Anti-apoptotic effect of insulin-like growth factor (IGF)-I and its receptor in porcine preimplantation embryos derived from in vitro fertilization and somatic cell nuclear transfer.

Insulin-like growth factor (IGF)-I is a receptor-mediated autocrine and/or paracrine growth and/or survival factor for mammalian embryo development. It is known to promote the growth and development of mouse preimplantation embryos. The present study was designed to investigate the effects of IGF-I (50 ng/ml), anti-IGF-I receptor antibody (50 ng/ml) and their combination on porcine preimplantation embryo development. Furthermore, the mechanism underlying the embryotropic effects of IGF-I was evaluated by monitoring the incidence of apoptosis and expression of apoptosis-related genes. In both in vitro fertilized (IVF) and somatic cell nuclear transfer (SCNT) embryos, culturing with IGF-I increased the rate of blastocyst formation and this embryotrophic effect was neutralized by culturing with IGF-I along with anti-IGF-I receptor (IGF-IR) antibody. Culturing IVF and SCNT embryos with IGF-I significantly increased the number of total cells in blastocysts and decreased the number of apoptotic nuclei. These effects of IGF-I were also neutralized by culturing with IGF-I along with anti-IGF-IR antibody. Expression of the anti-apoptotic Bcl-2 gene was increased, while expression of the pro-apoptotic Bax was decreased in both IVF and SCNT embryos cultured with IGF-I. In both IVF and SCNT embryos, anti-IGF-IR antibody along with IGF-I neutralized the effect of IGF-I on expression of Bcl-2 and Bax genes. In conclusion, the present study demonstrated that IGF-I through its specific receptors improved the developmental competence of IVF and SCNT embryos by decreasing the incidence of apoptosis and regulating apoptosis-related genes in porcine preimplantation embryos.

Kim S ，Lee SH ，Kim JH ，Jeong YW ，Hashem MA ，Koo OJ ，Park SM ，Lee EG ，Hossein MS ，Kang SK ，Lee BC ，Hwang WS ... -  《MOLECULAR REPRODUCTION AND DEVELOPMENT》

Embryotropic effect of insulin-like growth factor (IGF)-I and its receptor on development of porcine preimplantation embryos produced by in vitro fertilization and somatic cell nuclear transfer.

Kim S ，Lee GS ，Lee SH ，Kim HS ，Jeong YW ，Kim JH ，Kang SK ，Lee BC ，Hwang WS ... -  《MOLECULAR REPRODUCTION AND DEVELOPMENT》

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.

Biswas D ，Jung EM ，Jeung EB ，Hyun SH ... -  《-》

Expression of leptin ligand and receptor and effect of exogenous leptin supplement on in vitro development of porcine embryos.

Kim HS ，Lee GS ，Kim JH ，Kang SK ，Lee BC ，Hwang WS ... -  《THERIOGENOLOGY》

Effect of epidermal growth factor in preimplantation development of porcine cloned embryos.

In this study, we determined the expression of epidermal growth factor (EGF) and its receptor (EGFr) gene, and the effect of exogenous EGF supplementation on preimplantation development of porcine somatic cell nuclear transfer (SCNT) embryos. In vitro matured gilt oocytes were fertilized with frozen-thawed semen in vitro or reconstructed with fetal fibroblasts by SCNT. In Experiment 1, total RNA was isolated from oocytes, preimplantation SCNT, or in vitro fertilization (IVF) embryos. The expression of EGF and EGFr mRNA was determined using reverse transcription-polymerase chain reaction (RT-PCR). In SCNT and IVF embryos, the EGF mRNA was detected in oocytes, 2-cell, 4-cell, 8-cell, morulae, and blastocysts, while EGFr mRNA was detected in oocytes, 2-cell, morulae, and blastocysts. In Experiment 2, SCNT embryos at 1-cell stage were cultured in North Carolina State University (NCSU)-23 medium supplemented with different concentrations of EGF (0.1, 1, or 10 ng/ml). Supplementing with 10 ng/ml EGF improved cleavage rate (82.8% vs. 76.8%, P<0.05), but not the rate of blastocyst formation compared to the control. At all concentrations, EGF increased (P<0.05) the total cell number in blastocysts (range 50.5-53.7 vs. 43.9). In Experiment 3, EGF (10 ng/ml) was added to NCSU-23 medium at the morula stage. The EGF did not affect blastocyst formation, total cell number in blastocysts or the ratio of inner cell mass (ICM) to total cell number. In conclusion, we demonstrated that EGF and EGFr mRNA are expressed in porcine IVF and SCNT preimplantation embryos, and that EGF increased the quality of blastocysts by increasing total cell numbers in porcine SCNT embryos.

Lee GS ，Kim HS ，Hyun SH ，Jeon HY ，Nam DH ，Jeong YW ，Kim S ，Kim JH ，Kang SK ，Lee BC ，Hwang WS ... -  《MOLECULAR REPRODUCTION AND DEVELOPMENT》