Regulated von Willebrand factor (vWf) secretion is restored by pro-vWf expression in a transfectable endothelial cell line

Von Willebrand factor (vWf) is a glycoprotein involved in primary hemostasi s and synthesized in endothelial cells (EC). vWf is stored in secretory gra nules specific for EC called Weibel-Palade bodies (WPb). Studies on the mol ecular mechanisms of vWf storage and acute release are hampered by the limi tations of the available endothelial cell culture models. We created a suit able model by stable transfection of the vWf-negative ECV304 endothelial ce ll line with pro-vWf cDNA. Pro-vWf was normally cleaved to mature vWf and s tored in WPb. Acute vWf release occurred in response to the calcium ionopho re A23187. Thus, vWf expression is sufficient to restore functional secreto ry granules in ECV304 cells. We used this model to study the role of WPb in the storage of tissue-type plasminogen activator (t-PA), a key fibrinolyti c enzyme that is acutely released by EC. but whose intracellular storage co mpartment is still a matter of debate. We observed that restoration of WPb in ECV304 cells results in the targeting of t-PA to these storage granules. (C) 2000 Elsevier Science B.V. All rights reserved.