Nightingale, TD; McCormack, JJ; Grimes, W; Robinson, C; Lopes da Silva, M; White, IJ; Vaughan, A; ... Cutler, DF; + view all Nightingale, TD; McCormack, JJ; Grimes, W; Robinson, C; Lopes da Silva, M; White, IJ; Vaughan, A; Cramer, LP; Cutler, DF; - view fewer (2018) Tuning the endothelial response: differential release of exocytic cargos from Weibel-Palade Bodies. Journal of Thrombosis and Haemostasis , 16 (9) pp. 1873-1886. 10.1111/jth.14218.

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Abstract

BACKGROUND: Endothelial cells harbour specialised storage organelles, Weibel-Palade Bodies (WPBs). Exocytosis of WPB content into the vascular lumen initiates primary haemostasis, mediated by Von Willebrands factor (VWF) and inflammation, mediated by several proteins including P-selectin. During full fusion, secretion of this large haemostatic protein and smaller pro-inflammatory proteins are thought to be inextricably linked. OBJECTIVE: To determine if secretagogue-dependent differential release of WPB cargo occurs, and whether this is mediated by the formation of an actomyosin ring during exocytosis. METHODS: We used VWF string analysis, leukocyte rolling assays, ELISA, spinning disk confocal microscopy, high-throughput confocal microscopy and inhibitor and siRNA treatments to demonstrate the existence of cellular machinery that allows differential release of WPB cargo proteins. RESULTS: Inhibition of the actomyosin ring differentially effects two processes regulated by WPB exocytosis; it perturbs VWF string formation but has no effect on leukocyte rolling. The efficiency of ring recruitment correlates with VWF release; the ratio of release of VWF to small cargoes decreases when ring recruitment is inhibited. The recruitment of the actin ring is time-dependent; fusion events occurring directly after stimulation are less likely to initiate haemostasis than later events, and is activated by PKC isoforms. CONCLUSIONS: Secretagogues differentially recruit the actomyosin ring, thus demonstrating one mechanism by which the pro-thrombotic effect of endothelial activation can be modulated. This potentially limits thrombosis whilst permitting a normal inflammatory response. These results have implications for the assessment of WPB fusion, cargo-content release and the treatment of patients with von Willebrand disease.

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