Sialic acid and Galectins mediate Xenogenic Neutrophil-Endothelial Adhesion

Abstract

Abstract Sialic acids (Sias) play multifarious roles in immunity, and likely contribute to inflammation seen in xenograft failure. The ‘net’ cellular sialyation state depends on many things including transient loss of Sia moieties with exposure of subterminal galactosyl structures (mediated by sialyltransferases and sialidases [neuraminidases]), which can function as receptors for extracellular lectins, such as galectins. Here, we report the role of Sias in modulating adhesion of human neutrophils (NΦ’s) to pig endothelial cells. Human (hAECs) and pig (pAECs) aortic endothelial cells were pretreated with Clostridium perfringens neuraminidase (NA, to remove Sias), human tumor necrosis factor (hTNF, 25 ug/ml), or both. CalceinAM-labelled human NΦ’s were treated with rhIL-8, NA, or N-acetyllactosamine (LacNAc, a galectin inhibitor). NΦ adhesion was measured and expressed as percent adhesion. NA treatment of AECs increased NΦ adhesion to pig (36.3% vs 22.4%) more than to human (24.2% vs 20.7%) AECs. NΦ adhesion was similarly increased by TNF activation of AECs in static and flow conditions. hIL-8 dramatically increased NΦ adhesion to pAECs (76.5%, p<.001) and hAECs (45.3%, p<.001), with the highest adhesion observed with NA-treated pAECs and hIL-8-activated NΦ’s. Pre-exposure of the NΦ’s to LacNAc prevented their galectin-mediated adhesion to galactose moieties exposed on the NA-treated pAECs (13.7%, p<.0001). In summary, AEC de-sialylation promotes xenogenic NΦ adhesion, an effect completely inhibited by pre-incubation of Nf’s with LacNAc. These results suggest the inhibition of sialidase activity or galectin binding as promising therapeutic approaches to limit inflammation in xenogeneic organ injury.