Fig. 4: Microgravity exposure for the first 24 h of embryo development, reduces blastocyst formation and cell numbers in mice.

A Female mice were super ovulated and cumulus oocyte complexes recovered from the oviduct and placed at the end of the micro-channel slide. Mouse sperm was collected from the cauda epididymis and vas deferens and placed at the other end of the micro-channel slide. Sperm were required to traverse the micro-channel slide at either normal gravity or simulated microgravity to fertilize oocytes for 24 h. Following incubation all groups were cultured in standard clinical in vitro embryo culture conditions for a further 72 h for assessments of pre-implantation embryo quality. B Proportion of 2-cells, 24 h post insemination, C proportion of expanded blastocysts, hatching blastocysts and fully hatched blastocyst from 2-cells, 96 h post insemination, D representative images of blastocyst formation at 96 h post insemination following a 24 h insemination period at simulated microgravity or normal gravity control, E total blastocyst cell number (TCN), 96 h post insemination, F total blastocyst inner cell mass (ICM) cell number (Oct4 positive), 96 h post insemination, G total blastocyst epiblast cell number (Nanog positive), 96 h post insemination, H proportion of blastocyst ICM relative to TCN, I proportion of blastocyst epiblast cells relative to TCN, J proportion of blastocyst epiblast cells relative to ICM and K representative images of blastocyst cell numbers generated from 24 h insemination window at simulated microgravity and normal gravity control. Data are presented as mean ± SEM, with individual culture replicates indicated by unique symbols or individual embryos (circles). N = 187 embryos assessed for fertilisation and blastocyst development and N = 35 for cell numbers from 6 replicates for normal gravity control and N = 239 embryos assessed for fertilisation and blastocyst development and N = 23 for cell numbers from 6 replicates for simulated microgravity. Normal gravity controls denoted in white, and simulated microgravity denoted in dark blue. Statistical significance characterized as * P < 0.05, and ** P < 0.01. P-values between 0.05 and 0.1 are presented numerically. Parts of this image were created in BioRender. Mcpherson, N. (2026) https://BioRender.com/ix6duwb.