Analysis of compaction initiation in human embryos by using time-lapse cinematography

Abstract

Purpose: To analyze the initiation of compaction in human embryos in vitro by using time-lapse cinematography (TLC), with the goal of determining the precise timing of compaction and clarifying the morphological changes underlying the compaction process.

Methods: One hundred and fifteen embryos donated by couples with no further need for embryo-transfer were used in this study. Donated embryos were thawed and processed, and then their morphological behavior during the initiation of compaction was dynamically observed via time-lapse cinematography (TLC) for 5 days.

Results: Although the initiation of compaction occurred throughout the period from the 4-cell to 16-cell stage, 99 (86.1 %) embryos initiated compaction at the 8-cell stage or later, with initiation at the 8-cell stage being most frequent (22.6 %). Of these 99 embryos, 49.5 % developed into good-quality blastocysts. In contrast, of the 16 (13.9 %) embryos that initiated compaction prior to the 8-cell stage, only 18.8 % developed into good-quality blastocysts. Embryos that initiated compaction before the 8-cell stage showed significantly higher numbers of multinucleated blastomeres, due to asynchronism in nuclear division at the third mitotic division resulting from cytokinetic failure.

Conclusions: The initiation of compaction primarily occurs at the third mitotic division or later in human embryos. Embryos that initiate compaction before the 8-cell stage are usually associated with aberrant embryonic development (i.e., cytokinetic failure accompanied by karyokinesis).

Fig. 1

Time-lapse cinematography (TLC). The inverted microscope was equipped with a CCD digital camera (Roper Scientific Photometrics, USA) connected to a computer, and images were displayed via MetaMorph (Universal Imaging Co., USA). Digital images of the cultured embryos were acquired for approximately 40 h, with an exposure time of 50 ms. In total, 2,000 to 8,000 images were taken during the observation period

Fig. 2

Compaction in the human embryo. After several cell divisions (a–e), the blastomeres became flattened (f), and the intercellular boundaries became obscured (g–i), until they finally unified in one cluster (j, k). These morphological changes are called “compaction”, and blastulation occurs only after complete compaction of the embryo (l)

Fig. 3

Initiation of compaction in human embryos. The values above the histogram indicate the numbers of embryos that have commenced compaction at each cell stage. For the 115 embryos examined, the initiation of compaction was distributed from the 4-cell stage to the 16-cell stage, and 86.1 % (99/115) started compaction at or just after the 8-cell stage. In contrast, only 13.9 % (16/115) of the embryos started compaction before the 8-cell stage

Fig. 4

Cytokinetic failure and multinucleated blastomere formation. Still-frames of a representative embryo showing the process of cytokinetic failure and multinucleated blastomere (MNB) formation are shown. In each panel, the time from the start of imaging is displayed (hours: minutes). At the 4-cell stage, the embryo showed one nucleus in each blastomere (a). At about 22 h into the time-lapse recording (b), one of the blastomeres started to cleave, and gradually formed the cleavage furrow (blue arrow; c, d), but then failed to cleave (e–g). At 51 h into the recording, the blastomere returned to its original shape, but with multi-nucleation (yellow arrow; h)