Expression analysis of an α-1, 3-galactosyltransferase, an enzyme that creates xenotransplantation-related α-Gal epitope, in pig preimplantation embryos

a-1,3-Galactosyltransferase (a-GalT), an enzyme creating Gala1-3Gal (a-Gal) epitope on the cell surface in some mammalian species such as pigs, is known to be a key factor that causes hyperacute rejection upon transplantation from pigs to humans. To establish the RNA interference-based suppression of endogenous a-GalT messenger RNA (mRNA) synthesis in porcine preimplantation embryos, we determined the suitable embryonic stage at which stage such approach is possible by using the semi-quantitative RT-PCR (qRT-PCR) and the cytochemical method using a fluorescence-labeled Bandeiraea simplicifolia Isolectin B4 (BS-I-B4). Staining with BS-I-B4 demonstrated that a-Gal epitope expression was first recognized at the 8-cell stage, and increased up to the hatched blastocyst stage. Single embryo-based qRT-PCR also confirmed this pattern. These results indicate that creation of a-Gal epitope is proceeded by de novo synthesis of a-GalT mRNA in porcine preimplantation embryos with peaking at the blastocyst stage.