Kit Ligand enhances in vitro oocyte maturation and early embryonic development in sheep

Refinement of in vitro maturation (IVM) protocols is critical for enhancing sheep oocyte production efficiency and supporting genetic improvement in breeding programs. However, the developmental competence of in vitro-matured oocytes remains suboptimal partly because conventional culture systems fail to fully recapitulate the complex paracrine interactions within the ovarian follicular microenvironment. Kit Ligand (KL), a granulosa cell-derived growth factor, plays a pivotal role in mediating oocyte-cumulus cell communication during in vivo maturation, but its functional role and optimal supplementation conditions in sheep IVM systems remain unclear. In this study, we investigated the effects of recombinant KL supplementation during IVM on nuclear maturation, cytoplasmic maturation, intracellular homeostasis, and subsequent embryonic development in sheep oocytes. Supplementation with 30 ng/mL KL significantly increased the first polar body extrusion rate and enhanced cumulus expansion. KL treatment upregulated the expression of cumulus expansion-related genes (PTX3, HAS2, PTGS2, and TNFαIP6) and promoted cortical granule redistribution to the oocyte cortex. In addition, KL reduced intracellular reactive oxygen species levels, decreased the BAX/BCL-2 ratio, increased mitochondrial membrane potential, and improved mitochondrial distribution. Following in vitro fertilization, KL-treated oocytes exhibited higher cleavage and blastocyst formation rates compared with the control group. These findings suggest that KL supplementation improves oocyte developmental competence in sheep, potentially through regulation of cumulus cell activity and maintenance of intracellular homeostasis.