Glycoconjugate Journal (1996) 13:947-953 Simultaneous expression by porcine aorta endothelial cells of glycosphingolipidsbearing the major epitope for human xenoreactive antibodies (Galal-3Gal), blood group H determinant and N-glycolylneuraminic acid DANIELE BOUHOURS* CHRISTINE POURCEL and JEAN- FRANCOIS BOUHOURS [nstitut de Transplantation et de Recherche en Transplantation, 1NSERM • 437, Centre Hospitalier R~gional, F-44035 Nantes Cedex, France Received 16 November 1995, revised 6 February 1996 Glycosphingolipids were isolated from primary cultures of porcine endothelial cells labelled with 14C-galactose or ~4C-glucosamine. They were characterized by their mobility on thin layer chromatogram, their sensitivity to exoglycosidases, and their labelling with antibodies. In addition to the major gtycosphingolipids, globotetra- and globotriaosylceramide, minor ones were identified as penta- and heptaglycosylceramide of the neolactoseries terminated by either Gala l-3Gal- (xenoreactive epitope) or Fucal-2Gal- (H determinant). Two gangliosides were found, GM3 and GD3, and N-glycolylneuraminic acid was their major sialic acid. Therefore, porcine endothelial cells differ from human endothelial cells by expression of glycosphingolipids that are absent in man: two Gatal- 3Gal-terminated glycolipids recognized by human natural antibodies, and two N-glycotylneuraminic acid-terminated gangliosides which are potent immunogens. Keywords: porcine endothelial cells, glycosphingolipids, xenoantigens Abbreviations: HPTLC, high performance thin-layer chromatography; GSL, glycosphingolipid; NeuAc, N- acetylneuraminic acid; NeuGc, N-glycolylneuraminic acid; PAEC, porcine aorta endothelial celI. Introduction Xenotransplantation of porcine organs to man is con- sidered as a possible way to alleviate the present shortage of human organs, at least temporarily while waiting for a compatible human graft. A major obstacle to xenotrans- plantation is the occurrence of hyperacute rejection which is the consequence of the binding of naturally occurring antibodies of host plasma to antigens of the endothelimn of the graft and activation of the complement cascade leading to alteration of the endothelium integrity. There- fere, it is important to identify antigens of endothelial cells which can be recognized by human xenoreactive natural antibodies. It is also relevant to characterize immunogens which could trigger the host immune system *To whom correspondenceshould be addressed. 0282-0080 7) 1996 Chapman & Hall when hyperacute rejection is brought under control. Strong experimental evidences have suggested that the major target of human xenoreactive natural antibodies is the disaccharide determinant Galal-3Gal- [1-3] which is expressed in porcine endothelial cells [4]. It is a human blood group type determinant corresponding to an unfucosylated blood group B (afucoB). The current concept is that porcine endothelium expresses a3-galactosyltransferase and is devoid of a2- fucosyltransferase activity [5], whereas the reverse situation is observed in human endothelium which thus expresses blood group H, A or B determinant, depending on the individual ABO blood group [6]. Induction into transgenic pig of the expression of a2-fucosyltransferase competing for the same substrate as a3-galactosyltrans- ferase has appeared as a means for decreasing the expression of xenoreactive antigens and humanizing