Efficient blastomere biopsy for mouse embryo splitting for future applications in human assisted reproduction

Abstract

The objective of the current study was to establish a safe, efficient biopsy procedure for embryo splitting using the mouse model for future applications in human assisted reproduction. From mouse embryos at the 2-, 4-, 6- and 8-cell stage, half the number of blastomeres were microsurgically biopsied and transferred into empty mouse zonae pellucidae. Twin embryonic development was monitored during in-vitro culture. Blastocyst developmental rate using 2-, 4-, 6-, and 8-cell splitting was 74.4, 75.0, 66.7 and 38.4 respectively, with corresponding hatching rates of 94.9, 97.5, 92.7 and 83.8%. Blastocysts from 2-, 4-, and 6-cell splitting resulted in elevated hatching rates compared with non-operated blastocysts (87.5%), due to the Tyrode-assisted hatching effect. Blastocyst morphology was superior from 2- and 4-cell splitting when compared with 6- and 8-cell splitting. Furthermore, outgrowth of twin blastocysts from 2- and 4-cell splitting showed well-developed colonies with trophoblast cells and clusters of ICM cells, whereas those obtained from 6- and 8-cell splitting frequently formed small-sized colonies. Due to the high twinning success rate obtained under the experimental conditions employed in this study, it appears that with further modifications and proper safeguards, such embryo splitting efforts could have potential applications in humans.

Evaluation of Blastomere Biopsy Using a Mouse Model Indicates the Potential High Risk of Neurodegenerative Disorders in the Offspring

“…Many infertile couples benefit from PGD technology, and experimental studies have focused on evaluating procedure-related effects on the development of biopsied preimplantation embryos (30). However, it is clear that we must also recognize the potential risks to fetuses, neonates, and adults derived from this technology.…”

In Vitro Production of Embryos: Principles, Techniques and Applications

“…After removing one half of the bisected embryo an algae substance similar to the zonapellucida is added to the daughter demi-embryo. Embryo splitting at the 6-8 cell stage provided a much higher developmental efficiency compared to splitting at the 2-4 cell stage [14]. However, it is not preferable to bisect after 8-cell stage, because after this stage blastomeres start to differentiate into specialized cells.…”

“…Human embryo splitting has been reported recently [17]. However, the complex ethical, spiritual and legislative concerns should be closely observed when dealing with human embryo splitting [14].…”

Human embryo twinning with applications in reproductive medicine

“…From human triploid embryos at the 2-to 5-cell and 6-to 8-cell stage, blastomeres were biopsied following our previously published procedures (18,19). However, additional adaptations and technical refinements had first to be established for further improving blastomere biopsies on human embryos.…”

“…According to these recommendations, we have first established efficient blastomere biopsy for embryo splitting in the mouse as an experimental model system (18,19).…”