The effect of in vitro culture on cleavage rates and morphology of the in vivo- developed embryos in mice|
Mohammad Ali Khalili, Ph.D., Morteza Anvari
Background: Research studies on reproductive mechanism of laboratory animals are essential for further advancement of assisted reproductive techniques (ART). One of these studies includes the assessment of in-vitro development of pre-implantation embryos. The objective was to compare the cleavage rates and morphology of in-vivo formed 2 to 8 cell embryos and blastocysts with in-vitro culture of the same embryos for 24 h.
Materials and Methods: 6-8 weeks old female NMRI mice were superovulated with 8IU pregnant mare's serum gonadotropin (PMSG, ip). Two superovulated animals were caged with one male mouse for mating. Mated mice were killed by cervical dislocation at different time intervals to collect a total of 200 (50/ each) 2, 4, 8, and blastocyst embryos from uterine tubes and horns. Following morphological evaluation and cleavage rates, all embryos were incubated in Whittingham's T6 media+5% BSA for 24 h. Following incubation at 37ºC in 5% CO2, the cleavage rates as well as morphological feature of each embryo was re-evaluated and compared with the original embryos.
Results: The best quality embryos collected from uterine tubes were at 2-cells stage, which were reduced when compared with in-vivo developed 4-8 cells embryos. 88% and 52% of 2 and 8 cells embryos were respectively at grade A stage. 28 embryos out of 50 eight-cell embryos were at grades C and D after incubation. Following in vitro culture, the development of 16%, 24%, 24%, and 40% of the 2, 4, 8 cells, and blastocysts were arrested, respectively. Also, only 2 blastocysts (8%) reached the hatching stage which in comparison with in-vivo blstocysts were increased (P>0.05).
Conclusion: In-vitro culture of the in-vivo formed embryos reduced their cleavage rates and morphology, especially at more advanced stages. Therefore, it becomes necessary to improve the in-vitro culture condition and to transfer the embryos at early stage to consequently improve the implantation rates.
NMRI mice, Embryo morphology, Cleavage, Culture media.