In vitro production of bovine embryos derived from individual donors in the Corral® dish

Background: Since the identity of the embryo is of outmost importance during commercial in vitro embryo production, bovine oocytes and embryos have to be cultured strictly per donor. Due to the rather low yield of oocytes collected after ovum pick-up (OPU) per individual cow, oocyte maturation and embryo culture take place in small groups, which is often associated with inferior embryo development. The objective of this study was to improve embryonic development in small donor groups by using the Corral (R) dish. This commercial dish is designed for human embryo production. It contains two central wells that are divided into quadrants by a semi-permeable wall. In human embryo culture, one embryo is placed per quadrant, allowing individual follow-up while embryos are exposed to a common medium. In our study, small groups of oocytes and subsequently embryos of different bovine donors were placed in the Corral (R) dish, each donor group in a separate quadrant. Results: In two experiments, the Corral (R) dish was evaluated during in vitro maturation (IVM) and/or in vitro culture (IVC) by grouping oocytes and embryos of individual bovine donors per quadrant. At day 7, a significantly higher blastocyst rate was noted in the Corral (R) dish used during IVM and IVC than when only used during IVM (12.9% +/- 2.10 versus 22.8% +/- 2.67) (P < 0.05). However, no significant differences in blastocyst yield were observed anymore between treatment groups at day 8 post insemination. Conclusions: In the present study, the Corral (R) dish was used for in vitro embryo production (IVP) in cattle; allowing to allocate oocytes and/or embryos per donor. As fresh embryo transfers on day 7 have higher pregnancy outcomes, the Corral (R) dish offers an added value for commercial OPU/IVP, since a higher blastocyst development at day 7 is obtained when the Corral (R) dish is used during IVM and IVC

The growth of plant embryos in vitro. Preliminary experiments on the role of accessory substances

The aseptic culture of plant embryos isolated from the seed dates back to the work of Brown and Morris,(1) Hannig(2) and Dietrich.(3) More recent contributions to our knowledge concerning the culture in vitro of excised embryos have been made by Tukey,(4) Brunner(5) and LaRue,(6) among others. It has been recognized by, for example, Ray(7) that the embryo culture technique offers a useful tool for biochemical investigations, and it has also been recognized(4,8) that it may be used as a practical measure to circumvent the abortion of embryos. It has, however, been found that in general the growth of the excised embryo, even upon a medium containing essential inorganic materials and sugar, is far less than that of normal intact seedlings. This has led to the suggestion(5) that "accessory growth factors" which are needed in minute amounts, are required by the developing plant as they are by the developing animal organism. The present work, as well as that of Kogl and Haagen-Smit,(9) furnish final proof that this is the case; that these accessory substances, although normally furnished by the seed, may be replaced to some extent by pure compounds added in small amounts to the embryo culture medium. These investigations, taken up early in 1936, are concerned particularly with orienting experiments undertaken with an ultimate view toward the elucidation of the nature and mode of action of these accessory growth factors. The embryo culture technique is here to be used as a tool in the "hormonal" analysis of plant development

Embryo metabolism : what does it really mean?

The study of early embryo metabolism has fascinated researchers in the field for nearly a century. Herein, we give a brief account of the general features of embryo metabolism and some consideration of the research performed to reach such conclusions. It is becoming increasingly obvious that metabolism informs many fate decisions and outcomes beyond ATP generation, such as DNA methylation, Reactive Oxygen Species generation and cell signaling. We discuss the reasons for studying metabolism in the face of our current knowledge of the effect that the culture environment on the developing embryo and the downstream effects that can cause. The study of in vitro embryo metabolism can also give us insight into developmental perturbations in vivo. The strengths and limitations of the methods we use to study metabolism are reviewed with reference to species-specific fundamental biology and plasticity and we discuss what the future holds for metabolic studies and the unanswered questions that remain

Review: Role of tubal environment in preimplantation embryogenesis: application to co-culture assays

The culture of early preimplantation stage embryo is still delicate and the metabolic pathways of embryos are not completely understood. Embryo needs are evolutionary during the preimplantation development, consequently it is difficult to meet embryo needs in vitro. Culture conditions have to respect several physical and chemical equilibria: such as redox potential, pH, osmotic pressure, metabolic flux of energetic compounds, endogenous pools of amino acids and transcripts, etc. Embryo culture media are generally supplemented with amino acids, glucose, other energetic metabolites and antioxidant compounds, vitamin, and growth factors etc. Furthermore autocrine and paracrine regulation of embryo development probably exist. In fact embryo culture conditions have to be as non-toxic as possible. Various types of co-culture systems have been devised to overcome these problems. Complex interrelations exist between embryos and co-cultured cells. The beneficial effects of co-cultured cells may be due to continuous modifications of the culture medium, i.e. the elimination of toxic compounds and/or the supply of embryotrophic factor

Modelling aspects of oviduct fluid formation in vitro

© 2017 Society for Reproduction and Fertility. Oviduct fluid is the microenvironment that supports early reproductive processes including fertilisation, embryo cleavage and genome activation. However, the composition and regulation of this critical environment remain rather poorly defined. This study uses an in vitro preparation of the bovine oviduct epithelium to investigate the formation and composition of in vitro-derived oviduct fluid (ivDOF) within a controlled environment. We confirm the presence of oviduct-specific glycoprotein 1 in ivDOF and show that the amino acid and carbohydrate content resembles that of previously reported in vivo data. In parallel, using a different culture system, a panel of oviduct epithelial solute carrier genes and the corresponding flux of amino acids within ivDOF in response to steroid hormones were investigated. We next incorporated fibroblasts directly beneath the epithelium. This dual culture arrangement represents more faithfully the in vivo environment and impacts on ivDOF composition. Lastly, physiological and pathophysiological endocrine states were modelled and their impact on the in vitro oviduct preparation was evaluated. These experiments help clarify the dynamic function of the oviduct in vitro and suggest a number of future research avenues, such as investigating epithelial-fibroblast interactions, probing the molecular aetiologies of subfertility and optimising embryo culture media

Effect of Cell Density and Benzyl Amino Purine on the Growth of Somatic Embryo of Citrus Mandarin Batu 55 (Citrus Reticulata Blanco.) in Liquid Culture

Citrus mandarin Batu 55 (Citrus reticulata Blanco.) is one of Indonesian fruits commodities that have high economic value and consumers demand. The propagation of citrus mandarin by plant tissue culture generally was carried out on solid medium. The liquid culture system could increase cell multiplication therefore it became alternative method of plant propagation through somatic embryogenesis. The effect of initial cell density and Benzyl Amino Purin (BAP) concentration in liquid media were investigated. The initial cells density and right concentration of BAP given in media can increase cell proliferation of somatic embryo in liquid culture. Globular somatic embryo were cultured on Murashige and Tucker media with initial cell density 4, 6, 8 and 10 mgL-1 and BAP 0, 0.25, 0.5, and 0.75 mgL-1. Growth evaluation of somatic embryo were obtained by weighing fresh and dry weight every 2 weeks for 8 weeks for initial cell density treatment and 6 weeks of BAP treatment. The result of the research showed that cell density affect the growth of somatic embryo of citrus mandarin. Somatic embryo with low cell density showed slower growth compared than high cell density. Peak growth occured in 6th cultured with cell density 10 mgL-1. In addition to cell density, the growth of somatic embryo in liquid culture was affected by BAP. The growth of somatic embryo on the media containing BAP showed better results than without BAP. The highest BAP concentration on media showed fresh and dry weight of somatic embryo increased. In this research, growth of somatic embryo is not optimal yet because fresh and dry weights of somatic embryo still increase with high concentration 0.75 mgL-1 of BAP

Modeling mammalian gastrulation with embryonic stem cells

Understanding cell fate patterning and morphogenesis in the mammalian embryo remains a formidable challenge. Recently, in vivo models based on embryonic stem cells (ESCs) have emerged as complementary methods to quantitatively dissect the physical and molecular processes that shape the embryo. Here we review recent developments in using embryonic stem cells to create both two and three-dimensional culture models that shed light on mammalian gastrulation.Comment: 18 pages, 1 figur

Human embryos cultured in vitro to 14 days

We know a great deal about the development of the mammalian embryo until the time that the blastocyst implants into the uterus. With model organisms such as the mouse, we have also developed a considerable understanding of development immediately around gastrulation as embryos can be recovered at this stage for short-term in vitro culture. However, the intervening period of development remained a ‘black box’ because it takes place as the blastocyst is implanting into the uterus. Over the past 6 years, techniques pioneered and developed in Magdalena Zernicka-Goetz's laboratory for the in vitro culture of embryos through these implantation stages have opened up this box, affording the first glimpse of embryonic development through these previously hidden stages. Remarkably, the techniques developed with mouse embryos are equally applicable to human embryos, ushering the very first opportunities for studying our own development throughout this time. Here, I outline how the culture methods were developed, paving the way to culture of the human embryo to the point of gastrulation, an accomplishment recognized as the People's Choice for the Scientific Breakthrough of 2016 in Science magazine. I also discuss the new ethical challenges raised by the possibility of extending the time limits for human embryo culture

Embryo Donation: The Government Adopts a Cause

The disposition of cryopreserved supernumerary embryos has become a divisive issue that puts to test the tenets of the culture of life promoted by the Vatican and President George W. Bush. The Bush administration has spent millions of dollars to promote embryo adoptions while imposing restrictions on federal funding for embryonic stem cell research. On the other hand, contemporary Catholic moral theologians and philosophers disagree on the question of the morality of embryo rescue or adoption because the Church strongly opposes in vitro fertilization, the donation of gametes and embryo cryopreservation, as evidenced recently during the Italian fertility law referendum. President Bush has relied on ideologically charged culture of life rhetoric to promote embryo adoptions as the only alternative to dispose of cryopreserved ( frozen ) human embryos. In doing so, he has alienated an important segment of Christian pro-lifers who support embryonic stem cell research. From a Catholic perspective, the culture of life as conceived by Pope John Paul II vigorously opposes stem cell research, but embryo donation has not found its place within the Catholic culture of life, and substituting the word adoption for donation does not solve the perplexing dilemma

Embryo Donation: The Government Adopts a Cause

The disposition of cryopreserved supernumerary embryos has become a divisive issue that puts to test the tenets of the culture of life promoted by the Vatican and President George W. Bush. The Bush administration has spent millions of dollars to promote embryo adoptions while imposing restrictions on federal funding for embryonic stem cell research. On the other hand, contemporary Catholic moral theologians and philosophers disagree on the question of the morality of embryo rescue or adoption because the Church strongly opposes in vitro fertilization, the donation of gametes and embryo cryopreservation, as evidenced recently during the Italian fertility law referendum. President Bush has relied on ideologically charged culture of life rhetoric to promote embryo adoptions as the only alternative to dispose of cryopreserved ( frozen ) human embryos. In doing so, he has alienated an important segment of Christian pro-lifers who support embryonic stem cell research. From a Catholic perspective, the culture of life as conceived by Pope John Paul II vigorously opposes stem cell research, but embryo donation has not found its place within the Catholic culture of life, and substituting the word adoption for donation does not solve the perplexing dilemma