Endothelial cell storage and secretion of bioactive components: a crucial role in haemostasis and inflammation

Tematyka

It is essential that the response to vascular injury or infection is fast; this minimises loss of blood and spread of pathogens. As such, endothelial cells harbour specialised rod shaped storage organelles (WPB) that contain multiple pre-made pro-inflammatory and pro-haemostatic proteins. Within minutes of endothelial cell stimulation WPB are exocytosed and release their stored content into the vasculature thus starting the processes of both haemostasis and leukocyte recruitment. Pro-inflammatory content include P-selectin which on display at the cell surface allows initial rolling and recruitment of leukocytes to sites of tissue injury. The most important haemostatic component of WPB is the glycoprotein von Willebrands factor (VWF) that comprises 90% of stored protein. Upon exocytosis VWF is unfurled by the shear-force present in the blood vasculature to produce millimetre-long protein strings revealing multiple binding sites for platelets. Failure to secrete properly processed VWF result in bleeding disorders (Von Willebrand’s disease is the most common inherited bleeding disorder).
Endothelial cell storage and secretion of bioactive components: a crucial role in haemostasis and inflammation

My talk will describe a number of novel assays for monitoring WPB trafficking and content secretion including biochemical assays, high-throughput image analysis, high-speed live cell imaging and correlative light and electron microscopy. These approaches allow us to study the mechanics, both of WPB exocytosis and of subsequent, exocytosis related, trafficking events (such as additional compensatory endocytosis). Our research demonstrates that efficient secretion of VWF requires the formation of an actomyosin ring around the base of WPB that squeezes out the protein content as strings. We also describe some of the upstream signalling molecules required for ring recruitment and demonstrate that this ring is recruited by different physiological stimuli to varying extents both with respect to the timing and the number of exocytic events. Finally we show that following exocytosis an additional round of endocytosis is started. This limits any increase in cell size and helps to control exocytic mode to maintain single granule exocytosis. These processes allow the endothelium to precisely regulate the haemostatic and inflammatory response. Targeting such processes with therapeutics may provide new methods for controlling haemostasis and inflammation.