Exogenous pressure during in vitro follicle growth suppresses oocyte size expansion without impairing functional competence

Abstract Pressure stimuli, such as those arising from physical forces and hydrostatic pressure in vivo, are known to regulate cellular differentiation and growth. In the ovarian microenvironment, such cues are important regulators of folliculogenesis; however, their roles during in vitro follicle growth remain unclear. This study examined the effects of exogenous pressure on mouse preantral follicles cultured in a gas-pressurized system. Follicles were cultured under 0, 33, or 100 kPa for 8 days. Although exogenous pressure suppressed oocyte diameter expansion (0 kPa, 79.9 μm; 33 kPa, 74.7 μm; 100 kPa, 69.0 μm), the number of transzonal projection and the expression of Myo10 were comparable among groups, indicating that granulosa cell–oocyte communication was maintained. Following in vitro maturation and fertilization, oocytes from the 33 and 100 kPa groups showed comparable rates of maturation, fertilization, and blastocyst development to those of the 0 kPa group. Live offspring were successfully obtained in all groups. Further, the total RNA and expression levels of representative maternal transcripts in oocytes were comparable among groups. Additional analysis revealed that oocytes of similar diameters obtained after shorter culture periods under non-pressurized conditions showed significantly lower RNA content and developmental competence than those grown under exogenous pressure. These results indicate that oocyte diameter does not necessarily reflect cytoplasmic maturation and functional competence under exogenous pressurized conditions. Overall, our findings highlight the limitations of using oocyte diameter as an oocyte quality indicator and provide new insights into the role of pressure stimuli in folliculogenesis and in vitro follicle growth systems.