Effects of vascular endothelial growth factor on porcine preimplantation embryos produced by in vitro fertilization and somatic cell nuclear transfer.
This study examined the effects of vascular endothelial growth factor (VEGF) on porcine embryos produced by in vitro fertilization (IVF) and somatic cell nuclear transfer (SCNT) at different developmental stages. Four sets of experiments were performed. In the first, supplementation of the in vitro culture medium with 5 ng/mL VEGF was suitable for porcine IVF embryo development, and the blastocyst formation rate was significantly higher than the control and other groups (57.73 ± 6.78% (5 ng/mL VEGF) vs. 43.21 ± 10.22% (control), 42.16 ± 10.24% (50 ng/mL VEGF) and 41.91 ± 11.74% (500 ng/mL VEGF); P < 0.05). The total cell number after supplementation with 5 ng/mL VEGF was significantly higher than the control and other groups (151.85 ± 39.77 (5 ng/mL VEGF) vs. 100.00 ± 34.43 (control), 91.2 ± 31.51 (50 ng/mL VEGF), and 112.53 ± 47.66 (500 ng/mL VEGF); P < 0.05). In the second experiment, when VEGF was added at different developmental stages of IVF derived embryos (early stage, days 1-3, late stage, days 4-7), the blastocyst formation rate and total cell number were significantly higher at the late stage (47.71 ± 9.13% and 131.5 ± 20.70, respectively) than in the control (34.32 ± 7.44% and 85.50 ± 20.41, respectively) and at the early stage (33.60 ± 5.78% and 86.75 ± 25.10, respectively; P < 0.05). There was no significant difference in the blastocyst development rate or total cell number between the whole culture period (days 1-7) and the late stage culture period after supplementation with 5 ng/mL VEGF (P > 0.05). In the third experiment, the cleavage rate was significantly higher when SCNT embryos were cultured with VEGF during the whole culture period than in the late stage (63.56 ± 15.52% vs. 39.72 ± 4.94%; P < 0.05), but there was no significant difference between the control and the early stage culture period (P > 0.05). The blastocyst formation rate was significantly higher at the late stage culture period with VEGF than at the early stage culture period (34.40 ± 15.06% vs. (16.07 ± 5.01%; P < 0.05). There was no significant difference in the total cell number between the groups (P > 0.05). In experiment 4, using real-time PCR, VEGF mRNA expression was detected in all the developmental stages of IVF and SCNT embryos, but the expression level varied according to the developmental stage. VEGF receptor, KDR mRNA was detected in all stages IVF and SCNT embryos. However, flt-1 mRNA was not expressed in all embryonic stages of IVF and SCNT embryos. These data suggest that VEGF supplementation at the late embryonic developmental stage might improve the developmental potential of both IVF and SCNT preimplantation porcine embryos through its receptors.
Laboratory of Veterinary Embryology and Biotechnology, College of Veterinary Medicine, Chungbuk National University, Cheongju-361-763, Chungbuk, South Korea.
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Biswas
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Jung
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Jeung
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Hyun
D000818:Animals; D001755:Blastocyst / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D002478:Cells, Cultured; D046149:Embryo Culture Techniques / Q000379:methods*; D004622:Embryo, Mammalian; D047108:Embryonic Development / Q000187:drug effects* / Q000235:genetics; D005260:Female; D005307:Fertilization in Vitro; D018507:Gene Expression Regulation, Developmental; D008297:Male; D053652:Nuclear Transfer Techniques; D013552:Swine / Q000196:embryology*; D013997:Time Factors; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics / Q000378:metabolism; D042442:Vascular Endothelial Growth Factors / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*
D042442:Vascular Endothelial Growth Factors; D040301:Vascular Endothelial Growth Factor Receptor-2