Purpose:

Shortage of transplantable human organs motivates research into alternatives, including xenotransplantation. When porcine lungs are perfused with human blood, a majority of human neutrophils and platelets are sequestered by the lung within fifteen minutes. Here we explore whether E-selectin mediates these phenomena.

Methods:

Porcine aortic endothelial cells (PAECs) expressing human anti-inflammatory and anticoagulant genes were cultured to confluency on microfluidic channels and activated with human TNF-α. Four highly selective E-selectin antagonists were provided by Glycomimetics, Inc (GMI) and added during the last hour of TNF-α incubation and during neutrophil (PMN) perfusion. PMNs were isolated from fresh human blood and perfused over the PAEC monolayers. PMN rolling events and distances were tracked using Fluxion software.

Results:

Whereas PMNs did not roll on non-activated PAECs, PMNs perfused over TNF-α-activated PAECs exhibited frequent tethering and rolling (mean 63±2 rolling events/hpf; mean 811±22 [mu]m traveled). GMI 1271 completely eliminated rolling at concentrations of 100-1000[mu]M with dose-dependent inhibition of rolling and rolling distance at lower concentrations. The other three compounds were even more effective, as shown. With treatment, rolling almost always terminated with release of the neutrophil rather than adhesion and transmigration. However, during whole blood perfusion, platelet adhesion was prevalent and leukocyte adhesion was observed despite high doses of GMI 1271.

Conclusions:

The GMI compounds tested are potent antagonists of the primary binding of human PMNs to activated PAECs, which appears to be mainly E-selectin-driven. During whole blood perfusion, however, adhesion of platelets and secondary leukocyte binding appears to be E-selectin-independent, suggesting that an approach which additionally targets platelet adhesion may be needed to fully prevent neutrophil and platelet sequestration by porcine lung xenografts perfused with human blood.

CITATION INFORMATION: Laird C, French B, Burdorf L, Cheng X, Fogler W, Magnani J, Phelps C, Ayares D, Azimzadeh A, Pierson R. Binding of Human Neutrophils to Activated Porcine Endothelium Is Mediated by E-Selectin. Am J Transplant. 2017;17 (suppl 3).

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