P-161 Mechanical phenotyping of mammalian oocytes using Atomic Force Microscopy (AFM) to evaluate their developmental potential

Abstract

Abstract Study question Can we measure the mechanical properties of mammalian oocytes with AFM and use them as a biomarker to identify oocytes with the best development potential? Summary answer We characterized mammalian oocytes’ mechanical properties using AFM, correlated them with oocyte cortex organization, and explored these parameters in contexts critical for oocyte quality. What is known already Oocyte production is crucial for reproduction but prone to errors, yielding a significant number of poor-quality oocytes detrimental to fertility and offspring development. Indeed, oocyte quality determines embryo health post-fertilization. Current selection methods for oocytes rely on subjective morphological analyses. Remarkably, human and mouse oocyte quality correlates with mechanical properties, particularly cortical tension (CT). CT defects in oocytes are rather frequent and impair early embryonic development after fertilization. However, they do not induce any visible oocyte morphological alteration. Mechanical properties could, therefore, be used as a biomarker of oocyte quality. Study design, size, duration We designed an AFM protocol for the measurement and analysis of mammalian oocytes’ mechanical properties, including a new elasto-capillary model describing oocyte mechanics. Concomitantly, we analyzed oocyte cortex organization using spinning disk microscopy in live. We measured mouse oocytes at various stages of meiotic maturation (prophaseI, meiosisI, meiosisII), mutant mouse oocytes with known cortical tension defects, oocytes coming from young and aged mice, and human oocytes at various stages of meiotic maturation (prophaseI, meiosisI, meiosisII). Participants/materials, setting, methods Mouse oocytes were collected from 11-week-old and 44-56-week-old OF1 female mice. Human oocytes were collected from patients who underwent ovarian stimulation for Assisted Reproduction Technologies (ART) as part of their protocol. Only oocytes that were in prophaseI or meiosisI after ovarian punction and thus not suitable for the ART procedure were used in this study. All patients gave informed consent (agreement CNIL number 22_5725). Main results and the role of chance Our AFM approach allows measuring various mechanical parameters, including oocyte cortical tension, elasticity, and capillary indentation rate, quantifying whether the oocyte behaves predominantly as a droplet with surface tension or as an elastic material. We find that these parameters correlate to specific oocyte cortex organization, showing that a thin actin cortex enriched in myosin II is associated with high tension, high elastic modulus, and low capillary indentation rate. In contrast, a thick actin cortex with few myosin II is associated with low cortical tension, low elasticity, and high capillary indentation rate. We go further and show that these parameters can predict cortex organization. Indeed, oocytes from aged mice display altered mechanical parameters, reflecting specific changes in their cortex organization. Finally, we transferred our approach to human oocytes. We show that the mechanical parameters of mouse and human oocytes are in the same range. Still, in humans, they evolve differently from those of mice during meiotic maturation, again reflecting specific changes in the organization of their cortex. Thus, mechanical parameters correlate with oocytes quality, and can predict cortex organization in mouse and human. Limitations, reasons for caution AFM is hardly usable for clinical application, we are thus working in parallel on developing non-invasive devices to perform high-throughput measurements of oocyte mechanical properties in a non-invasive manner, to serve as a mechanical biomarker to identify oocytes with the best developmental potential for ART. Wider implications of the findings Ultimately, this project could improve ART procedures by increasing their success rate while decreasing treatment cycles and risks for the patient and newborn, and optimize protocols for oocyte freezing for fertility preservation. Trial registration number not applicable