Intrafollicular immature oocyte transfer (IFIOT) for in vivo maturation of vitrified bovine oocytes

Abstract

Oocyte cryopreservation is fundamental for the advancement of reproductive biotechnologies, but its efficiency in cattle is limited. This study evaluated whether in vivo maturation through the intrafollicular immature oocyte transfer (IFIOT) can mitigate damage from vitrification. In a 2 × 2 design, fresh and vitrified oocytes were subjected to in vitro or in vivo maturation. Vitrified oocytes had lower metaphase II (MII) rates (P < 0.05) compared to fresh. However, in vivo maturation of vitrified oocytes increased MII rates (69.4 % vs. 45.7 %; P < 0.05) and reduced chromatin abnormalities (18.6 % vs. 33.7 %) compared to in vitro maturation. Electron microscopy revealed a higher incidence of vacuoles, damage, and disorganization of organelles in vitrified oocytes. The concentration of reactive oxygen species (ROS) was lower (P < 0.05) in vitrified and fresh oocytes matured in vivo compared to those in vitro matured. Vitrification impaired embryonic development, reducing cleavage rates on day 2 and blastocyst rates on days 6 and 7. Embryos from fresh oocytes matured in vivo exhibited larger diameters, higher cell numbers, and lower proportions of apoptotic cells. Iron-reducing antioxidant activity did not differ among groups in the in vitro maturation medium and follicular fluid. In conclusion, IFIOT in vitrified oocytes improved MII rates and reduced chromatin abnormalities but did not alter intraoocyte ROS levels nor improve embryonic development. In fresh oocytes, IFIOT was effective, reducing ROS and producing blastocysts with faster development and better quality. These findings suggest that IFIOT can partially mitigate nuclear damage caused by vitrification, but additional studies are needed to explore cytoplasmic protection strategies.