Impact of a controlled temperature gradient from 35.0°- 37.5°C on mouse embryo development and morphokinetics

Abstract

To examine the impact of different in vitro culture temperatures on mouse embryo development and morphokinetic timings. Prospective study An embryo culture incubator with 6 individual chambers and time-lapse imaging (ESCO MIRI TLTM) was set to vary the temperature between chambers by 0.5°C to give a gradient that ranged from 35.0-37.5°C. Gas concentration was consistent between chambers (6% CO2, 5% O2, 89% N2). Frozen-thawed 1-cell mouse embryos (B6C3F1xB6D2F1, EmbryotechTM) were thawed, pooled and randomly distributed between 6 culture dishes in Global culture medium + 10% SPS. One dish containing individually cultured embryos was placed into each culture chamber/temperature for simultaneous culture and assessment. Embryos were cultured over 96h, recording blastocyst formation as well as morphokinetic timings for time to 2-cell, 3-cell, 4-cell, compaction and blastocoel formation. Experiments were repeated 3-times. Data were analyzed using ANOVA and Fisher Multiple-Comparison Test and are presented as the mean ± SEM. Statistically significance was measured at p<0.05. Changes in temperature during mouse embryo culture significantly impacted blastocyst formation and morphokinetic timings. Blastocyst formation rates were highest at 37.0°C and decreased at other temperatures examined. Cell division timings became more rapid as temperature increased and significantly differed between temperature treatments for most endpoints examined.Tabled 1Mouse morphokinetic timings and blastocyst development at different culture temperaturesTemperatureCell Division Timings (hrs)% Blastocystt-2cellt-3cellt-4cellt-Compactiont-Blastocyst35.0° C8.7±0.633.6±1.0a34.9±1.1a61.2±1.6a80.5±5.1a31.2%a35.5° C9.1±0.5a34.1±0.8a35.4±0.8a61.4±0.8a81.5±1.1a52.7%b36.0° C8.6±0.431.7±0.9ab33.4±0.7a57.9±1.3b77.5±1.6a55.5%b36.5° C7.7±0.430.3±0.7bc31.1±0.6b53.1±0.8c69.3±1.2b52.7%b37.0° C7.3±0.428.6±0.4c30.5±0.7bc53.5±1.3c70.7±3.0bc65.5%c37.5° C6.8±0.4b28.0±0.c28.6±0.7c49.2±0.9d62.1±0.6c56.7%bcDifferent superscripts within a column represent statistically significant differences between tempertaures p<0.05. Open table in a new tab Different superscripts within a column represent statistically significant differences between tempertaures p<0.05. New incubator technology offers the opportunity to examine the impact of temperature on embryo development while controlling for other culture variables. Changes in incubation temperature as little as 0.5°C have a significant impact on mouse embryo development, affecting blastocyst formation and morphokinetic timings. This data has potential implications for application of morphokinetic selection algorithms between facilities, as temperature between labs must be tightly controlled.