P- and E-selectin receptor antagonism prevents human leukocyte adhesion to activated porcine endothelial monolayers and attenuates porcine endothelial damage.

ABSTRACT

BACKGROUND: Alongside the need to develop more effective and less toxic immunosuppression, the shortage of human organs available for organ transplantation is one of the major hurdles facing the field. Research into xenotransplantation, as an alternative source of organs, has unveiled formidable challenges. Porcine lungs perfused with human blood rapidly sequester the majority of circulating neutrophils and platelets, which leads to inflammation and organ failure within hours, and is not significantly attenuated by genetic modifications to the pig targeted to diminish antibody binding and complement and coagulation cascade activation. METHODS: Here, we model the interaction of freshly isolated human leukocytes with xenotransplanted vasculature under physiologic flow conditions using microfluidic channels coated with porcine endothelial cells. Both isolated human neutrophils and whole human blood were perfused over transgenic pig aortic endothelial cells that had been activated with rhTNF-α or rhIL-4 using the BioFlux system. Novel compounds GMI-1271 and rPSGL1.Fc were tested as E- and P- selectin antagonists, respectively. Cellular adhesion and rolling events were tracked using FIJI (imageJ). RESULTS: Porcine endothelium activated with either rhTNF-α or rhIL-4 expressed high amounts of selectins, to which isolated human neutrophils readily rolled and tethered. Both E-and P-selectin antagonism significantly reduced the number of neutrophils rolling and rolling distance in a dose-dependent manner, with near total inhibition at higher doses (P < .001). Similarly, with whole human blood, selectin blocking compounds exhibited dose-dependent inhibition of prevalent leukocyte adhesion and severe endothelial injury (Untreated 394 ± 97 PMNs/hpf, 57 ± 6% loss EC; GMI1271+rPSGL1.Fc 23 ± 9 PMNs/hpf, 8 ± 6% loss EC P < .01). CONCLUSIONS: Selectin blockade may be useful as part of an integrated strategy to prevent neutrophil-mediated organ xenograft injury, especially during the early time points following reperfusion.