Cytokine Treatment of Endothelial Cells Increases Glycoprotein Ibα-Dependent Adhesion to von Willebrand Factor

Abstract

AbstractEndothelial cells (EC) possess at least two membrane receptors for von Willebrand factor (vWF ), the vitronectin receptor (VNR, αvβ3 ), which recognizes an Arg-Gly-Asp (RGD) sequence in the C-terminus of vWF, and glycoprotein Ibα (GP Ibα), which interacts with a region in the N-terminal A1 domain of vWF. In the absence of added cytokines, EC attachment to a vWF substratum is mediated largely through the αvβ3 , with a smaller contribution by GP Ibα. In the present study, we have examined the effect of cytokines on the receptor specificity of EC attachment to wild-type vWF (WT-vWF ) and to vWF, which had been mutated in the C-terminal RGDS sequence (RADS-vWF ). Exposure of human umbilical vein EC (HUVEC) to tumor necrosis factor-α (TNF-α) or to TNF-α in combination with interferon-γ (IFN-γ), but not to interleukin-1β (IL-1), increased attachment to RADS-vWF by about twofold. The TNF-α–induced increase in EC attachment was accompanied by an increase in cell surface GP Ibα expression; GP Ibα surface expression was not increased by IL-1. Attachment of untreated HUVEC to WT-vWF could be inhibited 60% to 70% by a monoclonal antibody (MoAb) (LM609) to the VNR and 30% to 40% by the A1 fragment of vWF (containing the GP Ibα binding domain). The pattern of inhibition of attachment to WT-vWF was largely unchanged after TNF-α treatment of HUVEC. In contrast, the attachment to WT-vWF of HUVEC, treated with TNF-α +IFN-γ was completely inhibited by vWF-A1 and inhibited only 35% by the anti-VNR antibody LM609. Two MoAbs to GP Ibα produced similar, but incomplete, inhibition. Pretreatment of HUVEC with the combination of TNF-α +IFN-γ produced a dramatic decrease in VNR expression, confirming previous findings of Defilippi et al. These results suggest that in the presence of the inflammatory cytokines TNF-α +IFN-γ, the endothelial GP Ib complex is a major determinant of HUVEC adhesion to surface-bound vWF.