Cytokine treatment of endothelial cells increases glycoprotein Ibα- dependent adhesion to von Willebrand factor

Abstract

Endothelial cells (EC) possess at least two membrane receptors for von Willebrand factor (vWF), the vitronectin receptor (VNR, α(v)β3), 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 α(v)β(3), 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 lb complex is a major determinant of HUVEC adhesion to surface-bound vWF.