Lipid-rich blastomeres in the two-cell stage of porcine parthenotes show bias toward contributing to the embryonic part

This study was designed to determine the fate of the blastomeres in two-cell porcine parthenotes that display uneven size (larger vs. smaller) or cytoplasmic brightness (darker vs. brighter) during development to the blastocyst stage. For the non-invasive tracing of cell lineage, lipophilic fluorescence dye DiI (red) and DiD (blue) was randomly microinjected into each of two different blastomeres in each embryo. In blastocysts derived from the two-cell parthenotes with unevenly-sized blastomeres, no biased contribution was found in the progeny of either blastomere. However, in the blastocysts derived from the two-cell parthenote having different cytoplasmic brightnesses, the progeny of darker (more lipid-rich cytoplasm) blastomeres were more than two-fold more likely to form the embryonic part (43.6%; 17/39) than they were to form the abembryonic part (17.9%; 7/39), while the contribution of brighter blastomeres (less lipid) was just the opposite. The expressions of four marker genes involved in lineage allocation (Cdx2, Tead4, Oct4 and Carm1) were also analyzed in darker and brighter blastomeres of two-cell parthenotes using quantitative RT-PCR. The expression of Carm1 that encodes arginine methyltransferase 1 and that promotes inner cell mass (ICM) differentiation was significantly higher (P < 0.05) in darker blastomeres. The ICM marker Oct4 also tended to be more highly expressed in the darker blastomeres, while Cdx2 and the TE marker Tead4 showed comparably higher expressions in the brighter blastomeres. However, in all cases, the marginal differences in the expression levels of Oct4, Cdx2 and Tead4 were not statistically significant (P > 0.05). Our findings indicate that expression of genes related to early differentiation, especially Carm1, are partially associated with lipid droplet distribution in the two-cell porcine parthenote and may lead to biased embryonal axis formation.