Abstract 535: Proteomics Identifies Endothelium-derived Clusterin as a Novel Regulator of Von Willebrand Factor Multimerization

Abstract

Background: Secretion of vascular endothelial cells (ECs) is considered to be the first line of defense against vascular injury. Vascular endothelial growth factor (VEGF) is one of the most important endothelial growth factors, involved in various endothelial functions including the regulatory secretion of ECs. However, the systematic analysis of the VEGF-induced secretory proteins has not yet been carried out. Objectives: To investigate the secretory proteins rapidly released from ECs under VEGF stimulation and their functions on the ECs. Methods: A non-serum culture system was established for analyzing the proteins secreted form human primary ECs in response to VEGF with mass spectrometry. Results: In the proteomic result, 362 proteins were identified and assigned to the VEGF-induced secretome, including the known von Willebrand factor (VWF), a multimeric protein mediating adhesion of platelets at vascular injury sites. Clusterin, a multifunctional protein involved in various diseases including Alzheimer disease and cancers, was selected for further study as a new VEGF-induced secretory protein. Clusterin was co-localized with VWF both in Weibel-Palade bodies (WPBs) and extracellular VWF strings of ECs. We showed that the localization of clusterin in pseudo-WPBs in 293T cells depends on the carboxy-terminal domains of VWF. Functional study showed that deficiency of clusterin in ECs increased the size of VWF multimers. Conclusions: We not only obtained a VEGF-induced secretome using primary ECs, but also determined clusterin as a novel component of WPBs, which is capable of maintaining the normal pattern of VWF multimers, and suggest its function on vascular thrombosis and haemostasis.