Sung-Kyu Chae

Ganglioside GM3 Induces Cumulus Cell Apoptosis through Inhibition of Epidermal Growth Factor Receptor-mediated PI3K/AKT Signaling Pathways during in vitro Maturation of Pig Oocytes†

Gangliosides are components of the mammalian plasma membrane that help regulate receptor signaling. Ganglioside GM3, for example, plays an important role in initiating apoptosis in cancer cells; however, physiological roles for GM3 in normal processes, such as during pig oocyte maturation, are not clear. The aim of this study was to investigate the functional link between GM3 and cellular apoptosis in porcine cumulus‐oocyte‐complexes (COCs) during in vitro maturation. Our results indicated that denuded oocytes possess less ST3GAL5, a GM3‐synthesizing enzyme, than cumulus cells or COCs after 44 hr of in vitro maturation. GM3 also affected the meiotic maturation of cultured pig oocytes, as evaluated by orcein staining. In vitro treatment of COCs with exogenous GM3 also reduced cumulus cell expansion, the proportion of meiotic maturation, and increased cumulus cell transcription of PTX3, TNFAIP6, and HAS2. Interestingly, GM3 treatment reduced the expression of Epidermal growth factor receptor (EGFR)‐mediated Phosphoinositide 3‐kinase/AKT signaling proteins in COCs in a concentration‐dependent manner, instead increasing the abundance of pro‐apoptotic factors such as AIF, activated Caspase 9, cleaved PARP1, and Caspase 3 were. Thus, GM3 might affect porcine oocyte maturation via suppression of EGFR‐mediated PI3K/AKT signaling and/or induction of apoptosis during in vitro maturation.

Ganglioside GD1a promotes oocyte maturation, furthers preimplantation development, and increases blastocyst quality in pigs

Gangliosides are key lipid molecules required for the regulation of cellular processes such as proliferation, differentiation, and cell signaling, including signaling of epidermal growth factor receptor (EGFR). Epidermal growth factor (EGF) has long been considered a potential regulator of meiotic and cytoplasmic maturation in mammalian oocytes. However, there is no report on the direct effect of ganglioside GD1a in porcine oocyte maturation. In this study, we first investigated a functional link between GD1a and meiotic maturation during in vitro maturation (IVM) of porcine embryos. Moreover, we confirmed the effect of exogenous GD1a treatment on blastocyst development, quality, and fertilization rate in early embryonic development. First, we observed that the protein level of ST3GAL2, a GD1a synthesizing enzyme, significantly increased (P < 0.01) in cumulus-oocyte-complexes (COCs) during IVM progress. The proportion of arrested germinal vesicles (GV) increased in oocytes treated with EGF+GD1a (41.6 ± 1.5%) at the IVM I stage. Upon completion of meiotic maturation, the proportion of metaphase II (M II) was significantly higher (P < 0.05) in the EGF+GD1a (89.9 ± 3.6%) treated group. After IVF, the percentage of penetrated oocytes was significantly higher (P < 0.05) in the EGF+GD1a (89.1 ± 2.3%) treated group than in the control group. Furthermore, exogenous GD1a treatment improved the developmental competence and quality of blastocysts during preimplantation embryo development stage. These results suggest that ganglioside GD1a may play an important role in IVM mechanisms of porcine maturation capacity. Furthermore, our findings will be helpful for better promoting the embryo development and blastocyst quality in pigs.

Changes of Ganglioside GM3 Expression in Porcine Oocyte Maturation and Early Embryonic Development In Vitro

Gangliosides exist in glycosphingolipid-enriched domains on the cell membrane and regulate various functions such as adhesion, differentiation, and receptor signaling. Ganglioside GM3 by ST3GAL5 enzyme provides an essential function in the biosynthesis of more complex ganglio-series gangliosides. However, the role of gangliosides GM3 in porcine oocytes during in vitro maturation and early embryo development stage has not yet understood clear. Therefore, we examined ganglioside GM3 expression patterns under apoptosis stress during maturation and preimplantation development of porcine oocytes and embryos. First, porcine oocytes cultured in the NCSU-23 medium for 44 h after H2O2 treated groups (0.01, 0.1, 1 mM). After completion of meiotic maturation, the proportion MII (44 h) was significantly different among control and the H2O2 treated groups (76.8±0.3 vs 69.1±0.4; 0.01 mM, 55.7±1.0; 0.1 mM, 38.2±1.6%; 1 mM, P<0.05). The expressions of ST3GAL5 in H2O2 treated groups were gradually decreased compared with control group. Next, changes of ST3GAL5 expression patterns were detected by using immunofluorescene (IF) staining during preimplantation development until blastocyst. As a result, we confirmed that the expressions of ST3GAL5 in cleaving embryos were gradually decreased (P<0.05) according to the early embryo development progress. Based on these results, we suggest that the ganglioside GM3 was used to the marker as pro-apoptotic factor in porcine oocyte of maturation and early embryo production in vitro, respectively. Furthermore, our findings will be helpful for better understanding the basic mechanism of gangliosides GM3 regulating in oocyte maturation and early embryonic development of porcine in vitro.