OBJECTIVE: To investigate the influence of epidermal growth factor (EGF) on preimplantation development, implantation, and expression of epidermal growth factor receptor (EGFR) itself in mouse embryos. MATERIALS AND METHOD: Eight-cell stage mouse embryos were cultured for 48 hours with EGF at concentrations of 0.1, 1.0, 10 and 100 ng/ml. Embryos not treated with EGF were served as control. The percentages of embryos which developed to the expanded, hatched blastocyst stage and in vitro implantation at 48 hours were determined. Reverse transcription-polymerase chain reaction (RT-PCR) has been used to examine the expression of EGFR in developed hatched blastocysts. Following reverse transcription, strategically designed nested primers, optimized for specificity, were used for amplification from the cDNA equivalent of a single embryo. The products were then verified by restriction enzyme digestion and sequence analysis. Results were analyzed with chi 2 test and Student's t-test as appropriate, and statistical significance was defined as p < 0.05. RESULTS: The percentages of fully expanded blastocysts at 48 hours in all the EGF treated group were not significantly different from the control. The percentages of hatched blastocysts were significantly higher in the EGF treatment group at 0.1 ng/ml (90.5 +/- 9.8%) compared to the control (82.1 +/- 7.2%), 1.0 ng/ml (82.2 +/- 12.7%), and 100 mg/ml (81.9 +/- 11.8%) (p < 0.05, p < 0.05, p < 0.05, respectively). The percentages of hatched blastocysts were significantly higher in the EGF treatment group at 10 ng/ml (89.4 +/- 7.5%) compared to the control, and 100 ng/ml (p < 0.05, p < 0.05, respectively). The percentages of attached blastocysts in vitro were significantly higher following incubation with EGF at concentrations of 0.1 ng/ml (37.0 +/- 17.0%), 1.0 ng/ml (32.0 +/- 14.3%), 10 ng/ml (21.3 +/- 7.2%) compared to the control (9.5 +/- 7.7%) (p < 0.05, p < 0.05, p < 0.05, respectively). The attachment rates in 0.1 ng/ml and 1.0 ng/ml EGF treatment groups were also significantly higher than those in other EGF treatment groups. Embryo development and attachment were not significantly inhibited or enhanced in cultures supplemented with 100 ng/ml EGF compared to the control. The mRNA concentration of EGFR in embryos treated with 0.1 ng/ml of EGF was significantly higher than those of the control and other EGF treatment groups. CONCLUSION: EGF may have a stimulatory role in later stage embryonic development, implantation and expression of EGFR in hatched blastocyst itself at the specific concentration.
OBJECTIVE: To investigate the influence of epidermal growth factor (EGF) on preimplantation development, implantation, and expression of epidermal growth factor receptor (EGFR) itself in mouse embryos. MATERIALS AND METHOD: Eight-cell stage mouse embryos were cultured for 48 hours with EGF at concentrations of 0.1, 1.0, 10 and 100 ng/ml. Embryos not treated with EGF were served as control. The percentages of embryos which developed to the expanded, hatched blastocyst stage and in vitro implantation at 48 hours were determined. Reverse transcription-polymerase chain reaction (RT-PCR) has been used to examine the expression of EGFR in developed hatched blastocysts. Following reverse transcription, strategically designed nested primers, optimized for specificity, were used for amplification from the cDNA equivalent of a single embryo. The products were then verified by restriction enzyme digestion and sequence analysis. Results were analyzed with chi 2 test and Student's t-test as appropriate, and statistical significance was defined as p < 0.05. RESULTS: The percentages of fully expanded blastocysts at 48 hours in all the EGF treated group were not significantly different from the control. The percentages of hatched blastocysts were significantly higher in the EGF treatment group at 0.1 ng/ml (90.5 +/- 9.8%) compared to the control (82.1 +/- 7.2%), 1.0 ng/ml (82.2 +/- 12.7%), and 100 mg/ml (81.9 +/- 11.8%) (p < 0.05, p < 0.05, p < 0.05, respectively). The percentages of hatched blastocysts were significantly higher in the EGF treatment group at 10 ng/ml (89.4 +/- 7.5%) compared to the control, and 100 ng/ml (p < 0.05, p < 0.05, respectively). The percentages of attached blastocysts in vitro were significantly higher following incubation with EGF at concentrations of 0.1 ng/ml (37.0 +/- 17.0%), 1.0 ng/ml (32.0 +/- 14.3%), 10 ng/ml (21.3 +/- 7.2%) compared to the control (9.5 +/- 7.7%) (p < 0.05, p < 0.05, p < 0.05, respectively). The attachment rates in 0.1 ng/ml and 1.0 ng/ml EGF treatment groups were also significantly higher than those in other EGF treatment groups. Embryo development and attachment were not significantly inhibited or enhanced in cultures supplemented with 100 ng/ml EGF compared to the control. The mRNA concentration of EGFR in embryos treated with 0.1 ng/ml of EGF was significantly higher than those of the control and other EGF treatment groups. CONCLUSION:EGF may have a stimulatory role in later stage embryonic development, implantation and expression of EGFR in hatched blastocyst itself at the specific concentration.