Xenoreactive natural antibodies (XNAs) and complement mediate hyperacute rejection of discordant xenografts. Inhibition of complement alone results in some prolongation of graft survival, but delayed xenograft rejection still precludes long-term graft survival. In vitro data provide evidence for the direct proinflammatory activation of endothelial cells (ECs) by XNAs. These antibodies are primarily directed against galactoseα(1-3)-galactose (α-gal), the major xenoantigen in the pig to primate xenotransplant model. Previous studies have shown EC activation by XNAs but failed to address the question of whether α-gal-specific ligands can induce EC activation.Background.
The aim of this study was to investigate whether agonist binding to the α-gal epitope by α-gal-specific lectins as compared with XNAs or elicited xenoreactive antibodies can directly elicit type II porcine aortic EC (PAEC) activation (i.e., activation that requires protein synthesis).Methods and Results.
The tetravalent, α-gal-bindingBandeiraea simplicifolia lectin I (BS-I), the wholly α-gal-specific BS-I isolectin B4, and elicited primate anti-pig xenoreactive antibodies(decomplemented cynomolgus monkey anti-porcine serum) induced E-selectin protein expression in PAECs. This induction was α-gal-specific, as preincubation with synthetic α-gal carbohydrate or adsorption of lectin or serum to rabbit, but not human, red blood cells removed the activating component. E-selectin expression, induced by BS-I, was inhibited in the presence of genistein, a tyrosine kinase inhibitor, and by mepacrine, an inhibitor of phospholipase A2. Human and primate XNAs lacked this activity when tested at relevant concentrations; however, stimulation of PAECs with affinity-purified human XNA (IgM and IgG) resulted in slightly increased interleukin-8 and P-selectin mRNA levels but had no apparent effects on E-selectin transcription. BS-I strongly induced E-selectin, P-selectin, intercellular adhesion molecule-1, and interleukin-8 mRNA in an NF-κB-dependent manner.Conclusions.
Several agonists that specifically bind to α-gal can evoke type II EC activation. Hence, anti-Gal antibodies may contribute directly to xenograft rejection in the absence of complement activation.