Classification of Morphological Changes Based on the Number of Cleavage Divisions in Bovine Embryos

Abstract

Abstract. Quantification based on cleavage division (CD) of bovine preimplantation embryos facilitates quantitative analyses of embryonic developmental processes because CD occurs roughly once each day for all blastomeres for up to at least 9 days after ovulation. Therefore, embryonic morphological changes during this period were classified according to CD number. In this study, embryos collected from superovulated donors 0-9 days after ovulation were first classified morphologically into 14 conventional developmental stages. The total cell numbers (TCN) of embryos were measured using the air-dry method. The respective CD numbers of the embryos were then determined using logarithmic transformation of the TCN. The CD numbers of embryos were increased 0-10th with 11 stages. The 0th CD corresponded to 1-cell stage embryos; the 1st CD corresponded to 2-cell stage embryos; the 2nd CD corresponded to 3-4-cell stage embryos; the 3rd CD corresponded to 5-8-cell stage embryos; the 4th CD corresponded to 9-16-cell stage embryos, the 5th CD corresponded to morulae (17-32-cell stage embryos); and the 6th CD corresponded to the compact morulae. Furthermore, the 7th CD included early blastocysts to blastocysts. The 8th CD included expanded, collapsed and hatching blastocysts. The 9th CD included hatched blastocysts. The 10th CD included expanding-hatched blastocysts. The relationship between the CD number and the morphological characteristics of the bovine embryos 0-9 days after ovulation was expressed using a linear equation, and this revealed a high degree of correlation (y=0.98x-0.96, r=0.99). These results suggest that morphological changes of bovine embryos can be classified accurately using an 11-stage classification system based on the number of cleavages.

A mathematical model of the interaction between bovine blastocyst developmental stage and progesterone-stimulated uterine factors on differential embryonic development observed on Day 15 of gestation

“…Developmental stage is defined by log 10 cell number before blastocyst hatching and by embryo length (mm) after blastocyst hatching, with embryo size increasing exponentially from Day 1 to blastocyst hatching on Day 9 (Ushijima et al, 2009). The cell doubling time from Day 5 is therefore inversely proportional to h(t) n (if n = 0.33, then doubling progesterone decreases the doubling time by 20%).…”

“…The number of cells and length of a blastocyst at the time of hatching have been estimated (Berg et al, 2010;Spencer, 2013) and we assume that L H = log 10 (0.3) log 10 mm and A H = log 10 (512) log 10 cells. The size of each transferred blastocyst was not measured, and representative sizes were based on criteria derived from superovulated embryos, where the blastocyst stages were early (range 63-101 cells, mean 78.2 cells), normal (range 97-129 cells, mean 117.6 cells), expanding (range 142-193 cells, mean 159.2 cells), hatching (range 195-235 cells, mean 219.8 cells; Ushijima et al, 2009). Normal and expanding blastocysts were further classified as small or large and this was assumed to be based on the midpoint of the log 10 cell number range (range 97-112 cells, mean 104.2; range 113-129 cells, mean 120.7; range 142-170 cells, mean 155.4; range 171-193 cells; mean 181.7).…”

A mathematical model of in vivo bovine blastocyst developmental to gestational Day 15

“…Embryo developmental stage is defined by log 10 cell number before blastocyst hatching and embryo length (mm) after blastocyst hatching. Embryo size increases exponentially from Day 1 to blastocyst hatching on Day 9 (Ushijima et al, 2009) and the rate of change in embryo developmental stage (A; log 10 cell number) before blastocyst hatching is…”

A new simple and reliable vitrification device based on Hollow Fiber Vitrification (HFV) method evaluated using IVP bovine embryos

“…Total number of cells was counted as described by Ushijima et al (2009) with minor modifications. Embryos were placed into hypotonic solution for 5-10 min.…”