Jing-Fei Dong

Platelets adhered to endothelial cell-bound ultra-large von Willebrand factor strings support leukocyte tethering and rolling under high shear stress

Summary.u2002 Leukocyte rolling on vascular endothelium is mediated by an interaction between P‐selectin expressed on endothelial cells and P‐selectin glycoprotein ligand‐1 on leukocytes. This interaction reduces the velocity of leukocyte movements to allow subsequent firm adhesion and transmigration. However, the interaction has so far been observed only under low venous shear stress and cannot explain the accumulation of monocytes in atherosclerotic plaques found in arteries, where shear stress is much higher. We have previously shown that newly released ultra‐large von Willebrand factor (ULVWF) forms extremely long string‐like structures to which platelets tether. Here, we investigated whether platelets adhered to ULVWF strings are activated and form aggregates. We also determined whether activated platelets on ULVWF strings can support leukocyte tethering and rolling under high shear stresses. We found that platelets adhered to ULVWF expressed P‐selectin and bound PAC‐1, suggesting their rapid activation. We also found that leukocytes tethered to and rolled on these platelet‐decorated ULVWF strings, but not directly on endothelial cells, under high shear stresses of 20 and 40 dyn/cm2 in a P‐selectin dependent manner. These results suggest that the endothelial cell‐bound ULVWF provide an ideal matrix to aggregate platelets and recruit leukocytes to endothelial cells under high shear stress. The observed phenomenon delineates a mechanism for leukocytes to be tethered to arterial endothelial cells under high shear, providing a potential link between inflammation and thrombosis.

ADAMTS‐13 cleaves long von Willebrand factor multimeric strings anchored to endothelial cells in the absence of flow, platelets or conformation‐altering chemicals

phosphatidylethanolamine-binding molecular probe. J Nucl Med 2008; 49: 1345–52. 12 Glauert AM. Practical Methods in Electron Microscopy. Amsterdam: North-Holland Pub. Co., 1998. 13 Bevers EM, Comfurius P, Dekkers DW, Zwaal RF. Lipid translocation across the plasma membrane of mammalian cells. Biochim Biophys Acta 1999; 1439: 317–30. 14 Tait JF, Gibson D. Phospholipid binding of annexin V: effects of calcium and membrane phosphatidylserine content. Arch Biochem Biophys 1992; 298: 187–91. 15 Williamson P, Schlegel RA. Transbilayer phospholipid movement and the clearance of apoptotic cells. Biochim Biophys Acta 2002; 1585: 53–63.

Sulfatides inhibit platelet adhesion to von Willebrand factor in flowing blood

Summary.u2002 Sulfatides are sulfated glycosphingolipids present on cell surfaces that bind to adhesive proteins such as von Willebrand factor (VWF), P‐selectin, laminin and thrombospondin. Previous studies have localized the sulfatide‐binding site of VWF to amino acid residues Gln626–Val646 in the A1 domain. The A1 domain also contains the binding site for platelet glycoprotein Ib (GP Ib), a site that has been reported to be distinct from the sulfatide‐binding site. In this study, we analyzed the interaction of sulfatides with VWF and its effect on GP Ib‐mediated platelet adhesion under flow conditions. Recombinant VWF A1 domain (rVWF‐A1) bound specifically and saturably to sulfatides (half‐maximal concentration of ∼12.5u2003µgu2003mL−1), binding that was blocked by dextran sulfate (IC50≈100u2003µgu2003mL−1) but not by heparin at concentrations up to 100 Uu2003mL−1. Furthermore, sulfatides (125u2003µgu2003mL−1) prevented the adhesion of platelets or glycocalicin‐coupled polystyrene beads to a rVWF‐A1‐coated surface under high shear stress. In addition, plasma VWF prebound to a sulfatide‐coated surface failed to support subsequent platelet adhesion. These results provide firm evidence that sulfatides bind the VWF A1 domain at a site overlapping the GP Ib‐binding site.

Effects of anti-β2GPI antibodies on VWF release from human umbilical vein endothelial cells and ADAMTS13 activity

Essentials The antiphospholipid syndrome predisposes to thrombosis due to activation of endothelium and blood components. The role of anti‐β2GPI antibodies in VWF release and ADAMTS13 function is not well understood. Some anti‐β2GPI antibodies induce endothelial release of soluble VWF but not VWF strings. An anti‐β2GPI antibody can decrease ADAMTS13 activity in vitro similar to ex vivo results.

The leucine-rich repeat region of GP Ib/spl alpha/ is important in the interaction of the platelet GP Ib-IX-V complex with immobilized von Willebrand factor under fluid shear stress

Platelets play key roles in physiology, such as the arrest of bleeding following vascular injury, and in pathology, such as mural arterial thrombosis. The initial step in both processes is platelet adhesion to von Willebrand factor (vWf). The adhesion molecule on the platelet surface responsible for this interaction is the platelet glycoprotein (GP) Ib-IX-V complex. This complex consists of four subunits: GP Ib/spl alpha/, GP Ib/spl beta/, GP IX, and GP V[1]. The GP Ib/spl alpha/ subunit contains the vWf binding site and seven structural motifs of a leucine-rich sequence. The purpose of this study was to investigate the importance of the leucine-rich repeat region of GP Ib/spl alpha/ in the interaction of the GP Ib-IX-V complex with immobilized vWf under fluid shear stress. Cells expressing GP Ib/spl alpha/ with various point mutations in its leucine-rich repeat region either failed to interact with immobilized vWf or rolled on the surface at higher velocities compared to cells expressing wild-type GP Ib/spl alpha/. These results illustrate the importance of the leucine-rich repeat region of GP Ib/spl alpha/ in the receptor-ligand interaction under fluid shear stress.