Francesca E Duncan

STUDY QUESTION Is there an association between morphokinetic variables of meiotic maturation and the severity of aneuploidy following in vitro maturation (IVM) in the mouse? SUMMARY ANSWER The severity of meiotic aneuploidy correlates with an extended time to first polar body extrusion (tPB1) and duration of meiosis I (dMI). WHAT IS KNOWN ALREADY Morphokinetic variables measured using time-lapse technology allow for the non-invasive evaluation of preimplantation embryo development within clinical assisted reproductive technology (ART). We recently applied this technology to monitor meiotic progression during IVM of mouse gametes. Whether there is a relationship between morphokinetic variables of meiotic progression and aneuploidy in the resulting egg has not been systematically examined at the resolution of specific chromosomes. Next-generation sequencing (NGS) is a robust clinical tool for determining aneuploidy status and has been reverse-translated in mouse blastocysts and oocytes. Therefore, we harnessed the technologies of time-lapse imaging and NGS to determine the relationship between the morphokinetics of meiotic progression and egg aneuploidy. STUDY DESIGN, SIZE, DURATION Cumulus–oocyte complexes (COCs) were collected from large antral follicles from hyperstimulated CD-1 mice. Cumulus cells were removed, and spontaneous IVM was performed in the absence or presence of two doses of Nocodazole (25 nM or 50 nM) to induce a spectrum of spindle abnormalities and chromosome segregation errors during oocyte meiosis. Comprehensive chromosome screening was then performed in the resulting eggs, and morphokinetic variables and ploidy status were compared across experimental groups (control, n = 11; 25 nM Nocodazole, n = 13; 50 nM Nocodazole, n = 23). PARTICIPANTS/MATERIALS, SETTING, METHODS We monitored IVM in mouse oocytes using time-lapse microscopy for 16 hours, and time to germinal vesicle breakdown (tGVBD), time to first polar body extrusion (tPB1), and duration of meiosis I (dMI) were analyzed. Following IVM, comprehensive chromosome screening was performed on the eggs and their matched first polar bodies via adaptation of an NGS-based preimplantation genetic testing for aneuploidy (PGT-A) assay. Bioinformatics analysis was performed to align reads to the mouse genome and determine copy number-based predictions of aneuploidy. The concordance of each polar body–egg pair (reciprocal errors) was used to validate the results. Ploidy status was categorized as euploid, 1–3 chromosomal segregation errors, or ≥ 4 chromosomal segregation errors. Additionally, aneuploidy due to premature separation of sister chromatids versus non-disjunction was distinguished. MAIN RESULTS AND THE ROLE OF CHANCE We applied and validated state-of-the-art NGS technology to screen aneuploidy in individual mouse eggs and matched polar bodies at the chromosome-specific level. By performing IVM in the presence of different doses of Nocodazole, we induced a range of aneuploidy. No aneuploidy was observed in the absence of Nocodazole (0/11), whereas IVM in the presence of 25 nM and 50 nM Nocodazole resulted in an aneuploidy incidence of 7.69% (1/13) and 82.60% (19/23), respectively. Of the