Effects Of Nitric Oxide On Mouse Embryos Cultured In Vitro

In vitro culture(IVC) technology is one of the most important procedures for embryo production in vitro.Recently,many studies demonstrated that nitric oxide(NO),as a signal molecular,also has its key role in the control of mammalian reproductive system including in vitro fertilization and embryo development.But there had been little reports about how much concentration of nitric oxide is needed for these two procedures.The objective of this study was to investigate effects of the concentration of nitric oxide on mouse IVF and preimplantation embryo development and the treatments were:NO donor of SNP(sodium nitroprusside ) at the concentrations of 0,0.001,0.01,0.1 and lmmol/L were added into M16 and HTF. Mouse oocytes can be fertilized and cultured up to the blastocyst stage.The cleavage rate,4-cell embryos rate,blastocysts rate and the number of total cells per blastocyst was observed.The results were:①There were no significant differences in terms of cleavage rate among 0.01 mmol/L SNP 0.001 mmol/L SNP and 0 mmol/L SNP(1＞0.05).For 4-cell embryos rate 0.01 mmol/L SNP was significantly higher than 0.001 mmol/L SNP and 0 mmol/L SNP(P＜0.05),whilst there were no significant differences between 0.001 mmol/L SNP and 0 mmol/L SNP(P＞0.05).There were no significant differences of the blastocysts rate among 0.01 mmol/L SNP,0.001 mmol/L SNP and 0 mmol/L(P＞0.05).However,the total cell number per blastocyst of 0.001 mmol/L SNP was significantly higher than that in 0.001 mmol/L SNP and 0 mmol/L SNP(P＜0.05) group.The rates of cleavage,4-cell embryos and blastocysts,and the total cell number per blastocyst in 0.01 mmol/L SNP,0.001 mmol/L SNP and 0 mmol/L SNP were all significantly higher than those in 0.1 mmol/L SNP and 1.0 mmol/L SNP(P＜0.01) groups.Also the 0.1 mmol/L SNP was significantly higer than 1.0 mmol/L SNP (P＜0.01).②Addition of SNP before 4-cell stage,there were no significant differences regarding to the blastocysts rate between 0.001 mmol/L SNP and 0 mmol/L SNP or 0.01 mmol/L SNP(P＞0.05),while 0.01 mmol/L SNP was significantly higher than 0 mmol/L SNP(P＜0.05),and 0.01 mmol/L SNP,0.001 mmol/L SNP and 0 mmol/L SNP were significantly higher than those in 0.1 mmol/L SNP and 1.0 mmol/L SNP(P＜0.01).Also in the the 0.1 mmol/L SNP group,its blastocyst rate was significantly higer than 1.0 mmol/L SNP(P＜0.01).③For the IVF, wo found the cleavage rate of embryos treated with 0.001 mmol/L SNP was significantly higher than that with 0.01 mmol/L SNP and 0 mmol/L SNP(P＜0.05),but there were no significant differences between 0.01 mmol/L SNP and 0 mmol/L SNP(P＞0.05),there were also no significant differences of the blastocysts rate of IVF embryos derived from treatment with 0 mmol/L SNP,0.01 mmol/L SNP,0.001 mmol/L SNP(P＞0.05),the total cell number per blastocyst of 0.001 mmol/L SNP group was higher than that of 0.01 mmol/L SNP and 0 mmol/L SNP(P＜0.05) groups, while there has no signification differences between 0.01 mmol/L SNP and 0 mmol/L SNP(P＞0.05),but the cleavage rate and the blastocysts rate of 0.01 mmol/L SNP,0 mmol/L SNP and 0.001 mmol/L SNP groups were significantly higher than those of 0.1 mmol/L SNP and 1.0 mmol/L SNP(P＜0.01) groups,moreover such parameters in 0.1 mmol/L SNP group were significantly higher than that of 1.0 mmol/L SNP(P＜0.01).④For the in vitro cultured embryos,we found 4-cell embryo rate in 0.01 mmol/L SNP group was significantly higher than that in 0.001 mmol/L SNP and 0 mmol/L SNP(P＜0.05) groups,however,there were no significant differences between 0.001 mmol/L SNP and 0 mmol/L SNP(P＞0.05).No significant differences were found,for the blastocysts rate of embryos,among 0.01 mmol/L SNP,0.001 mmol/L SNP and 0 mmol/L(P＞0.05) groups.All of the them were significantly higher than that in 0.1 mmol/L SNP and 1.0 mmol/L SNP(P＜0.01) treatments,the rate of blastocyt in 0.1 mmol/L SNP was significantly higher than that in 1.0 mmol/L SNP(P＜0.01).⑤for the quality of embryos evaluated mainly by total cell number, we found 0.1 mmol/L SNP and 1.0 mmol/L SNP could inhibit the development of early embryos in mouse.Taken together,our results showed that treatments by 0.01 mmol/L and/or 0.001 mmol/L SNP during in vitro fertilization and in vitro culture could promote early development of mouse embryos.But high concentrations of SNP would exert negative effects on preimplantational development of embryos in mouse.