Effect of culture system on the yield and quality of bovine blastocysts as assessed by survival after vitrification.
The aim of this study was to assess the effect of a bovine in vitro culture system on blastocyst yield and quality after vitrification. In Experiment 1, IVM/IVF zygotes were cultured in either synthetic oviduct fluid (SOF) in 5% CO2, 5% O2, 90% N2; or TCM199-granulosa cells (TCM199-GCM) in 5% CO2 in air. In vivo blastocysts were used as a control. Culture in SOF resulted in a significantly higher blastocyst yield on both Day 7 (31.3 vs 13.2%, P < 0.001) and 8 (36.8 vs 23.7%, P < 0.001) than did culture in TCM199-GCM. After vitrification, survival at 72 h of in vivo blastocysts was significantly higher than both in vitro groups, while significantly more blastocysts produced in TCM199-GCM survived compared to those produced in SOF (0, 43.5, 78.3% for SOF, TCM199-GCM and in vivo, respectively P < 0.01). In Experiment 2, SOF-GCM proved to be the best post-warming culture system of those tested and was adopted as the post-warming medium for all subsequent experiments. In Experiment 3, zygotes were cultured in SOF or SOF-GCM, in either 5% CO2 in air, or 5% CO2, 5% O2, 90% N2. In agreement with Experiment 1, culture in SOF in 5% O2 resulted in significantly more blastocysts at Day 7 (26.4 vs 17.3%, P < 0.01) and Day 8 (31.5 vs 23.2%, P < 0.01) than did culture in SOF-GCM. However, survival at 72 h post vitrification was significantly higher for SOF-GCM (44 vs 8.3%, P < 0.001). Increasing the O2 concentration to 20% significantly reduced the blastocyst eld from SOF (31.5 vs 17.3%, P < 0.001). In addition, the quality of blastocyst produced was reduced in terms of survival post vitrification (8.3 vs 0%, P < 0.05). In contrast, there was no difference in blastocyst yield (23.2 vs 25.2%) or survival (44.0 vs 36.9%) in SOF-GCM, irrespective of O2 concentration. Experiment 4 examined the duration of exposure to GCM necessary to acquire improved blastocyst quality. Zygotes were cultured in SOF; SOF until Day 3, followed by SOF-GCM for the remainder of the culture; SOF until Day 5, followed by SOF-GCM for the remainder of the culture; or SOF-GCM for the entire culture. Survival at 72 h post vitrification was significantly higher (P < 0.05) in Groups 2 (50.0%, 13/26) and 4 (55.3%, 26/47) than in Groups 1 (21.7%, 10/46) and 3 (10.8%, 4/37). In conclusion, culture system can affect blastocyst yield and quality and crytolerance is a useful indicator of blastocyst quality.
Rizos
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Department of Animal Science and Production, University College Dublin, Lyons Research Farm, Newcastle, County Dublin, Ireland.
Ward
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Boland
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MP
Lonergan
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D000818:Animals; D001755:Blastocyst / Q000502:physiology*; D002417:Cattle / Q000502:physiology*; D018920:Coculture Techniques / Q000662:veterinary; D015925:Cryopreservation / Q000379:methods / Q000662:veterinary*; D003470:Culture Media; D005260:Female; D005307:Fertilization in Vitro / Q000379:methods / Q000662:veterinary; D008297:Male; D009865:Oocytes / Q000502:physiology; D011247:Pregnancy; D013480:Superovulation
D003470:Culture Media