P218

Differences in human and porcine platelet oligosaccharides and differences in human and porcine ASGR1 influence phagocytosis by liver sinusoidal cells in vitro

Leela Paris, Ray Chihara, Emad Hakmi, Richard Sidner, Chris Burlak, A. Joseph Tector

Transplant Surgery, Indiana University Medical School, Indianapolis, IN, United States

Studies using pig-to-human ex vivo and pig-to-baboon in vivo liver transplant models revealed that acute thrombocytopenia is a limiting factor preventing successful porcine liver xenotransplantation in humans.  Recently, it was found that Asialoglycoprotein receptor 1 (ASGR1) and the Macrophage antigen complex-1 (Mac-1) on porcine liver sinusoidal cells initiate phagocytosis of human platelets. Both receptors have carbohydrate-binding motifs that recognize galactose and N-acetyl glucosamine in their bconformations. Analysis of surface carbohydrates on human and porcine platelets and receptor carbohydrate binding domains will help elucidate the mechanism of human platelet phagocytosis by porcine liver sinusoidal cells. A comparison of human and porcine ASGR1 binding domains revealed three amino acid differences. Flow cytometric analysis demonstrated that human platelets have significantly more Galb1-4GlcNac and bGlcNAc than domestic or a1,3-galactosyltransferase knockout (GTKO)/human Decay Accelerating Factor knock-in (hDAF) porcine platelets. Immunoblot analysis revealed that the carbohydrate motifs are present on multiple proteins including CD42. Treatment of human, domestic and GTKO/hDAF porcine platelets with sialidase increased the available Galb1-4GlcNac and bGlcNAc on each, but did not diminish the differences between human and porcine platelets. Desialylation of human platelets resulted in an increased uptake of human platelets by porcine liver sinusoidal endothelial cells and Kupffer cells. Slight variations between human and porcine receptor carbohydrate binding domains and differences in carbohydrate expression on porcine and human platelet surfaces appear to contribute to the phagocytosis of human platelets by porcine liver sinusoidal cells.