In vitro culture of pig embryos

Reed, Michael L.; Illera, M.; Petters, Robert M.

doi:10.1016/0093-691x(92)90249-q

Cited by 78 publications

(32 citation statements)

References 34 publications

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“…The normal formulation of NCSU23 contains three amino acids-glutamine, taurine, and hypotaurine-which are not present in the essential and nonessential amino acid formulations. Preimplantation pig embryos can use glutamine as an energy substrate (Petters et al, 1990), and taurine and hypotaurine have been shown to improve in vitro development either individually or more effectively, in combination (Reed et al, 1992). All three have been implicated in protecting the embryo from oxidative stress (Guérin et al, 2001;Reiger, 1992), and taurine and hypotaurine have also been shown to protect mouse embryos from osmotic stress (Dawson and Baltz, 1997).…”

Section: Energy Substrates and Amino Acids In Pig Culture 211mentioning

confidence: 97%

The Effect of Energy Substrate Concentration and Amino Acids on the In Vitro Development of Preimplantation Porcine Embryos

Beebe

McIlfactrick²,

Nottle³

2007

Cloning and Stem Cells

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As the pig becomes increasingly used for biomedical research, an effective and efficient in vitro culture system is essential. This study aimed to improve the commonly used porcine embryo culture medium, NCSU23, by altering the energy substrates and adding amino acids, using electrically activated diploid parthenotes from oocytes obtained from the ovaries of prepubertal and adult animals. Morphological development to day 6 and blastocyst cell number were examined. Glucose (5.56 mM) was replaced by pyruvate and lactate (0.2 mM and 5.7 mM, respectively) for either the entire culture period or for the first 48 h only. Blastocyst rates were not different between any of the treatments, and were similar for prepubertal and adult oocytes. When the embryos were cultured with pyruvate and lactate for the first 48 h and then glucose, there was a significant increase in blastocyst cell number compared to glucose only. Blastocysts produced using pyruvate and lactate for the entire time tended to have more cells than those exposed to glucose only and less than those who were cultured in pyruvate and lactate for the first 48 h and then glucose. Nonessential amino acids added for the first 48 h and nonessential and essential amino acids added for the remaining time significantly increased blastocyst cell number only when the embryos were grown in pyruvate and lactate followed by glucose. Blastocyst rates were not different between any of the treatments, and this result was the same when using sow or gilt oocytes. The modified medium was then tested using in vitro matured and fertilized embryos from sow oocytes. Blastocyst rates and cell number were significantly increased in the modified medium compared to those grown in unmodified NCSU23. This shows that altering energy substrates and adding amino acids can increase the quantity and cell number of IVP blastocysts compared with NCSU23.

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Section: Energy Substrates and Amino Acids In Pig Culture 211mentioning

confidence: 97%

The Effect of Energy Substrate Concentration and Amino Acids on the In Vitro Development of Preimplantation Porcine Embryos

Beebe

McIlfactrick²,

Nottle³

2007

Cloning and Stem Cells

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“…It is likely that 10 mM of glycine might be too high to support the development of embryos at the oviductal stage, but glycine could enhance the development of uterine stage embryos under the present culture conditions. Supplementation of media with taurine has been demonstrated to enhance in vitro development of mouse [5,26], hamster [17], rabbit [13], pig [24], and cattle embryos [16]. Most of these studies have performed embryo culture in a gas atmosphere of 5% CO 2 in air (approximately 20% …”

Section: Discussionmentioning

confidence: 99%

Effects of Glutamine, Glycine and Taurine on the Development of In Vitro Fertilized Bovine Zygotes in a Chemically Defined Medium.

Takahashi

Kanagawa

1998

The Journal of Veterinary Medical Science

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ABSTRACT. The effects of glutamine, glycine and taurine on the development of bovine zygotes derived from IVM/IVF were determined using a protein-free chemically defined medium. After the cumulus cells were removed at 18 hr post-insemination, the presumptive zygotes were cultured for 4 or 6 days (about 104 or 154 hr) under a gas atmosphere of 5% O 2 : 5% CO 2 : 90% N 2 . A modified synthetic oviduct fluid medium supplemented with 20 amino acids (1 mM glutamine, essential amino acids for basal medium Eagle and nonessential amino acids for minimum essential medium), insulin, and PVA was used as a basic medium (mSOFai). Omitting 1 mM glutamine from mSOFai did not affect the embryonic development after 4 and 6 days of culture. After 4 days of culture, no significant effects of glycine and taurine on the development of zygotes to the morula stage were observed. However, supplementation with glycine or taurine significantly (P<0.05) affected, with no interaction, the embryonic development to blastocysts after 6 days of culture. Addition of 5 mM glycine and 2 or 10 mM taurine significantly (P<0.05) increased the percentage of blastocysts. The mean cell number in the blastocysts was affected by the glycine level, and was increased by the addition of 10 mM glycine (P<0.001). These results demonstrate that glycine and taurine in a chemically defined medium containing a group of essential and non-essential amino acids improve the development of bovine zygotes to the blastocyst stage under 5% O 2 . -KEY WORDS: bovine embryo, culture, glutamine, glycine, taurine.

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“…Apart from glutamine, NCSU23 also contains two more amino acids, taurine and hypotaurine. Between them, these three amino acids are thought to protect the embryo from oxidative [15] as well as osmotic stress [16,17] and to improve the intracellular oxidative status [18]. We previously found that better embryonic development occurs in this medium when pyruvate (0.2 mM) and lactate (5.7 mM) are provided and the concentration of glucose is greatly reduced (0.6 mM) for the first 48 h post insemination, and glucose (5.6 mM) is provided exclusively for the remainder of the culture period.…”

mentioning

confidence: 99%

Adding Essential Amino Acids at a Low Concentration Improves the Development of In Vitro Fertilized Porcine Embryos

Beebe

Vassiliev

McIlfatrick

et al. 2009

Journal of Reproduction and Development

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Abstract. The aims of this study were to investigate improvements to the pig preimplantation embryo culture system using in vitro produced embryos. For experiment 1, the optimum time to change the medium from NCSU23 containing 0.6 mM glucose, 0.2 mM pyruvate, 5.7 mM lactate and nonessential amino acids to NCSU23 containing 5.6 mM glucose and both essential and nonessential amino acids was examined. There were no statistically significant differences in blastocyst rates or cell number when the medium was changed at 48, 72 or 96 h, although there was a consistent trend for the 96 h treatment to produce fewer blastocysts with fewer cells. For experiment 2, the addition of essential amino acids at either a 1:50 or a 1:100 dilution of the purchased stock solution for day 1 to 6 or for days 3 to 6 only was investigated. Adding essential amino acids at a 1:50 dilution for day 3 to 6 significantly reduced the blastocyst rate and adding them at a 1:50 dilution from day 1 to 6 significantly reduced both the blastocyst rate and blastocyst cell number compared to when it was added at a 1:100 dilution. Embryos were produced by IVF, cultured for 6 days and good quality blastocysts were transferred into 6 synchronized pseudopregnant recipients (24 to 35 blastocysts per recipient) resulting in 4 pregnancies and 21 live birth piglets. These results show that adding essential amino acids at a 1:100 dilution provided the best culture conditions and the blastocysts produced were able to attain full term development after transfer. Key words: Blastocysts, Essential amino acids, In vitro culture, Piglets, Porcine (J. Reprod. Dev. 55: [373][374][375][376][377] 2009) he pig is becoming increasingly important as a biomedical research tool [1] and shows great promise as a model for embryonic stem cell technologies. The capacity to reliably generate embryonic stem cell lines is essential to using the pig as such a model. While in vivo produced embryos are ideal for producing cell lines, collecting them is expensive and time consuming. An alternative is to use embryos produced entirely in vitro. Obviously, a critical component in generating in vitro embryos is the culture medium in which they are grown.Several reports have investigated improving two commonly used porcine embryo culture media, North Carolina State University 23 and 37 (NCSU23 and NCSU37;[2]) [3][4][5]. As originally formulated, these media contain only glucose as the primary energy substrate. These reports found that providing pyruvate and lactate for the early preimplantation period (approximately 48 to 73 h post insemination) and only glucose for the remainder results in improved development and embryonic quality.Amino acids have been grouped by Eagle [6] into essential (EAA) and non-essential (NEAA), and are normally added to culture media as prepared concentrated stock solutions at a dilution of 1:50 for EAA and 1:100 for NEAA. Early cleavage development was found to be stimulated by the addition of NEAA to the culture medium but inhibited by EAA in both the mouse [...

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In vitro culture of pig embryos

Cited by 78 publications

References 34 publications

The Effect of Energy Substrate Concentration and Amino Acids on the In Vitro Development of Preimplantation Porcine Embryos

The Effect of Energy Substrate Concentration and Amino Acids on the In Vitro Development of Preimplantation Porcine Embryos

Effects of Glutamine, Glycine and Taurine on the Development of In Vitro Fertilized Bovine Zygotes in a Chemically Defined Medium.

Adding Essential Amino Acids at a Low Concentration Improves the Development of In Vitro Fertilized Porcine Embryos

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