Ganglioside GM3 is known as an inhibition factor of cell differentiation and proliferation via inhibition of epidermal growth factor receptor (EGFR) phosphorylation. Our previous study showed that the exogenous ganglioside GM3 reduced the meiotic maturation of porcine oocytes and induced apoptosis at 44 h of in vitro maturation (IVM). However, the role of ganglioside GM3 in the relationship between EGFR signaling and apoptosis during porcine oocyte maturation has not yet been studied. First, porcine cumulus-oocyte complexes (COCs) were cultured in the NCSU-23 medium with exogenous ganglioside GM3 according to maturation periods (non-treated, only IVM I: 0 - 22 h, only IVM II: 22 - 44 h and IVM I & II: 0 - 44 h). We confirmed that the proportion of germinal vesicle breakdown (GVBD) increased significantly in the IVM I treated group than in the control group. We also confirmed that the meiotic maturation until M II stage and polar body formation decreased significantly in the only IVM I treated group. Cumulus cell expansion and mRNA levels of the expansion-related factors (HAS2, TNFAIP6 and PTX3) decreased significantly in the IVM I treated group than in the control group. Protein levels of EGFR, p-EGFR, ERK1/2, and p-ERK1/2 decreased significantly in the GM3-treated groups, during the IVM I period. In addition, cellular apoptosis, determined using TUNEL assay, and protein levels of Cleaved caspase 3, were increased significantly in the GM3-treated COCs during the IVM I period. Based on these results, ganglioside GM3 exposure of porcine COCs during the IVM I period reduced meiotic maturation and cumulus cell expansion via inhibition of EGFR activity in pigs.

Ganglioside GT1b, glycosphigolipids with three sialic acid, is known to play an important role in signal transduction such as epidermal growth factor receptor (EGFR). EGF is also known to induce resumption of meiosis and cumulus cells expansion during porcine oocyte maturation. Therefore, this study was conducted to evaluate the effects of ganglioside GT1b on resumption of meiosis and cumulus cells expansion in porcine oocyte maturation. First, porcine cumulus-oocyte complexes were cultured in NCSU-23 medium supplemented with GT1b (0, 1, 2 and 4 μM) at 44 h. We observed that the proportion of the metaphase II (M II) stage was significantly increased in the 2 μM GT1b (78.0 ± 2.3) treated group than in the other groups. Furthermore, expression of cumulus cells expansion factor genes (Has2, TNFAIP6, Ptx3) were significantly increased in the 2 μM GT1b treated group than in the other groups. Next, we investigated the meiotic maturation and the expressions of cumulus cells expansion factor genes after GT1b and/or EGF treatment. The proportion of the M II stage was significantly higher in the GT1b+EGF (90.1 ± 2.3) treated group than in the other groups. Moreover, expressions of cumulus cells expansion factor genes were significantly increased in the GT1b+EGF treated group than in the control group. After in vitro fertilization, fertilization rate, preimplantation development competence and quality of blastocyst were improved in oocytes derived from GT1b+EGF treated group. Taken together, these results suggest that exogenous ganglioside GT1b improving the developmental competence of porcine embryos via increase of resumption of meiosis and cumulus cells expansion during in vitro maturation of porcine oocytes.