Improved blastocyst development of single cow OPU-derived presumptive zygotes by group culture with agarose-embedded helper embryos

• Main Authors: Dey Shukla, Bang Jae, Choi Byung, Kwon Tae, Jin Jong, Deb Gautam, Cho In, Kong Il Keun
• Format: Article
• Language: English
• Published: BMC 2011-08-01
• Series: Reproductive Biology and Endocrinology
• Online Access: http://www.rbej.com/content/9/1/121

<p>Abstract</p> <p>Background</p> <p>The <it>in vitro </it>culture of presumed zygotes derived from single cow ovum pick-up (OPU) is important for the production of quality blastocysts maintaining pedigree. The aim of the present study was to evaluate the agar chip-embedded helper embryo coculture system for single cow OPU-derived zygotes by assessing embryo quality.</p> <p>Methods</p> <p>Cumulus oocyte complexes (COCs) were collected from Hanwoo cows with high genetic merit twice a week using the ultra-sound guided OPU technique and from slaughterhouse ovaries. The Hanwoo cow COCs and slaughterhouse ovaries were matured <it>in vitro</it>, fertilized <it>in vitro </it>with thawed Hanwoo sperm and cultured for 24 h. The presumed zygotes were subsequently placed in three different culture systems: (1) control OPU (controlOPU) with single cow OPU-derived presumed zygotes (2~8); (2) agar chip-embedded slaughterhouse helper embryo coculture (agarOPU) with ten presumed zygotes including all presumed zygotes from a cow (2~8) and the rest from agar chip-embedded slaughterhouse presumed zygotes (8~2); and (3) slaughterhouse <it>in vitro </it>embryo production (sIVP) with ten slaughterhouse ovary-derived presumed zygotes, each in 50 μL droplets. Day 8 blastocysts were assayed for apoptosis and gene expression using real time PCR.</p> <p>Results</p> <p>The coculture system promoted higher blastocyst development in OPU zygotes compared to control OPU zygotes cultured alone (35.2 vs. 13.9%; P < 0.01). Genes predicted to be involved in implantation failure and/or embryo resorption were down-regulated (P < 0.05) in control OPU zygotes (<it>CD9</it>, 0.4-fold; <it>AKRAB</it>1, 0.3-fold) and in cocultured zygotes (<it>CD9</it>, 0.3-fold; <it>AKRAB</it>1, 0.3-fold) compared to sIVP blastocysts (1.0-fold). Moreover, genes involved in implantation and/or normal calf delivery were up-regulated (P < 0.05 to P < 0.01) in control OPU zygotes (<it>PGSH</it>2, 5.0-fold; <it>TXN</it>, 4.3-fold; <it>PLAU</it>, 1.7-fold) and cocultured zygotes (<it>PGSH</it>2, 14.5-fold; <it>TXN</it>, 3.2-fold; <it>PLAU</it>, 6.8-fold) compared to sIVP (1.0-fold) blastocysts. However, the expression of <it>PLAC8, TGF-β1, ODC1</it>, <it>ATP5A1 </it>and <it>CASP3 </it>did not differ between the three culture groups.</p> <p>Conclusions</p> <p>Results show that the agar chip-embedded helper embryo coculture system enhances developmental competence and embryo quality in cultures of limited numbers of high pedigree single cow OPU presumed zygotes.</p>