Comparing The Effect Of 6-And 10- Days Old Chick Embryo Amniotic Fluid On Development Of Preimplantation Mouse Embryo



Introduction: Today, investigators pay attention to natural fluids (such as follicular and amniotic fluid) to use them as a medium or a supplement. Using human amniotic fluid (HAF) in some researches, not only improved in vitro development of preimplantation embryos but also significantly increased percentage of pregnancy. On the contrary, Gianaroli et al. have claimed there was no significant difference in fertilization, cleavage and pregnancy rates between HAF and control. Dorfmann et al. have also claimed HAF reduced pregnancy rate. Some investigators have worked on chick embryo amniotic fluid (CEAF). Blakwood et al and Ocampo et al. transferred preimplantion mammalian embryo in amniotic cavity of developing chick embryo (CEAm) and indicated better mammalian embryo development. However, using aspirated CEAF by Blakewood didn't show significant difference compare to control medium. Since CEAF is more available, its aspration is not dangerous and does not cost much in comparison to HAF and syntetic culture media, and there is also controversy on this pointe, decided: 1) To evaluate the possibility effects of 20%, 50%, 100% inactivated and 100% active CEAF on the development of mouse embryo. 2) To investigate the differences between 6- and 10- days old CEAF on the development of mouse embryo. 3) To introduce CEAF as natural medium at least for the mouse embryo culture.
Materials and methods: 6- and 10 - days old chick embryo amniotic fluid (6 - AF, 10- AF) were aspirated and two experiments were designed. Experiment one: 6 - AF was prepared as followed: Ham’s F - 10 + 20% inactivated AF (iAF20), Ham’s F - 10 +50% inactivated AF (iAF 50), inactivated AF (iAF 100), active AF (AF 100), Ham’s F-10 + 10% albuminar -5 (SA:as control). Two-cell mouse embryo (Rondom-Bred swiss white Mice strain) were assigned to these five groups randomly. Experiment two: All of these procedure were repeated for 10- AF, as well.
Results: Experiment one: In AF 100, iAF100 and iAF50 the percentage of total blastocyst (TB) was significantly higher than SA (87.5, 81.7, 82.1 vs 56.1 resp.)(p<0.001) Hatched blastocyst (HB) percentage was significantly higher in AF100 and iAF20compare to control(58,60.3 vs 22.5 resp.) iAF20 iAF50 iAF100 iAF100 SA TB 72.2 82.1* 81.7* 87.5* 56.1* HB 60.3* 50.8 52.2 58 22.5 *:P<0.001 Experiment two: percentage of TB was significantly higher in AFlOO and iAFlOO compare to SA(82.3,73.6 vs 51.9 resp.) (p<0.001). HB percentage in AF100,iAF100and iAF50 was significantly higher than SA(68,69.7,59.2 vs 30.3 resp.)(p<0.001). iAF20 iAF50 iAF100 iAF100 SA TB 96.2 65.9 73.6* 82.3* 51.9* HB 51.8* 59.2 69.7* 68* 30.3 *:p<0.001 No significant difference was found between two experimental groups, 6-AF and 10-AF,and between different densities of CEAF ,as well.
Conclusion: CEAF as a supplement or a natural medium could support the development of preimplantation mouse embryo.