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Yanina S Bogliotti, Marcela Vilarino, Pablo J Ross
Cytoplasmic microinjection into one-cell embryos is a very powerful technique. As an example, it enables the delivery of genome editing tools that can create genetic modifications that will be present in every cell of an adult organism. It can also be used to deliver siRNA, mRNAs or blocking antibodies to study gene function in preimplantation embryos. The conventional technique for microinjecting embryos used in rodents consists of a very thin micropipette that directly penetrates the plasma membrane when advanced into the embryo...
October 5, 2016: Journal of Visualized Experiments: JoVE
Ming Chen, Shun-Ping Chang, Gwo-Chin Ma, Wen-Hsian Lin, Hsin-Fu Chen, Shee-Uan Chen, Horng-Der Tsai, Feng-Po Tsai, Ming-Ching Shen
Preimplantation genetic diagnosis (PGD) is a powerful tool to tackle the transmission of monogenic inherited disorders in families carrying the diseases from generation to generation. It currently remains a challenging task, despite PGD having been developed over 25 years ago. The major difficulty is it does not have an easy and general formula for all mutations. Different gene locus needs individualized, customized design to make the diagnosis accurate enough to be applied on PGD, in which the quantity of DNA is scanty, whereas timely laboratory diagnosis is mandatory if fresh embryo transfer is desired occasionally...
2016: Thrombosis Journal
Tao Lin, Jae Eun Lee, Reza K Oqani, So Yeon Kim, Eun Seok Cho, Yong Dae Jeong, Jun Jong Baek, Dong Il Jin
The aim of this study is to investigate whether endoplasmic reticulum (ER) stress attenuation could improve porcine somatic cell nuclear transfer (SCNT) embryo developmental competence. We treated porcine SCNT embryos with TUDCA (tauroursodeoxycholic acid, an inhibitor of ER stress) and/or TM (tunicamycin, an ER stress inducer), and examined embryonic developmental potential, embryo quality, the levels of ER stress markers (XBP1 protein and mRNA) and apoptosis-related-genes (BAX and BCL2 mRNA). Immunostaining detected X-box-binding protein (XBP1), a key gene regulator during ER stress, at all stages of SCNT embryo development...
October 17, 2016: Reproductive Biology
Jae Gyu Yoo, Byeong-Woo Kim, Mi-Rung Park, Deug-Nam Kwon, Yun-Jung Choi, Teak-Soon Shin, Byung-Wook Cho, Jakyeom Seo, Jin-Hoi Kim, Seong-Keun Cho
Objective: The present study investigates pre- and post-implantation developmental competence of nuclear-transferred porcine embryos derived from male and female fetal fibroblasts. Methods: Male and female fetal fibroblasts were transferred to in vitro-matured enucleated oocytes and in vitro and in vivo developmental competence of reconstructed embryos was investigated. And, a total of 6,789 female fibroblast nuclear-transferred embryos were surgically transferred into 41 surrogate gilts and 4,746 male fibroblast nuclear-transferred embryos were surgically transferred into 25 surrogate gilts...
October 20, 2016: Asian-Australasian Journal of Animal Sciences
J Wu, A Platero Luengo, M A Gil, K Suzuki, C Cuello, M Morales Valencia, I Parrilla, C A Martinez, A Nohalez, J Roca, E A Martinez, J C Izpisua Belmonte
More than eighteen years have passed since the first derivation of human embryonic stem cells (ESCs), but their clinical use is still met with several challenges, such as ethical concerns regarding the need of human embryos, tissue rejection after transplantation and tumour formation. The generation of human induced pluripotent stem cells (iPSCs) enables the access to patient-derived pluripotent stem cells (PSCs) and opens the door for personalized medicine as tissues/organs can potentially be generated from the same genetic background as the patient recipients, thus avoiding immune rejections or complication of immunosuppression strategies...
October 2016: Reproduction in Domestic Animals, Zuchthygiene
Tong Zhang, Chunqiang Zhang, Xiaomei Fan, Ruilan Li, Jiaxin Zhang
C-type natriuretic peptide (CNP) has been considered as a physiological meiotic inhibitor that stimulates the cGMP production by cumulus cell natriuretic peptide receptor 2 (NPR2), which inhibits oocyte phosphodiesterase type 3 activity and increases cAMP. In this study, we explored the effect of CNP pretreatment on the in vitro maturation (IVM) of bovine oocytes by examining changes in cleavage rate, blastocyst formation, mitochondrial DNA (mtDNA) copy number, reactive oxygen species (ROS) level, glutathione (GSH) content, and redox state...
October 19, 2016: In Vitro Cellular & Developmental Biology. Animal
Lessly P Sepulveda-Rincon, Delphine Dube, Pierre Adenot, Ludivine Laffont, Sylvie Ruffini, Laurence Gall, Bruce K Campbell, Veronique Duranthon, Nathalie Beaujean, Walid E Maalouf
The first lineage specification during mammalian embryo development can be visually distinguished at blastocyst stage. Two cell lineages are observed on the embryonic/abembryonic axis of the blastocyst: the inner cell mass and the trophectoderm. The timing and mechanisms driving this process are still not fully understood. In mouse embryos, cells seem pre-patterned to become certain cell lineage; as the first cleavage plane has been related with further embryonic-abembryonic axis at blastocyst stage. Nevertheless, this possibility has been very debatable...
October 19, 2016: Biology of Reproduction
V Muller, M Makhmadalieva, I Kogan, I Fedorova, E Lesik, E Komarova, L Dzhemlikhanova, D Niauri, A Gzgzyan, E Ailamazyan
BACKGROUND: Ectopic pregnancy (EP) has been reported to occur in 1.4-5.4% of all clinical pregnancies resulting from in vitro fertilization (IVF) and embryo transfer (ET). Data on factors associated with abnormal embryo implantation following assisted conception are limited. MATERIALS AND METHODS: A systematic review and meta-analysis was performed to determine whether there is an association between the day (cleavage-stage, D3, versus blastocyst, D5) or the type (fresh versus frozen/thawed) of ET and EP rate...
October 2016: Gynecological Endocrinology
Denys deCatanzaro, Tyler J Pollock
Estradiol-17β (E2) plays critical roles in female maturation, sexual receptivity, ovulation, and fertility. In many mammals, contact with males can similarly affect these female parameters, while male excretions contain significant quantities of E2 We administered radiolabeled estradiol ([(3)H]E2) to male mice in doses representing a small fraction of their endogenous E2 These males were paired with sexually-receptive females, and radioactivity was traced into the females' systems. In Experiment 1, males were given [(3)H]E2 at 24 and 1 h prior to mating...
October 6, 2016: Journal of Endocrinology
Irsan E Kooi, Saskia E van Mil, David MacPherson, Berber M Mol, Annette C Moll, Hanne Meijers-Heijboer, Gertjan J L Kaspers, Jacqueline Cloos, Hein Te Riele, Josephine C Dorsman
Several murine retinoblastoma models have been generated by deleting the genes encoding for retinoblastoma susceptibility protein pRb and one of its family members p107 or p130. In Rb(-/-) p107(-/-) retinoblastomas, somatic copy number alterations (SCNAs) like Mdm2 amplification or Cdkn2a deletion targeting the p53-pathway occur, which is uncommon for human retinoblastoma. In our study, we determined SCNAs in retinoblastomas developing in Rb(-/-) p130(-/-) mice and compared this to murine Rb(-/-) p107(-/-) tumors and human tumors...
October 17, 2016: Genes, Chromosomes & Cancer
A L S Guimarães, S A Pereira, M N Diógenes, M A N Dode
The aim of this study was to evaluate the effect of adding a combination of insulin, transferrin and selenium (ITS) and l-ascorbic acid (AA) during in vitro maturation (IVM) and in vitro culture (IVC) on in vitro embryo production. To verify the effect of the supplements, cleavage and blastocyst rates, embryo size and total cell number were performed. Embryonic development data, embryo size categorization and kinetics of maturation were analyzed by chi-squared test, while the total cell number was analyzed by a Kruskal-Wallis test (P < 0...
October 17, 2016: Zygote: the Biology of Gametes and Early Embryos
Xiaofeng Zheng, Pengyi Yang, Brad Lackford, Brian D Bennett, Li Wang, Hui Li, Yu Wang, Yiliang Miao, Julie F Foley, David C Fargo, Ying Jin, Carmen J Williams, Raja Jothi, Guang Hu
Poly(A) tail length and mRNA deadenylation play important roles in gene regulation. However, how they regulate embryonic development and pluripotent cell fate is not fully understood. Here we present evidence that CNOT3-dependent mRNA deadenylation governs the pluripotent state. We show that CNOT3, a component of the Ccr4-Not deadenylase complex, is required for mouse epiblast maintenance. It is highly expressed in blastocysts and its deletion leads to peri-implantation lethality. The epiblast cells in Cnot3 deletion embryos are quickly lost during diapause and fail to outgrow in culture...
October 8, 2016: Stem Cell Reports
Enrico Papaleo, Luca Pagliardini, Valeria Stella Vanni, Diana Delprato, Patrizia Rubino, Massimo Candiani, Paola Viganò
A cost analysis covering direct healthcare costs relating to IVF freeze-all policy was conducted. Normal- and high- responder patients treated with a freeze-all policy (n = 63) compared with fresh transfer IVF (n = 189) matched by age, body mass index, duration and cause of infertility, predictive factors for IVF (number of oocytes used for fertilization) and study period, according to a 1:3 ratio were included. Total costs per patient (€6952 versus €6863) and mean costs per live birth were similar between the freeze-all strategy (€13,101, 95% CI 10,686 to 17,041) and fresh transfer IVF (€15,279, 95% CI 13,212 to 18,030)...
October 3, 2016: Reproductive Biomedicine Online
Kristin M Whitworth, Joshua A Benne, Lee D Spate, Stephanie L Murphy, Melissa S Samuel, Clifton N Murphy, Jürgen A Richt, Eric Walters, Randall S Prather, Kevin D Wells
The CRISPR/Cas9 genome editing tool has increased the efficiency of creating genetically modified pigs for use as biomedical or agricultural models. The objectives were to determine if DNA editing resulted in a delay in development to the blastocyst stage or in a skewing of the sex ratio. Six DNA templates (gBlocks) that were designed to express guide RNAs that target the transmembrane protease, serine S1, member 2 (TMPRSS2) gene were in vitro transcribed. Pairs of CRISPR guide RNAs that flanked the start codon and polyadenylated Cas9 were co-injected into the cytoplasm of zygotes and cultured in vitro to the blastocyst stage...
October 15, 2016: Transgenic Research
Amanda Souza Setti, Rita Cássia Sávio Figueira, Daniela Paes de Almeida Ferreira Braga, Matheus de Castro Azevedo, Assumpto Iaconelli, Edson Borges
OBJECTIVE: To study whether embryos derived from oocytes presenting a smooth endoplasmic reticulum cluster (SERC) are less likely to develop into blastocysts and implant. DESIGN: Transversal study. SETTING: Private university-affiliated in vitro fertilization (IVF) center. PATIENT(S): Total of 7,609 oocytes obtained from 743 intracytoplasmic sperm injection (ICSI) cycles. INTERVENTION(S): Oocytes split between the SERC-positive cycles (with at least one SERC-positive oocyte) and the SERC-negative cycles (only oocytes free of SERC)...
October 12, 2016: Fertility and Sterility
Konstantinos A Economou, Dimitra Christopikou, Erika Tsorva, Stephen Davies, Minas Mastrominas, Haris Cazlaris, Michael Koutsilieris, Panagoula Angelogianni, Dimitris Loutradis
PURPOSE: Artificial oocyte activation using calcium ionophores and enhancement of embryonic developmental potential by the granulocyte-macrophage colony-stimulating factor (GM-CSF) have already been reported. In this study, we evaluated the synergistic effect of these two methods on aged human unfertilized oocytes after intracytoplasmic sperm injection (ICSI). Then, we cultured the resulting embryos to the blastocyst stage and screened them for chromosomal abnormalities, to assess the safety of this protocol...
October 14, 2016: Journal of Assisted Reproduction and Genetics
Long Jin, Hai-Ying Zhu, Qing Guo, Xiao-Chen Li, Yu-Chen Zhang, Cheng-Du Cui, Wen-Xue Li, Zheng-Yun Cui, Xi-Jun Yin, Jin-Dan Kang
Cloning remains as an important technique to enhance the reconstitution and distribution of animal population with high-genetic merit. One of the major detrimental factors of this technique is the abnormal epigenetic modifications. MGCD0103 is known as a histone deacetylase inhibitor. In this study, we investigated the effect of MGCD0103 on the in vitro blastocyst formation rate in porcine somatic cell nuclear transferred (SCNT) embryos and expression in acetylation of the histone H3 lysine 9 and histone H4 lysine 12...
September 13, 2016: Theriogenology
Diting Qian, Zhiling Li, Yuting Zhang, Yue Huang, Que Wu, Gaizhen Ru, Man Chen, Bin Wang
Our study aimed to develop embryo models to evaluate the impact of oxidative stress on embryo development. Mouse zygotes, which stayed at G1 phase, were treated with prepared culture medium (containing 0.00, 0.01, 0.02, 0.03, 0.04, 0.05, or 0.1 mM hydrogen peroxide (H2O2)) for 30 min in experiment 1. The dose-effects of H2O2 on embryo development were investigated via comparisons of the formation rate at each stage (2- and 4-cell embryos and blastocysts). Experiment 2 was carried out to compare behaviors of embryos in a mild oxidative-stressed status (0...
2016: Oxidative Medicine and Cellular Longevity
Antonio Capalbo, Filippo Maria Ubaldi, Laura Rienzi, Richard Scott, Nathan Treff
Embryonic mosaicism, defined as the presence of karyotypically distinct cell lines within an embryo, has been frequently reported with a high incidence in preimplantation embryos derived from IVF and is thought to be one of the major biological limitations for the routine application of PGD for aneuploidies (PGD-A). The incidence of mosaicism in preimplantation embryos is in fact reported to be between 4 and 90%. However, these data are in sharp contrast with what is known from clinical pregnancies, where true foetal mosaicism is observed in less than 0...
October 13, 2016: Human Reproduction
L Antonio González-Grajales, Laura A Favetta, W Allan King, Gabriela F Mastromonaco
BACKGROUND: Successful development of iSCNT (interspecies somatic cell nuclear transfer) embryos depends on complex interactions between ooplasmic and nuclear components, which can be compromised by genetic divergence. Transfer of ooplasm matching the genetic background of the somatic cell in iSCNT embryos is a valuable tool to study the degree of incompatibilities between nuclear and ooplasmic components. This study investigated the effects of ooplasm transfer (OT) on cattle (Bos taurus) and plains bison (Bison bison bison) embryos produced by iSCNT and supplemented with or without ooplasm from cattle or plains bison oocytes...
October 13, 2016: BMC Developmental Biology
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