreover, more cumulus cells pretreated with Scriptaid were in G0/G1 phase compared with control group (84.22% vs. 75.96%, P<0.05. In conclusion, donor cumulus cells treated with Scriptaid is beneficial to early development of SCNT embryos, ascending acH3K9/ acH4K12 and G0/G1 phase cells proportion of cumulus cell. Scriptaid can be used to improve the efficiency of somatic cell nuclear transfer in ovine.

  17. Determining the status of non-transferred embryos in Ireland: a conspectus of case law and implications for clinical IVF practice.

    LENUS (Irish Health Repository)

    Sills, Eric Scott

    2009-01-01

    The development of in vitro fertilisation (IVF) as a treatment for human infertilty was among the most controversial medical achievements of the modern era. In Ireland, the fate and status of supranumary (non-transferred) embryos derived from IVF brings challenges both for clinical practice and public health policy because there is no judicial or legislative framework in place to address the medical, scientific, or ethical uncertainties. Complex legal issues exist regarding informed consent and ownership of embryos, particularly the use of non-transferred embryos if a couple separates or divorces. But since case law is only beginning to emerge from outside Ireland and because legislation on IVF and human embryo status is entirely absent here, this matter is poised to raise contractual, constitutional and property law issues at the highest level. Our analysis examines this medico-legal challenge in an Irish context, and summarises key decisions on this issue rendered from other jurisdictions. The contractual issues raised by the Roche case regarding informed consent and the implications the initial judgment may have for future disputes over embryos are also discussed. Our research also considers a putative Constitutional \\'right to procreate\\' and the implications EU law may have for an Irish case concerning the fate of frozen embryos. Since current Medical Council guidelines are insufficient to ensure appropriate regulation of the advanced reproductive technologies in Ireland, the report of the Commission on Assisted Human Reproduction is most likely to influence embryo custody disputes. Public policy requires the establishment and implementation of a more comprehensive legislative framework within which assisted reproductive medical services are offered.

  18. A comparative study between cleavage stage embryo transfer at day 3 and blastocyst stage transfer at day 5 in in-vitro fertilization/intra-cytoplasmic sperm injection on clinical pregnancy rates

    Directory of Open Access Journals (Sweden)

    Prabhleen Kaur

    2014-01-01

    Full Text Available Objective: To evaluate the efficacy of blastocyst transfer in comparison with cleavage stage transfer. Study Design: A randomized, prospective study was conducted in Infertility clinic, Department of Obstetrics and Gynecology, Mahatma Gandhi Hospital, Jaipur on 300 patients aged 25-40 years undergoing in-vitro fertilization (IVF/intra-cytoplasmic sperm injection (ICSI cycle from May 2010-April 2011. When three or more Grade-I embryos were observed on day 2 of culture, patients were divided randomly into two study groups, cleavage stage transfer and blastocyst transfer group having 150 patients each. Primary outcomes evaluated were, Clinical pregnancy rate and Implantation rate. The results were analyzed using proportions, standard deviation and Chi-square test. Results: Both the groups were similar for age, indication and number of embryos transferred. Clinical pregnancies after blastocyst transfer were significantly higher 66 (44.0% compared to cleavage stage embryo transfer 44 (29.33% (P < 0.01.Implantation rate for blastocyst transfer group was also significantly higher (P < 0.001. Conclusion: Blastocyst transfer having higher implantation rate and clinical pregnancy rate lead to reduction in multiple pregnancies.

  19. The impact of cryopreservation timing and embryo factors on clinical outcome of frozen-thawed embryo transfer cycles%冷冻时机和胚胎因素对冻融胚胎移植结局的影响

    Institute of Scientific and Technical Information of China (English)

    朱桂杰; 谭丽

    2013-01-01

    Objective To investigate the impact of cryopreservation timing and embryo quality on the clinical out -come oi irozen - thawed embryo transfer cycles ( FET). Methods A retrospective analysis was performed on 417 FET cycles. The patients were divided into different groups according to freezing time , embryo blastomere count and good - quality embryo count. Frozen - thawed effects and clinical outcomes in different cryopreservation timings and embryo qualities were observed. Results The good -quality embryo rate of D2 embryo was significantly higher than that of D3 embryo (56. 84% vs 48. 82% ), though with significantly lower clinical pregnancy rate (27. 61% vs 38. 52% ). However, there was no significant difference in the post - thaw embryo survival rate or abortion rate between D 2 embryo and D3 embryo ( 83. 50% vs 79. 44% , and 10. 81 % vs 10. 09% , P >0. 05) . In D2 embryo group , the post - thaw embryo survival rate and total blastomere survival rate were significantly higher in cells exceeded 4 - cell than those less than 4 - cell (89. 35% vs 79. 11% , and 42. 01% vs 30. 22% , respectively, P < 0. 05). In D3 embryo group, they were significantly higher in cells exceeded 6 -cell comparing with those less than 6 -cell (86.52% vs 72.43% , and 41. 13% vs 31.07% , respectively, P <0. 01). When more and more were transferred , The clinical pregnancy rate and multiple pregnancy rate were significantly positively correlated with the good - quality embryo count ( P < 0. 05). Conclusion The D3 embryo cryopreservation in FET provides better clinical outcome . The embryos quality and good - quality embryo count transferred is close correlated with FET outcomes , suggesting the key factors to FET cycles.%目的 探讨冷冻时机和胚胎因素对冻融胚胎移植(FET)结局的影响.方法 回顾性分析417个FET周期,按冷冻时机、冷冻时胚胎卵裂球数、FET优质胚胎数进行分组,观察不同冷冻时机和胚胎质量复苏

  20. 牛与小鼠胚胎在分化抑制培养系统中的行为比较%Comparison of the growth behavior between bovine and murine embryo in a differentiation inhibitory system

    Institute of Scientific and Technical Information of China (English)

    杨奇; 安立龙; 窦忠英; 高志敏; 雷安民; 杨春荣

    2001-01-01

    在自制培养基中,以原代小鼠胎儿成纤维细胞为饲养层,观察比较了牛与小鼠胚胎在体外分化抑制培养体系中的生长行为。结果表明,牛囊胚一般培养36~60 h后孵化脱带,96~120 h后贴壁,120~144 h为ICM传代的最佳时刻。小鼠囊胚一般培养12~24 h孵化脱带,24~48 h贴壁,72~96 h为传代的最佳时刻。牛胚胎贴附于饲养层上生长,极易从饲养层上剥离;小鼠胚胎镶嵌在饲养层中,滋养层细胞与饲养层细胞间连接紧密。牛ICM色深发黑,集落隆起程度较低;小鼠ICM呈暗黄色,有呈柱状增殖的趋势。牛滋养层呈网状,小鼠滋养层为较致密的单层薄膜。%The growth behavior differences between bovine embryos and murine embryos were studied on PMEF feeder layer using DMEM+15% NBS+0.1 mmol/L β-mercaptoetheanol+0.1 μmol/L Na2 SeO3+10 μg/mL LIF+10 μg/mL IGF-1 as the media.The results showed as follows:In the culture system,bovine blastocyst hatched affer being cultured for 36-60 h,and attached on the feeder layer after being cultured for96-120 h.After cultured for 120-144 h,the growing ICM were passaged at first.Morulae attached on PMEF feeder layer 12-24 h later than blastocyst for passing on blastula stage and hatching period.Murine blastocyst hatched after being cultured for 12-24 h,and attached on the feeder layer after cultured for 24-48 h.After cultured for 72-96 h,the growing ICM were passaged.Bovine embryo lived on the feeder layer and easy to be stripped from the feeder layer,while murine embryo implanted in the feeder layer,and murine trophoblastic cells were closely connected with feeder layer cells arrounding them.The color of bovine ICM was more black than murine ICM.However,the colonies of bovine ICM were lower compared with murine ICM colonies,and murine ICM tended to be column-like.Bovine trophoblast was net-like,and murine trophoblast formed a membranous structure.

  1. DNA methylation in porcine preimplantation embryos developed in-vivo or produced by in-vitro fertilization, parthenogenetic activation and somatic cell nuclear transfer

    DEFF Research Database (Denmark)

    Deshmukh, Rahul Shahaji; Østrup, Olga; Østrup, Esben;

    2011-01-01

    vitro fertilized (IVF), somatic cell nuclear transfer (SCNT) and parthenogenetically activated (PA) embryos were evaluated for DNA methylation quantification at different developmental stages. Fertilized (IV and IVF) one-cell stages lacked a substantial active demethylation of the paternal genome....... Embryos produced under in vitro conditions had higher levels of DNA methylation than IV. A lineage-specific DNA methylation (hypermethylation of the inner cell mass and hypomethylation of the trophectoderm) was observed in porcine IV late blastocysts, but was absent in PA- and SCNT-derived blastocysts...

  2. Promising system for selecting healthy in vitro-fertilized embryos in cattle.

    Science.gov (United States)

    Sugimura, Satoshi; Akai, Tomonori; Hashiyada, Yutaka; Somfai, Tamás; Inaba, Yasushi; Hirayama, Muneyuki; Yamanouchi, Tadayuki; Matsuda, Hideo; Kobayashi, Shuji; Aikawa, Yoshio; Ohtake, Masaki; Kobayashi, Eiji; Konishi, Kazuyuki; Imai, Kei

    2012-01-01

    Conventionally, in vitro-fertilized (IVF) bovine embryos are morphologically evaluated at the time of embryo transfer to select those that are likely to establish a pregnancy. This method is, however, subjective and results in unreliable selection. Here we describe a novel selection system for IVF bovine blastocysts for transfer that traces the development of individual embryos with time-lapse cinematography in our developed microwell culture dish and analyzes embryonic metabolism. The system can noninvasively identify prognostic factors that reflect not only blastocyst qualities detected with histological, cytogenetic, and molecular analysis but also viability after transfer. By assessing a combination of identified prognostic factors--(i) timing of the first cleavage; (ii) number of blastomeres at the end of the first cleavage; (iii) presence or absence of multiple fragments at the end of the first cleavage; (iv) number of blastomeres at the onset of lag-phase, which results in temporary developmental arrest during the fourth or fifth cell cycle; and (v) oxygen consumption at the blastocyst stage--pregnancy success could be accurately predicted (78.9%). The conventional method or individual prognostic factors could not accurately predict pregnancy. No newborn calves showed neonatal overgrowth or death. Our results demonstrate that these five predictors and our system could provide objective and reliable selection of healthy IVF bovine embryos. PMID:22590579

  3. Promising system for selecting healthy in vitro-fertilized embryos in cattle.

    Directory of Open Access Journals (Sweden)

    Satoshi Sugimura

    Full Text Available Conventionally, in vitro-fertilized (IVF bovine embryos are morphologically evaluated at the time of embryo transfer to select those that are likely to establish a pregnancy. This method is, however, subjective and results in unreliable selection. Here we describe a novel selection system for IVF bovine blastocysts for transfer that traces the development of individual embryos with time-lapse cinematography in our developed microwell culture dish and analyzes embryonic metabolism. The system can noninvasively identify prognostic factors that reflect not only blastocyst qualities detected with histological, cytogenetic, and molecular analysis but also viability after transfer. By assessing a combination of identified prognostic factors--(i timing of the first cleavage; (ii number of blastomeres at the end of the first cleavage; (iii presence or absence of multiple fragments at the end of the first cleavage; (iv number of blastomeres at the onset of lag-phase, which results in temporary developmental arrest during the fourth or fifth cell cycle; and (v oxygen consumption at the blastocyst stage--pregnancy success could be accurately predicted (78.9%. The conventional method or individual prognostic factors could not accurately predict pregnancy. No newborn calves showed neonatal overgrowth or death. Our results demonstrate that these five predictors and our system could provide objective and reliable selection of healthy IVF bovine embryos.

  4. A modified natural cycle results in higher live birth rate in vitrified-thawed embryo transfer for women with regular menstruation.

    Science.gov (United States)

    Guan, Yichun; Fan, Hongfang; Styer, Aaron K; Xiao, Zhiying; Li, Zhen; Zhang, Jianrui; Sun, Lijun; Wang, Xingling; Zhang, Zhan

    2016-10-01

    There is no consensus regimen for the optimal endometrial preparation for cryopreservation and vitrified-thawed embryo transfer cycles. This is largely caused by the lack of sufficient investigation and analyses on the respective pregnancy and perinatal outcomes by different regimens. This study aimed to compare both pregnancy and perinatal outcomes between the modified natural and artificial cycles in vitrified-thawed day three embryo transfer for women with regular menstruation. A total of 1,482 vitrified-thawed day three embryo transfer cycles were reviewed including 427 modified natural cycles (NC), 132 ovulation induction cycles (OC), 794 artificial cycles (AC), and 129 GnRH agonist artificial cycles (GAC). The primary outcome that was evaluated was live birth rate. The NC regimen demonstrated a higher rate of ongoing pregnancy (43.8% vs. 30.2%, P = 0.002) and a lower rate of late abortion (2.8% vs. 14.0%, P = 0.003) than the GAC regimen as well as a higher implantation rate (31.9% vs. 27.1%, P = 0.008) and live birth rate (43.1% vs. 34.1%, P = 0.002) than the AC regimen. A significantly higher peak endometrial thickness before transfer was observed in patients using the NC and GAC regimens (10.0 ± 1.7, 9.9 ± 2.4) compared to the AC regimens (9.2 ± 1.5, P = 0.000). Multivariate logistic regression showed that the NC protocol was associated with a higher live birth rate. There were no significant differences in rates of pregnancy complications, neonatal mortality, birth defects, mean birth weight, and other perinatal outcomes among the regimens. Modified natural cycle endometrial preparation regimen for vitrified-thawed day three embryo transfer is associated with superior live birth pregnancy outcomes compared to artificial cycles. Future studies are warranted to investigate the underlying biologic mechanisms of these findings. Abbreviations ART: assisted reproductive technology; BMI: body mass index; FET: frozen-thawed embryo transfer; HCG: human chorionic

  5. 对废弃胚胎行囊胚培养在体外受精-胚胎移植中的意义%Potential use of discarded embryos for in vitro fertilization and embryo transfer (IVF-ET)

    Institute of Scientific and Technical Information of China (English)

    王荣; 费小阳; 王丽卿; 邹立波

    2014-01-01

    Objective To explore the potential use of discarded embryos for in vitro fertilization- embryo transfer (IVF- ET). Methods The discarded embryos from d3 were cultured into blastula by the sequential culture method. Results A total of 1962 discarded embryos from 782 patients were cultured, in which 376 blastocysts were formed with a blastocyst rate of 19.2%. The blastocyst formation rates were significantly higher from 1PN, 0PN and>6 cells embryos. The clinical pregnancy rate in women with blastosphere formed was significantly higher than those without blastosphere formed. Sixty four blastocysts were formed for 2PN,10 of which were transferred while the others were frozen. The clinical pregnancy rate of blastocyst transfer was 66.7%. Thir-teen women got pregnant in 24 frozen- thawed blastocyst transfer cycles. A total of 18 healthy babies were born from blastocyst transfer. Conclusion The discarded embryos have different development potential.They can develop into biastospheres. Blas-tocyst from appropriate discarded embryos can improve the utilization rate of embryos.%目的:探讨对不可移植的废弃胚胎行囊胚培养在体外受精-胚胎移植(IVF- ET)中的意义。方法通过囊胚序贯培养法将d3废弃胚胎培养至囊胚期。比较不同来源胚胎及d3胚胎的卵裂球数与囊胚形成的关系;利用废弃胚胎囊胚形成情况对体外受精妊娠结局进行预测;并将获得的可移植囊胚行胚胎移植及冷冻。结果共收集782例患者的1962个废弃胚胎,经序贯培养,形成囊胚376枚(19.2%);1PN胚胎、0PN胚胎、d 3卵裂球数>6的胚胎囊胚形成率较高(均P<0.05);废弃胚胎中有囊胚形成者的临床妊娠率明显高于无囊胚形成者(P<0.05);由2PN胚胎形成囊胚共64枚,其中10枚囊胚行移植,剩余囊胚行玻璃化冷冻保存,囊胚移植后临床妊娠率66.7%。解冻移植囊胚24个周期,妊娠周期13个,妊娠率54.2%。囊

  6. Effect of leptin on oocyte maturation and subsequent pregnancy rate of cloned embryos reconstructed by somatic cell nuclear transfer in pigs

    Institute of Scientific and Technical Information of China (English)

    Hengxi Wei; Qiuyan Li; Jun Li; Yan Li; Yunping Dai; Yufang Ma; Kai Xue; Ning Li

    2008-01-01

    Cloning pigs by somatic cell nuclear transfer (SCNT) has wide applications in basic research,human medicine and agricultural production.To improve cloning efficiency,the effect of two basic maturation media,NCSU-23 and TCMI99,was compared,and TCM199 was selected for the following experiments with leptin.We systematically studied the effects of leptin supplementation on oocytes in vitro maturation (IVM),in vitro development of parthenogenetically activated (Phi) and SCNT embryos and/n vivo develop-ment of SCNT embryos after embryo transfer (ET).The results showed that supplementation of 100 or 200 ng/ml leptin into the mat-uration medium did not greatly affect nuclear maturation of oocytes,or cleavage rates of PA and SCNT (P<0.05).Blastocyst rates of PA and SCNT embryos were significantly improved when 100 or 200 ng/ml leptin was added to maturation medium,and the number of cells in PA blastocysts was also improved (P<0.05).The number of cells in blastocyst of SCNT was improved,when 100 ng/ml leptin was added (P<0.05).Furthermore,supplementation of 100 or 200 ng/ml leptin to the IVM medium may improve pregnancy rate and the delivery rate in pig cloning.

  7. 113例全胚胎冷冻患者首次冻融胚胎移植的临床分析%Study of embryo transfer in infertile patients with 113 cases of whole embryo freezing

    Institute of Scientific and Technical Information of China (English)

    阮永铭; 郑华; 吴重聪; 蔡桂丰; 杨桂艳; 曾伟荣

    2012-01-01

    Objective; To study the frozen - thawed embryo transfer (FET) as the first transplant, compared with the fresh embryo transplant in clinical pregnancy outcome. Methods: A database was analyzed retrospectively which covered 902 ART cycles from January 2010 to December 2010 in reproductive fertility center of ZhuHai Maternal and Children Health Hospital, including 789 fresh embryo transfer cycles and 113 FET cycles. Results; Compared with the control group, the FET group had no significant differences in the terms of the cause of infertility, the abortion rate, the number of oocytes, frozen embryos and transferred embryos. There were also no significant differences between the control group and the FET group in clinical pregnancy outcome, but the differences existed in the FET groups caused by different factors. Conclusions; The FET groups caused by different factors had differences in the clinical pregnancy outcome. Whether the patient was integrated to take FET, all aspects should be considered in order to obtain the optimal ART pregnancy outcome when the fresh embryo transfer was not suitable.%目的 探讨冻融胚胎移植(frozen - thawed embryo transfer,FET)作为首次移植的临床结局,比较其与新鲜胚胎移植的临床妊娠结局.方法 回顾性分析珠海市妇幼保健院生殖中心2010年1月至12月接受人类辅助生殖技术(assisted reproductive technology,ART)的共902例患者的临床资料.其中,冻融胚胎移植作为首次移植113例,按全胚胎冷冻的原因不同分为5组,即卵巢过度刺激综合征(OHSS)组、孕酮升高组、雌二醇升高组、胚胎发育迟缓组、其他特殊原因组;同期新鲜胚胎移植789例,为对照组.结果 FET组与对照组相比,无论在不孕原因、获卵数、冷冻胚胎数、移植胚胎数、流产率的差异均无显著的差异,而两组患者临床妊娠率(P<0.05)有统计学差异,但FET各组间存在差异.结论 不同因素所致FET的临床妊娠结局存在

  8. Estimating transfer of bovine virus-diarrhoea virus in Danish cattle by use of register data

    DEFF Research Database (Denmark)

    Alban, L.; Stryhn, H.; Kjeldsen, A.M.;

    2001-01-01

    To study how routinely recorded data (also called "register data") might be used in disease monitoring on a regional or national level, a database for bovine virus-diarrhoea virus (BVDV) was made from existing databases, covering the period January 1995-November 1999. This paper includes a general...... description of the database, including basic statistics for selected variables. Information was largely complete for cattle herds in the milk-recording scheme (MRS), but only partly available for other herds. A methodology was developed to identify when and how a herd initially was infected. For most herds...

  9. Increased circulating cell-derived microparticle count is associated with recurrent implantation failure after IVF and embryo transfer.

    Science.gov (United States)

    Martínez-Zamora, M Angeles; Tàssies, Dolors; Reverter, Juan Carlos; Creus, Montserrat; Casals, Gemma; Cívico, Salvadora; Carmona, Francisco; Balasch, Juan

    2016-08-01

    Cell-derived microparticles (cMPs) are small membrane vesicles that are released from many different cell types in response to cellular activation or apoptosis. Elevated cMP counts have been found in almost all thrombotic diseases and pregnancy wastage, such as recurrent spontaneous abortion and in a number of conditions associated with inflammation, cellular activation and angiogenesis. cMP count was investigated in patients experiencing unexplained recurrent implantation failure (RIF). The study group was composed of 30 women diagnosed with RIF (RIF group). The first control group (IVF group) (n = 30) comprised patients undergoing a first successful IVF cycle. The second control group (FER group) included 30 healthy women who had at least one child born at term and no history of infertility or obstetric complications. cMP count was significantly higher in the RIF group compared with the IVF and FER groups (P < 0.05 and P < 0.01, respectively) (RIF group: 15.8 ± 6.2 nM phosphatidylserine equivalent [PS eq]; IVF group: 10.9 ± 5.3 nM PS eq; FER group: 9.6 ± 4.0 nM PS eq). No statistical difference was found in cMP count between the IVF and FER groups. Increased cMP count is, therefore, associated with RIF after IVF and embryo transfer.

  10. The correlation between endometrial thickness and outcome of in vitro fertilization and embryo transfer (IVF-ET outcome

    Directory of Open Access Journals (Sweden)

    Al-Rejjal Rafat

    2008-09-01

    Full Text Available Abstract Background To evaluate the relationship between endometrial thickness on day of human chorionic gonadotrophin administration (hCG and pregnancy outcome in a large number of consecutive in vitro fertilization and embryo transfer (IVF-ET cycles. Methods A retrospective cohort study including all patients who had IVF-ET from January 2003–December 2005 conducted at a tertiary center. Results A total of 2464 cycles were analysed. Pregnancy rate (PR was 35.8%. PR increased linearly (r = 0.864 from 29.4% among patients with a lining of less than or equal to 6 mm, to 44.4% among patients with a lining of greater than or equal to 17 mm. ROC showed that endometrial thickness is not a good predictor of PR, so a definite cut-off value could not be established (AUC = 0.55. Conclusion There is a positive linear relationship between the endometrial thickness measured on the day of hCG injection and PR, and is independent of other variables. Hence aiming for a thicker endometrium should be considered.

  11. Twin pregnancy and partial hydatidiform mole following in vitro fertilization and embryos transfer: a novel case of placental mosaicism

    Institute of Scientific and Technical Information of China (English)

    SUN Cheng-juan; ZHAO You-ping; YU Song; FAN Ling; WU Qing-qing; LI Guang-hui; ZHANG Wei-yuan

    2012-01-01

    Twin pregnancy with mosaic partial hydatidiform mole (PHM) and survival of two healthy fetuses following in vitro fertilization and embryos transfer (IVF-ET) is a rare situation and is considered a challenge for management.A 32-year-old Chinese woman conceived twin pregnancy following IVF-ET.At 22 weeks' gestation,an additional intrauterine echogenic mass with features of PHM were shown by successive ultrasound examinations.At 35 weeks'gestation,two live male infants and two placentas were delivered by caesarean section (CS).Histologic examination of the abnormal placenta confirmed mosaic PHM.Genetic study showed the abnormal placental mosaicism (expressed in molar-69XXY and normal vili-46XY),co-existing with a hypospadia new-born (46XY) in one amniotic sac.However,the other one was normal.Serial serum β-hCG levels showed a declining trend and serum 3-human chorionic gonadotropin (hCG) were undetectable at 6 months after delivery.The case demonstrated that it is possible to prolonged gestation by PHM under close surveillance during the entire pregnancy.

  12. Evaluación retrospectiva de la tasa de preñez obtenida por transferencia de embriones en diferentes cruces bovinos en el municipio de Puerto Araujo, Santander, Colombia (Retrospective analysis of pregnancy rate after embryo transfer on F1 cows.

    Directory of Open Access Journals (Sweden)

    Ariza Luis Edgardo

    2006-04-01

    Full Text Available El presente trabajo analizó los datos obtenidos en los años 1999-2001 para evaluar el mejor cruce como receptora en un programa de Transferencia de Embriones (T.E., donde se usan principalmente animales F1 como Limbrah, Simbrah, Brabon, Brangus y Girolando. Se contó con una población de 1166 receptoras transferidas, de los cruces anteriormente señalados, de las cuales 577 se realizaron con embriones en fresco y 589 con embriones congelados en glicerol o etilenglicol. Después de transferidas, se evaluó la preñez mediante palpación rectal a los 60 días; ésta información se llevó a registros donde se cruzaron las variables: tipo de cruce de la receptora, morfología y calidad embrionaria y tipo de transferencia (en fresco o congelado. A esta información se le aplicó una prueba de Chi cuadrado, la cual arrojó que las mejores receptoras para un programa de T.E., corresponden a los cruces Brabon y Girolando con 39,58% y 39,56% de preñez respectivamente; así mismo, al comparar estos dos cruces, se pudo evidenciar que el cruce Girolando presenta mayores tasas de preñez al transferirle embriones en estados de desarrollo más tempranos. Abstrac.- The aim of this work was to evaluate the effect of bovine F1 recipients (Limousin X Brahman; Simmental x Brahman; Brahman x BON; Brahman x Angus and Gyr x Holstein on a program of embryo transfer. 1166 F1 cows transferred (577 of them with fresh embryos and 589 with frozen embryos between 1999 and 2001 were evaluated according with its pregnancy rate (rectal examination at 60 days. Chi square test was made to probe association between variables as F1 employed, embryo morphology and quality and embryo treatment (fresh Vs. frozen embryos. According with this work, the best F1 recipients were Brahman x BON and Gyr x Holstein each one showing pregnancy rates of 39,58% and 39,56% respectively. Additionally, these F1 is poorly influenced by the others variables evaluated, although seems to be that

  13. Occurrence of Campylobacter in the genitals of teaser bulls maintained at an embryo transfer center

    Directory of Open Access Journals (Sweden)

    Modolo J.R.

    2000-01-01

    Full Text Available Em central de transferência de embriões, após os procedimentos de reconhecimento do cio em 37 vacas receptoras, através de quatro rufiões vasectomizados, observou-se que 83% delas apresentavam retorno ao cio e algum corrimento serofibrinoso. Nos exames bacteriológicos realizados nos lavados prepuciais dos rufiões foi isolado o Campylobacter fetus subsp. venerealis em todos, fato que, analisado associadamente com o retorno ao cio das vacas receptoras, é indicativo da ocorrência de campilobacteriose no plantel. Essa ocorrência demonstra a necessidade de medidas eficazes de planejamento de saúde animal, pela utilização de rufiões com desvio lateral do pênis. Uma vez impossibilitado o contato sexual, seria impedida a transmissão do agente durante o coito. Torna-se imperioso consignar que a prática da prevenção racional de enfermidades continua sendo o procedimento mais econômico para uma produtividade animal mais rentável.

  14. Growth hormone gene family (GH, GHR, GHRH and Pit-1) polymorphisms and its association with superovulation response, ovulation rate and embryo quality in Embryo Transfer Station (BET) of Cipelang

    OpenAIRE

    Cece Sumantri; M. Imron; Sugyono; E. Andreas; M. Restu; A.B.L. Ishak

    2011-01-01

    The decrease in fertility is considered to be the main cause of reproductive loss in dairy cattle and beef industry. Many candidate genes that play an important role in fertility and embryonic development. The purpose of this study was to detect genetic variations of the growth hormone gene family (GH|MspI, GH|AluI, GHR| AluI, GHRH|HaeIII and Pit-1|HinfI) and its association with superovulation response, ovulation, fertilization and transferable embryos rate. A total of 45 blood samples taken...

  15. Intrauterine insemination of cultured peripheral blood mononuclear cells prior to embryo transfer improves clinical outcome for patients with repeated implantation failures.

    Science.gov (United States)

    Madkour, Aicha; Bouamoud, Nouzha; Louanjli, Noureddine; Kaarouch, Ismail; Copin, Henri; Benkhalifa, Moncef; Sefrioui, Omar

    2016-02-01

    Implantation failure is a major limiting factor in assisted reproduction improvement. Dysfunction of embryo-maternal immuno-tolerance pathways may be responsible for repeated implantation failures. This fact is supported by immunotropic theory stipulating that maternal immune cells, essentially uterine CD56+ natural killer cells, are determinants of implantation success. In order to test this hypothesis, we applied endometrium immuno-modulation prior to fresh embryo transfer for patients with repeated implantation failures. Peripheral blood mononuclear cells were isolated from repeated implantation failure patients undergoing assisted reproductive technology cycles. On the day of ovulation induction, cells were isolated and then cultured for 3 days and transferred into the endometrium cavity prior to fresh embryo transfer. This immunotherapy was performed on 27 patients with repeated implantation failures and compared with another 27 patients who served as controls. Implantation and clinical pregnancy were increased significantly in the peripheral blood mononuclear cell test versus control (21.54, 44.44 vs. 8.62, 14.81%). This finding suggests a clear role for endometrium immuno-modulation and the inflammation process in implantation success. Our study showed the feasibility of intrauterine administration of autologous peripheral blood mononuclear cells as an effective therapy to improve clinical outcomes for patients with repeated implantation failures and who are undergoing in vitro fertilization cycles. PMID:25613318

  16. Developmental stage on day-5 and fragmentation rate on day-3 can influence the implantation potential of top-quality blastocysts in IVF cycles with single embryo transfer

    Directory of Open Access Journals (Sweden)

    Devroey Paul

    2007-01-01

    Full Text Available Abstract Background In IVF-ICSI cycles with single embryo transfer (SET, embryo selection for transfer is of crucial importance. The present study aimed to define which embryo parameters might be related to the implantation potential of advanced blastocysts. Methods Overall, in 203 cycles with SET, developmental characteristics of 93 implanted (group A and 110 non-implanted (group B advanced blastocysts of good quality were compared. The following developmental parameters were assessed in the two groups: normal fertilization, developmental stage on day 5, number of blastomeres on day 2 and on day 3, fragmentation rate on day 3, compaction on day 4 and cleavage pattern on day 2 and day 3. Results Expanded blastocysts compared to full blastocysts have higher implantation potential (56.5% vs. 29.3%, p 10–50% fragments on day 3 showed a significant lower implantation (29.7% than those with ≤ 10%fragments (49.4%, P = 0.03. All the other parameters analysed were comparable for the two groups. Conclusion Developmental stage on day 5 and fragmentation rate on day 3 were related to the implantation potential of advanced blastocysts and should also be taken into account in the selection of the best advanced blastocyst for transfer.

  17. Gene expression, oocyte nuclear maturation and developmental competence of bovine oocytes and embryos produced after in vivo and in vitro heat shock.

    Science.gov (United States)

    Pavani, Krishna C; Baron, Erica; Correia, Pedro; Lourenço, Joana; Bettencourt, Bruno Filipe; Sousa, Madalena; da Silva, Fernando Moreira

    2016-10-01

    Three assays were performed. In assay 1, oocytes harvested during the winter months were subjected to kinetic heat shock by stressing the oocytes at 39.5°C (HS1) or at 40.5°C (HS2) for either 6, 12, 18 or 24 h and then matured at control temperature (38.5°C). The nuclear maturation rates (NMR) of all oocytes were recorded after 24 h. In assay 2, oocytes collected year-round maturated, were implanted via in vitro fertilization (IVF) and developed for 9 days. Gene expression analysis was performed on target genes (Cx43, CDH1, DNMT1, HSPA14) with reference to the two housekeeping genes (GAPDH and SDHA) in embryos. Similarly, in assay 3, genetic analysis was performed on the embryos produced from heat-stressed oocytes (from HS1 and HS2). In assay 1, the duration of heat stress resulted in a significant decline in NMR (P CDH1 genes (P < 0.05). Targeted gene expression was aberrant in embryo development, which can provide evidence on early embryo arrest and slowed embryo development.

  18. Association between mitochondrial DNA haplotype compatibility and increased efficiency of bovine intersubspecies cloning

    Institute of Scientific and Technical Information of China (English)

    Hao Yan; Zhonghai Yan; Qingwen Ma; Fei Jiao; Shuzhen Huang; Fanyi Zeng; Yitao Zeng

    2011-01-01

    Reconstructed embryos derived from intersubspecies somatic cell nuclear transfer (SCNT) have poorer developmental potential than those from intrasubspecies SCNT. Based on our previous study that Holstein dairy bovine (HD) mitochondrial DNA (mtDNA) haplotype compatibility between donor karyoplast and recipient cytoplast is crucial for SCNT embryo development, we performed intersubspecies SCNT using HD as donor karyoplast and Luxi yellow heifer (LY) as recipient cytoplast according to mtDNA haplotypes determined by polymerase chain reactionrestriction fragment length polymorphism (PCR-RFLP) analysis. The results demonstrated that intersubspecies mtDNA homotype SCNT embryos had higher pre- and post-implantation developmental competence than intrasubspecies mtDNA heterotype embryos as well as improved blastocyst reprogramming status, including normal H3K9 dimethylation pattern and promoter hypomethylation of pluripotent genes such as Oct4 and Sox2, suggesting that intersubspecies SCNT using LY oocytes maintains HD cloning efficiency and may reprogram HD nuclei to develop into a normal cloned animal ultimately. Our results indicated that karyoplast-cytoplast interactions and mtDNA haplotype compatibility may affect bovine intersubspecies SCNT efficiency. This study on bovine intersubspecies SCNT is valuable for understanding the mechanisms of mtDNA haplotype compatibility between karyoplast and cytoplast impacting the bovine SCNT efficiency, and provides an alternative and economic resource for HD cloning.

  19. [Use of an in vitro model in bovine to evidence a functional and molecular dialogue between preimplantation embryo and oviduct epithelial cells].

    Science.gov (United States)

    Cordova, A; Perreau, C; Schmaltz-Panneau, B; Locatelli, Y; Ponsart, C; Mermillod, P

    2013-09-01

    Beyond being a pipe between ovary and uterus, the oviduct is an active player in different aspects of early reproductive processes, in particular in the transport of embryos to the site of implantation and the regulation of its early development. Different studies evidenced a communication between oviduct and early embryo at the molecular and functional levels. Since the study of these interactions is difficult in vivo, different in vitro systems have been developed to mimic the maternal milieu during early development. These systems allowed to confirm the action of the cells on the quality of early development (blastocyst rate and viability). In turn, the embryos are producing signals that are able to modify and adapt the activity of maternal cells. PMID:23958329

  20. Effects of single dose GnRH agonist as luteal support on pregnancy outcome in frozen-thawed embryo transfer cycles: an RCT

    Directory of Open Access Journals (Sweden)

    Robab Davar

    2015-08-01

    Full Text Available Background: There is no doubt that luteal phase support is essential to enhance the reproductive outcome in IVF cycles. In addition to progesterone and human chorionic gonadotropin, several studies have described GnRH agonists as luteal phase support to improve implantation rate, pregnancy rate and live birth rate, whereas other studies showed dissimilar conclusions. All of these studies have been done in fresh IVF cycles. Objective: To determine whether an additional GnRH agonist administered at the time of implantation for luteal phase support in frozen-thawed embryo transfer (FET improves the embryo developmental potential. Materials and Methods: This is a prospective controlled trial study in 200 FET cycles, patients were randomized on the day of embryo transfer into group 1 (n=100 to whom a single dose of GnRH agonist (0.1 mg triptorelin was administered three days after transfer and group 2 (n=100, who did not receive agonist. Both groups received daily vaginal progesterone suppositories plus estradiol valerate 6 mg daily. Primary outcome measure was clinical pregnancy rate. Secondary outcome measures were implantation rate, chemical, ongoing pregnancy rate and abortion rate. Results: A total of 200 FET cycles were analyzed. Demographic data and embryo quality were comparable between two groups. No statistically significant difference in clinical and ongoing pregnancy rates was observed between the two groups (26% versus 21%, p=0.40 and 21% versus 17%, p=0.37, respectively. Conclusion: Administration of a subcutaneous GnRH agonist at the time of implantation does not increase clinical or ongoing pregnancy.

  1. Förster resonance energy transfer between pyrene and bovine serum albumin: Effect of the hydrophobic pockets of cyclodextrins

    Science.gov (United States)

    Maity, Arnab; Mukherjee, Puspal; Das, Tarasankar; Ghosh, Prasun; Purkayastha, Pradipta

    The phenomenon of Förster resonance energy transfer (FRET) between pyrene and bovine serum albumin (BSA) protein in presence of cyclodextrins (CDs) is explored in the present work. CDs provide hydrophobic environment and thus the aromatic molecules get encapsulated in them depending on the relative size and space. In this work we revealed that along with pyrene monomer, the side chains of amino acids in BSA can get trapped partly in the hydrophobic cavities of CDs if space permits. While being encapsulated by β-CD as pyrene monomer, it can interact with the BSA tryptophan moiety exposed toward the aqueous environment to form a dimer through π-π interaction. This, in turn, affects the energy transfer process by reducing the efficiency. On the other hand, pyrene excimer gets encapsulated in a γ-CD molecule due to availability of enough space. The excimer shows a new band at a higher wavelength. This further reduces FRET efficiency due to scarcity of acceptor for the tryptophan moieties in BSA.

  2. Comparison between lignocaine hydrochloride and ropivacaine hydrochloride as lumbosacral epidural anaesthetic agents in goats undergoing laparoscopy assisted embryo transfer

    Directory of Open Access Journals (Sweden)

    Anubhav Khajuria

    2014-10-01

    Full Text Available Goats (n=12 undergoing laparoscopy assisted embryo transfer were randomly allotted to two groups (I and II and injected lignocaine hydrochloride (4mg/kg or ropivacaine hydrochloride (1mg/kg at the lumbosacral epidural space. The animals were held with raised hind quarters for first three minutes following injection. Immediately after induction of regional anaesthesia, they were restrained in dorsal recumbency in Trendelenburg position in a cradle. Laparoscopy was performed after creating pneumoperitoneum using filtered room air. The mean (± S.E induction time in animals of group I was significantly shorter (5.33 ± 0.61 min than those belonging to group II (12.66 ±1.99 min. Complete analgesia developed throughout the hind quarters and abdomen for 30 min and 60 min in group I and II animal’s respectively. Unlike animals of group I, group II goats continued to show moderate analgesia for 180 minutes. The motor activity returned after a lapse of 130.00 ± 12.64 min and 405.00 ± 46.31 min respectively. Occasional vocalization and struggling was noticed in two goats one from each group irrespective of the surgical manipulations during laparoscopy. The rectal temperature and respiration rates showed only non-significant increase, but the heart rate values were significantly higher (P < 0.5 up to 150 min in animals of both the groups when compared to their baseline values. From this study, it was concluded that both anaesthetic agents produced satisfactory regional anaesthesia in goats undergoing laparoscopy. However, considering the very long delay in regaining the hind limb motor activity, the use of ropivacaine may not be recommended for this purpose. Supplementation of sedative/tranquilizer with lumbosacral epidural anaesthesia needs evaluation.

  3. In vitro embryo outgrowth is a bioassay of in vivo embryo implantation and development

    OpenAIRE

    Binder, Natalie K.; Hannan, Natalie J; David K. Gardner

    2015-01-01

    Objective: To determine the efficacy of embryo outgrowth on fibronectin as a low cost, high throughput alternative to embryo transfer to model embryo attachment and the initial stages of implantation. Methods: Following in vitro embryo culture, embryo quality was assessed via embryo transfer or embryo outgrowth with metabolic assessment. Results: This study shows that blastocysts attach to fibronectin at the same rate that they implant in vivo, and that the carbohydrate utilisation of e...

  4. Bed rest following embryo transfer might negatively affect the outcome of IVF/ICSI: a systematic review and meta-analysis.

    Science.gov (United States)

    Craciunas, Laurentiu; Tsampras, Nikolaos

    2016-04-01

    The majority of patients undergoing in vitro fertilization (IVF) and intracytoplasmatic sperm injection (ICSI) treatment will reach the stage of embryo transfer (ET), but only a small proportion of transferred embryos implant. Bed rest following ET has been recommended as a way to prevent embryo expulsion by gravity. We performed a systematic review and meta-analysis of randomized controlled trials (RCTs) published prior to May 2014 reporting the effect of bed rest following ET, and irrespective of language, country of origin, blinding or sample size. Four RCTs, including 757 women met the inclusion criteria. Bed rest following ET did not improve clinical pregnancy and live birth rates, but reduced the implantation rate. The quality of the trials included was moderate because of attrition bias and possible reporting bias. The findings of this systematic review and meta-analysis are concordant with previously published literature and suggest that bed rest is not beneficial following ET. Moreover, it might negatively affect the outcome of IVF/ICSI cycles via stress/anxiety mechanisms. PMID:26986834

  5. Bed rest following embryo transfer might negatively affect the outcome of IVF/ICSI: a systematic review and meta-analysis.

    Science.gov (United States)

    Craciunas, Laurentiu; Tsampras, Nikolaos

    2016-04-01

    The majority of patients undergoing in vitro fertilization (IVF) and intracytoplasmatic sperm injection (ICSI) treatment will reach the stage of embryo transfer (ET), but only a small proportion of transferred embryos implant. Bed rest following ET has been recommended as a way to prevent embryo expulsion by gravity. We performed a systematic review and meta-analysis of randomized controlled trials (RCTs) published prior to May 2014 reporting the effect of bed rest following ET, and irrespective of language, country of origin, blinding or sample size. Four RCTs, including 757 women met the inclusion criteria. Bed rest following ET did not improve clinical pregnancy and live birth rates, but reduced the implantation rate. The quality of the trials included was moderate because of attrition bias and possible reporting bias. The findings of this systematic review and meta-analysis are concordant with previously published literature and suggest that bed rest is not beneficial following ET. Moreover, it might negatively affect the outcome of IVF/ICSI cycles via stress/anxiety mechanisms.

  6. Simplification of bovine somatic cell nuclear transfer by application of a zona-free manipulation technique

    DEFF Research Database (Denmark)

    Booth, P J; Tan, S J; Reipurth, R;

    2001-01-01

    Contemporary nuclear transfer techniques often require the involvement of skilled personnel and extended periods of micromanipulation. Here, we present details of the development of a nuclear transfer technique for somatic cells that is both simpler and faster than traditional methods......, and (c) establish any potential embryotoxic effects of PHA-P. The initial data indicated that, of calcium ionophore A23187, ionomycin, and electropulse treatments as primary activation agents, the two former were equally efficient even with reduced exposure times. WOW-culture of zona-free versus zona.......8% of cultured oocytes). Subsequent application of the optimized technique for nuclear transfer using nine different granulosa cell primary cultures (cultured in 0.5% serum for 5-12 days) generated 37.6 +/- 3.9% (11 replicates; range, 16.4-58.1 blastocysts per successfully fused and surviving reconstructed...

  7. Vitrificación de ovocitos bovinos y su uso en el desarrollo partenogenético de embriones Vitrification of bovine oocytes and its use in the parthenogenetic development of embryos

    Directory of Open Access Journals (Sweden)

    J Ruiz

    2010-01-01

    Full Text Available El objetivo de este trabajo fue evaluar el efecto de la vitrificación en la viabilidad de ovocitos activados químicamente para la producción de embriones partenogenéticos bovinos. Ovocitos bovinos aspirados de ovarios obtenidos en el matadero fueron madurados in vitro por 20-22 horas y se distribuyeron en los siguientes grupos. I (n=76: ovocitos vitrificados/descongelados, II (n=119: ovocitos expuestos a crioprotectantes sin vitrificación y III (n=142: ovocitos control. Los ovocitos fueron vitrificados en microgotas sobre un papel de aluminio preenfriado flotando en nitrógeno líquido, utilizando una solución de equilibrio con 4% de etilenglicol y una solución de vitrificación con 35% de etilenglicol 5% de polivinilpirrolidona y 0,4 M de trehalosa. Las microgotas vitrificadas fueron almacenadas en nitrógeno líquido y fueron descongeladas 1-3 días después del almacenamiento. Los tres grupos de ovocitos se activaron partenogenéticamente por exposición de 4 minutos a 5 µM de ionomicina de Ca a temperatura ambiente seguido de una incubación por 5 horas en 6-dimetilaminopurina a 38,5 ºC en una atmósfera húmeda con 5% CO2. Los embriones se cultivaron en medio mSOF durante 8-9 días. Las tasas de ovocitos sobrevivientes fueron 55,1% y 93,7% para ovocitos vitrificados/descongelados (I y expuestos (II respectivamente. Las tasas de segmentación de 55,3%, 72,3% y 74,6%, y de desarrollo hasta blastocistos fueron 7,1%, 17,4% y 21,7% en los grupos I, II y III respectivamente. Estos resultados demuestran que la técnica de vitrificación ha quedado establecida y permite la producción de embriones partenogenéticos bovinos.The aim of this study was to evaluate vitrification effects on the viability of chemically activated oocytes in order to produce parthenogenetic bovine embryos. Bovine oocytes retrieved from ovaries obtained in a slaughterhouse were matured in vitro for 20-22 hours and then assigned to the following groups: I (n=76

  8. Decreased Sperm Motility Retarded ICSI Fertilization Rate in Severe Oligozoospermia but Good-Quality Embryo Transfer Had Achieved the Prospective Clinical Outcomes

    Science.gov (United States)

    Zheng, Jufeng; Lu, Yongning; Qu, Xianqin; Wang, Peng; Zhao, Luiwen; Gao, Minzhi; Shi, Huijuan; Jin, Xingliang

    2016-01-01

    Introduction Spermatozoa motility is the critical parameter to affect the treatment outcomes during assisted reproductive technologies (ART), but its reproductive capability remains a little informed in condition of severe male factor infertility. This retrospective cohort study aimed to evaluate the effects of reduced sperm motility on the embryological and clinical outcomes in intra-cytoplasmic sperm injection (ICSI) treatment of severe oligozoospermia. Patients and Methods 966 cycles (812 couples) of severe oligozoospermia diagnosed by spermatozoa count ≤ 5 × 106/mL and motile spermatozoa ≤ 2 × 106/mL were divided into four groups in according to the number of motile spermatozoa in one ejaculate on the day of oocyte retrieval (Group B—E). The control (Group A) was 188 cycles of moderate oligozoospermia with spermatozoa count > 5 × 106/mL and motile spermatozoa > 2 × 106/mL. All female partners were younger than 35 years of age. Logistic regression analyzed embryological outcomes (the rates of fertilization, cleavage and good-quality embryo) and clinical outcomes (the rates of pregnancy, implantation, early miscarriage and live birth). Quality of embryo transfer (ET) was divided into three classes as continuous factor to test the effects of embryo quality on clinical outcomes. Results The reduction in the number of motile sperm in four groups of severe oligozoospermia gave rise to comparable inability of the fertilization (p < 0.001) and a decreased rate of good-quality embryo at Day 3 (p < 0.001) by compared to the control. The cleavage rate of the derived zygotes was similar to the control. ET classes significantly affected the clinical outcomes (p < 0.001). Class I ET gave rise to similar rates of clinical outcomes between five groups, but Class II and Class III ET retarded the rates of pregnancy, implantation and live birth and this particularly occurred in Group C, D and E. The rate of early miscarriage was not comparably different between groups

  9. Morphological and Gene Expression Changes in Cattle Embryos from Hatched Blastocyst to Early Gastrulation Stages after Transfer of In Vitro Produced Embryos.

    Directory of Open Access Journals (Sweden)

    Jessica van Leeuwen

    Full Text Available A detailed morphological staging system for cattle embryos at stages following blastocyst hatching and preceding gastrulation is presented here together with spatiotemporal mapping of gene expression for BMP4, BRACHYURY, CERBERUS1 (CER1, CRIPTO, EOMESODERMIN, FURIN and NODAL. Five stages are defined based on distinct developmental events. The first of these is the differentiation of the visceral hypoblast underlying the epiblast, from the parietal hypoblast underlying the mural trophoblast. The second concerns the formation of an asymmetrically positioned, morphologically recognisable region within the visceral hypoblast that is marked by the presence of CER1 and absence of BMP4 expression. We have termed this the anterior visceral hypoblast or AVH. Intra-epiblast cavity formation and the disappearance of the polar trophoblast overlying the epiblast (Rauber's layer have been mapped in relation to AVH formation. The third chronological event involves the transition of the epiblast into the embryonic ectoderm with concomitant onset of posterior NODAL, EOMES and BRACHYURY expression. Lastly, gastrulation commences as the posterior medial embryonic ectoderm layer thickens to form the primitive streak and cells ingress between the embryonic ectoderm and hypoblast. At this stage a novel domain of CER1 expression is seen whereas the AVH disappears. Comparison with the mouse reveals that while gene expression patterns at the onset of gastrulation are well conserved, asymmetry establishment, which relies on extraembryonic tissues such as the hypoblast and trophoblast, has diverged in terms of both gene expression and morphology.

  10. Effect of freeze-thaw cycles on load transfer between the biomineral and collagen phases in bovine dentin

    International Nuclear Information System (INIS)

    Stabilization of biological materials by freezing is widespread in the fields of medicine and biomaterials research and yet, in the case of hard biomaterials such as dentin, there is not a good understanding of how such treatments might affect the mechanical properties. The freezing and thawing may have a number of different effects on dentin including formation of cracks in the microstructure and denaturation of the collagen. Using high-energy synchrotron X-ray diffraction, the apparent moduli of bovine dentin samples were measured before and after various numbers of freeze-thaw cycles. It was determined that repeated freezing and thawing has no measurable effect on the hydroxyapatite or fibrillar apparent moduli up to 10 cycles. This confirms that the use of low temperature storage for stabilization of dentin is reasonable in cases where stiffness is a property of importance. Highlights: → Studied the effect of freezing on the load transfer of HAP and fibrils in dentin. → X-ray scattering measured HAP and fibril apparent moduli vs. freezing cycles. → Apparent moduli did not vary significantly between 0 and 10 freeze thaw cycles. → Residual strains imply no widespread cracking due to volumetric expansion of water. → Dentin can be freeze-thawed with no significant effects on elastic properties.

  11. Obesity does not aggravate vitrification injury in mouse embryos: a prospective study

    Directory of Open Access Journals (Sweden)

    Ma Wenhong

    2012-08-01

    Full Text Available Abstract Background Obesity is associated with poor reproductive outcomes, but few reports have examined thawed embryo transfer in obese women. Many studies have shown that increased lipid accumulation aggravates vitrification injury in porcine and bovine embryos, but oocytes of these species have high lipid contents (63 ng and 161 ng, respectively. Almost nothing is known about lipids in human oocytes except that these cells are anecdotally known to be relatively lipid poor. In this regard, human oocytes are considered to be similar to those of the mouse, which contain approximately 4 ng total lipids/oocyte. To date, no available data show the impact of obesity on vitrification in mouse embryos. The aim of this study was to establish a murine model of maternal diet-induced obesity and to characterize the effect of obesity on vitrification by investigating the survival rate and embryo developmental competence after thawing. Methods Prospective comparisons were performed between six–eight-cell embryos from obese and normal-weight mice and between fresh and vitrified embryos. Female C57BL/6 mice were fed standard rodent chow (normal-weight group or a high-fat diet (obese group for 6 weeks. The mice were mated, zygotes were collected from oviducts and cultured for 3 days, and six–eight-cell embryos were then selected to assess lipid content in fresh embryos and to evaluate differences in apoptosis, survival, and development rates in response to vitrification. Results In fresh embryos from obese mice, the lipid content (0.044 vs 0.030, Pvs.9.3%, Pvs. 93.1%, P Conclusions This study demonstrated that differences in survival and developmental rates between embryos from obese and normal-weight mice were eliminated after vitrification. Thus, maternal obesity does not aggravate vitrification injury, but obesity alone greatly impairs pre-implantation embryo survival and development.

  12. Evaluation of reciprocal differences in Bos indicus x Bos taurus backcross calves produced through embryo transfer: II. Postweaning, carcass, and meat traits.

    Science.gov (United States)

    Amen, T S; Herring, A D; Sanders, J O; Gill, C A

    2007-02-01

    Angus (A) x Bos indicus (B; Brahman or Nellore) reciprocal backcross, embryo transfer calves belonging to 28 full-sib families were evaluated for differences in feedyard initial BW, feedyard final BW, carcass weight, LM area, adjusted fat thickness, intramuscular fat, and Warner-Bratzler shear force. Two methods of analysis were investigated; method I made no distinction between how the F(1) parents were produced, whereas method II distinguished the 2 types of F(1) parents (AB vs. BA, corresponding to A x B vs. B x A, respectively). No significant reciprocal differences for these weight and carcass traits were detected under method I analyses, although the same trend existed for subsequent BW rankings as for birth weight and weaning weight. For each weight phase, the cross that involved a larger proportion of B in the sire in relation to the amount in the dam (F(1) x A and B x F(1)) ranked heavier than the respective reciprocal cross (A x F(1) and F(1) x B). As a whole, A backcross calves had larger (P carcass (P < 0.001), and had larger LM area (P < 0.05) with less adjusted fat (P < 0.001). No difference existed between the sexes for Warner-Bratzler shear force or marbling. No interactions involving sex, sire type, and dam type were observed for any of these traits. The results were similar under methods I and II analyses, with the exception that a significant sire type x dam type interaction was observed for initial feedyard BW. Results from this study suggest that for weight-related traits, both the breed constitution of the embryo transfer calf and the cross that produces the calf play an important role in its ultimate performance for B crossbred calves. For body composition and meat-related traits, it appears that the breed makeup of the embryo transfer calf itself is more important to animal performance than the specific cross used to produce the calf.

  13. Evaluation of reciprocal differences in Bos indicus x Bos taurus backcross calves produced through embryo transfer: II. Postweaning, carcass, and meat traits.

    Science.gov (United States)

    Amen, T S; Herring, A D; Sanders, J O; Gill, C A

    2007-02-01

    Angus (A) x Bos indicus (B; Brahman or Nellore) reciprocal backcross, embryo transfer calves belonging to 28 full-sib families were evaluated for differences in feedyard initial BW, feedyard final BW, carcass weight, LM area, adjusted fat thickness, intramuscular fat, and Warner-Bratzler shear force. Two methods of analysis were investigated; method I made no distinction between how the F(1) parents were produced, whereas method II distinguished the 2 types of F(1) parents (AB vs. BA, corresponding to A x B vs. B x A, respectively). No significant reciprocal differences for these weight and carcass traits were detected under method I analyses, although the same trend existed for subsequent BW rankings as for birth weight and weaning weight. For each weight phase, the cross that involved a larger proportion of B in the sire in relation to the amount in the dam (F(1) x A and B x F(1)) ranked heavier than the respective reciprocal cross (A x F(1) and F(1) x B). As a whole, A backcross calves had larger (P x dam type interaction was observed for initial feedyard BW. Results from this study suggest that for weight-related traits, both the breed constitution of the embryo transfer calf and the cross that produces the calf play an important role in its ultimate performance for B crossbred calves. For body composition and meat-related traits, it appears that the breed makeup of the embryo transfer calf itself is more important to animal performance than the specific cross used to produce the calf. PMID:17235022

  14. In vitro fertilization in Japan — Early days of in vitro fertilization and embryo transfer and future prospects for assisted reproductive technology —

    Science.gov (United States)

    SUZUKI, Masakuni

    2014-01-01

    Assisted reproductive technology (ART) such as in vitro fertilization (IVF) and embryo transfer (ET) has been essential in the treatment of infertility. The world’s first IVF-ET baby was born in 1978 based on the technique developed by Dr. Robert Edwards and Dr. Patrick Steptoe.1) In Japan, the first IVF-ET birth was reported in 1983 by Prof. Masakuni Suzuki at Tohoku University School of Medicine.2,3) IVF-ET is a procedure used to achieve pregnancy that consists of extracting oocytes from an infertile woman, fertilizing them in vitro, and transferring fertilized eggs into the patient’s uterine cavity (Fig. 1). Since the first report of successful IVF-ET, numerous techniques related to ART, such as cryopreservation of oocytes and embryos, gamete intrafallopian transfer (GIFT), and microinsemination, have been developed and refined (Table 1). Herein we describe the history of basic research in IVF-ET that led to human applications, how the birth of the first IVF-ET baby was achieved in Japan, the current status of ART in Japan, issues related to ART, and future prospects for ART. PMID:24814992

  15. In vitro fertilization in Japan - early days of in vitro fertilization and embryo transfer and future prospects for assisted reproductive technology.

    Science.gov (United States)

    Suzuki, Masakuni

    2014-01-01

    Assisted reproductive technology (ART) such as in vitro fertilization (IVF) and embryo transfer (ET) has been essential in the treatment of infertility. The world's first IVF-ET baby was born in 1978 based on the technique developed by Dr. Robert Edwards and Dr. Patrick Steptoe. In Japan, the first IVF-ET birth was reported in 1983 by Prof. Masakuni Suzuki at Tohoku University School of Medicine. IVF-ET is a procedure used to achieve pregnancy that consists of extracting oocytes from an infertile woman, fertilizing them in vitro, and transferring fertilized eggs into the patient's uterine cavity (Fig. 1). Since the first report of successful IVF-ET, numerous techniques related to ART, such as cryopreservation of oocytes and embryos, gamete intrafallopian transfer (GIFT), and microinsemination, have been developed and refined (Table 1). Herein we describe the history of basic research in IVF-ET that led to human applications, how the birth of the first IVF-ET baby was achieved in Japan, the current status of ART in Japan, issues related to ART, and future prospects for ART. PMID:24814992

  16. 人类配子及胚胎转移伦理问题的探讨%Discussion on the Transfer of Human Gamete and Embryos

    Institute of Scientific and Technical Information of China (English)

    李红英; 葛建一; 侯建全; 徐溢涛; 张拥军; 陈瑞华

    2012-01-01

    This paper analyzed the reasons for the transfer of cryopreservation gamete and embryos, such as information factor, social movement of personnel, surrogacy requirements, scientific research requirements, business or personal purpose, and so forth. Based on the clinical required transfer and scientific research required transfer, the proposals of Ethical Committee were made. This paper also proposed that, should set up and perfect the management mechanism for cryopreservation gametes and embryos, fully play the role of Ethical Committee, provide full understanding and so forth, in order to eliminate some violations.%通过对要求进行冻存配子、胚胎转移的原因进行分析,即信息知讯、社会人员流动、代孕要求、相关科研需要、商务或个人目的等.从医疗需要的转移和科研需要的转移方面给出伦理委员会的指导性意见,并提出建立健全冻存配子、胚胎的管理机制,充分发挥伦理委员会职能,提高全员认识等建议,以杜绝一些违规行为的发生.

  17. In vitro fertilization in Japan — Early days of in vitro fertilization and embryo transfer and future prospects for assisted reproductive technology —

    OpenAIRE

    SUZUKI, Masakuni

    2014-01-01

    Assisted reproductive technology (ART) such as in vitro fertilization (IVF) and embryo transfer (ET) has been essential in the treatment of infertility. The world’s first IVF-ET baby was born in 1978 based on the technique developed by Dr. Robert Edwards and Dr. Patrick Steptoe. 1) In Japan, the first IVF-ET birth was reported in 1983 by Prof. Masakuni Suzuki at Tohoku University School of Medicine. 2,3) IVF-ET is a procedure used to achieve pregnancy that consists of extracting oocytes from ...

  18. A Case of Unicornuate Uterus with Atypical Located Hyperstimulated Ovary after in Vitro Fertilization Pre-Embryo Transfer (IVF-ET

    Directory of Open Access Journals (Sweden)

    Mariya Angelova Angelova

    2015-06-01

    Full Text Available The authors describe a case of a congenital Mullerian anomaly, uterus unicornis with missing right fallopian tube. An in Vitro Fertilization Pre-Embryo Transfer (IVF-ET procedure was done and presently is known that the patient has left fallopian tube and left ovary, two kidneys, and right ovary is missing. No diagnostic laparoscopy and hysteroscopy were done, only hysterosalpingography (HSG before the IVF procedure.  Several days after the follicular puncture of the left ovary the patient was urgently admitted to the hospital for specialized gynaecology in Varna. Transabdominal ultrasonography showed right ovary atypically located immediately next to the liver and with emerging theca-lutein cysts.

  19. Resurgence of Minimal Stimulation In Vitro Fertilization with A Protocol Consisting of Gonadotropin Releasing Hormone-Agonist Trigger and Vitrified-Thawed Embryo Transfer

    Directory of Open Access Journals (Sweden)

    Zhang John

    2016-07-01

    Full Text Available Minimal stimulation in vitro fertilization (mini-IVF consists of a gentle controlled ovarian stimulation that aims to produce a maximum of five to six oocytes. There is a misbelief that mini-IVF severely compromises pregnancy and live birth rates. An appraisal of the literature pertaining to studies on mini-IVF protocols was performed. The advantages of minimal stimulation protocols are reported here with a focus on the use of clomiphene citrate (CC, gonadotropin releasing hormone (GnRH ago- nist trigger for oocyte maturation, and freeze-all embryo strategy. Literature review and the author’s own center data suggest that minimal ovarian stimulation protocols with GnRH agonist trigger and freeze-all embryo strategy along with single embryo transfer produce a reasonable clinical pregnancy and live birth rates in both good and poor responders. Additionally, mini-IVF offers numerous advantages such as: i. Reduction in cost and stress with fewer office visits, needle sticks, and ultrasounds, and ii. Reduction in the incidence of ovarian hyperstimulation syndrome (OHSS. Mini-IVF is re-emerging as a solution for some of the problems associated with conventional IVF, such as OHSS, cost, and patient discomfort.

  20. Gestation-related gene expression and protein localization in endometrial tissue of Suffolk and Cheviot ewes at gestation Day 19, after transfer of Suffolk or Cheviot embryos.

    Science.gov (United States)

    Sequeira, M; Pain, S J; de Brun, V; Meikle, A; Kenyon, P R; Blair, H T

    2016-10-01

    The objective of this study was to investigate the gene expression of progesterone and estrogen receptor α (PR, ERα), insulin-like growth factor (IGF) 1, IGF-2, their receptor (IGFR1), IGF-binding proteins (BP) 1 to 6, insulin receptor, adiponectin receptors (AdipoR1/2), cyclooxygenase 2 (PTGS2), mucin 1 and to localize PR, ERα, IGF-1, IGFR1, PTGS2, and proliferating cellular nuclear antigen (PCNA) in the endometrium of pregnant (Day 19) Suffolk and Cheviot ewes carrying Suffolk and Cheviot embryos transferred within and reciprocally between breeds. Gene expression was determined by real-time quantitative polymerase chain reaction (RT-qPCR), and antigen determination was measured by immunohistochemistry in the luminal epithelium (LE), superficial and deep glands (SG, DG, respectively) and superficial and deep stroma. Gene expression of PR, IGF-1, IGFBP2, and IGFBP5 was higher in Suffolk than that in Cheviot ewes (P interaction between ewe and embryo breed affected PTGS2 staining (P protein expression in the endometrium of Suffolk and Cheviot ewes is affected by both ewe and embryo breed at Day 19 of pregnancy. PMID:27325575

  1. Efeito do número da passagem e do gênero das células doadoras de núcleo no desenvolvimento de bovinos produzidos por transferência nuclear Effect of culture time and gender of nuclei donor cells on bovine development produced by nuclear transfer

    Directory of Open Access Journals (Sweden)

    Giovana Krempel Fonseca Merighe

    2010-10-01

    Full Text Available Objetivou-se com este trabalho avaliar o efeito do número da passagem e do sexo das células doadoras de núcleo no desenvolvimento embrionário e fetal após transferência nuclear. Para isso, oócitos bovinos foram maturados, enucleados e reconstruídos com células somáticas de animal adulto. Após a fusão e ativação química, os zigotos reconstituídos foram cultivados em Charles Rosenkranz 2 (CR2 com monocamada de células da granulosa a 38,8ºC em atmosfera umidificada a 5% de CO2 em ar, durante sete dias, e transferidos para receptoras sincronizadas. As taxas de clivagem e desenvolvimento a blastocisto de embriões reconstruídos com células cultivadas por tempo maior foram inferiores às obtidas com os demais tempos de cultivo. Além disso, os blastocistos produzidos não resultaram no desenvolvimento de uma gestação a termo. Embora a taxa de clivagem em embriões fêmeas tenha sido maior, o número de embriões que atingiram o estádio de blastocisto foi maior nos embriões machos. No período gestacional, fêmeas apresentaram maior taxa de aborto entre 90 e 120 dias de gestação. Esses resultados indicam que células doadoras de núcleos cultivados por longos períodos dificultam a produção de blastocistos e aumentam as chances de perdas durante a gestação. Embriões clonados machos têm maior competência para se desenvolver a blastocisto e resultam em menor taxa de perda gestacional.The objective of this study was to evaluate the effects of culture time and sex of nuclei donor cells on embryo and fetal development after nuclear transfer. Thus, bovine oocytes were matured, enucleated and reconstructed with somatic cells from an adult animal. After fusion and chemical activation, the reconstituted zygotes were cultured in Charles Rosenkranz 2 (CR2 on a granular monolayer cell at 38.8ºC in a humidified atmosphere 5% CO2 in air for seven days, and transferred to synchronized receptors. Cleavage rates and development to

  2. Production of transgenic cashmere goat embryos expressing red fluorescent protein and containing IGF1 hair-follicle-cell specific expression cassette by somatic cell nuclear transfer

    Institute of Scientific and Technical Information of China (English)

    BOU; ShorGan

    2009-01-01

    In the present study, cashmere goat fetal fibroblasts were transfected with pCDsR-KI, a hair-follicle-cell specific expression vector for insulin-like growth factor 1 (IGF1) that contains two markers for selection (red fluorescent protein gene and neomycin resistant gene). The transgenic fibroblasts cell lines were obtained after G418 selection. Prior to the somatic cell nuclear transfer (SCNT), the maturation rate of caprine cumulus oocytes complexes (COCs) was optimized to an in vitro maturation time of 18 h. Parthenogenetic ooctyes were used as a model to investigate the effect of two activation methods, one with calcium ionophore IA23187 plus 6-DMAP and the other with ethanol plus 6-DMAP. The cleavage rates after 48 h were respectively 88.7% and 86.4%, with no significant difference (P>0.05). There was no significant difference between the cleavage rate and the blastocyst rate in two different media (SO- Faa and CR1aa; 86.3% vs 83.9%, P>0.05 and 23.1% vs 17.2%,P>0.05). The fusion rate of a 190 V/mm group (62.4%) was significantly higher than 130 V/mm (32.8%) and 200 V/mm (42.9%), groups (P<0.05). After transgenic somatic cell nuclear transfer (TSCNT) manipulation, 203 reconstructed embryos were obtained in which the cleavage rate after in vitro development (IVD) for 48 h was 79.3% (161/203). The blastocyst rate after IVD for 7 to 9 d was 15.3% (31/203). There were 17 embryos out of 31 strongly ex- pressing red fluorescence. Two of the red fluorescent blastocysts were randomly selected to identify transgene by polymerase chain reaction. Both were positive. These results showed that: (i) RFP and Neor genes were correctly expressed indicating that transgenic somatic cell lines and positive trans- genic embryos were obtained; (ii) one more selection at the blastocyst stage was necessary although the donor cells were transgenic positive, because only partially transgenic embryos expressing red fluorescence were obtained; and (iii) through TSCNT manipulation and

  3. Production of transgenic cashmere goat embryos expressing red fluorescent protein and containing IGF1 hair-follicle-cell specific expression cassette by somatic cell nuclear transfer

    Institute of Scientific and Technical Information of China (English)

    GUO XuDong; YANG DongShan; Ao XuDong; WU Xia; LI GuangPeng; WANG LingLing; BAO MingTao; XUE Lian; BOU ShorGan

    2009-01-01

    In the present study, cashmere goat fetal flbroblasta were transfected with pCDsR-KI, a hair-follicle-cell specific expression vector for insulin-like growth factor 1(IGF1) that contains two markers for selection (red fluorescent protein gene and neomycin resistant gene). The transgenic fibroblasta cell lines were obtained after G418 selection. Prior to the somatic cell nuclear transfer (SCNT), the maturation rate of caprine cumulus oocytes complexes (COCs) was optimized to an in vitro maturation time of 18 h.Parthenogenetic ooctyes were used as a model to Investigate the effect of two activation methods, one with calcium ionophore IA23187 plus 6-DMAP and the other with ethanol plus 6-DMAP. The cleavage rates after 48 h were respectively 88.7% and 86.4%, with no significant difference (P>0.05). There was no significant difference between the cleavage rate and the blastocyst rate in two different media (SO-Faa and CR1aa; 86.3% va 83.9%, P>0.05 and 23.1% vs 17.2%, P>0.05). The fusion rate of a 190 V/mm group (62.4%) was significantly higher than 130 V/mm (32.8%) and 200 V/mm (42.9%), groups (P<0.05).After transgenic somatic cell nuclear transfer (TSCNT) manipulation, 203 reconstructed embryos were obtained in which the cleavage rate after in vitro development (IVD) for 48 h was 79.3% (161/203). The blastocyst rate after IVD for 7 to 9 d was 15.3% (31/203). There were 17 embryos out of 31 strongly ex-pressing red fluorescence. Two of the red fluorescent blastocysta were randomly selected to identify transgene by polymeraee chain reaction. Both were positive. These results showed that: (i) RFP and Neo genes were correctly expressed indicating that transgenlc somatic cell lines and positive trans-genic embryos were obtained; (ii) one more selection at the blastocyst stage was necessary although the donor cells were transgenic positive, because only partially transgenic embryos expressing red fluorescence were obtained; and (iii) through TSCNT manipulation and

  4. Efeito de diferentes meios de cultivo no desenvolvimento e proporção do sexo de embriões bovinos produzidos in vitro Effect of different culture media on development and sex ratio of bovine embryos fertilized in vitro

    Directory of Open Access Journals (Sweden)

    S.G.T. Gilardi

    2004-10-01

    Full Text Available Avaliou-se o efeito da suplementação de meios de cultivo sobre o desenvolvimento e proporção do sexo de embriões bovinos fertilizados in vitro. Complexos cumulus-oócitos obtidos de ovários de matadouro foram maturados e fertilizados in vitro. Os zigotos (n= 484 foram distribuídos aleatoriamente em meio CR2aa, contendo soro fetal bovino (SFB (T1, albumina sérica bovina (BSA (T2 ou BSA mais insulina:transferrina:selênio e vitaminas (BSA+ (T3, no cultivo embrionário in vitro, a uma atmosfera de 5% CO2 a 38,8ºC em ar. A taxa de clivagem foi observada 72-76 horas pós-fertilização (PF e a taxa de blastocistos com sete e oito dias PF. Os blastocistos (n= 63 foram sexados pela técnica de reação em cadeia de polimerase. A taxa de clivagem em T2 foi maior (P0,05 entre T2 e T3, porém menor (P0,05 entre os tratamentos. O T1 influenciou o desenvolvimento de blastocistos, mas não teve efeito sobre a proporção do sexo.The effect of culture media on the development and on the sex ratio of bovine embryos fertilized in vitro was studied. Cumulus oocyte-complexes from slaughterhouse ovaries were matured and fertilized in vitro. Zygotes (n= 484 were randomly allotted to different culture media and cultured with their cumulus cells in CR2aa medium and an atmosphere of 5% CO2 in air at 38.8ºC. The fetal calf serum (FCS, bovine seric albumin (BSA or BSA plus insulin:transferrin:selenium and vitamins (BSA+ supplementation effect on embryo culture was evaluated. Cleavage rate was assessed at 72-76h post-fertilization (PF and blastocyst rate on days 7 and 8 PF. The blastocysts (n= 63 were also sexed using polymerase chain reaction. Cleavage rate for BSA medium supplemented was higher (P0.05, but lower (P<0.01 than FCS. Culture medium FCS supplemented affected blastocyst development but not the sex ratio.

  5. Effects of exposure to the xenoestrogen octylphenol and subsequent transfer to clean water on liver and gonad ultrastructure during early development of Zoarces viviparus embryos.

    Science.gov (United States)

    Jespersen, Ase; Rasmussen, Tina H; Hirche, Majken; Sørensen, Kristine J K; Korsgaard, Bodil

    2010-08-01

    Female eelpouts (Zoarces viviparus L.) are exposed during early pregnancy to nominal concentrations of 100 microg/L of 4-tert-octylphenol (OP) or 0.5 microg/L of 17beta-estradiol (E2). Effects on maternal metabolism and on liver and gonad development in embryos were examined and compared with controls (C) during exposure and after transfer to clean water (depuration). In the mother fish, significantly higher concentrations of plasma vitellogenin (vtg) and calcium were found in the two exposed groups, when compared with the C group after exposure and depuration. When compared, however, with the respective values after exposure, vtg had decreased significantly after depuration. The hepatosomatic index was normalized after depuration. In both exposed groups, the hepatocytes were rounded and not distinctly polygonal as in the controls. The amount of glycogen was considerably less while the number of mitochondria increased, and the rER significantly proliferated after exposure as well as after depuration. The gonads of nine of more than 28 embryos in the group treated with OP exhibited a number of abnormalities as compared with the normal gonad development in both sexes. Feminization of the male gonads in the exposed specimens and a number of histopathological features were observed in all the abnormal gonads, whereas reliable male features, such as formation of seminiferous tubules or spermioduct, were not observed. This study showed that 4t-tert-OP and 17beta-estradiol exert estrogenic effects during very early development of the embryos and that depuration had a positive effect on the motherfish and her embryos.

  6. Growth hormone gene family (GH, GHR, GHRH and Pit-1 polymorphisms and its association with superovulation response, ovulation rate and embryo quality in Embryo Transfer Station (BET of Cipelang

    Directory of Open Access Journals (Sweden)

    Cece Sumantri

    2011-06-01

    Full Text Available The decrease in fertility is considered to be the main cause of reproductive loss in dairy cattle and beef industry. Many candidate genes that play an important role in fertility and embryonic development. The purpose of this study was to detect genetic variations of the growth hormone gene family (GH|MspI, GH|AluI, GHR| AluI, GHRH|HaeIII and Pit-1|HinfI and its association with superovulation response, ovulation, fertilization and transferable embryos rate. A total of 45 blood samples taken from cows that have been superovulated Angus, Brahman, HF, Limousin and Simmental. DNA was extracted with phenol-chloroform protocol followed by polymerase chain reaction technique (PCR using specific primers for GH, GHR, GHRH and Pit-1 gene. PCR product was cut with restriction enzyme MspI, AluI, HaeIII and HinfI and electrophoresed on agarose gel and stained with ethidium bromide (EtBr. Superovulation is done by injecting a totally of 20 ml FSH for 4 days. Injecting the prostaglandin hormone (PGF2α was performed on the eleventh day of CIDR implantation. Artificial insemination (AI performed two or three days after the injection of PGF2α and Flushing was done on the seventh day after the AI. The results showed that the Angus, Limousin, Brahman and Simental GH loci diversity of GH|MspI, GH|AluI, GHR|AluI, GHRH|HaeIII and Pit-1|HinfI was not associated with superovulation response, ovulation, fertilization and transferable embryo rate. In HF dairy cattle, genotype on Pit-1|HinfI AA has higher percentage of superovulation response (P < 0.05 when compared to AB genotype, but did not differ to BB genotype. Dairy cattle HF AA genotype also had higher ovulation rate (P < 0.05 when compared to AB and BB genotypes, but AB and BB have the same ovulation rate.

  7. 性控精液在牛体外性控胚胎生产中的应用%Application of Sexed Semen on the in vitro Bovine Sexed Embryos

    Institute of Scientific and Technical Information of China (English)

    孙凤俊; 张佳谊; 韩广文

    2013-01-01

    Use of sexed semen in conjunction with in vitro embryo production is a potentially efficient means of obtaining offspring of predetermined sex. The ability to sort individual sperm cells into viable X- and Y-chromosome-bearing fractions made producers' sex selection dreams reality in the 1990s and now semen can be sexed with greater than 90% accuracy with use of a flow cytometric cell sorter. However, there are some drawbacks to implementing the extensive use of sexed semen technology include the apparent lower fertility of sorted sperm, the lower survival of sorted sperm after cryopreservation and the reduced number of sperm that could be separated in a specified time period. The issues of In vitro production of bovine embryos using sexed semen and its application prospect are discussed in this review.%利用性控精液结合体外胚胎生产技术是获得预知性别后代的一种有潜在效率的方法.自20世纪90年代,利用流式细胞仪将X精子和Y精子分离开来,使畜牧生产性别选择的梦想变为现实,性控精液得到了极大的推广应用.本文重点阐述利用性控精液进行胚胎体外生产的研究和生产状况.

  8. A bipartição como alternativa para melhorar os índices de gestação na transferência de embriões eqüinos The embryo splitting as an alternative to improve gestation indices from embryo transfer in horses

    Directory of Open Access Journals (Sweden)

    Leonardo Luiz da Silveira

    2005-04-01

    Full Text Available A transferência de embriões (TE já vem sendo utilizada em eqüinos há pelo menos duas décadas, sempre a partir de ovulação simples. Para aumentar a eficiência reprodutiva, este estudo avaliou a bipartição embrionária como uma alternativa para melhorar os índices de gestação na transferência de embriões em eqüinos. Foram utilizadas 21 éguas de diferentes padrões raciais, com idade variando entre 4 e 15 anos de idade, pesando entre 270 a 480kg. A partir da identificação do cio (rufiação, os animais foram monitorados através de exames ultra-sonográficos trans-retais até o momento da ovulação, sendo as receptoras, uma vez ao dia e as doadoras três vezes ao dia. As receptoras utilizadas ovularam um dia antes ou até três dias depois das doadoras. As doadoras foram coletadas entre 144 e 156 horas após a ovulação (D0. Foram recuperados 20 embriões (mórulas em 29 coletas (68,96%, sendo que 10 embriões foram transferidos inteiros (T1, e 10 embriões foram bipartidos (T2, originando 20 hemi-embriões e transferidos para 20 receptoras. Não houve diferença na taxa de prenhez entre os grupos, T1, 70% (7/10, e T2, 50% (10/20 (P>0,05. Em relação ao número inicial de embriões em cada grupo (10, houve diferença na taxa de prenhez entre os grupos, T1, 70% (7/10 e T2, 100% (10/10 (PEmbryo transfer (ET has been used in horses for at least twenty years, always from a single ovulation. In order to increase reproductive efficiency this study evaluated embryo splitting as an alternative, to improve pregnancy rates in horse embryo transfer. Twenty-one mares of different genetic groups, aged between 4 and 15 yrs, and weighing 270 to 480kg were used. Estrus was identified using a teaser and the animals were monitored with transrectal ultrasonography exams until ovulation (donors were examined three times a day and recipients once a day. The recipients ovulated one day prior, or up to 3 days after, the donors. Donors were

  9. An immediate assessment of serum progesterone levels in frozen embryo transfer%在冻融胚胎移植当日即时评估血清孕酮的价值

    Institute of Scientific and Technical Information of China (English)

    邵小光; 史艳彬; 佟玉

    2013-01-01

    An important influencing factor for pregnancy rate and pregnancy outcome in frozen embryo transfer cycles is the difficulty handling endometrial receptivity status. Appropriate endometrial progesterone(P) level is essential to endometrial receptivity. Progesterone is one of the most basic hormones to maintain endometrial receptivity in luteal phase. It is generally believed that progesterone threshold for endometrium luteinized is 3. 18-31. 8 nmol/L in mid-luteal phase. The serum progesterone level less than 47. 7 nmol/L in mid-luteal phase is often diagnosed as luteal phase defect. The data of the frozen-thawed embryo transfer cycles in our center from 2011 to 2012 suggested that it is of predictive value to detect progesterone level on the embryo transfer day. When P<15. 9 nmol/L on the embryo transfer day, the pregnancy rate was significantly decreased and abortion rate significantly increased in the patients with normal endometrium, and it would be better to cancel the transfer. When 15. 9 nmol/L≤P<47. 7 nmol/L on the transfer day,the pregnancy rate was lower and abortion rate higher than those in the patients with P≥47. 7 nmol/L, and the embryos could be transferred, but it needs to increase the amount of progesterone for luteal support to achieve pregnancy or improve the pregnancy outcome. The pregnancy rates was the highest and the abortion rate was the lowest in the patients with P≥47. 7 nmol/L on the embryo transfer day, but it also needs amount of progesterone to maintain the original luteal support.

  10. Effect of zona pellucida removal on early development of in vitro produced bovine embryos Efecto de la remoción de la zona pelúcida sobre el desarrollo temprano de embriones bovinos producidos in vitro

    Directory of Open Access Journals (Sweden)

    AE Velásquez

    2013-01-01

    Full Text Available During early embryo development the zona pellucida acts as a barrier against polyspermia and guarantees communication between blastomeres before and during compaction. However, the development of new technologies of embryo production such as "Handmade Cloning" demands removal of this membrane. The aim of this study was to determine the effect of zona pellucida removal on in vitro bovine embryo development. First, the consequences of zona pellucida removal was assessed by comparing the percentage of first cleavage, percentage of blastocysts and cell number among zona-included and zona-free embryos; either through the removal of the zona pellucida immediately after IVF or parthenogenesis. Embryo development was also evaluated when zona pellucida was removed before parthenogenesis. In a second set of experiments, the gene expression levels of BAX, BCL2, CASP3, CDH1, OCT4 and SOX2 were evaluated in zona-free and zona-included IVF-derived embryos. No significant differences were found in the percentage of first cleavage, percentage of blastocyst and cell number on IVF-embryos cultured with or without zona. Parthenogenetic embryos followed the same general pattern even when the zona pellucida was eliminated before activation; however there was a significant increase in the rate of first cleavage when the zona pellucida was removed after activation but this did not impact upon further development. Furthermore, no significant differences in gene expression level were found between zona-free and zona-included IVF-embryos for the studied genes. We concluded that the lack of zona pellucida did not affect the early development when an appropriate system is used for embryo culture to ensure blastomeric contact and normal compaction.Durante el desarrollo embrionario temprano, la zona pelúcida actúa como barrera contra la polispermia y además garantiza la comunicación entre blastómeras y la compactación de estas. Sin embargo, el desarrollo de nuevas

  11. RNA-Seq analysis uncovers transcriptomic variations between morphologically similar in vivo- and in vitro-derived bovine blastocysts

    Directory of Open Access Journals (Sweden)

    Driver Ashley M

    2012-03-01

    Full Text Available Abstract Background A valuable tool for both research and industry, in vitro fertilization (IVF has applications range from gamete selection and preservation of traits to cloning. Although IVF has achieved worldwide use, with approximately 339,685 bovine embryos transferred in 2010 alone, there are still continuing difficulties with efficiency. It is rare to have more than 40% of fertilized in vitro cattle oocytes reach blastocyst stage by day 8 of culture, and pregnancy rates are reported as less than 45% for in vitro produced embryos. To investigate potential influences in-vitro fertilization (IVF has on embryonic development, this study compares in vivo- and in vitro-derived bovine blastocysts at a similar stage and quality grade (expanded, excellent quality to determine the degree of transcriptomic variation beyond morphology using RNA-Seq. Results A total of 26,906,451 and 38,184,547 fragments were sequenced for in vitro and in vivo embryo pools, respectively. We detected expression for a total of 17,634 genes, with 793 genes showing differential expression between the two embryo populations with false discovery rate (FDR Conclusions Thus, our results support that IVF may influence at the transcriptomic level and that morphology is limited in full characterization of bovine preimplantation embryos.

  12. Heterotopic Pregnancy After In Vitro Fertilization and Embryo Transfer After Bilateral Total Salpingectomy/Tubal Ligation: Case Report and Literature Review.

    Science.gov (United States)

    Xu, Ying; Lu, Yingli; Chen, Huiling; Li, Dandan; Zhang, Jingwen; Zheng, Lianwen

    2016-01-01

    Heterotopic pregnancy is defined as the simultaneous occurrence of intrauterine and ectopic pregnancy, either of which may be single or multiple. It occurs in up to 1% of pregnancies after in vitro fertilization and embryo transfer. This article reports 2 rare cases of heterotopic pregnancy after in vitro fertilization and presents a literature review. In the first case, a 28-year-old woman had previous laparoscopic bilateral total salpingectomy for a right tubal pregnancy and a left hydrosalpinx. However, she had ovarian heterotopic pregnancy after a third in vitro fertilization cycle. Emergency laparotomy was performed. The synchronous intrauterine pregnancy continued with no further complications and ended in the delivery of a singleton term pregnancy. The second case combined interstitial and intrauterine pregnancies after bilateral tubal ligation for hydrosalpinges followed by in vitro fertilization and frozen embryo transfer. The possibility of heterotopic pregnancy after bilateral total salpingectomy/tubal ligation, although extremely rare, should also be considered by gynecologists when they treat an in vitro fertilization patient even though an intrauterine pregnancy has been confirmed.

  13. Production of transgenic blastocyst by nuclear transfer from different types of somatic cells in cattle

    Institute of Scientific and Technical Information of China (English)

    GONG Guochun; LI Rong; LI Ning; DAI Yunping; FAN Baoliang; ZHU Huabing; WANG Haiping; WANG Lili; FANG Changge; WAN Rong; LIU Ying

    2004-01-01

    The present study examined the effects of genetic manipulation to the donor cell and different types of transgenic donor cells on developmental potential of bovine nuclear transfer (NT) embryos. Four types of bovine somatic cells, including granulosa cells, fetal fibroblasts, fetal oviduct epithelial cells and fetal ovary epithelial cells, were transfected with a plasmid (pCE-EGFP-Ires-Neo-dNdB) containing the enhanced green fluorescent protein (EGFP) and neomycin-resistant (Neor) genes by electroporation. After 14 days selection with 800 μg/mL G418, transgenic cell lines from each type of somatic cells were obtained. Nontransgenic granulosa cells and all 4 types of transgenic somatic cells were used as nuclear donor to produce transgenic embryos by NT. There was no significant difference in development rates to the blastocyst stage for NT embryos from transgenic and nontransgenic granulosa cells (44.6% and 42.8%, respectively), and transfer of NT embryos derived from transgenic and nontransgenic granulosa cells to recipients resulted in similar pregnancy rates on day 90 (19% and 25%, respectively). The development rates to the blastocyst stage of NT embryos were significantly different among different types of transgenic donor cells (P<0.05). Blastocyst rates from fetal oviduct epithelial cell and granulosa cell (49.1% and 44.6%, respectively) were higher than those from fetal fibroblast (32.7%) and fetal ovary epithelial cell (22.5%). These results suggest that (i) genetic manipulation to donor cells has no negative effect on in vitro and early in vivo developmental competence of bovine NT embryos and (ii) granulosa and fetal oviduct epithelial cells can be used to produce transgenic bovine NT embryos more efficiently. In addition, GFP can be used to select transgenic NT embryos as a non-invasive selective marker.

  14. Comparative genomics and the role of lateral gene transfer in the evolution of bovine adapted Streptococcus agalactiae

    OpenAIRE

    Richards, Vincent P; Lang, Ping; Bitar, Paulina D. Pavinski; Lefebure, Tristan; Schukken, Ynte H.; Zadoks, Ruth N.; Michael J Stanhope

    2011-01-01

    In addition to causing severe invasive infections in humans, Streptococcus agalactiae, or group B Streptococcus (GBS), is also a major cause of bovine mastitis. Here we provide the first genome sequence for S. agalactiae isolated from a cow diagnosed with clinical mastitis (strain FSL S3-026). Comparison to eight S. agalactiae genomes obtained from human disease isolates revealed 183 genes specific to the bovine strain. Subsequent polymerase chain reaction (PCR) screening for the presence/abs...

  15. 供体细胞来源和TSA处理对牦牛iSCNT胚胎重编程的影响%Effects of donor cell source and TSA treatment on reprogramming of yakbovine iSCNT embryos

    Institute of Scientific and Technical Information of China (English)

    熊显荣; 高川; 符梅; 李键; 字向东; 钟金城

    2012-01-01

    【Objective】The present study was to investigate the effect of different sources of donor cells and TSA treatment on in vitro development of yak interspecies somatic cell nuclear transfer(iSCNT)embryos.【Method】With cattle oocytes as recipient,yak fetal fibroblasts of 50-day-old,6 and 18-month-old adult fibroblasts as the donor nuclear,yak iSCNT embryos were reconstructed and then the effect of donor cell source on the development of cloned embryos evaluated.We compared the effect of different TSA treated concentrations(0,20,40,60 and 80 nmol/L)and times(0,6,8 and 12 h)on the development of yak cloned embryos,respectively,then determined the best program for treatment,and treated donor cells and embryos with this program for detecting the effect of TSA on the relative expression level of HDAC2 by qRT-PCR.【Result】With fetal fibroblast cells as donor cells,yak interspecies cloning embryos blastocyst rate was the highest,but there was no significant difference(P〉0.05).Treatment donor cell with 40 nmol/L TSA for 6 h significantly increased the blastocyst rate(30.6%),and significant difference existed with the control group(P〈0.05).Treatment of reconstructed embryos with 40 nmol/L TSA for 12 h was helpful for embryos development.However,the blastocyst formation rate decreased along with the treated time and concentration of TSA.The expression level of HDAC2 was reduced significantly after donor cell and embryos treated with TSA.【Conclusion】There was no significant effect of donor cells source on the development of yak iSCNT embryos.Treated donor cells for 6 h and reconstructed embryos for 12 h with 40 nmol/L TSA can significantly improve in vitro development of yak iSCNT embryos and reprogramming.%【目的】探讨不同来源的供体细胞和TSA处理对牦牛异种体细胞核移植(Interspecies nuclear transfer,iSCNT)胚胎的影响。【方法】以黄牛卵母细胞为受体,分别以牦牛50日龄胎儿及6和18月龄个体成纤维细

  16. Analysis of impact factors for post-thaw embryo survival rate and clinical pregnancy rate of frozen-thawed embryo transfer program%影响冻融胚胎复苏率和移植成功率因素分析

    Institute of Scientific and Technical Information of China (English)

    姚宁; 郑菊芬; 向祖琼; 赵磊文; 赵晓明; 孙赞; 洪燕; 陈佩

    2009-01-01

    Objective To explore the impact factors for post-thaw embryo survival rate and clinical pregnancy rate in frozen-thawed embryo transfer program. Methods The clinical data of 573 cycles of frozen-thawed embryo transfers were retrospectively analysed. Groups were divided according to the pre-freeze embryo quality, pre-freeze embryonic developmental stage, frozen-thawed embryo quality and cryopreservation technique, respectively, and post-thaw embryo survival rates and/or clinical pregnancy rates were compared among groups. Results The clinical pregnancy rate of high quality pre-freeze embryo was significantly higher than that of low quality pre-freeze embryo (31.8% vs 20.0%) (P0.05). The clinical pregnancy rates of the transfer cycles only with fully intact embryos and with mixed embryos were significantly higher than that only with partially damaged embryos(36.7% vs 24.1% and 29.2% vs 24.1%, respectively)(P0.05).解冻后完全移植100%卵裂球完整者和混合移植者的临床妊娠率显著高于完全移植部分卵裂球损伤者(36.7% vs 24.1%和29.2% vs 24.1%)(P<0.05).冷冻方法改进后的胚胎复苏率和全胚复苏率显著高于冷冻方法改进前(82.0% vs 66.3%和50.0% vs 27.5%)(P<0.05).结论 胚胎质量与胚胎复苏率和临床妊娠率密切相关,冷冻胚胎解冻后全胚复苏率与临床妊娠率正相关,冷冻过程中的操作细节是胚胎实验室保证冻融胚胎复苏、移植成功的重要因素.

  17. Pregnancies and improved early embryonic development with bovine oocytes matured in vitro with 9-cis-retinoic acid

    OpenAIRE

    Hidalgo, C.O. (Carlos); Díez, Carmen; Duque, Paloma; Facal, Nieves; Gómez, Enrique

    2010-01-01

    Retinoids have an important role in cell growth, morphogenesis and differentiation. In the present study the developmental potential of bovine oocytes was examined after in vitro maturation in the presence of 9-cis-retinoic acid, a vitamin A metabolite, at 5 nmol l(-1) in chemically defined conditions. Experiments studied early in vitro development, blastocyst differential cell counts and the capacity of embryos to establish pregnancy after transfer to recipients. After in vitro fertilization...

  18. Efficacy of local aspiration in the conservative treatment of live interstitial pregnancy coexisting with live intrauterine pregnancy after in vitro fertilization and embryo transfer

    Institute of Scientific and Technical Information of China (English)

    WANG Yang; MA Cai-hong; QIAO Jie; CHEN Xin-na; LIU Ping

    2012-01-01

    Background Heterotopic pregnancy (HP) is defined as a pregnancy in which one or more embryos is viably implanted in the uterus while the other is implanted elsewhere as an ectopic pregnancy.The occurrence of HP rises dramatically with the increased use of assisted reproductive technology.HP of interstitial pregnancy is one special situation which needs more concern.Here we evaluate the efficacy of local aspiration and instillation of hyperosmolar glucose in the treatment of live interstitial pregnancy complicated with live intrauterine pregnancy after in vitro fertilization and embryo transfer.@@Methods Five female patients were diagnosed with live interstitial pregnancies complicated with intrauterine pregnancies.They were treated with transvaginal ultrasound-guided aspiration of interstitial pregnancy and instillation of hyperosmolar glucose at the Center for Reproductive Medicine of Peking University Third Hospital from January 1st,2008 to May 30th,2011.@@Results Gemmule embryos in all 5 cases were aspirated successfully and there was no abdominal hemorrhage,threatened abortion or infection in any of the cases.The sac of interstitial pregnancy continued to progress after aspiration and stopped growing between 11 to 20 weeks.By the 30th week of pregnancy,80% of the interstitial masses had disappeared.Four cases have delivered and one is still in on-going pregnancy.All of the four cases underwent cesarean section and there were nothing special detected in the corner of the uterus.@@Conclusion Local aspiration and instillation of hyperosmolar glucose may be an effective way to treat live interstitial pregnancy when coexisting with a live intrauterine pregnancy.

  19. Construction of Bovine (Bos taurus) Transgenic Cloned Embryos with Lysostaphin and Endolysin Genes by Electronic Transfection%电转染法制备奶牛转溶葡球菌酶(Lysostaphin)和内溶素(Endolysin)基因胚胎

    Institute of Scientific and Technical Information of China (English)

    杨林; 杜卫华; 郝海生; 刘岩; 秦彤; 赵学明; 王栋; 朱化彬; 王宗礼

    2013-01-01

    Lysostaphin is a single chain protease containing zinc which can kill staphylococcus aureus effectively. Endolysin which is the peculiar of the double-stranded DNA bacteriophages is a murein hydrolytic enzyme, it has a wide range of antibacterial effect. Lysostaphin and Endolysin have the high synergistic effect. In this study, the vectors pBCl-seq2 +seq3-EGFP-neo containing Endolysin and Lysostaphin genes and two other marker genes of enhenced green fluorescent protein (EGFP) andneomycin (neo) were transfected into bovine (Bos taurus) fetal fibroblast by electroporation and nucleofector of AMAXA. Stable transfected monoclonal cells which were identified to be the positive-cells in the way of PCR technique were obtained through fluorescence and G418 selection. Using transfected cells as the donor, transfected embryos were produced with somatic cell nuclear transfer, we used different conditions of AMAXA nuleofecor(A-023,V-013, V-023 and T-016) to transfect bovine fetal fibroblast, the results showed the suitable program was T-016. There were 5 times transfection efficient of AMAXA nuleofecor (20.11%) than it of electroporation. The blastocyst developed normally and the rate was of it 20.08%. In our study, we built up bovine fetal fibroblast cell line, sought out transfection parameter of high transfection efficiency, and acquired transgenic cell lines and transgenic blastocyst containing Lysostaphin and Endolysin genes, In conclusion, the results can provide technology supporting for producing anti-mastitis transgenic bovine and searching the new therapy way of mastitis.%溶葡球菌酶(Lysostaphin)是一种含锌的单链蛋白酶,能有效地杀灭金黄色葡萄球菌.内溶素(Endolysin)是双链DNA噬菌体所特有,是一类胞壁质水解酶,具有广泛的抗菌效果.内溶素与抗生素之间有高效的协同作用.本研究通过BTX电转染和AMAXA核转染的方法将含有溶葡球菌酶(Lysostaphin)和内溶素(Endolysin)两个目的基因(Seq2

  20. Morbidity-mortality and performance evaluation of Brahman calves from in vitro embryo production

    Directory of Open Access Journals (Sweden)

    Pimenta-Oliveira Andreza

    2011-12-01

    Full Text Available Background The use of bovine in vitro embryo production (IVP increases the reproductive potential of genetically superior cows, enabling a larger scale of embryo production when compared with other biotechnologies. However, deleterious effects such as abnormal fetal growth, longer gestation period, increased birth weight, abortion, preterm birth and higher rates of neonatal mortality have been attributed to IVP. The aim of this study was to compare the influence of in vitro embryo production and artificial insemination (AI on gestation length, complications with birth, birth weight, method of feeding colostrum, passive transfer of immunity, morbidity-mortality, and performance in Brahman calves. Results Whilst gestation length and birth weight were significantly increased in IVP-derived calves, no difference in weaning weight was observed between groups. The passive transfer of immunity (PT, was assessed in IVP (n = 80 and AI (n = 20 groups 24 hours after birth by determination of gamma-glutamyl transferase (GGT and gammaglobulin activity as well as by quantification of the concentration of total protein in serum. No differences in passive transfer or incidences of dystocia and diseases at weaning were observed between groups. Birth weight, method of feeding colostrum and dystocia were not correlated with PT in either group. Conclusions In this study, in vitro embryo production did not affect the health status, development, or passive transfer of immunity in Brahman calves.

  1. Genomic selection strategies in dairy cattle breeding programmes: Sexed semen cannot replace multiple ovulation and embryo transfer as superior reproductive technology

    DEFF Research Database (Denmark)

    Pedersen, Louise Dybdahl; Kargo, Morten; Berg, Peer;

    2012-01-01

    semen. However, when all young bull candidates were born following MOET, the results showed that the use of Y-semen in the breeding nucleus tended to decrease the rate of inbreeding as it enabled GS to increase within-family selection. This implies that the benefit from using sexed semen in a modern......The aim of this study was to test whether the use of X-semen in a dairy cattle population using genomic selection (GS) and multiple ovulation and embryo transfer (MOET) increases the selection intensity on cow dams and thereby the genetic gain in the entire population. Also, the dynamics of using...... different types of sexed semen (X, Y or conventional) in the nucleus were investigated. The stochastic simulation study partly supported the hypothesis as the genetic gain in the entire population was elevated when X-semen was used in the production population as GS exploited the higher selection intensity...

  2. Comparison of passive transfer of immunity in neonatal dairy calves fed colostrum or bovine serum-based colostrum replacement and colostrum supplement products

    Science.gov (United States)

    Poulsen, Keith P.; Foley, Andrea L.; Collins, Michael T.; McGuirk, Sheila M.

    2011-01-01

    Objective To compare serum total protein (sTP) and serum IgG (sIgG) concentrations in neonatal calves administered colostrum or a bovine serum-based colostrum replacement (CR) product followed by a bovine serum-based colostrum supplement (CS) product. Design Randomized controlled clinical trial. Animals 18 Jersey and 269 Holstein neonatal heifer calves. Procedures 141 calves were given 4 L of colostrum in 1 or 2 feedings (first or only feeding was provided ≤ 2 hours after birth; when applicable, a second feeding was provided between 2 and 12 hours after birth). Other calves (n = 146) were fed 2 L of a CR product ≤ 2 hours after birth and then 2 L of a CS product between 2 and 12 hours after birth. Concentrations of sTP and sIgG were measured 1 to 7 days after birth. Data from cohorts on individual farms and for all farms were analyzed. Results Mean sTP and sIgG concentrations differed significantly between feeding groups. In calves fed colostrum and calves fed CR and CS products, mean ± SD sTP concentration was 5.58 ± 0.67 g/dL and 5.26 ± 0.54 g/dL, respectively, and mean sIgG concentration was 1,868 ± 854 mg/dL and 1,320 ± 620 mg/dL, respectively. The percentage of calves that had failure of passive transfer of immunity (ie, sIgG concentrations < 1,000 mg/dL) was not significantly different between groups. Conclusions and Clinical Relevance Results suggested that sequential feeding of bovine serum-based CR and CS products to neonatal calves is an alternative to feeding colostrum for achieving passive transfer of immunity. PMID:20946083

  3. Couples with unexplained subfertility and unfavorable prognosis: a randomized pilot trial comparing the effectiveness of in vitro fertilization with elective single embryo transfer versus intrauterine insemination with controlled ovarian stimulation

    NARCIS (Netherlands)

    I.M. Custers; T.E. König; F.J. Broekmans; P.G.A. Hompes; E. Kaaijk; J. Oosterhuis; M.H. Mochtar; S. Repping; M. van Wely; P. Steures; F. van der Veen; B.W.J. Mol

    2011-01-01

    To evaluate the effectiveness of IVF with elective single embryo transfer (IVF-eSET) vs. IUI with controlled ovarian stimulation (IUI-COS) as an alternative treatment to reduce the risk for a multiple pregnancy. Randomized pilot trial. Three academic and six teaching hospitals in the Netherlands. Co

  4. Prevention of multiple pregnancies in couples with unexplained or mild male subfertility : randomised controlled trial of in vitro fertilisation with single embryo transfer or in vitro fertilisation in modified natural cycle compared with intrauterine insemination with controlled ovarian hyperstimulation

    NARCIS (Netherlands)

    Bensdorp, A. J.; Tjon-Kon-Fat, R. I.; Bossuyt, P. M. M.; Koks, C. A. M.; Oosterhuis, G. J. E.; Hoek, A.; Hompes, P. G. A.; Broekmans, F. J. M.; Verhoeve, H. R.; de Bruin, J. P.; van Golde, R.; Repping, S.; Cohlen, B. J.; Lambers, M. D. A.; van Bommel, P. F.; Slappendel, E.; Perquin, D.; Smeenk, J. M.; Pelinck, M. J.; Gianotten, J.; Hoozemans, D. A.; Maas, J. W. M.; Eijkemans, M. J. C.; van der Veen, F.; Mol, B. W. J.; van Wely, M.

    2015-01-01

    Objectives To compare the effectiveness of in vitro fertilisation with single embryo transfer or in vitro fertilisation in a modified natural cycle with that of intrauterine insemination with controlled ovarian hyperstimulation in terms of a healthy child. Design Multicentre, open label, three arm,

  5. Prevention of multiple pregnancies in couples with unexplained or mild male subfertility : Randomised controlled trial of in vitro fertilisation with single embryo transfer or in vitro fertilisation in modified natural cycle compared with intrauterine insemination with controlled ovarian hyperstimulation

    NARCIS (Netherlands)

    Bensdorp, A. J.; Tjon-Kon-Fat, R. I.; Bossuyt, P. M M; Koks, C. A M; Oosterhuis, G. J E; Hoek, A.; Hompes, P. G A; Broekmans, F. J M; Verhoeve, H. R.; De Bruin, J. P.; Van Golde, R.; Repping, S.; Cohlen, B. J.; Lambers, M. D A; Van Bommel, P. F.; Slappendel, E.; Perquin, D.; Smeenk, J. M.; Pelinck, M. J.; Gianotten, J.; Hoozemans, D. A.; Maas, J. W M; Eijkemans, M. J C; Van Der Veen, F.; Mol, B. W J; Van Wely, M.

    2015-01-01

    Objectives: To compare the effectiveness of in vitro fertilisation with single embryo transfer or in vitro fertilisation in a modified natural cycle with that of intrauterine insemination with controlled ovarian hyperstimulation in terms of a healthy child. Design: Multicentre, open label, three arm

  6. Couples with unexplained subfertility and unfavorable prognosis : a randomized pilot trial comparing the effectiveness of in vitro fertilization with elective single embryo transfer versus intrauterine insemination with controlled ovarian stimulation

    NARCIS (Netherlands)

    Custers, Inge M.; Konig, Tamar E.; Broekmans, Frank J.; Hompes, Peter G. A.; Kaaijk, Eugenie; Oosterhuis, Jur; Mochtar, Monique H.; Repping, Sjoerd; van Wely, Madelon; Steures, Pieternel; van der Veen, Fulco; Mol, Ben W. J.

    2011-01-01

    Objective: To evaluate the effectiveness of IVF with elective single embryo transfer (IVF-eSET) vs. IUI with controlled ovarian stimulation (IUI-COS) as an alternative treatment to reduce the risk for a multiple pregnancy. Design: Randomized pilot trial. Setting: Three academic and six teaching hosp

  7. Embryo splitting: a role in infertility?

    Science.gov (United States)

    Wood, C

    2001-01-01

    Embryo splitting may be used to increase the potential fertility of couples requiring IVF. Using cattle as a model, it is possible to increase pregnancy rates from 70% per transfer of good quality in-vivo-produced embryos, to 110% by transferring the two demi-embryos resulting from the bisection of one embryo. The 30-40% greater chance of conception would reduce costs for the government, health authorities and patients, and reduce stress, time and complications for women having IVF treatment. Embryo splitting may also provide donor embryos for infertile couples that cannot conceive naturally or with IVF. The shortage of children for adoption and donor embryos may be overcome by the production of demi-embryos.

  8. Rol de la mitocondria y el estrés oxidativo en el bloqueo del desarrollo de embriones bovinos producidos in vitro Mitochondrial rol and oxidative stress in the developmental blockade of in vitro produced bovine embryos

    Directory of Open Access Journals (Sweden)

    AM Tarazona

    2010-01-01

    mediator of physiological and pathological states. Over the past years it has been shown that hydrogen peroxide (H2O2 is a pivoting molecule able to trigger cell death by different mechanisms that may or may not involve the transcription factors such as NFκB-p53, and is executed by effector caspases. It is believed that mitochondria may play an important role as a producer or as a target of H2O2, and as a mediator in apoptotic death of embryos. The purpose of this review is to present the state of the art about apoptosis triggered by oxidative stress and mediated by mitochondria in in vitro produced bovine embryos, as part of the explanation for the low efficiency in this process.

  9. 牛输卵管上皮细胞转人胶原蛋白cDNA基因及转基因克隆胚胎%Bovine oviduct epithelial cells transfected with human collagen cDNA gene and transgenic cloning embryo

    Institute of Scientific and Technical Information of China (English)

    吕自力; 王亮; 刘婷婷; 石国庆

    2012-01-01

    Isolated bovine oviduct epithelial ceils by trypsin digestion from oviduct tissue of a 2-year-old dairy cow. Oviduct epithelial cells grow well in DMEM/F12 medium. The first generation oviduct epithelial cells were used as target cells to carry out electroporation transfection. Molecular size of the transfected genes is 31 085 bp. The gene is a plasmid that carry β-casein promoter,human collagen eDNA and EGFP,Neor as double marker. Electroporatioa experiment found that bovine oviduct epithelial cells can get positive-transfectant in hypotonic buffer. Among them,90 mOsm/kg is the best. For voltage,800 V is the best. High and low voltage are not conducive to the success of electroporation. Successfully transfected cells were screened with 800ug/ml of G418. on day 10, a larger cluster of positive cell clones were obtained. The cluster of positive clone cells were expanded to obtain a more pure cell line. Detected by flow cytomerty,the purity was 81.6% . This cell as a nuclear transfer donor carried out gene transfer experiments. The results showed that in transgenic cells and nontransgenic cells, their fusion rate of reconstructed embryos obtained significant difference(51.9 % vs 63.2 %), their morula/blastocyst rate of reconstructed embrys were not significantly different (20.3 % vs 25.5% ). DNA of the embryos displaying green fluorescence was analyzed,The results showed that the embryos was successfully transferred to the foreign gene and the genetic structure of the transfer was complete.%以2岁奶牛输卵管为材料,用胰酶消化法分离得到了牛输卵管上皮细胞。输卵管上皮细胞在DMEM/F12培养基中生长良好。用一代牛输卵管上皮细胞为靶细胞,对其进行了电穿孔转染,转染对象是分子大小为31085bp的以β-casein启动子为基础的含有人胶原蛋白cDNA基因和EGFP、Neor双标记基因的质粒。电穿孔试验发现,牛输卵管上皮细胞在低渗缓冲液中电转染可以获

  10. In vitroembryo outgrowth is a bioassay ofin vivo embryo implantation and development

    Institute of Scientific and Technical Information of China (English)

    Natalie K. Binder; Natalie J. Hannan; David K. Gardner

    2015-01-01

    Objective:To determine the efficacy of embryo outgrowth on fibronectin as a low cost, high throughput alternative to embryo transfer to model embryo attachment and the initial stages of implantation.Methods:Following in vitro embryo culture, embryo quality was assessedviaembryo transfer or embryo outgrowth with metabolic assessment.Results:This study shows that blastocysts attach to fibronectin at the same rate that they implantin vivo, and that the carbohydrate utilisation of embryos that successfully outgrow is comparable to those that are able to develop into a fetus.Conclusions:Embryo outgrowth is a suitable alternative endpoint to embryo transfer.

  11. Effect of insulin-like growth factor-1 during in vitro oocyte maturation and in vitro culture of bovine embryos Efeito do fator de crescimento semelhante à insulina-1 durante a maturação in vitro dos oócitos e cultivo in vitro de embriões bovinos

    OpenAIRE

    M.D. Quetglas; L.A Coelho; Garcia, J M; E.B. Oliveira Filho; C.R. Esper

    2001-01-01

    The effects of insulin-like growth factor-I (IGF-I) on in vitro maturation (IVM) (experiment I) and on in vitro embryo development (experiment II) of bovine oocytes fertilized in vitro, were evaluated in terms of cleavage (CR), blastocyst (BR) and hatching (HR) rates. For IVM, immature cumulus-oocyte complexes were cultured in TCM-199 medium supplemented with Hepes, sodium bicarbonate, sodium pyruvate, additives, fetal calf serum (B-199 medium) and gonadotropins (14 U/ml PMSG and 7 U/ml hCG)....

  12. Combining use of embryo sexing and cloning within closed mixed MOETs for selection on dairy cattle

    OpenAIRE

    Colleau, J Jacques

    1992-01-01

    Given the same overall number of transferred embryos, a comparison was carried out between adult mixed (ie with bull progeny-testing) MOET (multiple ovulation and embryo-transfer) schemes with embryo sexing only versus embryo sexing plus cloning of female embryos. In the former schemes, natural and ET (embryo transfer) animals were allowed to breed. In the latter schemes, each category of females (single born or cloned) was allowed to give birth either to new single animals (natural calving...

  13. Factors influencing the outcome of embryo freezing and Ihawing program

    Institute of Scientific and Technical Information of China (English)

    叶英辉; 金帆; 徐晨明; 邢兰凤

    2002-01-01

    Objective: To investigate the factors that might influence the sucess of an ernbryo freezing and thawing program.Method: The relationship between the pregnancy rate in 73 cycles of embryo freezing and thewing program and the following factors was analyzed;matermal age,E2 level at the time of HCG trigger,embryo storage time,number of thawed embryos transferred,presence of sponsoring embryos and intact embryos.And the suvival rate of thawed embryos with different morphology,cell stage and storage time was evaluated.Result:Transfer with three of more than three thawed embryos resulted in pragnancy rates of 38.5% and 35.7%,respectively.compared with 5.3% for transfer of fewer than three embryos.The presence of sponsoring embryos and intact embryos significantly incresses pregnancy rate in embryo freezing and thawing program .No other factor examined had any effect on pregnancy outcome.The survival rate of good morphology embryos was higher than poor ones,but was not influenced by cell stage and storage time.Conclusion:Embryo morphology before freezing , number of thawed embryos transferred and the presence of intact embryos are important to the outcome of embryo freezing and thawing program.

  14. Factors influencing the outcome of embryo freezing and thawing program

    Institute of Scientific and Technical Information of China (English)

    叶英辉; 金帆; 徐晨明; 邢兰凤

    2002-01-01

    Objective: To investigate the factors that might influence the succ ess of an embryo freezing and thawing program. Method: The relationship betwee n the pregnancy rate in 73 cycles of embryo freezing and thawing program and the following factors was analyzed: maternal age, E2 level at the time of HCG trigg er, embryo storage time, number of thawed embryos transferred, presence of spons oring embryos and intact embryos. And the survival rate of thawed embryos with d ifferent morphology, cell stage and storage time was evaluated. Result: Tra nsfer with three or more than three thawed embryos resulted in pregnancy rates o f 38.5% and 35.7%, respectively, compared with 5.3% for transfer of fewer th an t hree embryos. The presence of sponsoring embryos and intact embryos significantl y increases pregnancy rate in embryo freezing and thawing program. No other fact or examined had any effect on pregnancy outcome. The survival rate of good morph ology embryos was higher than poor ones, but was not influenced by cell stage an d storage time. Conclusion: Embryo morphology before freezing, number of thaw ed embryos transferred and the presence of intact embryos are important to the o utcome of embryo freezing and thawing program.

  15. 玻璃化冷冻对冻融卵裂期胚胎移植周期结局的影响%Influence of vitrification method on clinical outcome of human frozen-thawed cleaved embryo transfer

    Institute of Scientific and Technical Information of China (English)

    柳倩茹; 黄绮云; 池霖生; 张静雯; 朱莉

    2014-01-01

    Objective To investigate the influence of vitrification method on clinical outcome of human frozen-thawed cleaved embryo transfer.Methods In the in vitro fertilization and embryo transfer cycles,supernumerary embryos were cryopreserved by vitrification or slow-freezing sequencing method.The embryo survival rate,intact survival rate,pregnant rate and implantation rate were compared after thawing.Results In 248 vitrified embryo thaw cycles,746 vitrified embryos were thawed,and 643(86.2%) of them survived and 594(79.4%) of them intactly survived after the thawing.In 244 women,632 embryos were transferred,and 107(43.9%) of them got clinical pregnancy,the implantation rate was 26.1% (165/632).As to slow-freezing method,in 225 embryo thaw cycles,818 embryos were thawed,598 (73.1%) of them survived,436(53.3%) of them intactly survived,and 594 embryos were transferred in 225 cycles,and 74 (32.9%) of them got clinical pregnancy,the implantation rate was 15.2%(90/594).The embryo survival rate,intact survival rate,pregnant rate,implantation rate were higher in vitrification group than those in slow-freezing group (P < 0.05).Conclusion Vitrification is a more effective method of cryopreserving the human embryo than conventional slow freezing.%目的 探讨玻璃化冷冻法对卵裂期冻融胚胎移植周期结局的影响.方法 将接受体外受精-胚胎移植患者剩余的可利用胚胎进行玻璃化冷冻或者慢速程序化冷冻,比较胚胎复苏后的复苏率、胚胎存活率、完整存活率、种植率、妊娠率等指标.结果 玻璃化冷冻复苏248个周期、746个胚胎,存活643个(86.2%),完整存活594个(79.4%),移植244个周期,临床妊娠107个周期(43.9%),种植率26.1%,流产9个周期(8.5%).慢速程序冷冻复苏225个周期、818个胚胎,存活598个(73.1%),完整存活436个(53.3%),移植225个周期,临床妊娠74个周期(32.9%),种植率15.2%,流产7个周期(9.5%).玻璃化法冷冻后的

  16. 体外受精-胚胎移植的助孕效果:胚胎体外培养和低温保存%Reproductive effects of in vitro fertilization-embryo transfer:in vitro culture and cryopreservation of embryos

    Institute of Scientific and Technical Information of China (English)

    王芳; 陈绍威

    2014-01-01

    BACKGROUND:Since the first successful birth of a"test tube baby", in vitro fertilization-embryo transfer technology has been greatly developed within 20 years. OBJECTIVE:To retrospectively analyze the assisted reproductive effects of in vitro fertilization-embryo transfer, and to explore the culture of embryos in vitro and the application of cryopreservation technology. METHODS:A total of 131 infertile patients received in vitro fertilization-embryo transfer or intracytoplasmic sperm injection in Department of Reproductive Medicine, Affiliated Hospital of Luzhou Medical Col ege, China. There were 147 oocyte retrieval cycles and 124 transfer cycles, the mean age of the patients was 31.32 years. Standard long-protocol ovulation induction was given to 131 oocyte retrieval cycles, super-long-protocol ovulation induction was given to 4 oocyte retrieval cycles, and super-short-protocol ovulation induction was given to two oocyte retrieval cycles;93 cases were transplanted with fresh embryos and 28 cases with frozen embryos. The clinical pregnancy rate, embryo implantation rate and abortion rate of patients receiving different ovulation induction protocols and fresh or frozen embryo transfer were observed. RESULTS AND CONCLUSION:In the patients receiving standard long, super-long and super-short protocols for ovulation induction, the clinical pregnancy rate was respectively 34.7%, 50%, 20%;the embryo implantation rates were 19.1%, 22.2%, 30%;and abortion rate was 21.7%, 50%, 33.3%;in those patients, no case exhibited natural cycle of ovulation. In the patients receiving fresh embryo transfer, the clinical pregnancy rate was 33.6%, embryo implantation rate was 18.1%, and abortion rate was 20.1%;in frozen embryo transferred patients, the clinical pregnancy rate was 53.8%, embryo implantation rate was 22.3%, and abortion rate was 20.7%. Experimental findings indicate that, standard long-protocol ovulation induction has good effects, the efficacy of fresh embryo transfer is

  17. Aplicação da ultrassonografia colorida doppler em programas de transferência de embriões equinos Potential use of doppler ultrasound in equine embryo transfer programs

    Directory of Open Access Journals (Sweden)

    Jair Camargo Ferreira

    2011-06-01

    distúrbios de fluxo sanguíneo de trato reprodutivo ainda se fazem necessários.Embryo transfer in mares has been commercially used for more than three decades and it is one of most frequently applied biotechnologies on equine reproduction. Ultrasonic doppler exam of donors during the pre and post-breeding and recipients evaluation at the embryo transfer moment is essential for embryo transfer program success. Doppler ultrasonography is a non-invasive technic that allows real-time evaluation of the reproductive system hemodynamics in large animal. For providing anatomical details and blood-flow physiologic information of vessels and tissues, doppler exam can be used for ovulation prognostication, and to evaluate uterine and luteal functional status. Additionally, it can serve as a diagnostic aid of hemodynamic disturbance in reproductive system. Vascularity changes of future ovulatory follicles precede the diameter deviation. Based on this, it is possible to estimate the beginning of the breeding season and the best moment to initiate superovulatory treatments. Moreover, follicular blood flow can be used to decide the most appropriated moment for ovulation induction and breeding of donors mares. Uterine and luteal evaluation using Doppler-mode in mares can be useful to select embryo recipients with satisfactory progesterone production and adequate uterine vascularity for embryo development and maintenance of pregnancy. The maternal recognition of pregnancy also can be evaluated. Although the applicability of the Doppler technology in embryo transfer programs, additional studies are necessary to determine the standards of normality and to characterize different blood-flow disturbances of the reproductive system.

  18. Evaluation of reciprocal differences in Bos indicus x Bos taurus backcross calves produced through embryo transfer: I. Birth and weaning traits.

    Science.gov (United States)

    Amen, T S; Herring, A D; Sanders, J O; Gill, C A

    2007-02-01

    Angus (A) and Bos indicus (B; Brahman or Nellore) reciprocal backcross, embryo transfer calves, belonging to 28 full-sib families, were evaluated for differences in birth weight, gestation length, and weaning weight. Two methods were investigated; method I made no distinction between how the F(1) parents were produced, whereas method II distinguished between the 2 types of F(1) parents (AB vs. BA corresponding to A x B vs. B x A, respectively). Bos indicus backcross calves had a 4.3 d longer (P x F(1) calves had a 5.2 d longer (P = 0.01) gestation length than F(1) x B calves (290.5 vs. 285.3, respectively). Under method II analysis, there was a consistent trend for gestation length, in which BA F(1) parents produced calves that ranked greater than calves from AB F(1) parents, as sires and dams. Crosses with a greater proportion of B in the sire in relation to the amount in the dam had a heavier (P x A and B x F(1); 38.1 and 38.4 kg, respectively) than their respective reciprocal crosses (A x F(1) and F(1) x B; 34.3 and 33.5 kg, respectively). The F(1) x A and B x F(1) crosses showed a large difference in birth weight between males and females (5.3 and 4.1 kg, respectively), whereas A x F(1) and F(1) x B crosses showed a small difference (P > 0.10) in birth weight between males and females (1.5 and 1.1 kg, respectively). Further examination within each sex showed a difference between male reciprocals that was generally much larger than that between female reciprocals. Calves with a greater percentage of B in the sire compared with the proportion in the dam ranked heavier for weaning weight as for birth weight, though these differences were not significant. In breeding systems involving B x Bos taurus crosses, even when using embryo transfer, not only does the breed composition of the calves affect their preweaning performance, but the particular cross that produces the calves also should be considered in making breeding decisions. PMID:17235021

  19. 非优良胚胎形成的囊胚与卵裂期优良胚胎的冻融胚胎移植的临床结局比较%Comparison of clinical outcomes of blastocysts derived from non-top quality embryos and cleavage-stage high-quality embryos in frozen-thawed embryo transfer cycles

    Institute of Scientific and Technical Information of China (English)

    许丽娟; 陈薪; 田小龙; 刘玉东; 王楠; 叶德盛; 郭萍萍; 陈士岭

    2015-01-01

    目的:探讨不同发育天数胚胎的发育潜能,为D3非优良胚胎进行囊胚培养及其冻融移植提供依据。方法回顾性分析687例复苏周期胚胎移植患者的资料,根据胚胎冷冻时间不同,分为3组:D5冷冻组(n=87)、D6冷冻组(n=111)和D3冷冻组(n=489),采用外源性雌孕激素或自然周期准备内膜,比较各组间的临床妊娠率、流产率、种植率等指标。结果每组移植周期临床妊娠率、流产率、种植率分别为:D5冷冻组58.6%、9.8%、42.9%;D6冷冻组32.4%、19.4%、23.3%;D3冷冻组44.9%、16.4%、26.9%。D5冷冻组的临床妊娠率和种植率均明显高于另外两组,且差异均具有统计学意义(P<0.05)。结论 D3的非优良胚胎继续培养,若能够形成囊胚,所形成的D5囊胚冻融移植的临床结局优于D3的优良胚胎和非优良胚胎形成的D6囊胚冻融移植的临床结局。因此D3的非优良胚胎的囊胚培养及其冻融移植具有临床应用价值,且在冻融胚胎移植周期若有不同冷冻时间的胚胎可供选择时,可以优先选择非优良胚胎来源的D5囊胚,其次选择D3的卵裂期胚胎。%Objective To explore the developmental potential of embryos at different developmental days and provide evidence for blastocyst culture of non-top quality cleavage stage embryos in frozen-thawed embryo transfer (FET) cycles. Methods The clinical data of 687 FET cycles were retrospectively analyzed. According to the embryo freezing time, the patients were divided into day 5 (D5) blastocyst group (n=87), day 6 (D6) blastocyst group (n=111) and day 3 cleavage-stage embryo (D3) group (n=489) with hormone replacement cycles or natural cycles for endometrial preparation. The clinical pregnancy rates, miscarriage rates, and implantation rates were compared between the 3 groups. Results The clinical pregnancy rate, miscarriage rate and implantation rate per transfer were 58.6%, 9.8%, and 42.9% in D

  20. Consequences of manipulating gametes and embryos of ruminant species.

    Science.gov (United States)

    McEvoy, T G; Ashworth, C J; Rooke, J A; Sinclair, K D

    2003-01-01

    During the past 12 years, ruminants have provided a focus for some significant advances in mammalian reproductive biotechnologies. Lambs were the first offspring generated after nuclear transfer of fetal or adult cells to enucleated oocytes, and many calves of pre-determined gender are today the result of commercialized semen sexing. In 1990, the birth of one calf provided living proof that even 'dead' spermatozoa can be paternal, whereas, more recently, a short-lived gaur calf and viable mouflon lamb represented a novel option for conservation of endangered species. As well as highlights, hazards have emerged, resulting in setbacks or developmental anomalies, such as those associated with the large offspring syndrome which encompasses a range of adverse fetal, placental and post-natal phenomena expressed in ruminants. In this review, the developmental and other consequences of applying manipulative procedures, such as assisted fertilization, semen sexing, cloning and gene transfer, to gametes and embryos from bovine, ovine and caprine species are considered. Although assisted fertilization techniques can overcome mammalian infertility, they also usurp natural gamete selection safeguards, but not always with impunity. In the case of manipulations such as cloning, and to a lesser extent gene transfer, it is evident that nuclear-cytoplasmic interactions and nuclear-mitochondrial DNA interdependences are at least partially damaged or destroyed with a view to reconstruction. Therefore, among surviving zygotes and embryos it is inevitable that the legacy is frequently one of altered genetic, epigenetic or cellular programmes and processes. PMID:14635934

  1. Comparison of the Effects of Nest PCR System Established by Sry Gene and Y Chromosome Repeated Sequence for Sex Determination of Bovine Embryo%Sry基因和Y染色体重复序列在牛早期胚胎性别鉴定中应用效果的比较

    Institute of Scientific and Technical Information of China (English)

    张伟; 王世银; 张兆旺; 赵兴绪

    2011-01-01

    Sty gene and Y chromosome repeated sequence were selected as male-specific gene for sex determination of bovine embryo in this study. Eight pairs of primers were designed and a nest PCR system was established in order to compare the amplification effect. The results showed that the system established by Y chromosome repeated sequence was more sensitive and stable than the one established by Sty gene when the PCR template was the DNA from one embryonic ceil, so it was more suitable for sex determination of bovine embryo.%本试验以牛Sty基因和Y染色体重复序列作为雄性特异性基因,分别设计引物,建立多重巢式PCR体系,比较二者在牛早期胚胎性别鉴定中的应用效果。试验结果表明,当扩增体系中的模板量为一个胚胎细胞的DNA量时,以Y染色体重复序列构建的扩增体系比Sry基因具有更高的灵敏度和稳定性,更适合用于牛早期胚胎性别鉴定。

  2. Clinical observation on preventing ovarian hyperstimulation syndrome by frozen-thawed embryo transfer after all embryo freezing%全胚冷冻后行冻融胚胎移植预防卵巢过度刺激综合征的临床观察

    Institute of Scientific and Technical Information of China (English)

    黄永刚; 黄朝霞; 侯晓红; 肖宇; 吕杰强

    2012-01-01

    Objective: To compare the clinical pregnancy outcomes of frozen - thawed embryo transfer (FET) after all embryo freezing and routine in -vitro fertilization and embryo transfer (IVF - ET) , and explore the clinical application for prevention of ovarian hyperstimulation syndrome ( OHSS) . Methods: The clinical data of 124 patients receiving FET to prevent moderate and severe OHSS and 987 patients receiving routine long protocol of IVF — ET were analyzed retrospectively, ihe embryo implantation rates and clinical pregnancy rates in the two groups were compared. Results: There was no statistically significant difference in age, duration of infertility, body mass index, the number of transferred embryos, embryo implantation rate, and clinical pregnancy rate between the two groups ( P > 0. 05 ) , After IVF -ET, 25 patients in patients receiving IVF, went to hospital for treatment because of severe OHSS, and no one in patients receiving FET went to hospital. Conclusion: FET after all embryo freezing can achieve a good pregnancy outcome, which is a safe clinical application scheme for preventing moderate and severe OHSS.%目的:比较全胚冷冻后行冻融胚胎移植(FET)和常规体外受精-胚胎移植(IVF-ET)的临床妊娠结局,探讨预防卵巢过度刺激综合征(OHSS)的临床应用.方法:回顾性分析1VF-ET过程中为防止中、重度OHSS而行FET的124例患者和常规长方案IVF-ET987例患者的临床资料,比较其胚胎种植率、临床妊娠率.结果:2组的年龄、不孕年限、体重指数差异无统计学意义(P>0.05),移植胚胎个数、胚胎种植率、临床妊娠率差异亦无统计学意义(P>0.05).IVF组移植后有25例患者因重度OHSS发生而住院治疗,冷冻组无一例住院治疗.结论:全胚冷冻后行冻融胚胎移植可以获得良好的妊娠结局,是一项安全的预防中、重度卵巢过度刺激综合征的临床应用方案.

  3. The relationship of the number of embryo transferred and IVF-ET outcome%胚胎移植的数量和IVF-ET结局的关系

    Institute of Scientific and Technical Information of China (English)

    杨晓丽; 张烁

    2009-01-01

    目的 探讨体外受精-胚胎移植(in vitro fertilization and embryo transfer, IVF-ET)过程中胚胎移植数从3个降至2个对IVF-ET结局的影响.方法 对161例35岁以下的不孕患者共161个IVF-ET周期的资料进行回顾性分析.将患者分为两组,其中移植2个胚胎的为第1组,移植3个胚胎的为第2组,从而比较不同胚胎移植数对临床妊娠率和多胎妊娠发生率的影响.结果 两组的临床妊娠率差异无统计学意义(P>0.05),但移植3个胚胎组的多胎妊娠率显著高于移植2个胚胎组(P<0.05).结论 35岁以下接受IVF的妇女,移植2个优质胚胎不影响妊娠的成功,同时可以降低多胎发生率.

  4. Assessment of placental transfer and the effect on embryo-fetal development of a humanized monoclonal antibody targeting lymphotoxin-alpha in non-human primates.

    Science.gov (United States)

    Wang, Hong; Schuetz, Chris; Arima, Akihiro; Chihaya, Yutaka; Weinbauer, Gerhard F; Habermann, Gunnar; Xiao, Jim; Woods, Cynthia; Grogan, Jane; Gelzleichter, Thomas; Cain, Gary

    2016-08-01

    An enhanced embryo-fetal development study was conducted in cynomolgus monkeys using pateclizumab, a humanized IgG1 monoclonal antibody (mAb) targeting lymphotoxin-alpha. Pateclizumab administration between gestation days (GD) 20 and 132 did not induce maternal or developmental toxicities. The ratio of fetal-to-maternal serum concentration of pateclizumab was 0.73% on GD 50 and 61% by GD 139. Decreased fetal inguinal lymph node-to-body weight ratio was present in the high-dose group without microscopic abnormalities, a change attributable to inhibition of lymphocyte recruitment, which is a pharmacologic effect of pateclizumab during late lymph node development. The effect was observed in inguinal but not submandibular or mesenteric lymph nodes; this was attributed to differential susceptibility related to sequential lymph node development. Placental transfer of therapeutic IgG1 antibodies; thus, begins during the first trimester in non-human primates. Depending on the potency and dose levels administered, antibody levels in the fetus may be pharmacologically or toxicologically relevant. PMID:27211603

  5. In vitro developmental competence of pig nuclear transferred embryos: effects of GFP transfection, refrigeration, cell cycle synchronization and shapes of donor cells.

    Science.gov (United States)

    Zhang, Yun-Hai; Pan, Deng-Ke; Sun, Xiu-Zhu; Sun, Guo-Jie; Liu, Xiao-Hui; Wang, Xiao-Bo; Tian, Xing-Hua; Li, Yan; Dai, Yun-Ping; Li, Ning

    2006-08-01

    The present study was designed to evaluate the feasibility of producing pig transgenic blastocysts expressing enhanced green fluorescent protein (GFP) and to examine the effects of shape and preparation methods of donor cells on in vitro developmental ability of pig nuclear transferred embryos (NTEs). In experiment 1, the effect of GFP transfection on development of pig NTEs was evaluated. The cleavage and blastocyst rates showed no significant difference between NTEs derived from transfected and non-transfected donors. In experiment 2, the effect of different nuclear donor preparation methods on in vitro development of NTEs was examined. The cleavage rate showed no statistically significant differences among three preparation methods. The blastocyst rates of donor cells treated once at -4 degrees C and those of freshly digested cells were similar to each other (26.3% vs 17.9%). The lowest blastocyst rates (5.88%) were observed when cells cryopreserved at -196 degrees C were used as donors. In experiment 3, the effect of different cell cycle synchronization methods on the in vitro development potential of pig NTEs was evaluated. The cleavage rate of NTEs derived from cycling cells was much better than that of NTEs derived from serum-starved cells (64.4% vs 50.5%, p refrigerated pig GFP-transfected cells could be used as donors in nuclear transfer and these NTEs could be effectively developed to blastocyst stage; (ii) serum starvation of GFP-transfected cells is not required for preimplantation development of pig NTEs; and (iii) a rough surface of GFP-transfected donor cells affects fusion rate negatively but has no influence on the cleavage rate or blastocyst rate of pig NTEs. PMID:16822335

  6. Replication of somatic micronuclei in bovine enucleated oocytes

    Directory of Open Access Journals (Sweden)

    Canel Natalia

    2012-11-01

    Full Text Available Abstract Background Microcell-mediated chromosome transfer (MMCT was developed to introduce a low number of chromosomes into a host cell. We have designed a novel technique combining part of MMCT with somatic cell nuclear transfer, which consists of injecting a somatic micronucleus into an enucleated oocyte, and inducing its cellular machinery to replicate such micronucleus. It would allow the isolation and manipulation of a single or a low number of somatic chromosomes. Methods Micronuclei from adult bovine fibroblasts were produced by incubation in 0.05 μg/ml demecolcine for 46 h followed by 2 mg/ml mitomycin for 2 h. Cells were finally treated with 10 μg/ml cytochalasin B for 1 h. In vitro matured bovine oocytes were mechanically enucleated and intracytoplasmatically injected with one somatic micronucleus, which had been previously exposed [Micronucleus- injected (+] or not [Micronucleus- injected (−] to a transgene (50 ng/μl pCX-EGFP during 5 min. Enucleated oocytes [Enucleated (+] and parthenogenetic [Parthenogenetic (+] controls were injected into the cytoplasm with less than 10 pl of PVP containing 50 ng/μl pCX-EGFP. A non-injected parthenogenetic control [Parthenogenetic (−] was also included. Two hours after injection, oocytes and reconstituted embryos were activated by incubation in 5 μM ionomycin for 4 min + 1.9 mM 6-DMAP for 3 h. Cleavage stage and egfp expression were evaluated. DNA replication was confirmed by DAPI staining. On day 2, Micronucleus- injected (−, Parthenogenetic (− and in vitro fertilized (IVF embryos were karyotyped. Differences among treatments were determined by Fisher′s exact test (p≤0.05. Results All the experimental groups underwent the first cell divisions. Interestingly, a low number of Micronucleus-injected embryos showed egfp expression. DAPI staining confirmed replication of micronuclei in most of the evaluated embryos. Karyotype analysis revealed that all Micronucleus-injected embryos had

  7. New techniques on embryo manipulation.

    Science.gov (United States)

    Escribá, M J; Valbuena, D; Remohí, J; Pellicer, A; Simón, C

    2002-01-01

    For many years, experience has been accumulated on embryo and gamete manipulation in livestock animals. The present work is a review of these techniques and their possible application in human embryology in specific cases. It is possible to manipulate gametes at different levels, producing paternal or maternal haploid embryos (hemicloning), using different techniques including nuclear transfer. At the embryonic stage, considering practical, ethical and legal issues, techniques will be reviewed that include cloning and embryo splitting at the cleavage stage, morula, or blastocyst stage.

  8. Enhanced or reduced fetal growth induced by embryo transfer into smaller or larger breeds alters post-natal growth and metabolism in pre-weaning horses.

    Directory of Open Access Journals (Sweden)

    Pauline Peugnet

    Full Text Available In equids, placentation is diffuse and nutrient supply to the fetus is determined by uterine size. This correlates with maternal size and affects intra-uterine development and subsequent post-natal growth, as well as insulin sensitivity in the newborn. Long-term effects remain to be described. In this study, fetal growth was enhanced or restricted through ET using pony (P, saddlebred (S and draft (D horses. Control P-P (n = 21 and S-S (n = 28 pregnancies were obtained by AI. Enhanced and restricted pregnancies were obtained by transferring P or S embryos into D mares (P-D, n = 6 and S-D, n = 8 or S embryos into P mares (S-P, n = 6, respectively. Control and experimental foals were raised by their dams and recipient mothers, respectively. Weight gain, growth hormones and glucose homeostasis were investigated in the foals from birth to weaning. Fetal growth was enhanced in P-D and these foals remained consistently heavier, with reduced T3 concentrations until weaning compared to P-P. P-D had lower fasting glucose from days 30 to 200 and higher insulin secretion than P-P after IVGTT on day 3. Euglycemic clamps in the immediate post-weaning period revealed no difference in insulin sensitivity between P-D and P-P. Fetal growth was restricted in S-P and these foals remained consistently lighter until weaning compared to S-D, with elevated T3 concentrations in the newborn compared to S-S. S-P exhibited higher fasting glycemia than S-S and S-D from days 30 to 200. They had higher maximum increment in plasma glucose than S-D after IVGTT on day 3 and clamps on day 200 demonstrated higher insulin sensitivity compared to S-D. Neither the restricted nor the enhanced fetal environment affected IGF-1 concentrations. Thus, enhanced and restricted fetal and post-natal environments had combined effects that persisted until weaning. They induced different adaptive responses in post-natal glucose metabolism: an early insulin-resistance was

  9. Prevention of multiple pregnancies in couples with unexplained or mild male subfertility : Randomised controlled trial of in vitro fertilisation with single embryo transfer or in vitro fertilisation in modified natural cycle compared with intrauterine insemination with controlled ovarian hyperstimulation

    OpenAIRE

    Bensdorp, A J; Tjon-Kon-Fat, R. I.; Bossuyt, P.M.M.; Koks, C A M; Oosterhuis, G. J E; van Hoek, A.; Hompes, P.G.A.; Broekmans, F.J.M.; Verhoeve, H R; De Bruin, J. P.; van Golde, R.; Repping, S.; Cohlen, B.J.; Lambers, M D A; van Bommel, P F

    2015-01-01

    Objectives: To compare the effectiveness of in vitro fertilisation with single embryo transfer or in vitro fertilisation in a modified natural cycle with that of intrauterine insemination with controlled ovarian hyperstimulation in terms of a healthy child. Design: Multicentre, open label, three arm, parallel group, randomised controlled non-inferiority trial. Setting: 17 centres in the Netherlands. Participants: Couples seeking fertility treatment after at least 12 months of unprotected inte...

  10. Genetic Polymorphism of the Lactoferrin Gene in Dairy and Beef Cattles at National Artificial Insemination and Embryo Transfer Stations

    Directory of Open Access Journals (Sweden)

    Anneke Anggraeni

    2016-12-01

    Full Text Available Lactoferrin (LTF adalah gen pengontrol komponen protein susu dan memiliki karakteristik sebagai antimikrobial. LTF pada susu berfungsi untuk mencegah diare, sedangkan pada sapi laktasi untuk mencegah mastitis pada ambing. Mempertimbangkan peran penting dari gen LTF, maka perlu dilakukan peningkatan kadar LTF dalam susu melalui seleksi pada taraf DNA. Polymorfisme genetik dari gen LTF diidentifikasi pada sapi perah dan potong dengan metoda Polymerase Chain Reaction - Restricsion Fragment Length Polymorphism (PCR-RFLP, dengan enzim restriksi EcoRI. Genotyping dilakukan pada sapi perah Friesian Holstein (FH total sejumlah 89 ekor, meliputi dari Balai Inseminasi Buatan Lembang (BIB Lembang untuk 17 pejantan, Balai Besar IB Singosari (BBIB Singosari untuk 32 pejantan, dan Balai embrio Transfer Cipelang (BET Cipelang pada 40 dara. Genotyping dilakukan pula pada sapi potong dara berasal dari empat bangsa, meliputi Limousin (14 ekor, Angus (5 ekor, Simmental (13 ekor dan Brahman (5 ekor dari BET Cipelang. Gen LTF|EcoRI pada sapi perah dan potong pengamatan menghasilkan dua tipe alel, yaitu alel A dan B. Kedua jenis sapi tersebut menghasilkan hanya dua genotipe, yaitu genotipe AA dan AB, tanpa genotipe BB. Ini dapat menjadi hal yang baik karena genotipe AA dan AB dipertimbangkan berasosiasi dengan ketahan pada mastitis. Nilai-nilai dari heterozygositas observasi (Ho dari gen ini lebih tinggi dibandingkan heterozigositas ekspektasi (He. Disimpulkan bahwa gen LTF|EcoRI memiliki variasi yang baik pada sapi perah dan sapi potong dari ketiga balai bibit nasional tersebut.

  11. Cytogenetics and immature embryo culture at Embrapa Trigo breeding program: transfer of disease resistance from related species by artificial resynthesis of hexaploid wheat (Triticum aestivum L. em. Thell

    Directory of Open Access Journals (Sweden)

    Maria Irene Baggio de Moraes Fernandes

    2000-12-01

    to facilitate gene flow between wheat and related species. Since the environment at the center of origin of wheat in Southern Asia is quite different from subtropical environments, Brazilian breeding programs overcome more challenges to adapt wheat crop to biotic and abiotic stresses than some other countries. The germplasm bank of Embrapa Trigo has about 1000 registered entries of Triticum relatives, Aegilops, Secale and Agropyron species supplied from several germplasm banks distributed over the world which were multiplied and/or selected for naturally occurring or artificially inoculated fungal diseases. Since Aegilops squarrosa L. entries showed very good performance, the genetic variability observed in this species was firstly exploited. It is reported here the strategy used for transferring useful genes from Ae. squarrosa (DD, 2n = 14: crossing with tetraploid species (AABB, 2n = 28, rescue and in vitro culture of immature embryos for regeneration of the trihaploid (ABD, 2n = 21 hybrid, and colchicine treatment for genome duplication resulting in the artificial synthesis of hexaploid wheat lines (AABBDD, 2n = 42. Results of 10,739 artificial pollinations involving 28 cross combinations amongst eight T. durum L., T. dicoccum and T. cartlicum tetraploid entries used as female parents and ten selected Ae. squarrosa sources of resistance as male parents are presented here. Immature embryos from 18 cross combinations were recovered and cultured in vitro. Green plantlets from 13 combinations were regenerated. Fertile amphiploids were recovered only from crosses among entries of tetraploid T. durum and diploid Ae. squarrosa. They originated 11 fertile synthetic amphiploid lines from seven different combinations. Useful stem and leaf rust as well as powdery mildew resistance for future use in breeding programs were obtained.

  12. Efeito do citrato e taurina em meio CR2aa no desenvolvimento de embriões bovinos fecundados in vitro Effect of citrate and taurine added to CR2aa medium on the development of in vitro-fertilized bovine embryos

    Directory of Open Access Journals (Sweden)

    L.S.A. Camargo

    2009-02-01

    Full Text Available Avaliou-se o efeito do citrato em meio CR2aa suplementado com soro fetal bovino (SFB ou livre de proteínas séricas e sua associação com taurina no desenvolvimento de embriões bovinos fecundados in vitro. Embriões foram cultivados em CR2aa contendo 0, 0,5, 1,0 e 3,0mM citrato, suplementado com 10% SFB (experimento 1 ou com álcool polivinil (PVA; experimento 2. No terceiro experimento, embriões foram cultivados em meio com 0,5mM citrato, ou 7mM taurina, ou com a associação de ambos, suplementado com SFB. Os cultivos foram realizados com células do cumulus em ambiente a 38,8ºC com 5% de CO2 em ar atmosférico. Melhora no desenvolvimento embrionário foi observado no cultivo de embriões em CR2aa com 0,5 e 1,0mM citrato na ausência de SFB (P0,05 a produção de embriões ou o número de células. Citrato em meio CR2aa pode ser uma alternativa para cultivo embrionário em condições atmosféricas com 5% de CO2 em ar na ausência de proteína sérica.The effect of citrate added to CR2aa medium supplemented with fetal calf serum (FCS or serum-proteinfree and its association with taurine on the development of in vitro-fertilized bovine embryos was evaluated. Embryos were cultured with 0, 0.5, 1.0, and 3.0mM citrate, in CR2aa supplemented with 10% FCS (experiment 1, or polyvinyl alcohol (PVA; experiment 2. In experiment 3, embryos were cultured with 0.5mM citrate, 7.0mM taurine or with association of both, in medium supplemented with FCS. Embryo culture was performed with cumulus cells at 38.8ºC in 5% CO2 under air for all experiments. Positive effect on embryo development was only observed with 0.5 and 1.0mM citrate in FCS-free CR2aa (P0.05 embryo rate nor total cell number. Citrate in CR2aa medium can be an alternative for serumfree embryo culture under 5% CO2 in air, absence of serum protein.

  13. I型牛疱疹病毒通用型转移载体的构建%Construction of a Universal Bovine Herpesvirus I Transfer Vector

    Institute of Scientific and Technical Information of China (English)

    李继昌; 童光志; 仇华吉; 周艳君; 张桂红; 王柳; 刘忠贵

    2001-01-01

    将I型牛疱疹病毒(BHV-1)LA株DNA HindIII A片段中的SalI-SalI亚片段(含TK基因)克隆到载体质粒pBluescript SK中,再用BglII和SacI切去347 bp,获得含TK基因部分缺失的重组质粒pSdTK,然后用HindIII和XbaI切去其中的多克隆位点;将来源于pCR3-Uni的CMV启动子、多克隆位点和BGH polyA信号插入pSdTK的XhoI位点上,构建了BHV-1通用转移载体pSdTK-CMB,此载体可用来表达牛其它病毒的抗原基因,为开发二价或多价基因工程疫苗提供基础。%Bovine herpes irus-1 strain LA DNA HindIII A was digested by the restriction endonuclease SalI,The SalI subfragment of 2.7kb containing thymidine kinase(TK)gene was cloned into pBluescript SK,resulting in a recombinant pSTK,Then it was digested by BglII and SacI,the 5.31kb fragment was recovered and self-ligated,resulting in a recombinant pSdTK,the multiple colning sites were removed by HindIII and XbaI;A fragment containing the immediate early promoter of cytomegalovirus,multiple cloning sites and bovine growth hormone polyadenylation signal derived from pCR3-Uni was amplified and inserted into the XhoI site of pSdTK.The resulting transfer vector pSdTK-CMB can be used to expess genes from other bovine viruses(i.e. BPIV3、BRSV、BVDV).

  14. Characterisation of bovine epiblast-derived outgrowth colonies

    DEFF Research Database (Denmark)

    Østrup, Esben; Gjørret, Jakob; Schauser, Kirsten Hallundbæk;

    2010-01-01

    The aim of the present study was to characterise bovine epiblast-derived outgrowth colonies (OCs) with respect to the embryonic origin of their cellular components. Epiblasts were isolated mechanically from bovine Day 12 embryos. Epiblasts were cultured on feeder layers of SNL cells (neomycin...

  15. Synergistic Effect of Insulin on in vitro Development of Immature Bovine Oocytes

    OpenAIRE

    Mojtaba Dashtizad; Abd W. Haron; Rosnina Yusoff; Morteza Daliri; Hadi Hajarian; Mehdi Najari; Yap K. Chee; Abas M. Othman

    2010-01-01

    Problem statement: Development of efficient culture system to support embryonic development would be valuable when quality of produced embryos was important. However, the rate of bovine embryo production in vitro was still lower than expected. Present study, including of three experiments, was carried out to investigate the effect of insulin on nuclear maturation and subsequent development of immature bovine oocytes and in vitro fertilized embryos. Approach: Grade one cumulus-oocyte-complexes...

  16. Applications of Tol2 Transposon-Mediated Gene Transfer for Stable Integration and Conditional Expression of Electroporated Genes in Chicken Embryos

    Science.gov (United States)

    Sato, Yuki; Takahashi, Yoshiko

    Because of the high accessibility to developing embryos, avian embryos (chicken and quail) have long been used as a good model animal to study embryogenesis in vertebrates, especially amniotes (reviewed in Wolpert, 2004). The techniques used for “classical” avian embryology included tissue transplantations, tissue ablations, and cell-labeling by vital dye. At the end of the last century, the in ovo electropora tion technique was developed by Nakamura and his colleagues, and this modern method opened a way to study the roles of developmental genes directly in living embryos (Funahashi et al., 1999) reviewed in (Nakamura et al., 2004; Yasuda et al., 2000; Yasugi and Nakamura, 2000). This powerful technique allows us to introduce genes (DNA, RNA, morpholino) into embryos in a tissue-specific way by targeting a restricted area of embryonic tissues. Thus, the electroporation technique using chickens has provided numerous novel insights into the understanding of early development in vertebrates, making the chicken a unique model animal.

  17. Cryopreservation of manipulated embryos: tackling the double jeopardy.

    Science.gov (United States)

    Dinnyes, A; Nedambale, T L

    2009-01-01

    The aim of the present review is to provide information to researchers and practitioners concerning the reasons for the altered viability and the medium- and long-term consequences of cryopreservation of manipulated mammalian embryos. Embryo manipulation is defined herein as the act or process of manipulating mammalian embryos, including superovulation, AI, IVM, IVF, in vitro culture, intracytoplasmic sperm injection, embryo biopsy or splitting, somatic cell nuclear transfer cloning, the production of sexed embryos (by sperm sexing), embryo cryopreservation, embryo transfer or the creation of genetically modified (transgenic) embryos. With advances in manipulation technologies, the application of embryo manipulation will become more frequent; the proper prevention and management of the resulting alterations will be crucial in establishing an economically viable animal breeding technology.

  18. Production of transgenic embryos through nuclear transfer using ovine fetal fibroblasts transferred with foreign genes%体细胞核移植生产绵羊转hALR基因囊胚

    Institute of Scientific and Technical Information of China (English)

    马玉珍; 任宇; 周雪原; 刘东军; 旭日干

    2011-01-01

    扩增人肝细胞再生增强因子(human augmenter of liver regeneration,ALR)基因,利用质粒pIRES2-EGFP 构建新霉素(Neo)、增强绿色荧光蛋白(enhanced green fluorescence protein,EGFP)双标记基因且EGFP和ALR基因为双顺反子的真核表达载体.LipofectAMINETM介导其转染体外培养的绵羊胎儿成纤维细胞(sheep fetal fibroblast cells,sFFCs);经G418筛选转基因细胞;激光共聚焦显微镜挑选绿色荧光单克隆细胞.PCR、RT-PCR和免疫组织化学方法进一步检测ALR基因及其表达;稳定表达外源基因的sFFCs作供体,移入去核的绵羊卵母细胞中,进行体细胞核移植.通过激光共聚焦显微镜和ALR抗体检测EGFP、ALR基因在胚胎水平上的表达,其结果表明:由IRES连接的EGFP和ALR基因可在绵羊胎儿成纤维细胞内同时表达,由此细胞核移植产生的转基因胚胎在发育的各阶段均可见绿色荧光;囊胚中所有细胞表达EGFP基因;发绿色荧光的胚胎中ALR基因同时存在.因此,由IRES连接标记基因和目的基因,以标记基因指示目的基因的表达,可简化检测目的基因的繁琐手段;用筛选的转基因早期胚胎进行移植,可提高制备转基因动物的效率.%Human ALR gene sequence was amplified by PCR from human total DNA and inserted into PIRES2-EGFP vector. The bicistronic eukaryotic expression vector, pIRES-EGFP/ALR, expressing EGFP, Neo and ALR genes was constructed. Sheep fetal fibroblast cells (sEFCs) were transfected with pIRES-EGFP/AZJ? By the induction of lipofectAMINE?. The positive cell clones were selected with medium containing G418 (800 ug/mL). The fluorescence of transgenic cells was examined with a confocal laser scanning microscope. The expression of ALR gene was tested by PCR, RT-PCR and immuno-histochemical staining. The transgenic cells were used as donors for nuclear transfer to enucleated ovine oocytes. Transgenic embryos were tested by confocal laser scanning microscope and

  19. Analysis of Related Factors Influencing the Cleavage Stage Outcome of Frozen Thawed Embryo Transfer%探析卵裂期胚胎解冻移植结局相关影响原因

    Institute of Scientific and Technical Information of China (English)

    王琦; 陈蔚清; 张程

    2015-01-01

    目的:探析人卵裂期胚胎解冻移植结局的主要影响因素。方法研究对象是2012年2月1日~2014年2月1日在我院接受冻融胚胎移植的患者,解冻周期205个。结果玻璃化冷冻组的可移植胚胎率、胚胎复苏卵裂球存活率、临床妊娠率显著高于程序化冷冻组,两组有显著差别(0.05). Conclusion The method of embryo freezingsuitable is the main factor af ecting the cleavage stagefrozen thawed embryo transfer outcomes.

  20. 胚胎移植方法和受体母猪因素对克隆猪生产效率的影响%Effect of Recipient Status and Embryo Transfer Methods on Production of Cloned Pigs

    Institute of Scientific and Technical Information of China (English)

    卫恒习; 李秋艳; 高凤磊; 李燕; 张守全; 李宁

    2012-01-01

    [Objective] In order to establish an efficient embryo transfer technology and improve the production efficiency of cloned pigs, the present study was investigated different embryo transfer methods, gestational age and synchronization status of recipients on cloning pigs. [Method] Cloned embryos ware transferred with different methods to different status recipient, the delivery rates and the cloning efficiency were compared. The optimal synchronization time of recipient was determined by using co-transfer of different kinds of cloned embryos with similar developmental ability at different development stages. [Result] The method of transferring embryos through oviduct umbrella receive higher delivery rate and cloning efficiency (2.2% vs 0.4%, P<0.01) than the method of oviduct puncture, and insemination after embryos transfer had a negative effect on cloning efficiency (0.6% vs 2.2%, P<0.01). The litter size and cloning efficiency were higher when using sows as recipient than gilts ((5.5±0.7) vs (2.7±0.3), P <0.05 and 3.0% vs 0.8%, P<0.01 respectively). Higher cloning efficiency was found in the group of recipient estrous time posterior of 12-36 h to the embryo activation when compared to the meanwhile and prior of 12-24 h groups (2.0% vs 0.5% vs 0%, P<0.05), and the optimal recipient synchronization time is the time of estrous posterior 24 h to embryo activation, and the cloning efficiency reached 3.0%. [ Conclusion ] An efficient embryos transfer technology was established in pig cloning by transferring embryos through oviduct umbrella and by using natural estrous sows beginning heat posterior 24 h to embryo activation as recipient.%[目的]建立高效的胚胎移植技术体系,以提高克隆猪的生产效率.[方法]比较不同移植方法和不同受体母猪状况的胚胎移植分娩率和克隆猪的出生效率,利用体外发育能力相似的不同品种和发育阶段的克隆胚胎混合移植确定最佳的受体母猪发情同期时间.[结果]

  1. Lowering storage temperature during ovary transport is beneficial to the developmental competence of bovine oocytes used for somatic cell nuclear transfer.

    Science.gov (United States)

    Wang, Y S; Zhao, X; Su, J M; An, Z X; Xiong, X R; Wang, L J; Liu, J; Quan, F S; Hua, S; Zhang, Y

    2011-03-01

    The objective of this study was to determine the effect of storage temperature during ovary transport on the developmental competence of bovine oocytes for use in somatic cell nuclear transfer (SCNT). Ovaries obtained from a slaughterhouse were stored in physiological saline for 3-4h at one of the three temperatures: 15 °C, 25 °C, or 35 °C. The developmental competence of oocytes used for SCNT was ascertained by cleavage and blastocyst formation rate, total cell number, apoptosis index, and the relative abundance of Bax and Hsp70.1 in day 7 blastocysts. Ovaries stored at 35 °C for 3-4h reduced the recovery rate of grade I and II oocytes compared with those stored at 25 °C or 15 °C (45.1±0.7% vs. 76.7±1.2% or 74.8±2.0%, Povaries stored at 15 °C, however, produced blastocysts with higher cell numbers (97.3±8.6 vs. 80.2±10.8 or 77.4±11.7; Povaries stored at 15 °C was lower than those stored at 25 °C or 35 °C (Pquality and developmental competence of oocytes used for SCNT due to the alleviation of stresses on the oocytes compared with those subjected to storage temperatures of 25 °C or 35 °C. PMID:21333472

  2. Contributions of the maternal uterine environment and piglet genotype on weaning survivability potential: I. Development of neonatal piglets after reciprocal embryo transfers between Meishan and White crossbred gilts.

    Science.gov (United States)

    Miles, J R; Vallet, J L; Ford, J J; Freking, B A; Cushman, R A; Oliver, W T; Rempel, L A

    2012-07-01

    In commercial pigs, the greatest susceptibility for pre-weaning mortality occurs in low birth-weight piglets. Despite their overall decreased birth weight, Meishan (MS) piglets have decreased pre-weaning mortality rates compared with contemporary Western breeds. The objective of the current study was to determine the contributions of the maternal uterine environment, piglet genotype, and their interaction on the development of neonatal piglets pertaining to pre-weaning survivability using reciprocal embryo transfer between MS and White crossbred (WC) pigs. Twenty-five successful pregnancies were produced from 2 farrowing seasons, generating litters of maternal uterine environment (MUE) by piglet genotype (PigG) combinations; MS × MS (n = 4 litters), MS × WC (n = 7 litters), WC × MS (n = 7 litters), and WC × WC (n = 7 litters). At approximately 24 h of age (Day 1), piglets (n = 173) were weighed and a blood sample was taken. Hematocrit, hemoglobin, glucose, plasma urea nitrogen, albumin, NEFA, lactate, and cortisol were measured in all blood samples. Representative piglets (n = 46) from each litter were harvested and body measurements (i.e., organ weights, tissue glycogen content, and body composition) were determined. Piglet data were analyzed by ANOVA using MIXED model procedures. Both MUE (P piglet BW, illustrating that piglets gestated in WC gilts were heavier than piglets gestated in MS gilts, and WC piglets were heavier than MS piglets. Serum albumin concentrations were increased (P piglets compared with WC piglets, indicating greater liver maturity. Significant MUE × PigG interactions were observed for hematocrit and hemoglobin, in which the greatest concentrations were observed in MS piglets gestated in MS and WC gilts, and the lowest concentrations were observed in WC piglets gestated in WC gilts, demonstrating increased oxygen-carrying capability. The percentage of fat and nitrogen, as well as the GE of the body, were greater (P piglets, indicating

  3. Cats cloned from fetal and adult somatic cells by nuclear transfer.

    Science.gov (United States)

    Yin, X J; Lee, H S; Lee, Y H; Seo, Y I; Jeon, S J; Choi, E G; Cho, S J; Cho, S G; Min, W; Kang, S K; Hwang, W S; Kong, I K

    2005-02-01

    This work was undertaken in order to study the developmental competence of nuclear transfer (NT) into cat embryos using fetal fibroblast and adult skin fibroblast cells as donor nuclei. Oocytes were recovered by mincing the ovaries in Hepes-buffered TCM199 and selecting the cumulus oocyte complexes (COCs) with compact cumulus cell mass and dark color. Homogenous ooplasm was cultured for maturation in TCM199+10% fetal bovine serum (FBS) for 12 h and used as a source of recipient cytoplast for exogenous somatic nuclei. In experiment 1, we evaluated the effect of donor cell type on the reconstruction and development of cloned embryos. Fusion, first cleavage and blastocyst developmental rate were not different between fetal fibroblasts and adult skin cells (71.2 vs 66.8; 71.0 vs 57.6; 4.0 vs 6.1% respectively; P < 0.05). In experiment 2, cloned embryos were surgically transferred into the oviducts of recipient queens. One of the seven recipient queens was delivered naturally of 2 healthy cloned cats and 1 stillborn from fetal fibroblast cells of male origin 65 days after embryo transfer. One of three recipient queens was delivered naturally of 1 healthy cloned cat from adult skin cells of female origin 65 days after embryo transfer. The cloned cats showed genotypes identical to the donor cell lines, indicating that adult somatic cells can be used for feline cloning. PMID:15695619

  4. COMPARAÇÃO ENTRE DOIS MEIOS PARA TRANSFERÊNCIA DE EMBRIÕES EM ÉGUAS DA RAÇA MANGALARGA MARCHADOR OMPARISON BETWEEN TWO MEDIUM FOR EMBRYO TRANSFER IN MARES FROM MANGALARGA MARCHADOR BREED

    Directory of Open Access Journals (Sweden)

    José Renato Costa Caiado

    2009-09-01

    Full Text Available

    Diferentes tampões são utilizados nos meios de lavagem e manutenção embrionária durante os procedimentos de transferência de embriões em equinos (TEE. Os mais usados são carbonato, fosfato e zwitteriônico. Durante os procedimentos de transferência é comum que o embrião permaneça por um tempo maior que o previsto em espera no meio de manutenção utilizado. Este experimento foi realizado com o objetivo de comparar o custo–benefício de dois meios contendo tampões diferentes DPBS (fosfato e Embriocare® (zwitteriônico, na transferência de embriões em éguas da raça Mangalarga Marchador. Dividiram-se os embriões aleatoriamente em dois tratamentos – zwitteriônico e fosfato –, permanecendo em descanso por 0, 30, 60 ou 120 minutos em ambos os meios. Os resultados indicam que esses tampões são igualmente eficientes (P > 0,05 nos processos de rasteamento, lavagem, manutenção à temperatura ambiente por diferentes períodos de tempo (0 a 120 minutos, sendo que as taxas de prenhez obtidas em ambiente tropical foram de 65% e de 69,7%, respectivamente. Porém, obteve-se a maior taxa de prenhez encontrada com DPBS com um tempo de manutenção do embrião por sessenta minutos no meio (P < 0,05. No tampão zwitteriônico, a maior taxa de prenhez foi alcançada com trinta minutos no meio (P < 0,05. Como a aquisição do Embriocare® foi mais onerosa, o DPBS apresentou o melhor custo–benefício.

    PALAVRAS-CHAVES: Equino, Mangalarga Marchador, transferência de embriões, tampão fosfato e zwitteriônico.

    Palavras-chave: eqüino, Mangalarga Marchador, transferência de embriões, tampão fosfato e zwitteriõnico. Different buffers are used in the solution for washing and embryo maintenance during the procedures of equine embryo transfer

  5. Cryopreservation of Embryos and Oocytes in Human Assisted Reproduction

    Directory of Open Access Journals (Sweden)

    János Konc

    2014-01-01

    Full Text Available Both sperm and embryo cryopreservation have become routine procedures in human assisted reproduction and oocyte cryopreservation is being introduced into clinical practice and is getting more and more widely used. Embryo cryopreservation has decreased the number of fresh embryo transfers and maximized the effectiveness of the IVF cycle. The data shows that women who had transfers of fresh and frozen embryos obtained 8% additional births by using their cryopreserved embryos. Oocyte cryopreservation offers more advantages compared to embryo freezing, such as fertility preservation in women at risk of losing fertility due to oncological treatment or chronic disease, egg donation, and postponing childbirth, and eliminates religious and/or other ethical, legal, and moral concerns of embryo freezing. In this review, the basic principles, methodology, and practical experiences as well as safety and other aspects concerning slow cooling and ultrarapid cooling (vitrification of human embryos and oocytes are summarized.

  6. Laser confers less embryo exposure than acid tyrode for embryo biopsy in preimplantation genetic diagnosis cycles: a randomized study

    Directory of Open Access Journals (Sweden)

    Valle Marcelo

    2011-04-01

    Full Text Available Abstract We compared two methods of zona pellucida drilling. 213 embryos were biopsied with acid Tyrode. Each biopsy took 3 minutes and the entire procedure ~29 minutes. 5% of blastomeres lysed, 49% of embryos became blastocyst and 36% of patients became pregnant. 229 embryos were biopsied with laser. Each biopsy took 30 seconds and the entire procedure ~7 minutes. 2.5% of blastomeres lysed, 50.6% of embryos became blastocyst and 47% of patients became pregnant. We can conclude that laser can be used for embryo biopsy. Reduction of embryo exposure and of removed blastomeres is associated with increased blastocysts available for transfer and a better clinical outcome.

  7. Impact of Cryopreservation Timing on Clinical Outcomes of Frozen-thawed Embryo Transfer Cycles%冷冻时机对冻融胚胎移植结局的影响

    Institute of Scientific and Technical Information of China (English)

    邓朝晖; 高士友; 柳朝华; 李昀

    2015-01-01

    Objective]To explore the impact of cryopreservation timing on the clinical outcomes of frozen‐thawed embryo transfer cycles (FET ) .[Methods] A retrospective analysis was performed for 420 FET cy‐cles .The patients were divided into different groups according to freezing times .And frozen‐thawed effects and clinical outcomes of differential cryopreservation timing were observed .[Results] The post‐thaw embryo survival rate ,implantation rate and clinical pregnancy rate of d4 embryo were higher than those of d2 and d3 embryos (83 .45% vs 77 .91% & 75 .12% ;37 .90% vs 27 .98% & 33 .33% ;61 .70% vs 50 .00% &52 .02% ) .The abortion rate of d3 embryo was lower than that of d2 and d4 embryos (4 .85% vs 12 .24% , 10 .34% ) .However ,no significant difference existed in those rates among d2 ,d3 and d4 embryos ( P >0 .05) .[Conclusion] No significant differences existed in post‐thaw embryo survival rate ,implantation rate , clinical pregnancy rate or abortion rate of different frozen‐thawed embryo development stages .And cryopreser‐vation timing should be selected flexibly according to demand and patient status .%【目的】探讨冷冻时机对冻融胚胎移植(FET )结局的影响。【方法】回顾性分析420个FET周期,按冷冻时机(d2~d4)进行分组,比较不同冷冻时机时的复苏效果和移植结局。【结果】d4的复苏率、种植率、临床妊娠率分别为83.45%、397.90%、61.70%,d2分别为77.91%、27.98%、50.00%),d3 为75.12%、33.33%、52.02%,但各组之间的复苏率、种植率、临床妊娠率、流产率比较无统计学差异( P > 0.05)。【结论】胚胎发育不同时机进行冷冻,解冻后胚胎的复苏率、种植率、临床妊娠率及流产率无明显差异,提示可根据工作需要及病人情况灵活选用冷冻时间。

  8. Embryo technologies in the horse.

    Science.gov (United States)

    Squires, E L; Carnevale, E M; McCue, P M; Bruemmer, J E

    2003-01-01

    Recent studies demonstrated that zwitterionic buffers could be used for satisfactory storage of equine embryos at 5 degrees C. The success of freezing embryos is dependent upon size and stage of development. Morulae and blastocysts transfer. The majority of equine embryos are collected from single ovulating mares, as there is no commercially available product for superovulation in equine. However, pituitary extract, rich in FSH, can be used to increase embryo recovery three- to four-fold. Similar to human medicine, assisted reproductive techniques have been developed for the older, subfertile mare. Transfer of in vivo-matured oocytes from young, healthy mares into a recipient's oviduct results in a 70-80% pregnancy rate compared with a 30-40% pregnancy rate when the oocytes are from older, subfertile mares. This procedure can also be used to evaluate in vitro maturation systems. In vitro production of embryos is still quite difficult in the horse. However, intracytoplasmic sperm injection (ICSI) has been used to produce several foals. Cleavage rates of 60% and blastocyst rates of 30% have been reported after ICSI of in vitro-matured oocytes. Gamete intrafallopian tube transfer (GIFT) is a possible treatment for subfertile stallions. Transfer of in vivo-matured oocytes with 200,000 sperm into the oviduct of normal mares resulted in a pregnancy rate of 55-82%. Oocyte freezing is a technique that has proven difficult in most species. However, equine oocytes vitrified in a solution of ethylene glycol, DMSO, and Ficoll and loaded onto a cryoloop resulted in three pregnancies of 26 transfers and two live foals produced. Production of a cloned horse appears to be likely, as several cloned pregnancies have recently been produced. PMID:12499026

  9. 心理干预对体外受精-胚胎移植临床妊娠率影响的Meta分析%Influence of psychological intervention on pregnancy rate of in vitro fertilization-embryo transfer

    Institute of Scientific and Technical Information of China (English)

    苏霞; 丛云凤

    2013-01-01

    Objective To evaluate the influence of psychological intervention on pregnancy rate of in vitro fertilization-embryo transfer.Methods Using the key words,including in vitro fertilization-embryo transfer and psychological,the data were retrieved from CNKI,CBM,VIP,Wanfang and PubMed.The quality of eligible studies was evaluated by two reviews independently and Meta-analysis was conducted on studies.Results Totally thirteen Chinese studies and three foreign studies were included.Conclusions Psychological intervention using in the process of in vitro fertilization-embryo transfer can improve the pregnancy rate,which is worthy of clinical application.%目的 评价心理干预对体外受精-胚胎移植临床妊娠率的影响.方法 通过计算机检索中文文献数据库,包括中国知网(CNKI)、中国生物医学文献数据库(CBM)、维普、万方等,关键词为心理护理、体外受精;外文数据库Pubmed,关键词包括in vitro fertilization-embryo transfer、psychological.获取文献之后按照入选标准要求筛选相关文献,选择合适文献纳入进行相关数据的Meta分析,探讨心理干预对体外受精-胚胎移植临床妊娠率的影响.结果 共有13篇中文文献,3篇外文文献纳入,进行Meta分析.结论 心理干预应贯穿于体外受精-胚胎移植的过程之中,可以提高体外受精-胚胎移植患者的妊娠成功率,是值得推广应用的护理措施.

  10. Role of hydrogen-bonding and photoinduced electron transfer (PET) on the interaction of resorcinol based acridinedione dyes with Bovine Serum Albumin (BSA) in water

    Energy Technology Data Exchange (ETDEWEB)

    Kumaran, Rajendran, E-mail: kumaranwau@rediffmail.com [Department of Chemistry, Dwaraka Doss Goverdhan Doss, Vaishnav College (Autonomous), 833, Gokul Bagh, E.V.R. Periyar Road, Arumbakkam, Chennai 600106, Tamil Nadu (India); Vanjinathan, Mahalingam [Department of Chemistry, Dwaraka Doss Goverdhan Doss, Vaishnav College (Autonomous), 833, Gokul Bagh, E.V.R. Periyar Road, Arumbakkam, Chennai 600106, Tamil Nadu (India); Ramamurthy, Perumal [National Centre for Ultrafast Processes, University of Madras, Taramani Campus Chennai 600113, Tamil Nadu (India)

    2015-08-15

    Resorcinol based acridinedione (ADDR) dyes are a class of laser dyes and have structural similarity with purine derivatives, nicotinamide adenine dinucleotide (NADH) analogs. These dyes are classified into photoinduced electron transfer (PET) and non-photoinduced electron transfer dyes, and the photophysical properties of family of these dyes exhibiting PET behavior are entirely different from that of non-PET dyes. The PET process in ADDR dyes is governed by the solvent polarity such that an ADDR dye exhibits PET process through space in an aprotic solvent like acetonitrile and does not exhibit the same in protic solvents like water and methanol. A comparison on the fluorescence emission, lifetime and nature of interaction of various ADDR dyes with a large globular protein like Bovine Serum Albumin (BSA) was carried out in aqueous solution. The interaction of PET based ADDR dyes with BSA in water is found to be largely hydrophobic, but hydrogen-bonding interaction of BSA with dye molecule influences the fluorescence emission of the dye and shifts the emission towards red region. Fluorescence spectral studies reveal that the excited state properties of PET based ADDR dyes are largely influenced by the addition of BSA. The microenvironment around the dye results in significant change in the fluorescence lifetime and emission. Fluorescence enhancement with a red shift in the emission results after the addition of BSA to ADDR dyes containing free amino hydrogen in the 10th position of basic acridinedione dye. The amino hydrogen (N–H) in the 10th position of ADDR dye is replaced by methyl group (N–CH{sub 3}), a significant decrease in the fluorescence intensity with no apparent shift in the emission maximum was observed after the addition of BSA. The nature of interaction between ADDR dyes with BSA is hydrogen-bonding and the dye remains unbound even at the highest concentration of BSA. Circular Dichroism (CD) studies show that the addition of dye to BSA results in

  11. Birth of live calves by in vitro embryo production of slaughtered cows in a commercial herd in South Africa

    Directory of Open Access Journals (Sweden)

    T. Arlotto

    2001-07-01

    Full Text Available In vitro fertilisation (IVF has become a useful breeding tool in most of the developed world. In this paper the success of bovine IVF and the birth of live calves under typical South African conditions is reported. Oocytes for IVF were collected from the ovaries of 6 slaughtered Bovelder beef cows. On average, 36.2 oocytes per donor were retrieved. From these oocytes, 43 blastocysts were produced from 5 of the donors by IVF with frozen Bovelder semen. The best 11 of these embryos were transferred into oestrous, synchronised Bovelder recipients in the same herd. As a result, 7 calves were born (a 64 %calving rate from 4 of the original donors. The calves had a normal birth mass, but the mean gestation length of the male calves was significantly longer than the herd average (291.6 versus 285.2 days respectively. No calving difficulties were encountered. In summary, it was shown that IVF for bovine embryo production and transfer is possible on a commercial basis in South Africa.

  12. Patients with polycystic ovary syndrome have successful embryo arrest

    OpenAIRE

    Yin, Baoli; Hao, Haoying; Wei, Duo; Song, Xiaobing; Xie, Juanke; Zhang, Cuilian

    2015-01-01

    In this retrospective study, we investigate the relationship between embryo arrest and polycystic ovary syndrome (PCOS) during in vitro fertilization-embryo transfer (IVF-ET). In this study, 667 subjects were enrolled, including 330 patients with PCOS and 337 subjects without PCOS. The subjects underwent in vitro fertilization/intracytoplasmic sperm injection and embryo transfer (IVF/ICSI-ET) cycles at the Reproductive Medical Centre of Henan Provincial Hospital from January 2009 to December ...

  13. Melatonin in maturation media fails to improve oocyte maturation, embryo development rates and DNA damage of bovine embryos Melatonina no meio de maturação não melhorou as taxas de maturação dos ovócitos, de desenvolvimento embrionário e a fragmentação do DNA dos embriões bovinos

    Directory of Open Access Journals (Sweden)

    Luciana Takada

    2010-08-01

    Full Text Available Melatonin (MEL acts as a powerful scavenger of free radicals and direct gonadal responses to melatonin have been reported in the literature. Few studies, however, have evaluated the effect of MEL during in vitro maturation (IVM on bovine embryos. This study tested the addition of MEL to maturation medium (MM with no gonadotropins on nuclear maturation and embryo development rates and the incidence of DNA damage in resulting embryos. Cumulus-oocyte complexes were aspirated from abattoir ovaries and cultured in MM (TCM-199 medium supplemented with 10% fetal calf serum - FCS at 39ºC and 5% CO2 in air. After 24 hours of culture in MM with 0.5 µg mL-1 FSH and 5.0 µg mL-1 LH; 10-9 M MEL or 10-9 M MEL, 0.5 µg mL-1 FSH and 5.0 µg mL-1 LH, the oocytes were stained with Hoechst 33342 to evaluate nuclear maturation rate. After in vitro fertilization and embryo culture, development rates were evaluated and the blastocysts were assessed for DNA damage by Comet assay. There was no effect of melatonin added to the MM, alone or in combination with gonadotropins, on nuclear maturation, cleavage and blastocyst rates. These rates ranged between 88% to 90%, 85% to 88% and 42% to 46%, respectively. The extent of DNA damage in embryos was also not affected by MEL supplementation during IVM. The addition of 10-9 M MEL to the MM failed to improve nuclear maturation and embryo development rates and the incidence of DNA damage in resulting embryos, but was able to properly substitute for gonadotropins during IVM.Melatonin (MEL atua como um potente redutor de radicais livres. Efeito direto da MEL na função gonadal também foi observado. Existem poucos estudos relacionados ao efeito da MEL durante a maturação no desenvolvimento embrionário in vitro. Avaliou-se a adição de MEL no meio de maturação (sem gonadotrofinas nas taxas de maturação nuclear e de desenvolvimento embrionário e na incidência de fragmentação do DNA nos embriões produzidos in vitro

  14. 冻融胚胎和冻融囊胚对移植周期和分娩结局影响的比较%Effects of frozen thawed embryos versus frozen thawed blastocysts on transfer period and delivery outcomes

    Institute of Scientific and Technical Information of China (English)

    牟联俊; 李楠; 韦继红; 唐永梅

    2014-01-01

    BACKGROUND:Since the first frozen embryo transplantation succeeded in 1983, embryo cryotechnique has been an important component in human assisted reproductive techniques. It is controversial which embryos and blastocysts after cryopreservation is selected. OBJECTIVE:To compare the birth outcomes and neonatal status of frozen thawed embryos and blastocysts. METHODS:In frozen thawed embryo group (n=1 273) and frozen thawed blastocyst group (n=471), we compared pregnancy rate, abortion rate, ectopic pregnancy rate, premature delivery rate, average premature gestational weeks, term yield, average ful-term gestational age, newborn sex, birth weight, birth defects and so on. RESULTS AND CONCLUSION:There were frozen thawed blastocyst thaw cycles in 478 cases, 471 cases of transplantation period (including 7 cases without blastocyst transfer were canceled), 236 cases of pregnancy, 201 cases of delivery. Delivery number was 251, including 140 boys and 111 girls. The third day embryo thawing cycle of freezing and thawing appeared in 1 280 cases, 1 273 cases of transplantation period (including 7 cases of no embryo transplantation were canceled), 415 cases of pregnancy, 343 cases of delivery. The delivery number was 431, including 225 boys and 206 girls. Rate of pregnancy was significantly higher in frozen thawed blastocysts compared with frozen thawed embryos. No significant differences were detected in rate of miscarriage, ectopic pregnancy rate, premature delivery rate, average premature gestational weeks, term yield, average ful-termgestational age, newborn sex, and birth weight between frozen thawed embryos and blastocysts. There was no increase in birth defects between frozen thawed embryos and blastocysts. Results suggested that no significant difference was detected in birth outcomes and neonatal status of frozen thawed embryos and blastocysts. Nevertheless, pregnancy outcome is better in frozen thawed blastocysts than frozen thawed embryos.%背景:自从1983

  15. 不同脱颗粒细胞时间对体外受精胚胎移植的影响%Effects of differentdegranulation cells time on in vitro fertilization and embryo transfer

    Institute of Scientific and Technical Information of China (English)

    刘英; 秦文松; 杨铭; 林慧; 段金良

    2014-01-01

    目的:探讨短时受精后不同脱颗粒时间对体外受精胚胎移植周期的影响。方法:将行常规 IVF治疗患者的同周期卵子分为两组(除去完全受精失败以及受精率低下的治疗周期),即短时受精即刻脱颗粒细胞组(A 组:精卵孵育4h 后即刻脱颗粒细胞)和短时受精并次日晨脱颗粒细胞组(B 组:精卵孵育4h 并受精后17~18h 脱颗粒细胞),比较两组的受精率(2 PN 率)、多精受精率(多 PN 率)、卵裂率、优胚率、胚胎利用率、临床妊娠率。结果:两组的受精率(2 PN 率)、多精受精率(多 PN 率)、卵裂率、优胚率、胚胎利用率、临床妊娠率之间差异无统计学意义。结论:精卵孵育4h 后去除颗粒细胞对受精、胚胎发育没有显著影响。%Objectives:To explore the effects of different degranulation cells time on in vitro fertilization and embryo transfer cycles after short -term fertilization.Methods:The oocytes in the same IVF -ET cycle were divided into two groups (cycles with fertilization failure or low fertilization rate were not included).Group A (short-term fertilization and immediate degranulation,gamete co -incubating for 4 hours,then removing all degranula-tion cells),group B (short -term fertilization and degranulation in the morning on the next day,gamete co -incu-bating for 4 hours,then removing all degranulation cells 17 -18 hours after insemination).The fertilization rate (two pronucleus rate),the polyspermy rate (multiple pronucleus rate),the cleavage rate,the good quality embryo rate,the embryo utilization rate and the pregnancy rate of the two groups were compared.Results:There was no statistically difference in the fertilization rate (two pronucleus rate),the polyspermy rate (multiple pronucleus rate),the cleavage rate,the good quality embryo rate,the embryo utilization rate and the pregnancy rate among the two groups.Conclusion:Degranulation cells removal after

  16. Preimplantation embryo programming: transcription, epigenetics, and culture environment.

    Science.gov (United States)

    Duranthon, Veronique; Watson, Andrew J; Lonergan, Patrick

    2008-02-01

    Preimplantation development directs the formation of an implantation- or attachment-competent embryo so that metabolic interactions with the uterus can occur, pregnancy can be initiated, and fetal development can be sustained. The preimplantation embryo exhibits a form of autonomous development fueled by products provided by the oocyte and also from activation of the embryo's genome. Despite this autonomy, the preimplantation embryo is highly influenced by factors in the external environment and in extreme situations, such as those presented by embryo culture or nuclear transfer, the ability of the embryo to adapt to the changing environmental conditions or chromatin to become reprogrammed can exceed its own adaptive capacity, resulting in aberrant embryonic development. Nuclear transfer or embryo culture-induced influences not only affect implantation and establishment of pregnancy but also can extend to fetal and postnatal development and affect susceptibility to disease in later life. It is therefore critical to define the basic program controlling preimplantation development, and also to utilize nuclear transfer and embryo culture models so that we may design healthier environments for preimplantation embryos to thrive in and also minimize the potential for negative consequences during pregnancy and post-gestational life. In addition, it is necessary to couple gene expression analysis with the investigation of gene function so that effects on gene expression can be fully understood. The purpose of this short review is to highlight our knowledge of the mechanisms controlling preimplantation development and report how those mechanisms may be influenced by nuclear transfer and embryo culture.

  17. DNA methylation at a bovine alpha satellite I repeat CpG site during development following fertilization and somatic cell nuclear transfer.

    Directory of Open Access Journals (Sweden)

    Christine Couldrey

    Full Text Available Incomplete epigenetic reprogramming is postulated to contribute to the low developmental success following somatic cell nuclear transfer (SCNT. Here, we describe the epigenetic reprogramming of DNA methylation at an alpha satellite I CpG site (αsatI-5 during development of cattle generated either by artificial insemination (AI or in vitro fertilization (IVF and SCNT. Quantitative methylation analysis identified that SCNT donor cells were highly methylated at αsatI-5 and resulting SCNT blastocysts showed significantly more methylation than IVF blastocysts. At implantation, no difference in methylation was observed between SCNT and AI in trophoblast tissue at αsatI-5, however, SCNT embryos were significantly hyper-methylated compared to AI controls at this time point. Following implantation, DNA methylation at αsatI-5 decreased in AI but not SCNT placental tissues. In contrast to placenta, the proportion of methylation at αsatI-5 remained high in adrenal, kidney and muscle tissues during development. Differences in the average proportion of methylation were smaller in somatic tissues than placental tissues but, on average, SCNT somatic tissues were hyper-methylated at αsatI-5. Although sperm from all bulls was less methylated than somatic tissues at αsatI-5, on average this site remained hyper-methylated in sperm from cloned bulls compared with control bulls. This developmental time course confirms that epigenetic reprogramming does occur, at least to some extent, following SCNT. However, the elevated methylation levels observed in SCNT blastocysts and cellular derivatives implies that there is either insufficient time or abundance of appropriate reprogramming factors in oocytes to ensure complete reprogramming. Incomplete reprogramming at this CpG site may be a contributing factor to low SCNT success rates, but more likely represents the tip of the iceberg in terms of incompletely reprogramming. Until protocols ensure the epigenetic

  18. Effect of arachidonic acid supplementation and cyclooxygenase/lipoxygenase inhibition on the development of early bovine embryos Influência do ácido araquidónico e da inibição da ciclo-oxigenase ou lipo-oxigenase no desenvolvimento inicial de embriões bovinos

    Directory of Open Access Journals (Sweden)

    Rosa Maria Pereira

    2006-04-01

    Full Text Available The effect of arachidonic acid (AA cascade on bovine embryo development in a granulosa cell co-culture system was studied. Arachidonic acid (100 µM was supplemented from 1-cell to 8-16 cell block stage (first three days of co-culture and from 1-cell to hatching. Specific cyclooxygenase (indomethacin, 28 µM and lipoxygenase (nordihydroguaiaretic acid - NDGA, 28 µM inhibitors were used from 1-cell to 8-16 cell block stage with AA. Embryo development was assessed by cleavage, day 7-day 8 and hatched embryo rates and by measuring growth rates through development stages found in days 7-10 of culture (day 0 = insemination day. Embryo quality was scored at day 8. A 6.5-10.4% increase on cleavage rate after AA supplementation was found. This AA supplementation from 1-cell to hatching delayed embryo growth rate beyond day 7 and a reduction on hatching rate was detected. When AA supplementation was restricted to the first three days of co-culture those negative effects were overcome. Also, indomethacin and NDGA prevented the positive effect of AA and induced a significant reduction on cleavage, respectively. NDGA further decreased day 7 embryo rate and quality. Results suggest that AA has a two-phase action on bovine embryos, promoting early development and impairing embryo growth from day 7 onwards and hatching rates. Both cyclooxygenase and lipoxygenase were found to be important pathways to promote cleavage.Estudou-se a influência da cascata do ácido araquidónico (AA no desenvolvimento de embriões bovinos produzidos in vitro em co-cultura com células da granulosa. Os embriões foram suplementados com AA (100 µM desde o estádio de 1 célula até 8-16 células (primeiros três dias de co-cultura ou até a eclosão. Introduziram-se inibidores específicos da ciclo-oxigenase (indometacina, 28 µM e da lipo-oxigenase (ácido nordihidroguaiarético - NDGA, 28 µM, juntamente com o ácido araquidónico, desde o estádio de 1 célula até 8-16 c

  19. Accuracy of a combined score of zygote and embryo morphology for selecting the best embryos for IVF

    Institute of Scientific and Technical Information of China (English)

    Yu-li QIAN; Ying-hui YE; Chen-ming XU; Fan JIN; He-feng HUANG

    2008-01-01

    Objective:To evaluate the accuracy of a scoring system combining zygote and embryo morphology in predicting the outcome of in vitro fertilization(IVF)treatment.Methods:In a study group,117 consecutive IVF or intracytoplasmic sperm injection(ICSI) cycles with embryo transfer were carried out and 312 embryos were scored Using a combmed scoring system(CSS)of zygote and embryo morphology before transplantation.In a control group,a total of 420 IVF or ICSI cycles were carried out and 1176 embryos were scored using a cumulative embryo score(CES).The effects of the combined scoring system on the embryo implantation rate and pregnancy rate per cycle were analyzed.Results:Using the combined scoring system,the embryo implantation rate(27.6%)and the clinical pregnancy rate(48.7%)were significantly higher than those in the control group(20.8%and 38.6%,respectively).Also,the implantation rate of embryos scoring≥70 (38.5%:82 sacs/213 embryos)was significantly higher (P<0.001)than that of embryos scoring<70(4%:4 sacs/99 embryos).The pregnancy rate of patients with embryos scoring≥70 using the combined scoring system(66.7%)Was significantly higher(P<0.001)than that of patients with embryos scoring≥20 using the cumulative embryo score(59.0%).Conclusion:The results suggest that selecting embryos with a high Score(≥70)using the combined scoring system could inerease the implantation rate and pregnancy rate,and that using a scoring system combining assessments of human zygotes and pre-implantation embryos might predict IVF outcomes more accurately than using a cumulafive embryo score.

  20. Global transcriptome profiles of Italian Mediterranean buffalo embryos with normal and retarded growth.

    Directory of Open Access Journals (Sweden)

    Maria Strazzullo

    Full Text Available The transcriptome profiles were compared for buffalo embryos with normal growth and embryos with retarded growth on Day 25 after mating. Embryos with retarded growth on Day 25 after mating have a reduced likelihood of undergoing attachment to the uterine endometrium and establishing a pregnancy. Italian Mediterranean buffaloes were mated by AI and on Day 25 underwent trans-rectal ultrasonography to ascertain embryo development. Embryos with an embryonic width (EW>2.7 mm were classed as normal embryos and embryos with an EW<2.7 mm were classed as retarded embryos. Three buffaloes with embryos of the largest EW (3.7, 3.7 and 3.9 mm and three buffaloes with embryos of the smallest EW (1.5, 1.6 and 1.9 mm were slaughtered on Day 27 to recover embryos for transcriptome analysis using a bovine custom designed oligo array. A total of 1,047 transcripts were differentially expressed between embryos with normal growth and embryos with retarded growth. Retarded embryos showed 773/1,047 (74% transcripts that were down-regulated and 274/1,047 (26% transcripts that were up-regulated relative to normal embryos; in silico analyses focused on 680/1,047 (65% of the differentially expressed transcripts. The most altered transcripts observed in retarded embryos were associated with membrane structure and function and with metabolic and homeostasis maintenance functions. Other notable functions altered in retarded embryos were developmental processes and in particular nervous system differentiation and function. Specific biochemical pathways such as the complement cascade and coagulation were also altered in retarded embryos. It was concluded from the findings that buffalo embryos with retarded growth on Day 25 after mating show altered gene expression compared with normal embryos, and some de-regulated functions are associated with attachment to the uterine endometrium.

  1. Sourcing human embryos for embryonic stem cell lines: Problems & perspectives

    Directory of Open Access Journals (Sweden)

    Rajvi H Mehta

    2014-01-01

    Full Text Available The ability to successfully derive human embryonic stem cells (hESC lines from human embryos following in vitro fertilization (IVF opened up a plethora of potential applications of this technique. These cell lines could have been successfully used to increase our understanding of human developmental biology, transplantation medicine and the emerging science of regenerative medicine. The main source for human embryos has been ′discarded′ or ′spare′ fresh or frozen human embryos following IVF. It is a common practice to stimulate the ovaries of women undergoing any of the assisted reproductive technologies (ART and retrieve multiple oocytes which subsequently lead to multiple embryos. Of these, only two or maximum of three embryos are transferred while the rest are cryopreserved as per the decision of the couple. In case a couple does not desire to ′cryopreserve′ their embryos then all the embryos remaining following embryo transfer can be considered ′spare′ or if a couple is no longer in need of the ′cryopreserved′ embryos then these also can be considered as ′spare′. But, the question raised by the ethicists is, "what about ′slightly′ over-stimulating a woman to get a few extra eggs and embryos? The decision becomes more difficult when it comes to ′discarded′ embryos. As of today, the quality of the embryos is primarily assessed based on morphology and the rate of development mainly judged by single point assessment. Despite many criteria described in the literature, the quality assessment is purely subjective. The question that arises is on the decision of ′discarding′ embryos. What would be the criteria for discarding embryos and the potential ′use′ of ESC derived from the ′abnormal appearing′ embryos? This paper discusses some of the newer methods to procure embryos for the derivation of embryonic stem cell lines which will respect the ethical concerns but still provide the source material.

  2. Clonal propagation of primate offspring by embryo splitting.

    Science.gov (United States)

    Chan, A W; Dominko, T; Luetjens, C M; Neuber, E; Martinovich, C; Hewitson, L; Simerly, C R; Schatten, G P

    2000-01-14

    Primates that are identical in both nuclear and cytoplasmic components have not been produced by current cloning strategies, yet such identicals represent the ideal model for investigations of human diseases. Here, genetically identical nonhuman embryos were produced as twin and larger sets by separation and reaggregation of blastomeres of cleavage-stage embryos. A total of 368 multiples were created by the splitting of 107 rhesus embryos with four pregnancies established after 13 embryo transfers (31% versus 53% in vitro fertilization controls). The birth of Tetra, a healthy female cloned from a quarter of an embryo, proves that this approach can result in live offspring.

  3. Evaluation of Sheep Embryo Quality by Morphologic Methods – Advantages and Disadvantages

    Directory of Open Access Journals (Sweden)

    S. Angela

    2006-01-01

    Full Text Available The present study is focused on one of the most important steps in embryo technology evaluating embryo quality. During three years, we have conducted embryo transfer experiments on sheep. The evaluation of embryo quality was made using morphologic methods. We have evaluated embryos immediately after recovery (surgical methods recovery or after throwing of frozen embryo under a optical microscope were used for evaluation. International Embryo Transfer Society standards for embryo quality classification. There are four grades of quality: excellent, good, medium and poor. The morphological method’s advantages are: it is very fast, not so expensive and does not require excessive embryo manipulation. Disadvantages of this method are: grading is subjective, depending on the experience of the evaluator, metabolic, genetic or epigenetic disorders of embryos are not detectable.

  4. In vitro manipulation techniques of porcine embryos

    DEFF Research Database (Denmark)

    Liu, Ying; Li, Juan; Løvendahl, Peter;

    2015-01-01

    insemination. Therefore, a review of the overall efficiency for the developmental competence of embryos produced by these in vitro methods would be useful in order to obtain a more thorough overview of this growing area with respect to its development and present status. In this review a meta-analysis was used...... to analyse data collected from all published articles with a focus on zygotes and embryos for transfer, pregnancy, full-term development and piglets born. It was generally concluded that an increasing level of in vitro manipulation of porcine embryos decreased the overall efficiency for production of piglets...

  5. 不同黄体大小对供体超排和受体移植成功率的影响%Effect of Different Size of Corpora Iutea on the success ratio of Superorulation and Embryo Transfer

    Institute of Scientific and Technical Information of China (English)

    李强; 肖锐; 牛志宏; 张建军; 刘水涛; 张光伟; 张健

    2011-01-01

    本试验分析了291头14~15月龄荷斯坦青年母牛不同黄体大小对供体超排的效果.比较了924头受体母牛不同黄体大小对胚胎移植后妊娠率的影响.结果表明,埋栓前,黄体1.2cm组超排效果和有效胚胎数较高.黄体较小的组超排效果稍差一些.1.5cm以上黄体组的结果居中.受体母牛在移植胚胎时,0.8cm以上黄体的受体均获得了较高的妊娠率.但1.5cm以上黄体的受体妊娠率更高一些.由此结论提出:在超排前,黄体的大小与母牛体内孕酮浓度的高低和超排效果有关.受体在移植胚胎前,只要黄体的大小能保证最低妊娠所需的孕酮浓度,均能保证较好的妊娠率.黄体较大时,妊娠率有增加的趋势.%Two hundred and ninety-one Holstein heifers with 14-15 months age were superovulated and 924 received frozen embryos, in which heifers of donors and recipients had the different size of corpora lutea.The results showed that before CIDR inserion the heifers with 1.2 com of corpora lutea had a better superovulation response and higher transferable embryo yield.But the group below 1.0cm of corpora lutea had a poor superovulation response.And meanwhile an acceptable pregnancy rate of the recipients was obtained in heifers/cows with corpora lutea over 0.8cm.The recipients had a higher pregnancy rate with over 1.5 cm of corpora lutea.It can be conduded that the size of corpora lutea of donors was associated with superovulation response, progesterone concentrations and high embryo yield.Before the recipients receive embryos, an acceptable pregnancy rate would be obtained if the corpora lutea were over 0.8cm with a good progesterone concentration for maintaining pregnancy.Furthermore, the higher pregnancy rate had an increasing trend when the corpora lutea size was increased.

  6. 多囊卵巢综合征行体外受精-胚胎移植助孕结局分析%The Outcome Analysis of Polycystic Ovary syndrome in Vitro Fertilization and Embryo Transfer Pregnancy

    Institute of Scientific and Technical Information of China (English)

    吴成平; 王芳

    2015-01-01

    目的:探讨多囊卵巢综合征(PCOS)伴不孕患者采用体外受精-胚胎移植(IVF-ET)助孕治疗的疗效及特点。方法:回顾分析2013年52周期PCOS不孕因素IVF-ET病历资料作为A组,同期102周期输卵管因素IVF-ET病例作对照作为B组。结果:PCOS组与输卵管因素组比较分析:患者年龄、不孕年限、BMI、Gn天数、获卵数、可移植胚胎数、OHSS风险取消移植率比较,差异有统计学意义(P0.05)。结论:PCOS伴不孕患者采用IVF-ET助孕可获得与输卵管因素IVF-ET助孕相似的疗效,但要注意OHSS的风险预防。%Objective:To evaluate the polycystic ovary syndrome (PCOS) with infertility were treated by in vitro fertilization-embryo transfer (IVF-ET) curative effect and characteristics of assisted reproduction treatment. Method: 52 cycles PCOS infertility factor in IVF-ET medical records in 2013 year were analysed retrospectively as group A, and 102 cycles tubal factor IVF-ET cases in the same period as controls of group B.Result:Compared PCOS group and tubal factor group analysis: age, infertility time, BMI, the number of eggs, portable designed the Gn days, embryo number and risk of OHSS cancel graft rate comparison, the difference was statistically significant(P<0.05). PCOS patients compared with tubal factor group more young, shorter infertility time, higher body mass index (BMI),longer Gn time,more egg number, more corresponding portable embryos and OHSS risk increased significantly. But two groups of designed the Gn dosage, endometrium thickness, 2 PN fertilization rate, cleavage rate, good embryo rate and the clinical pregnancy rate were no statistically significant difference.Conclusion:PCOS patients of infertility with the IVF-ET assist reproduction can be obtained IVF-ET assist similar curative effect with the tubal factor, but should pay attention to prevent the risk of OHSS.

  7. First successful pregnancies following embryo selection using Time-lapse technology in Iran: Case report

    OpenAIRE

    Azita Faramarzi; Mohammad Ali Khalili; Mehrdad Soleimani

    2015-01-01

    Background: Embryo selection is a vital part of in vitro fertilization (IVF) programs, with morphology-based grading systems having been widely used for decades. Time-lapse imaging combined with embryo morph kinetics may proffer a non-invasive means for improving embryo selection. We report the first ongoing and chemical pregnancies using Time-lapse embryo scope to select best embryos for transfer in Iran. Cases: A case with tubal factor infertility was admitted to IVF program with normozo...

  8. Transfer

    DEFF Research Database (Denmark)

    Wahlgren, Bjarne; Aarkrog, Vibe

    Bogen er den første samlede indføring i transfer på dansk. Transfer kan anvendes som praksis-filosofikum. Den giver en systematisk indsigt til den studerende, der spørger: Hvordan kan teoretisk viden bruges til at reflektere over handlinger i situationer, der passer til min fremtidige arbejdsplads?...

  9. Comparison of the major malformation rate of children conceived from cryopreserved embryos and fresh embryos

    Institute of Scientific and Technical Information of China (English)

    LI Hong-zhen; QIAO Jie; CHI Hong-bin; CHEN Xin-na; LIU Ping; MA Cai-hong

    2010-01-01

    Background Cryopreserved embryo transfer has become indispensable in reproductive technology. More and more children are conceived from frozen-thawed embryo transfer (FET). The risk of birth defects associated with frozen-thawed embryo transfer has been evaluated and conflict results are obtained. The aim of this study was to compare the rate of major malformations in children conceived from cryopreserved embryos with that of children from fresh embryos. Methods A retrospective analysis was performed on children conceived from frozen-thawed embryos and fresh embryos between January 2005 and December 2008 at the Reproduction Center of the Third Hospital, Peking University.The major malformation rates were compared between two groups for all children, as well as singletons or twins,separately. The frequencies of different subtypes of malformations classified according to different organ system were also compared.Results Thirty-four of 3125 children from cryopreserved embryos had a major malformation. The malformation rate was 1.09%, which was comparable to that for children after fresh embryos transfer (1.53%(55/3604), OR:0.71, 95% CI; 0.46-1.09). The malformation rate was also similar when the analysis was limited to children from cryopreserved embryos resulted from in vitro fertilization (IVF)(1.39%)and fresh IVF(1.3%). However, children from cryopreserved embryos resulted from intracytoplasmic sperm injections (ICSI) had much lower malformation rate than from fresh ICSI(0.63% vs.1.83%, OR: 0.34, 95% CI: 0.16-0.75). No difference was found in the incidence of major malformations in singletons from cryo ICSI (0.73%) and fresh ICS1(1.9%), or from cryo IVF(1.49%) and fresh IVF(1.67%). Similar malformation rate was found in multiples from cryo ICSI(0.52%) and fresh ICSI(1.76%), or cryo IVF(1.30%) and fresh IVF(0.90%). The distribution and risk of the subtype of malformations, such as cardiovascular, gastrointestinal, neural tube, urogenital, rnusculoskeletal and facial

  10. Relationship between Pronuclear Scoring and Embryo Quality and Implantation Potential in IVF-ET

    Institute of Scientific and Technical Information of China (English)

    Qun LIU; Guijin ZHU; Juan HU; Yulan WEI; Xinling REN; Hanwang ZHANG; Yufeng LI; Lei JIN; Jing YUE

    2008-01-01

    To assess the relationship between pronuclear scoring and day-3 embryo quality and pregnancy outcome and to determine the clinical value of pronuclear stage scoring system in human in vitro fertilization-embryo transfer (WF-ET) program, a pronuclear scoring system was used to score zygotes 16-20 h after insemination during conventional WF or intracytoplasmic sperm injec- tion (ICS1). The embryos were classified into groups Z1, Z2, Z3 and Z4. Comparisons were made of the rates of arrested embryos and excellent embryos on day 3. Comparisons of pregnancy outcome were made only in those patients in whom cohorts of similarly Z-scored embryos were transferred. The results showed that there were less arrested embryos and more excellent embryos on day 3 in groups Z1 and Z2 than those in group Z3 and Z4. More embryos arrested and less excellent embryos developed in group Z4 than group Z3. The clinical pregnancy rates resulting from the transfer of single pronuclear score homologous embryo types were similar among groups Z1, Z2 and Z3. Implanta- tion rates of group Z1 were higher (P<0.05) than that of group Z3. These findings suggests that pro- nuclear scoring can predict developmental ability on day 3 and implantation potential. A evaluation that combines the Z-score and day 3 embryo morphology is useful in the determination of the most viable embryos and the number of embryos for transfer.

  11. In vivo versus in vitro produced bovine ova: similarities and differences relevant for practical application

    DEFF Research Database (Denmark)

    Holm, Peter; Callesen, Henrik

    1998-01-01

    - Abstract This present review describes some differences and similarities between bovine embryos produced in vivo and in vitro. The first part outlines the respective environments during maturation, fertilisation and early embryonic development of the two types of embryos and compares their mor-...

  12. Embryo disposition and the new death scene

    Directory of Open Access Journals (Sweden)

    Ellison, David

    2011-01-01

    Full Text Available In the IVF clinic - a place designed principally for the production and implantation of embryos - scientists and IVF recipients are faced with decisions regarding the disposition of frozen embryos. At this time there are hundred of thousands of cryopreserved embryos awaiting such determinations. They may be thawed for transfer to the woman herself, they may be donated for research or for use by other infertile couples, they may remain in frozen storage, or they may variously be discarded by being allowed to 'succumb', or 'perish'. Where the choice is discard, some IVF clients have chosen to formalise the process through ceremony. A new language is emerging in response to the desires of the would-be-parents who might wish to characterise the discard experience as a ‘good death’. This article examines the procedure known as ‘compassionate transfer’ where the embryo to be discarded is placed in the woman’s vagina where it is clear that it will not develop further. An alternate method has the embryo transferred in the usual manner but without the benefit of fertility-enhancing hormones at a point in the cycle unreceptive to implantation. The embryo destined for disposal is thus removed from the realm of technological possibility and ‘returned’ to the female body for a homely death. While debates continue about whether or not embryos constitute life, new practices are developing in response to the emotional experience of embryo discard. We argue that compassionate transfer is a death scene taking shape. In this article, we take the measure of this new death scene’s fabrication, and consider the form, significance, and legal complexity of its ceremonies.

  13. Embryo quality and impact of specific embryo characteristics on ongoing implantation in unselected embryos derived from modified natural cycle in vitro fertilization

    NARCIS (Netherlands)

    Pelinck, Marie-Jose; Hoek, Annemieke; Simons, Arnold H. M.; Heineman, Maas Jan; van Echten-Arends, Janny; Arts, Eus G. J. M.

    2010-01-01

    Objective: To study the implantation potential of unselected embryos derived from modified natural cycle IVF according to their morphological characteristics. Design: Cohort study. Setting: Academic department of reproductive medicine. Patient(S): A series of 449 single embryo transfers derived from

  14. Outcome analysis on the two embryo transfer policies in high-risk OHSS patients%OHSS高危患者两种移植方案的临床结局分析

    Institute of Scientific and Technical Information of China (English)

    谢鑫; 曹云霞; 周平; 吴欢; 张媛媛; 刘迎春

    2013-01-01

    目的 比较研究控制性超促排卵(COH)周期患者移植日前有卵巢过度刺激综合征(OHSS)高危因素时行全胚胎冷冻以及行新鲜周期移植的两种方案的临床结局.方法 回顾性分析接受COH治疗且取卵前出现OHSS高危因素的患者267例,随机分为A、B两组.A组:全胚胎冷冻并行首次解冻胚胎移植的122例;B组:行新鲜周期胚胎移植的145例.对两组患者的年龄、不孕年限、不孕因素、身体质量指数(BMI)、COH情况[促性腺激素(Gn)启动剂量、超排天数、Gn总量、注射人绒毛膜促性腺激素(HCG)日内膜、HCG日雌二醇(E2)水平、HCG日卵巢大小]、获卵数、优质胚胎数、重度OHSS发生率、新鲜周期及复苏周期的生化妊娠率、临床妊娠率进行比较分析.结果 两组患者年龄、不孕年限、不孕因素、BMI、COH情况、获卵数、优质胚胎数差异无统计学意义.A组患者重度OHSS的发生率(2.46%)显著低于B组(8.28%),差异有统计学意义(P<0.05).两组的生化妊娠率(50.0% vs 47.6%)、临床妊娠率(36.9% vs 33.8%),差异无统计学意义.结论 与新鲜周期移植比较,OHSS高危患者行全胚胎冷冻后复苏周期移植不影响患者的临床结局,但其明显降低了重度OHSS的发生率,使安全性提高并降低了医疗费用.%Objective To compare the outcome of two transfer policies among high-risk ovarian hyperstimulation syndrome( OHSS ) patients with whole embryo freezing and thawing 2 ~ 3 cycles after egg retrieval and fresh embryo transfer. Methods The age, infertility years, reasons of infertility, body mass index( BMI ), situation of COH [ the levels of initial dosage of gonadotropins( Gn ), days of COH, total dosage of Gn, thickness of membrane, level of es-tradiol( E2 ), size of the ovary on day of human chofinonic gonadotropin( HCG ) ], number of retrieved oocytes, number of good embryos, incidence of significant OHSS, biochemistry and clinical pregnancy rates were compared

  15. Cryopreservation of Oocytes and Embryos in Human Assisted Reproduction

    Directory of Open Access Journals (Sweden)

    Konc J

    2005-01-01

    Full Text Available Cryopreservation has become an integral component of assisted reproductive technology. The ability to cryopreserve, thaw, and establish pregnancies with supernumerary preimplantation embryos has become an important tool in fertility treatment. Human oocyte cryopreservation has practical application in preserving fertility for individuals prior to cancer treatments. While the efficiency of oocyte and embryo freezing technology has increased over time, there is still room for improvement, since even under ideal circumstances the clinical pregnancy rate from frozen embryo transfer is approximately two-thirds of that from the fresh transfer of embryos. Thus, studies connected with cryopreservation of human oocytes and embryos are very important to the expansion of effective clinical services. This review gives a summary of the theoretical and technical aspects of oocyte and embryo cryopreservation.

  16. MODELO TEÓRICO PARA EXPLICAR LA ACUMULACIÓN DE GOTAS LIPÍDICAS EN EMBRIONES BOVINOS MACHOS O HEMBRAS PRODUCIDOS in vitro Theoretical Model For Explaining Accumulation Of Fat Drops In In Vitro Produced Bovine Embryos

    Directory of Open Access Journals (Sweden)

    OMAR CAMARGO

    Full Text Available La glucosa 6-fosfato deshidrogenasa (G6PD, codificada por un gen ubicado en el cromosoma X, es la enzima limitante de vía de las pentosas fosfato (PF. La entrada de la glucosa así como su flujo y el rendimiento metabólico de esta vía están determinados tanto por los mismos niveles glucosa así como por la actividad de la G6PD. Por esta vía, la glucosa regula la trascripción de varios genes lipogénicos. En algunos embriones hembra producidos in vitro, se registra un retardo en la normal inactivación de uno de sus cromosomas X, lo cual se traduce en una doble actividad de los genes allí ubicados, si se compara con los embriones macho producidos in vitro. Se postula entonces que, la sobre-regulación de la vía PF a consecuencia de la doble dosis de su enzima limitante (G6PD y en presencia de elevados niveles de glucosa (mayores a 2,5 mM en el medio de cultivo, conllevaría a un dimorfismo sexual en relación con la transcripción de los genes Acetil CoA Carboxilasa Alfa (en adelante ACACA, símbolo oficial de la acetyl-Coenzyme A carboxylase alpha, y la Sintetasa de Ácidos Grasos (en edelante FASN, símbolo oficial de la fatty acid synthase que corriente abajo codifican para las enzimas limitantes en la síntesis de lípidos. Este dimorfismo sexual para el fenotipo metabolismo de lípidos, derivaría en una mayor acumulación citoplasmática de gotas lipídicas en los embriones hembra en comparación con los embriones machos que, de ser así, tendría efectos expansivos sobre el metabolismo general, la actividad transcripcional de otros genes y sobre la resistencia a la criopreservación.The encoding gene for glucose 6-phosphate dehydrogenase (G6PD is located on chromosome X. This enzyme regulates the entrance of glucose into the pentose phosphate pathway (PPP. Besides, throughout this route, glucose regulates the transcription of some lipogenic genes. Compared with in vitro produced male embryos, and due to a delaying in X

  17. Blastocyst rate of in vitro embryo production in sheep is affected by season.

    Science.gov (United States)

    Mara, L; Sanna, D; Casu, S; Dattena, M; Muñoz, I M Mayorga

    2014-08-01

    It has been reported that the number and quality of in vitro produced embryos is season related. This study was conducted to assess the effect of season on cleavage, blastocyst and lambing rates of in vitro produced ovine embryos during 3 years of collection data. Ovaries of Sarda sheep were collected from a slaughterhouse. In total, 5035 oocytes were recovered and matured in TCM-199 with 4 mg/ml bovine serum albumin (BSA), 100 μM cysteamine, 0.3 mM Na pyruvate, 0.1 UI/ml recombinant follicle-stimulating hormone (r-FSH), 0.1 UI/ml recombinant luteinising hormone (r-LH), and 1 μg/ml estradiol-17β. Matured oocytes were fertilized with fresh semen in synthetic oviductal fluid (SOF) with 20% heat inactivated estrous sheep serum. The presumptive zygotes were cultured for 6-7 days (blastocyst stage) in SOF medium supplemented with 1% Basel Medium Eagle (BME), 1% Minimum Essential Medium, 1 mM glutamine and 8 mg/ml fatty acid-free BSA. The embryos produced were vitrified and a total of 165 blastocysts (80 from the breeding season and 85 from the anoestrous season) were transferred in pairs into recipient ewes during the reproductive period. There were no significant differences in cleavage rates between seasons in any of the 3 years examined (84% versus 83%, 81% versus 80% and 80% versus 79%, respectively). The blastocyst rate varied significantly between seasons in 2005 and 2007 (P < 0.05), and in 2006 (P < 0.001). There were no differences in pregnancy and lambing rates between embryos during anoestrous versus during the breeding season. In conclusion, only the blastocyst rate appeared to have been affected by season, possibly due to variation in the number of developmentally competent oocytes. PMID:23458093

  18. Oocyte activation and preimplantation development of bovine embryos obtained by specific inhibition of cyclin-dependent kinases Ativação oocitária e desenvolvimento pré-implantação de embriões bovinos obtidos com o uso de inibidores específicos das quinases dependentes de ciclina

    Directory of Open Access Journals (Sweden)

    F. Perecin

    2007-04-01

    Full Text Available The efficiency of bohemine and roscovitine in combination with ionomycin on parthenogenetic activation and initial embryonic development of bovine oocytes was studied. Two experiments were performed: in the first, different concentrations (0, 50, 75 or 100µM and different exposure periods (2, 4 or 6 hours to bohemine or roscovitine were tested for activation rates of in vitro matured (IVM bovine oocytes, which were pre-exposed to ionomycin. The best treatments, 75µM bohemine and 50µM roscovitine, both for 6h, were used in the second experiment, in which IVM bovine oocytes were exposed to ionomycin, followed or not by bohemine or roscovitine treatment, and evaluated for nuclear status, activation rate and blastocyst development were assessed. The combined treatments (ionomycin + cyclin-dependent kinases inhibitors - CDKIs showed better results for activation rates (77.3% and initial embryonic development (35.2% than the single ionomycin treatment (69.4% for activation and 21.9% for development; and also lead to a more uniform activation (nearly 90% single pronucleus development. The results showed that CDKIs improve the effects of ionomycin on parthenogenetic activation and blastocyst development in bovine oocytes and could help to achieve more efficient activation protocols, increasing the developmental competence of embryos obtained by reproductive biotechniques.Realizaram-se dois experimentos para avaliar a eficiência da bohemina e roscovitina associadas à ionomicina para ativação partenogenética e desenvolvimento embrionário inicial de bovinos. No primeiro, foram testadas diferentes concentrações (0, 50, 75 ou 100µM e diferentes tempos de exposição (2, 4 ou 6 horas à bohemina ou à roscovitina na ativação de oócitos bovinos maturados in vitro (MIV pré-expostos à ionomicina. Os melhores tratamentos, bohemina 75µM e roscovitina 50µM, ambos por seis horas, foram utilizados no segundo experimento, no qual oócitos bovinos

  19. Cryopreservation of human embryos and its contribution to in vitro fertilization success rates.

    Science.gov (United States)

    Wong, Kai Mee; Mastenbroek, Sebastiaan; Repping, Sjoerd

    2014-07-01

    Cryopreservation of human embryos is now a routine procedure in assisted reproductive technologies laboratories. There is no consensus on the superiority of any protocol, and substantial differences exist among centers in day of embryo cryopreservation, freezing method, selection criteria for which embryos to freeze, method of embryo thawing, and endometrial preparation for transfer of frozen-thawed embryos. In the past decade, the number of frozen-thawed embryo transfer cycles per started in vitro fertilization (IVF) cycle increased steadily, and at the same time the percentage of frozen-thawed embryo transfers that resulted in live births increased. Currently, cryopreservation of human embryos is more important than ever for the cumulative pregnancy rate after IVF. Interestingly, success rates after frozen-thawed embryo transfer are now nearing the success rates of fresh embryo transfer. This supports the hypothesis of so called freeze-all strategies in IVF, in which all embryos are frozen and no fresh transfer is conducted, to optimize success rates. High-quality randomized controlled trials should be pursued to find out which cryopreservation protocol is best and whether the time has come to completely abandon fresh transfers.

  20. Extended Culture of Early Stage Embryos in Frozen-thawed Cycles

    Institute of Scientific and Technical Information of China (English)

    Hong-bo WANG; Yan-hui LI

    2009-01-01

    Objective To investigate the impact of extended culture of early stage embryos on pregnancy outcome of frozen embryo transfer (FET).Methods The survival rates of embryos after thawing and pregnancy outcome following FET were compared retrospectively between zygote and cleavage embryos which cultured to cleavage stage or extended cultured to blastocysts. Results A total of 425 zygote embryos in 67 cycles were thawed. After thawing, the survival rate was 94.4% and with an average transfer of 2.8 embryos, the clinical pregnancy rate was 55.2% (37/67). In 222 FET cycles, totally 1 270 cleavage stage embryos were thawed and the overall survival rates were 80.3%. With an average transfer of 2.7 embryos, the clinical pregnancy rate was 55.4% (123/222). A significantly lower percentage of degenerated embryos were found for zygotes (5.6%) than that for cleavage stage embryos (19.7%) (P0.05). Conclusion Although the clinical pregnancy rate was not different between patients with freeze-thaw zygote and cleavage stage embryo transfer, higher survival rate for zygote was shown compared with that for cleavage stage embryo. However, the present studies did not demonstrate that extended culture thawing embryos to blastocyst could achieve favor clinical outcome.

  1. High in vitro development after somatic cell nuclear transfer and trichostatin A treatment of reconstructed porcine embryos

    DEFF Research Database (Denmark)

    Li, J.; Østrup, Olga; Villemoes, Klaus;

    2008-01-01

    Abnormal epigenetic modification is supposed to be one of factors accounting for inefficient reprogramming of the donor cell nuclei in ooplasm after somatic cell nuclear transfer (SCNT). Trichostatin A (TSA) is an inhibitor of histone deacetylase, potentially enhancing cloning efficiency. The aim...

  2. DNA methylation in porcine preimplantation embryos developed in vivo and produced by in vitro fertilization, parthenogenetic activation and somatic cell nuclear transfer

    DEFF Research Database (Denmark)

    Deshmukh, Rahul Shahaji; Østrup, Olga; Østrup, Esben;

    2011-01-01

    DNA demethylation and remethylation are crucial for reprogramming of the differentiated parental/somatic genome in the recipient ooplasm upon somatic cell nuclear transfer. Here, we analyzed the DNA methylation dynamics during porcine preimplantation development. Porcine in vivo developed (IV), i...

  3. Antiviral effects of bovine interferons on bovine respiratory tract viruses.

    OpenAIRE

    Fulton, R W; Downing, M M; Cummins, J M

    1984-01-01

    The antiviral effects of bovine interferons on the replication of bovine respiratory tract viruses were studied. Bovine turbinate monolayer cultures were treated with bovine interferons and challenged with several bovine herpesvirus 1 strains, bovine viral diarrhea virus, parainfluenza type 3 virus, goat respiratory syncytial virus, bovine respiratory syncytial virus, bovine adenovirus type 7, or vesicular stomatitis virus. Treatment with bovine interferons reduced viral yield for each of the...

  4. Targeting exogenous GDNF gene to the bovine somatic cell beta-casein locus for the production of transgenic bovine animals.

    Science.gov (United States)

    Zhang, X M; Luo, F H; Ding, H M; Li, B; Zhang, J J; Wu, Y J

    2015-01-01

    Considerable attention is currently being directed toward methods for producing recombinant human proteins in the mammary glands of genetically modified transgenic livestock. However, the expression of inserted genes in transgenic animals is variable and often very low because of the randomness of the site of transgene integration. One possible strategy to avoid the expression problem associated with random integration is to use site-specific integration by targeting integration to a high expression locus and, thereby, to improve expression of the transferred gene. In the present study, we focused on glial cell line-derived neurotrophic factor (GDNF), a novel type of neurotrophic factor first cloned in 1993. Research has shown that GDNF may have potential applications in the treatment of Parkinson's disease and other diseases of the central nervous system since it acts as a protective factor for central dopaminergic neurons. Here, we constructed a gene targeting vector to knock-in the human GDNF gene at the bovine beta-casein gene locus as a first step to producing transgenic animals with a high level of expression of human GDNF protein in their mammary glands. Bovine fetal fibroblast cells were transfected with linearized pNRTCNbG by electroporation. Three cell clones were identified with successful targeting to the beta-casein locus; and were confirmed using both polymerase chain reaction analysis and sequencing. Gene-targeted cells were used as nuclear donors; a total of 161 embryos were reconstructed, 23 of which developed to the blastocyst stage. These blastocysts were transferred to 8 recipient cows, but no offspring were obtained. PMID:26634460

  5. Expression of antiapoptosis gene survivin in luteinized ovarian granulosa cells of women undergoing IVF or ICSI and embryo transfer: clinical correlations

    Directory of Open Access Journals (Sweden)

    Varras Michail

    2012-09-01

    Full Text Available Abstract Background The purpose of the study was to determine the incidence of survivin gene expression in human granulosa cells during ovarian stimulation in Greek women with normal FSH levels, undergoing IVF or ICSI and to discover any correlation between levels of gene expression and clinical parameters, efficacy of ovulation or outcomes of assisted reproduction. Methods Twenty nine women underwent ovulation induction for IVF or ICSI and ET with standard GnRH analogue-recombinant FSH protocol. Infertility causes were male and tubal factor. Cumulus–mature oocyte complexes were denuded and the granulosa cells were analyzed for each patient separately using quantitative reverse transcription polymerase chain reaction analysis for survivin gene expression with internal standard the ABL gene. Results The ABL and survivin mRNA were detected in granulosa cells in 93.1%. The expression levels of survivin were significantly lower in normal women (male infertility factor compared to women with tubal infertility factor (p = 0.007. There was no additional statistically significant correlation between levels of survivin expression and estradiol levels or dosage of FSH for ovulation induction or number of dominant follicles aspirated or number of retrieved oocytes or embryo grade or clinical pregnancy rates respectively. Conclusions High levels of survivin mRNA expression in luteinized granulosa cells in cases with tubal infertility seem to protect ovaries from follicular apoptosis. A subpopulation of patients with low levels of survivin mRNA in granulosa cells might benefit with ICSI treatment to bypass possible natural barriers of sperm-oocyte interactions.

  6. Study on hormone replacement therapy for patients with poor endometrial growth to freezing-thawing embryo transfer%激素替代方案应用于子宫内膜生长不良患者冻胚移植内膜准备

    Institute of Scientific and Technical Information of China (English)

    朱鹏云; 林妍; 刘芸

    2012-01-01

    Objective To investigate the effects of hormone replacement therapy on endometrial thickness of different dosages and different duration of the patients with poor endometrial growth receiving oral estrogen in frozen-thawed embryo transfer cycles. Methods Retrospective analysis was performed on 97 frozen-thawed embryo transfer cycles of patients with poor endometrial growth treated with estrogen. Patients were divided into implantation group and non-implantation group based on clinical outcome to analyze the relation between embryo implantation outcome and endometrial thickness. Furthermore, the duration and dosage of estrogen were analyzed to detect the effects of different durations and dosages of estrogen on endometrial thickness. Results ( 1 )lt was observed that there was no statistical difference of the endometrial thickness in the estrogen application time and the day of deciding embryo transfer between the embryo implantation group and non-implantation group. However, the endometrial thickening value( thickness value of the day of deciding embryo transfer minus the value of the 12th day of estrogen treatment )of implantation group was significantly higher than that of non-implantation group( P 0. 05 ). ( 3 )There was no significant different on the endometrial thickness and endometrial thickening value between the groups of taking 6 mg estrogen and 9 mg estrogen on the day of deciding embryo transfer( P > 0. 05 ). Conclusions For patients with poor endometrial growth treated with estrogen for preparing endometria in freezing-thawing embryo transfer, the endometrial thickening value was one of the important factors for outcome of frozen-thawed embryo transfer. Taking the estrogen within 19-21 days was favorable for endometrial growth and embryo implantation, but increasing the dosage had no effect on endometrial growth.%目的 探讨子宫内膜生长不良患者行激素替代方案冻融胚胎移植周期中口服雌激素的不同时间及剂量

  7. Gonadotropin stimulation regimens for follicular aspiration and in vitro embryo production from calf oocytes.

    Science.gov (United States)

    Armstrong, D T; Irvine, B J; Earl, C R; McLean, D; Seamark, R F

    1994-01-01

    Crossbred beef x dairy calves were randomly allocated at 3 wk of age to different gonadotropin treatment regimens for stimulation of follicle development and induction of oocyte maturation in vivo. Follicular responses were assessed laparoscopically, and oocytes were aspirated for assessment of maturational state or for in vitro fertilization (IVF) and culture to determine developmental capacity. Follicle-stimulating Hormone (FSH), administered in a single subcutaneous injection together with a low dosage of PMSG, was as effective as the same total dosage of FSH administered in 6 injections over a 3-d period. Without accompanying PMSG, this dose of FSH was ineffective in stimulating follicle development. The mean number of preovulatory follicles (> 5mm, with hyperemic appearance) doubled with each successive stimulation at 3-wk intervals, reaching 35 follicles per calf at 9 wk of age. Oocyte yields ranged from 55 to 81% of follicles aspirated, and did not differ significantly among age, FSH regimen and oocyte maturation stimulus. A combination of LH + FSH was more effective in stimulating cumulus cell expansion than LH by itself (73 vs 22% of recovered oocyte-cumulus cell complex (OCC) respectively; Poocytes (cumulus unexpanded) subjected to in vitro maturation (IVM) and IVF, 30% developed to blastocysts during co-culture with bovine oviduct epithelial cells, which was not significantly different from 25% of 36 oocytes from adult ovaries which reached the blastocyst stage under similar conditions. The results indicate that follicle responses of calf ovaries to FSH stimulation increase progressively from 3 to 9 wk of age, and that oocytes recovered laparoscopically from these follicles produce blastocysts in culture at rates similar to oocytes from adult cattle ovaries collected at slaughter. The approach offers promise for embryo production from donor calves of superior genetic merit for embryo transfer, thereby enhancing the rate of genetic gain above that

  8. Analysis of 86 ectopic pregnancy cases after in vitro fertilization and embryo transfer%体外受精-胚胎移植后异位妊娠86例临床分析

    Institute of Scientific and Technical Information of China (English)

    田婷; 韩丹; 刘书华; 周平; 曹云霞

    2015-01-01

    Objective To investigate the incidence, affecting factors, early diagnosis, prevention and treatment methods of ectopic pregnancy( EP) in assisted reproductive technology. Methods A retrospective review was done on the clinical data of 86 patients with EP after in vitro fertilization and embryo transfer( IVF-ET) , intracytoplasmic sperm injection( ICSI) , preimplantation genetic diagnosis( PGD) and frozen-thawing embryo transfer( F-ET) . Re-sults A total of 2 903 clinical pregnancies were obtained, and 86 ectopic pregnancies, the rate of EP was 2. 96%, of which 4 cases were heterotopic pregnancy(4. 65%) . It appeared to be no relationship in these aspects, that was, age, duration of infertility, body mass index(BMI), ovulation dysfunction, infertility caused by male, chromosome abnormality, history of childbirth, the average follicle numbers, the average graftage embryonic num-ber and the depth of transplant tube into the uterine cavity. However, tubal and pelvic diseases, endometriosis or endometriosis postoperatively was significantly associated with the occurrence of EP( OR=11. 457, P=0. 000;OR=3. 479, P=0. 002). When endometrial thickness was more than or equal to 7. 5 mm, the risk of tubal pregnancy decreased obviously(OR=10. 463, P=0. 002). Conclusion The main correlative factors with EP are tutal path-ological and pelvis changes, endometriosis or endometriosis postoperatively and the thickness of endometrium in the day of embryo transfer. In addition, sera hCG level tests and transvaginal ultrasonography are recommended for ear-ly diagnosis.%目的:探讨辅助生殖技术中异位妊娠( EP )的发生率、影响因素和早期诊疗、防治的方法。方法回顾性分析体外受精-胚胎移植( IVF-ET )、卵胞浆内单精子显微注射( ICSI)、胚胎植入前遗传学诊断( PGD)和冻融胚胎移植( F-ET)治疗后86例EP患者的临床资料。结果共有2903个周期获临床妊娠,86例EP,发生率2.96%,其中

  9. Contributions of the maternal uterine environment and piglet genotype on weaning survivability potential: II. Piglet growth, lactation performance, milk composition, and piglet blood profiles during lactation following reciprocal embryo transfers between Meishan and White crossbred gilts.

    Science.gov (United States)

    Miles, J R; Vallet, J L; Ford, J J; Freking, B A; Oliver, W T; Rempel, L A

    2015-04-01

    Crossbreeding studies between Meishan (MS) and Large White (LW) pigs have illustrated that increased piglet growth before weaning is attributed to the maternal genotype of LW dams. The objective of this study was to determine the contributions of the maternal uterine environment (MUE), piglet genotype (PigG), piglet age (PA), and their interactions on piglet growth, lactation performance, milk composition, and piglet blood profiles during lactation following reciprocal embryo transfers between MS and White crossbred (WC) gilts. Twenty-five successful pregnancies were generated by embryo transfer in 2 farrowing years representing all MUE × PigG combinations: MS × MS (n = 4 litters), MS × WC (n = 7 litters), WC × MS (n = 7 litters), and WC × WC (n = 7 litters). At d 1 and 10 and at weaning, piglets (n = 147, n = 96, and n = 94, respectively) were weighed and blood samples were collected and measured for hematocrit, hemoglobin, glucose, nitrogen, NEFA, albumin, lactate, and cortisol. In addition, sows were manually milked from a medial mammary gland to determine milk composition. All data were analyzed by ANOVA using MIXED model procedures with the fixed effects of MUE, PigG, PA, and their interactions. Piglet weight was greater (P piglets from WC dams compared to MS dams at d 10 and weaning but not at d 1. In addition, ADG were greater (P piglets from WC dams compared to MS dams throughout lactation. However, milk composition was greater (P piglets, irrespective of MUE, at d 1 of lactation and in MS piglets from MS dams at d 10 of lactation. Blood glucose was greater (P = 0.01) at d 1 in piglets from WC dams regardless of PigG but, at weaning, glucose was greater (P = 0.01) in WC piglets regardless of MUE. Serum NEFA levels were greater (P = 0.02) in piglets from MS dams throughout the lactation period. This study demonstrated that WC dams were superior to MS dams for piglet growth during lactation, in agreement with previous crossbreeding studies. However

  10. 子宫内膜轻创术对体外受精-胚胎移植治疗结局的影响%The effect of endometrial local injury on the outcome of in vitro fertilization-embryo transfer

    Institute of Scientific and Technical Information of China (English)

    刘杰; 郑洁; 雷亚兰; 孙虹; 李薇; 夏敏; 章汉旺

    2012-01-01

    Objective To analyze the effect of endometrial local injury on the incidence of successful pregnancy in patients undergoing in vitro fertilization-embryo transfer (IVF-ET). Methods Two hundred and thirty-seven patients were studied who were with endometrial disorder or thickness diagnosed by ultrasound on the day of gonadotropin was used. Two hundred and twenty patients (treatment group) were operated local injury to regain normal endometrium or remove thickened endometrium at the same day of gonadotropin was used. The remaining 17 patients were not treated and used as control. The outcome of IVF- ET in the two groups was evaluated by the rates of clinical pregnancy, embryo implantation and live births. Results There were no significant differences between the two groups in the age of the patients, duration for infertility, response to hormone stimulation, the numbers of oocytes and embryo, and the rate of fertilization. However, the rates of clinical pregnancy, embryo implantation and live births in the treatment group were 52.27%, 36.56% and 45.00%, respectively, which were significantly higher than those in the control group (23.53%, 13.51% and 23.53%). Conclusion Local injury to the endometrium significantly improves the successful pregnancy in patients undergoing in vitro fertilization.%目的:探讨子宫内膜轻创术对体外受精-胚胎移植(IVF-ET)临床妊娠率、胚胎种植率和活产率的影响.方法:回顾性分析2007 年1 月至2009 年6 月在我院生殖中心行IVF-ET 治疗的不孕症患者,促性腺激素(Gn)启动日B 超显示子宫内膜回声紊乱或内膜厚度> 5 mm 的患者共237 例,其中220 例行子宫内膜轻创术者为研究组,17 例未行轻创术者为对照组,比较两组患者临床妊娠率、胚胎种植率和活产率的差异.结果:研究组和对照组患者平均年龄、不育年限、Gn 使用天数、Gn 总量、HCG 日E2水平、获卵数、受精数、可移植胚胎数和移植胚

  11. A clinical analysis of 19 cases of ectopic pregnancy after in vitro fertilization and embryo transfer%体外受精-胚胎移植后异位妊娠19例临床分析

    Institute of Scientific and Technical Information of China (English)

    李银玲; 王兴玲; 孙丽君; 王雪梅; 张彩霞; 郝大勇

    2011-01-01

    目的 探讨体外受精-胚胎移植后异位妊娠的发病率、致病因素、临床表现、早期诊断及治疗.方法 回顾性分析体外受精-胚胎移植后19例异位妊娠的临床资料.结果 同期新鲜周期和冻胚复苏移植周期共1 398个,临床妊娠498例,临床妊娠率为35.62%,其中异位妊娠19例,异位妊娠发生率为1.26%,包括宫内外同时妊娠3 例,单侧输卵管双妊娠1例.主要临床表现阴道不规则出血和下腹疼痛.致病因素主要是输卵管病变(特别是一侧梗阻伴另一侧炎症或积水),其次为排卵障碍和慢性盆腔炎.结论 体外受精-胚胎移植后异位妊娠的发病因素主要有输卵管病变、异位妊娠病史、排卵障碍和慢性盆腔炎等;阴道超声检查结合血HCG值测定是其最有效的诊断措施;腹腔镜下输卵管切除术是治疗未破裂单纯异位妊娠的较好方法.%Objective To investigate incidence rate, etiological factors, clinical manifestation , early diagnosis and treatment of ectopic pregnancy(EP) after in vitro fertilization and embryo transfer (IVF-ET). Methods The clinical data of 19 patients with EP after IVF-ET were retrospectively analyzed. Results Totally, 1398 embryos ( 944 fresh embryoes and 454 frozen embryos were transplanted in a IVF-ET cycle. Eventually, 498 women conceived after IVF-ET and clinical pregnancy rate was 35.62%. Of which, 19 women had EP, and incidence rate of EP was 1.26% , including 3 cases of heterotopic pregnancy(HP) ,1 case of double pregnancy in single oviduct. The main clinical manifestations of women with EP were vaginal abnormal bleeding and lower abdomen distending pain. Tubal diseases (especially obstruction of one side Follopian tube associated with inflammation or hydrosalpinx in the other side of tube), ovulation failure and chronic pelvic inflammation were risk factors of EP. Conclusion Tubal diseases, history of EP, ovulation failure and chronic pelvic inflammation are main risk

  12. Study on the Adaptability of the Transferred Montbeliard Embryos Bulls in Northern Region of China%胚胎移植蒙贝利亚种公牛在北方地区适应性研究

    Institute of Scientific and Technical Information of China (English)

    钟代彬; 张晓霞; 乔绿; 刘玉; 张海涛; 薛建华; 张胜利

    2012-01-01

    本研究于2008年7月-2011年3月,对进口胚胎移植出生并经选育的9头蒙贝利亚种公牛的环境适应性、生长发育、抗病能力、繁殖性能等进行了观察、测定和分析研究。结果显示,30月龄时平均体高151.89cm,睾丸周径42.11cm,体重922.67kg,平均日增重0.959kg。10月龄开始初次采精,24月龄正式采精;平均每次射精量6.20raL,鲜精活力0.732,精子密度13.083qL个/mL,平均利用率82.8%,均产冻精40606剂;冻精活力≥0.4,精子畸形率〈15%。精液质量达到牛冷冻精液国家标准(GB4143-2008)。适应性研究表明,胚胎移植蒙贝利亚种公牛对北方地区的环境条件较为适应,生长发育状况良好,抗病力强,具有良好的繁殖生产性能。%From July 2008 to March 2011, the environmental adaptability, growth, disease resistance, reproductive performance of nine Montbeliard breeding bulls derived from imported embryo transplant were measured and analyzed. Results showed that the average height and average testicular circumference in the age of 30 months were 151.89cm and 42.11 cm, respectively. Average weight and average daily gain were 922.67kg and 0.959kg, respectively. Initial semen collection was in 10 months old and formal semen collection in 24 months old. The average volume per ejaculate was 6.20mL, fresh sperm viability of 73.2%, density of 13.08~108/mL, average utilization rate of 82.8%, average frozen semen of 40606. Semen energy exceeded 0.4, rate of abnormal sperm was less than 15%. The quality of bovine frozen semen could meet the national standard GB4143-2008. Results showed that Montbeliard breeding bulls from imported embryo transplant had a good adaptability, strong disease resistance and breedin~ performance in northern region

  13. 未成熟卵体外成熟技术在卵巢高反应患者IVF-ET中的应用%In vitro maturation of human immature oocyte in in vitro fertilization and embryo transfer

    Institute of Scientific and Technical Information of China (English)

    黄绘; 洪焱; 冯玉蓉; 刘敏利; 胡皓睿; 潘世春

    2012-01-01

    目的:探讨未成熟卵体外成熟(IVM)技术在卵巢高反应患者体外受精-胚胎移植(IVF - ET)中的应用价值.方法:在IVF - ET促排治疗中,对双卵巢卵泡数过多,有可能发生卵巢过度刺激综合征(OHSS)或继续治疗可能发生重度OHSS的患者,根据其意愿即刻停药,全部取卵改行IVM治疗12个周期(A组)或取部分小卵泡改行IVM治疗,同时保留部分卵泡继续行IVF - ET常规治疗18个周期(B组).小卵泡体外培养成熟后,通过卵胞浆内单精子注射(ICSI)获得受精卵并行胚胎移植或冷冻.统计分析未成熟卵的成熟率、卵母细胞的受精率、胚胎的发育情况及临床结局.结果:两组30个取卵周期,共获未成熟卵240个,经IVM、ICSI和体外培养后,成熟率、受精率、正常卵裂率及优质胚胎率分别为61.25% (147/240),77.55% (114/147),92.98% (106/114)和29.25%(31/106).A组8例行IVM新鲜胚胎移植(8周期)4例临床妊娠,A、B两组有8例行IVM解冻胚胎移植(9周期)3例临床妊娠,已有3例分娩.A组12例无OHSS发生,促性腺激素用量少于B组,B组18例中3例有OHSS风险而取消胚胎移植.结论:对常规IVF促排周期中卵巢高反应患者及时改行IVM,可以避免周期取消及OHSS的发生,减少促排卵药物的使用量,同时获得较好的临床妊娠率.%Objective ; To explore the clinical application of in vitro oocyte maturation (IVM) technique in in vitro fertilization - embryo transfer (IVF - ET). Methods; In the treatment of IVF - ET, women who canceled the treatment cycle due to high risk of developing ovarian hyperstimulation syndrome ( OHSS) were divided into two groups according to their choices. Women in group A (n = 12) underwent oocyte retrieval guided by transvaginal ultrasound for all small follicles and treated with IVM technique for 12 cycles. Women in group B (n = 18) were performed oocyte retrieval for some small follicles treated with IVM technique and kept on IVF - ET with some

  14. 慢速冷冻胚胎TET周期当天与解冻后培养1d移植结果分析%Outcome of Frozen-thawed Embryo Transfer between Different Culture Time In Vitro in Natural Cycles and Artificial Cycles

    Institute of Scientific and Technical Information of China (English)

    李瑞岐; 王文军; 麦美琪; 欧阳能勇; 马芸; 陈静华; 张宁锋

    2011-01-01

    [Objective] To assess the outcome of slow-frozen/thawed embryo transfer (TET) between different culture time of the embryos frozen at day 3(D3) in vitro in natural and artificial cycles. [Methods] A database was analyzed retrospectively concerning TET cycles covered 552 cycles, in which the number of thawed embryos ^1 and ^3 from February 2009 to December 2010. According to the different culture time after embryo thawing, they were divided into group A(ET at the thawing day) and group B (ET at the next day ofter embryo thawed). And these two groups were further divided into nature cycle and artificial cycle. [Results] Although there were not significantly different between the couples age, the number of embryos transferred and the embryos scale (P > 0.05).The implantation rates of D3 frozen embryos in group B were significantly higher than those in group A in both of the nature cycles and the artificial cycles (17.9 vs 12.3,P< 0.05; 17.3% vs 8.7%,P< 0.05), respectively.[Conclusion] The effect of extending incubation time of thawed embryos does not significantly effect the outcome of D2 freezing embryos. But in the TET cycles of D3 frozen embryos, ET at the next day of embryo thawed will be better for the embryo implantation.%[目的]探讨第3天(D3)慢速冷冻胚胎解冻后当天行胚胎移植(ET)和体外培养1d后ET在自然周期和人工周期中对临床结局的影响.[方法]回顾性分析本中心2009年1月至2010年12月期间的解冻胚胎数≥1,且≤3的552个卵裂期冷冻胚胎TET周期的临床结果,根据解冻后体外培养时间分为A组(当天ET)和B组(培养1d后ET),这两个组再分为自然周期组和人工周期组.[结果]D3冷冻胚胎在自然周期和人工周期,A、B两组女方年龄、平均移植胚胎数、胚胎评分以及妊娠率均没有统计学意义(P>0.05),但是,在自然周期和人工周期中B组种植率较A组均有统计学意义(17.9 vs 12.3,P< 0.05;17.3%vs 8.7%,P< 0.05).[结

  15. Genetic and physical mapping of the bovine X chromosome.

    Science.gov (United States)

    Yeh, C C; Taylor, J F; Gallagher, D S; Sanders, J O; Turner, J W; Davis, S K

    1996-03-01

    Three hundred eighty reciprocal backcross and F(2) full sib progeny from 33 families produced by embryo transfer from 77 Angus (Bos taurus), Brahman (Bos indicus), and F1 parents and grandparents were used to construct genetic maps of the bovine X and Y chromosomes. Ml individuals were scored for 15 microsatellite loci, with an average of 608 informative meioses per locus. The length of the bovine X chromosome genetic map was 118.7 cM (female only) and of the pseudoautosomal region was 13.0 cM (male only). The 15-marker framework map in Kosambi centimorgans is [BM6017-6.1 -TGLA89-35.8-TEXAN13-3.4-TGLA128-1.3 -BM2713 -21.1 -BM4604-2.4-BR215 - 12.9-TGLA68-10.0-BM4321 - 1.0-HEL14-4.9-TGLA15-2.3-INRA12O- 12.5-TGLA325- 1.6-MAF45-3.2-INRA3O], with an average interval of 7.91 cM. Clones containing pseudoautosomal or sex-linked microsatellites were isolated from a bovine bacterial artificial chromosome library and were physically mapped to bovine metaphase chromosomes by fluorescence in situ hybridization to orient the X and Y chromosome maps. BAC57, containing the pseudoautosomal microsatellite INRA3O, mapped to the distal end of the long arm of the X chromosome at q42-ter and to the short arm of the Y chromosome at p13-ter. This confirms the published assignment of this region to Ypl2-ter, but challenges the published assignment of Xpl4-ter and thus reorients the X chromosome physical map. BAC2O4, containing the X-linked microsatellite BM4604, mapped to the middle of the long arm of the X chromosome at q26-q31. The position of the physically mapped markers indicates either a lack of microsatellite markers for a large (30 to 50 cM) region of the short arm of the X chromosome or heterogeneity of recombination along the X chromosome. PMID:8833151

  16. Genetic and physical mapping of the bovine X chromosome

    Energy Technology Data Exchange (ETDEWEB)

    Yeh, Chen Chen; Taylor, J.F.; Sanders, J. O. [Texas A& M Univ., College Station, TX (United States)] [and others

    1996-03-01

    Three hundred eighty reciprocal backcross and F{sub 2} full sib progeny from 33 families produced by embryo transfer from 77 Angus (Bos taurus), Brahman (Bos indicus), and F{sub 1} parents and grandparents were used to construct genetic maps of the bovine X and Y chromosomes. All individuals were scored for 15 microsatellite loci, with an average of 608 informative meioses per locus. The length of the bovine X chromosome genetic map was 118.7 cM (female only) and of the pseudoautosomal region was 13.0 cM (male only). The 15-marker framework map in Kosambi centimorgans is (BM6017-6.1-TGLA89-35.8-TEXAN13-3.4-TGLA128-1.3-BM2713-21.1-BM4604-2.4-BR215-12.9-TGLA68-10.0-BM4321-1.0-HEL14-4.9-TGLA15-2.3-INRA120-12.5-TGLA325-1.6-MAF45-3.2-INRA30), with an average interval of 7.91 cM. Clones containing pseudoautosomal or sex-linked microsatellites were isolated from a bovine bacterial artificial chromosome library and were physically mapped to bovine metaphase chromosomes by fluorescence in situ hybridization to orient the X and Y chromosome maps. BAC57, containing the pseudoautosomal microsatellite INRA30, mapped to the distal end of the long arm of the X chromosome at q42-ter and to the short arm of the Y chromosome at p13-ter. This confirms the published assignment of this region to Yp12-ter, but challenges the published assignment of Xp14-ter and thus reorients the X chromosome physical map. BAC204, containing the X-linked microsatellite BM4604, mapped to the middle of the long arm of the X chromosome at q26-q31. The position of the physically mapped to the middle of the long arm of the X chromosome at q26-q31. The position of the physically mapped markers indicates either a lack of microsatellite markers for a large (30 to 50 cM) region of the short arm of the X chromosome or heterogeneity of recombination along the X chromosome. 46 refs., 2 figs., 3 tabs.

  17. Effect of insulin-like growth factor-1 during in vitro oocyte maturation and in vitro culture of bovine embryos Efeito do fator de crescimento semelhante à insulina-1 durante a maturação in vitro dos oócitos e cultivo in vitro de embriões bovinos

    Directory of Open Access Journals (Sweden)

    M.D. Quetglas

    2001-04-01

    Full Text Available The effects of insulin-like growth factor-I (IGF-I on in vitro maturation (IVM (experiment I and on in vitro embryo development (experiment II of bovine oocytes fertilized in vitro, were evaluated in terms of cleavage (CR, blastocyst (BR and hatching (HR rates. For IVM, immature cumulus-oocyte complexes were cultured in TCM-199 medium supplemented with Hepes, sodium bicarbonate, sodium pyruvate, additives, fetal calf serum (B-199 medium and gonadotropins (14 U/ml PMSG and 7 U/ml hCG. For embryo development, the oocytes/zygotes were cultured in B-199 medium with bovine oviduct epithelial cells in suspension under silicon oil. Treatments for in vitro culture conditions for both experiments were: 1- B-199 + 200 ng/ml IGF-I; 2- B-199 + 100 ng/ml IGF-I; 3- B-199 + 50 ng/ml IGF-I; 4- B-199 + 10 ng/ml IGF-I; 5- B-199 + 0 ng/ml IGF-I. All cultures were performed at 38.5ºC in 5% CO2 in air and the data were analyzed by chi-square test. In experiment I, there were no differences (P>0.05 among treatments for CR, BR or HR. In experiment II, the addition of IGF-I to the embryo culture medium (ECM resulted in a significant increase in CR while for BR and HR this effect was not observed. The addition of 200 ng/ml IGF-I to ECM increased CR (71.1% when compared to 100 ng/ml IGF-I (57.6% or control (56.7% groups, however, there were no differences when compared to 50 (69.4% or 10 ng/ml (73.1% groups. There was no beneficial effect of the addition of IGF-I in the IVM or ECM media on the in vitro development of embryos produced from oocytes matured and fertilized in vitro.Avaliaram-se o efeito do IGF-I na maturação in vitro (MIV (experimento I e no desenvolvimento embrionário (DE (experimento II de oócitos bovinos fecundados in vitro, quanto às taxas de clivagem (TC, de blastocistos (TB e de eclosão (TE. Para MIV, complexos cumulus-oócitos imaturos foram cultivados em meio TCM-199 suplementado com HEPES, bicarbonato e piruvato de sódio, aditivos, soro

  18. A microfluidic system supports single mouse embryo culture leading to full-term development

    NARCIS (Netherlands)

    Esteves, Telma Cristina; Rossem, van Fleur; Nordhoff, Verena; Schlatt, Stefan; Boiani, Michele; Le Gac, Séverine

    2013-01-01

    The present study demonstrates the feasibility of application of a microfluidic system for in vitro culture of pre-implantation mouse embryos, with subsequent development to full-term upon embryo transfer. Specifically, embryos cultured in groups in nL volume chambers achieve pre-implantation develo

  19. Progestin implants can rescue demi-embryo pregnancies in goats: a case study.

    Science.gov (United States)

    Beckett, D M; Oppenheim, S M; Moyer, A L; BonDurant, R H; Rowe, J D; Anderson, G B

    1999-06-01

    Survival after transfer of demi-embryos (i.e., half-embryos produced by embryo splitting) to recipients usually is lower than survival after transfer of intact embryos. Reduced survival after demi-embryo transfer could be due to loss of viability after splitting, failure of a viable demi-embryo to prevent corpus luteum (CL) regression in the recipient female, or a combination of factors. From a retrospective analysis of pregnancy and embryo survival rates after demi-embryo transfer in sheep and goats, we report the rescue of caprine demi-embryo pregnancies in which CL regression occurred at the end of diestrus despite the presence of a viable conceptus in the uterus with progestin implants. Day 5 or 6 morulae and blastocysts were flushed from superovulated ewes and does and split into demi-embryos of approximately equal halves. Demi-embryos were either transferred fresh to synchronized recipients of the homologous species or frozen in liquid nitrogen. Approximately half of the recipient does and ewes were treated with norgestomet implants on Day 10 of the embryo transfer cycle and again 2 wk later. Serum collected on Day 25 from recipients with implants was assayed for progesterone to determine if a CL of pregnancy had been maintained. Pregnancy was diagnosed by ultrasonography on Day 35 of gestation. Corpus luteum regression occurred despite the presence of a viable conceptus in the uterus in 6 of 55 progestin-treated caprine demi-embryo recipients and in 0 of 66 ovine demi-embryo recipients. Five of the caprine pregnancies were maintained to term with norgestomet implants and produced 5 live kids. The sixth fetus, which was carried by a progestin implant-treated 8-mo-old doeling, died at approximately 50 d of gestation. These results suggest that, at least in goats, some demi-embryos may provide inadequate signaling for maternal recognition of pregnancy, and such pregnancies can be rescued with progestin treatment to the doe.

  20. Aberrant DNA methylation in 5'regions of DNA methyltransferase genes in aborted bovine clones

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    High rate of abortion and developmental abnormalities is thought to be closely associated with inefficient epigenetic reprogramming of the transplanted nuclei during bovine cloning.It is known that one of the important mechanisms for epigenetic reprogramming is DNA methylation.DNA methylation is established and maintained by DNA methyltransferases(DNMTs),therefore,it is postulated that the inefficient epigenetic reprogramming of transplanted nuclei may be due to abnormal expression of DNMTs.Since DNA methylation can strongly inhibit gene expression,aberrant DNA methylation of DNMT genes may disturb gene expression.But presently,it is not clear whether the methylation abnormality of DNMT genes is related to developmental failure of somatic cell nuclear transfer embryos.In our study,we analyzed methylation patterns of the 5' regions of four DNMT genes including Dnmt3a,Dnmt3b,Dnmtl and Dnmt2 in four aborted bovine clones.Using bisulfite sequencing method,we found that 3 out of 4 aborted bovine clones(AF1,AF2 and AF3)showed either hypermethylation or hypomethylation in the 5' regions of Dnmt3a and Dnmt3b.indicating that Dnmt3a and Dnmt3b genes are not properly reprogrammed.However,the individual AF4 exhibited similar methylation level and pattern to age-matched in vitro fertilized (IVF)fetuses.Besides,we found that tle 5'regions of Dnmtl and Dnmt2 were nearly completely unmethylated in all normal adults.IVF fetuses,sperm and aborted clones.Together,our results suggest that the aberrant methylation of Dnmt3a and Dnmt3b 5' regions is probably associated with the high abortion of bovine clones.

  1. Cryopreservation of Arachis pintoi (leguminosae) somatic embryos.

    Science.gov (United States)

    Rey, H Y; Faloci, M; Medina, R; Dolce, N; Engelmann, F; Mroginski, L

    2013-01-01

    In this study, we successfully cryopreserved cotyledonary somatic embryos of diploid and triploid Arachis pintoi cytotypes using the encapsulation-dehydration technique. The highest survival rates were obtained when somatic embryos were encapsulated in calcium alginate beads and precultured in agitated (80 rpm) liquid establishment medium (EM) with daily increasing sucrose concentration (0.50, 0.75, and 1.0 M). The encapsulated somatic embryos were then dehydrated with silica gel for 5 h to 20% moisture content (fresh weight basis) and cooled either rapidly (direct immersion in liquid nitrogen, LN) or slowly (1 degree C per min from 25 degree C to -30 degree C followed by immersion in LN). Beads were kept in LN for a minimum of 1 h and then were rapidly rewarmed in a 30 degree C water-bath for 2 min. Finally, encapsulated somatic embryos were post-cultured in agitated (80 rpm) liquid EM with daily decreasing sucrose concentration (0.75 and 0.5 M) and transferred to solidified EM. Using this protocol, we obtained 26% and 30% plant regeneration from cryopreserved somatic embryos of diploid and triploid cytotypes. No morphological abnormalities were observed in any of the plants regenerated from cryopreserved embryos and their genetic stability was confirmed with 10 isozyme systems and nine RAPD profiles. PMID:24441368

  2. Blastocyst development potential of D3 low quality embryos

    Institute of Scientific and Technical Information of China (English)

    Zhen Jing-ran; Sun Zhen-yi; Yu Qi; Deng Cheng-yan; Zhou Yuan-zheng; He Fang-fang

    2011-01-01

    Objective:To investigate the potential of blastocyst development of D3 low quality embryos for blastocyst culturing and freezing.Methods:Two thousand one hundred and eighty embryos of 398 IVF/ICSI patients' were observed after D3 embryo transfer.The low-quality embryos were cultured by sequential micro-drops and continuously observed blastocyst formation from D3 to D6.Results:(1) A total of 1,546 low-quality embryos were collected,and 426 blastocysts formed in DS-D6.Total blastocyst formation rate (BFR) was 27.56%,of which embryos with grade 6 Ⅲ/Ⅳ were 40.87 % (318/778),5 Ⅰ /Ⅱ 28.85% (30/104),4Ⅰ/Ⅱ 8.88% (16/180),4Ⅲ/IV 19.72% (56/284),2-3 Ⅰ/Ⅱ 3% (6/200).(2) Embryos with more blastomeres had a higher blastocyst formation rate.The lower qualities of embryos resulted in lower blastocyst formation rate.Conclusions:Low-quality embryos still have the potential to develop into blastocysts.Therefore,they should not be discarded on D3.They should be continually cultured to D6 in order to reduce embryo wastage and get good clinical results.

  3. Study on the birth situation of in vitro fertilization and embryo transfer and its derivative technique%体外受精-胚胎移植及其衍生技术出生情况的研究

    Institute of Scientific and Technical Information of China (English)

    梁莹; 亓蓉; 孟繁玉; 周莉; 付蕾

    2011-01-01

    目的:探讨体外受精-胚胎移植及其衍生技术的出生情况.方法:回顾性分析151个生产周期,对剖宫产比率、畸形比率、男女出生比率、早产率、新生儿出生体重等进行分析研究.结