Effects Of Insulin-like Growth Factor-Ⅰ (IGF-1)on The Development And Apoptosis Of Preimplantation Buffalo Embryos

This study was to explore the effects of insulin-like growth factor-I (IGF-1) on the development and apoptosis of IVF/SCNT preimplantation buffalo embryos, and investigate the influence mechanisms of IGF-Ion embryonic development and apoptosis, in order to provide scientific basis for improving the quality of embryos and the efficiency of embryos in vitro culture system.Five experiments were designed in this study. The experiment1, the mRNA expression level of IGF-1R on preimplantation buffalo IVF/SCNT embryos was investigated by Real-time quantitative PCR. The results indicated that IGF-1R were present in all development stage of preimplantation buffalo embryos, and the mRNA expression level of IGF-1R was higher in early embryos (2-cell in IVF,2-8cell in SCNT), which was no significant difference with oocytes (P>0.05), then gradually decreased, and the expression level was lowest in blastocyst stage. The experiment2, the embryos from IVF were respectively cultured in the medium containing with0,5,10,50, and100ng/mL IGF-I, there was no significant difference in the cleavage rate among each groups (P>0.05), but the blastocyst development rate of the50ng/mL IGF-I group was significantly increased than that of0ng/mL group (35.12%vs22.97%, P<0.05). Comparing with the control group (Ong/mL IGF-1), more embryos could develop to blastocyst stage when SCNT embryos cultured in the medium supplementing with50ng/mL IGF-I (46.43%vs34.18%, P<0.05), but the cleavage rate was not significant different (P>0.05). The experiment3, the result of Real-time quantitative PCR analyzing showed that treating IVF or SCNT embryos with50ng/mL IGF-I significantly up-regulated the mRNA expression level of IGF-IR comparing to untreated control group (P<0.05). The experiment4, the apoptosis and cell number of IVF/SCNT buffalo blastocysts were detected by TUNEL, it was found that relatively control group (0ng/mL IGF-I), the cell number of IVF/SCNT blastocysts was significantly increased and the apoptotic index was obviously decreased as the embryos cultured in the medium with50ng/mL IGF-I (P<0.05). The experiment5, the mRNA expression level of the anti-apoptotic bcl-2gene was distinctly enhanced (P<0.05), while the mRNA expression level of the pro-apoptotic bax gene was remarkably reduced (P<0.05) after IVF/SCNT embryos culturing with50ng/mL IGF-I.In conclusion, the present study demonstrated that IGF-IR mRNA were present in all development stage of preimplantation buffalo embryos and supplementing with50ng/mL IGF-1into the culture medium could improve the developmental competence of buffalo embryos and increase the cell number of blastocyst, as well as decrease the apoptotic index. Moreover, addition of IGF-I could promote the expression level of its specific receptors(IGF-1R), down-regulate the mRNA of pro-apoptotic bax gene and up-regulate the mRNA of anti-apoptotic bcl-2gene to influence the development and apoptosis of embryos.