Acutely secreted VWF forms extended strings under fluid shear stress. (A) Confluent HUVECs were stimulated with 100 μM histamine for 30 minutes, washed with DPBS, and fixed with 2% paraformaldehyde without permeabilization. Cell-surface VWF was stained with a polyclonal antibody (A082; Dako North America) and an Alexa Fluor 594–conjugated secondary antibody. Bar represents 10 μm. (B) HUVECs in a parallel plate flow chamber were perfused with medium 199 supplemented with 2% BSA, 100 μM histamine, and fluorescent VWF polyclonal antibody, at a shear stress of 10 dyne/cm2. Images were captured 5 minutes after the initiation of flow. Bar represents 10 μm. (C) Perfusion assays were conducted at different values of fluid shear stress. The bars indicate the numbers (mean ± SEM) of VWF strings more than 20 μm long formed 5 minutes after the initiation of flow from 10 fields per experiment. Each experiment was performed at least 3 times.
