cfDNA is detectable on days 3 and 5, but more accurate on day 5. Although our data suggest moderate concordance rates, PGT-A with the use of cfDNA must be further optimized before clinical implementation.
A lower MC at the blastocyst stage was associated with euploid status and blastocyst formation, indicating better embryo quality compared to those with a higher MC. Higher MC in aneuploid and arrested embryos may be explained by slower cell division or degradation of genomic DNA over time. Blastulation timing may be helpful for selection of higher quality embryos. Combining blastulation timing and MC along with morphologic grading and euploid status may offer a new direction in embryo selection.
BACKGROUND: Debate exists about the ideal temperature for embryo culture. The temperature of the female reproductive tract may be different than the commonly used human core body temperature of 37 degrees Celsius ( C).OBJECTIVE: This study aimed to evaluate the effects of various embryo culture temperatures on embryo development.MATERIALS AND METHODS: One-cell stage mouse embryos (n¼ 615; B6C3F1xB6D2F1 strain) were thawed and randomly divided into 4 culture temperature treatment groups (35.5 C (n¼161), 36.5 C (n¼166), 37.5 C (n¼134), and 38.5 C (n¼154)). Embryos were cultured to blastocyst in Continuous Single Culture Complete (CSCC, Irvine Scientific) in an incubator (Heracell 160i) with 5%O2, 8%CO2 and pH of 7.3. Embryo development was scored daily for 5 days. Forty embryos from each treatment group that reached blastocyst were fixed in 2% formalin and labeled for cell counts with Hoechst dye and CDX2 antibody to differentiate trophectoderm (TE) from inner cell mass (ICM). Cells with nuclei that were contracted and dense or fragmented were classified as pyknotic (dead) cells and used as a marker of embryo quality. Chi square and ANOVA (with post-hoc analysis) were used for comparison between the groups with a P-value <0.05 considered significant.RESULTS: 73% (449/615) of all embryos cultured reached blastocyst. Embryos cultured at 36.5 C had the largest proportion developed to blastocyst compared to the other culture temperatures, (88%, p<0.001). Forty blastocysts from each temperature group were stained for cellular growth. The ICM cell counts were significantly reduced at 35.5 C compared to all other temperatures, p< 0.001. The TE and total cell count were significantly reduced at 35.5 C (p<0.001), and significantly increased at 38.5 C (p<0.01), compared to all other temperature groups. There were no significant differences of TE and total cell counts between the 36.5 C and 37.5 C groups. Staining for pyknotic cells, revealed the highest proportion of pyknotic cells 14.7% (p<0.001) were seen at culture temperature of 35.5 C.CONCLUSIONS: Culture of mouse embryos at temperatures of 36.5 C confers the greatest proportion of blastocyst formation compared to other temperatures studied. Total number of cells was highest at 38.5 C, despite lower blastulation rates. Embryos cultured at temperatures of 35.5 C lead to overall poor embryo growth outcomes (reduced blastocyst development, lower cell counts, and increased rate of apoptotic cells). More studies are needed to assess the effects of various temperatures on the growth of human embryos.BACKGROUND: Although PGT-A with Next Generation Sequencing (NGS) is becoming the standard of care, there are a large number of blastocysts that were frozen prior to having embryo biopsy and genetic analysis. As a result, there are an increasing number of patients requesting genetic information of their previously undiagnosed frozen blastocysts. It is important to understand how these embryos perform following warming (thaw), biopsy and revitrification (refreeze) for PGT-A analysis...
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