New insight into how endothelial cells control storage and release of secretory proteins

Cells from patients with rare disease help to uncover new mechanism

Ellie Karampini, PhD student at the research group of Molecular and Cellular Hemostasis, uncovered a novel mechanism into how vascular endothelial cells regulate secretion of hemostatic and inflammatory proteins from so-called Weibel-Palade bodies (WPBs). This helps to understand why people with defects in this process suffer from frequent bleeding and infections. The latest developments are published in Haematologica.

Ellie KarampiniEllie Karampini (photo), from the Bierings and Voorberg labs, studies the intracellular pathways that control the formation and exocytosis of secretory organelles such as WPBs or alpha- and dense granules in platelets. To dissect how WPBs are made and released she uses blood outgrowth endothelial cells (BOECs), which are endothelial cells that can be derived from peripheral blood.

Maaike SchillemansTogether with her colleague Maaike Schillemans (photo), Ellie has turned this into a powerful ex vivo model system by using patient-derived BOECs. With these cells she studied disease mechanisms in the endothelial context and established a method to introduce mutations in BOECs using CRISPR/Cas9 gene editing. 

Rare disease

In this study, Ellie used BOECs from a patient with the storage pool disorder Hermansky-Pudlak Syndrome type 2 (HPS-2), a rare disease,  to investigate protein trafficking to newly formed WPBs. Platelets from HPS patients, but also their immune cells such as neutrophils and T-cells, have absent or malformed secretory organelles because they are unable to transfer proteins from endosomes to maturing secretory organelles. 

By carefully studying trafficking of components of the WPB in HPS-2 BOECs, Ellie found that this same trafficking step is also responsible for sorting of a number of membrane proteins to WPBs. Interestingly, this also includes the protein VAMP8, which is a member of the SNARE complex that controls membrane fusion and exocytosis. She showed that WPBs that lack this key component of their SNARE machinery, such as in HPS-2 BOECs or in CRISPR-edited VAMP8 knock out BOECs, are less able to undergo exocytosis and secrete less VWF. 

Von Willebrand factor

Endothelial cells form the inner lining of the blood vessels and act as a semi-permeable barrier between blood and underlying tissues. They also have an ingenious self-repair mechanism that immediately secretes large quantities of the hemostatic protein Von Willebrand factor (VWF) when the blood vessel wall is damaged, for instance during an injury. VWF is stored along with a cocktail of inflammatory mediators in Weibel-Palade bodies, endothelial-specific secretory organelles that within seconds of activation can undergo exocytosis to deliver their content into the circulation. 

Blood platelets will adhere to the long strings of secreted VWF, which promotes the formation of a platelet plug. Inflammatory proteins that are simultaneously released from WPBs will direct leukocytes to the site of vascular injury. This process is crucial to prevent excessive blood loss and to minimize the risk of infections. 

This new study generated new insight into how endothelial cells control storage and release of secretory proteins such as VWF. It shows that storage pool disorders such as HPS-2 also impact the secretory function of the blood vessel wall, which may contribute to the bleeding complications seen in patients with storage pool disorders.

Read the full article in Haematologica.

This work is financed by the Landsteiner Foundation of Blood Transfusion Research (LSBR) and the European Hematology Association (EHA).