Endothelial cells form a monolayer, which lines blood vessels. They are crucially involved in maintaining blood fluidity and providing controlled vascular hemostasis at sites of injury. Thereby endothelial cells facilitate multiple mechanisms, including both procoagulant and anticoagulant, which must be kept in balance. Under physiological conditions, endothelial cells constitute a nonadhesive surface preventing activation of platelets and the coagulation cascade. Multiple fibrinolytic and antithrombotic properties act on their cell surface contributing to the maintenance of blood fluidity. These include platelet inhibition, the heparin-antithrombin III system, tissue factor pathway inhibition, thrombomodulin/protein C system, and fibrinolytic qualities. At sites of vascular damage, platelets react immediately by adhering to the exposed extracellular matrix, followed by platelet-platelet interactions to form a clot that effectively seals the injured vessel wall to prevent excessive blood loss. For solid thrombus formation, functional platelets are essential. In this process, endothelial cells serve as a support surface for formation of procoagulant complexes and clotting. This review gives an overview about the central role of the endothelium as a dynamic lining which controls the complex interplay of the coagulation system with the surrounding cells.

Septins are GTP-binding and membrane-interacting proteins with a highly conserved domain structure involved in various cellular processes, including cytoskeleton organization, cytokinesis, and membrane dynamics. To date, 13 different septin genes have been identified in mammals (SEPT1 to SEPT12 and SEPT14), which can be classified into four distinct subgroups based on the sequence homology of their domain structure (SEPT2, SEPT3, SEPT6 and SEPT7 subgroup). The family members of these subgroups have a strong affinity for other septins and form apolar tri-, hexa- or octameric complexes consisting of multiple septin polypeptides. The first characterized core complex is the hetero-trimer SEPT2-6-7. Within these complexes single septins can be exchanged in a subgroup-specific manner. Hexamers contain SEPT2 and SEPT6 subgroup members and SEPT7 in two copies each whereas the octamers additionally comprise two SEPT9 subgroup septins. The various isoforms seem to determine the function and regulation of the septin complex. Septins self-assemble into higher-order structures, including filaments and rings in orders, which are typical for different cell types. Misregulation of septins leads to human diseases such as neurodegenerative and bleeding disorders. In non-dividing cells such as neuronal tissue and platelets septins have been associated with exocytosis. However, many mechanistic details and roles attributed to septins are poorly understood. We describe here some important mammalian septin interactions with a special focus on the clinically relevant septin interactions.

Septins constitute a group of GTP binding proteins that assemble into homo- and hetero-oligomeric complexes and filaments. These higher order septin structures are thought to function like scaffolds and/or diffusion barriers serving as spatial localizers for many proteins with key roles in cell polarity and cell cycle progression. In this study, we extensively characterized septin interaction partners using yeast two-hybrid and three-hybrid systems in addition to precipitation analyses in platelets. As a result, we identified human hetero-trimeric septin complexes on a large scale, which had been only postulated in the past. In addition, we illustrated roles of SEPT9 that might contribute to hetero-trimeric septin complex formation. SEPT9 can substitute for septins of the SEPT2 group and partially for SEPT7. Mutagenic analyses revealed that mutation of a potential phosphorylation site in SEPT7 (Y318) regulates the interaction with other septins. We identified several septin-septin interactions in platelets suggesting a regulatory role of diverse septin complexes in platelet function.

Gray platelet syndrome (GPS) is a rare recessive disorder caused by biallelic variants in NBEAL2 and characterized by bleeding symptoms, the absence of platelet α-granules, splenomegaly, and bone marrow (BM) fibrosis. Due to the rarity of GPS, it has been difficult to fully understand the pathogenic processes that lead to these clinical sequelae. To discern the spectrum of pathologic features, we performed a detailed clinical genotypic and phenotypic study of 47 patients with GPS and identified 32 new etiologic variants in NBEAL2. The GPS patient cohort exhibited known phenotypes, including macrothrombocytopenia, BM fibrosis, megakaryocyte emperipolesis of neutrophils, splenomegaly, and elevated serum vitamin B12 levels. Novel clinical phenotypes were also observed, including reduced leukocyte counts and increased presence of autoimmune disease and positive autoantibodies. There were widespread differences in the transcriptome and proteome of GPS platelets, neutrophils, monocytes, and CD4 lymphocytes. Proteins less abundant in these cells were enriched for constituents of granules, supporting a role for Nbeal2 in the function of these organelles across a wide range of blood cells. Proteomic analysis of GPS plasma showed increased levels of proteins associated with inflammation and immune response. One-quarter of plasma proteins increased in GPS are known to be synthesized outside of hematopoietic cells, predominantly in the liver. In summary, our data show that, in addition to the well-described platelet defects in GPS, there are immune defects. The abnormal immune cells may be the drivers of systemic abnormalities such as autoimmune disease.

Background: The number of implanted ventricular assist devices ( VAD s) has increased significantly recently. Bleeding, the most frequent complication, cannot be solely attributed to anticoagulation therapy. Acquired von Willebrand syndrome ( AVWS ) caused by increased shear stress is frequent in VAD patients and can increase the bleeding risk. The HeartMate III ( HM III ) is a novel left VAD featuring potential improvements over the HeartMate II . Methods and Results: In this study, we investigated the prevalence and onset of AVWS in 198 VAD patients. To our knowledge, this is the largest cohort of VAD patients whose longitudinal data on AVWS have been collected. We also analyzed whether AVWS is less severe in HM III patients than in HeartMate II patients. Because platelet dysfunction can raise the bleeding risk, we investigated platelet function in a subset of patients. In total, 198 VAD patients and 60 patients with heart transplants as controls were included in this study. The ratio of von Willebrand factor collagen binding capacity to von Willebrand factor:antigen, multimer analyses, and platelet function (especially secretion of α‐ and δ‐granules) were investigated. All 198 VAD patients developed AVWS . As soon as the VAD was explanted, the AVWS disappeared within hours. AVWS was less severe in the HM III patients than in the HeartMate II patients. The HM III patients had fewer bleeding symptoms. In addition, VAD patients exhibited a platelet α‐ and δ‐granule secretion defect. Conclusions: AVWS develops in VAD patients and may increase the bleeding risk. The HM III device causes less severe AVWS . Platelet secretion defects should be investigated in VAD patients because they also raise the bleeding risk. Clinical Trial Registration: https://www.drks.de/drks_web . Unique identifier: DRKS 00000649.

Patients with ventricular assist devices (VADs) and extracorporeal membrane oxygenation (ECMO) suffer from an increased risk for thromboembolic events as well as for hemorrhages. High shear stress in the mechanical device results in acquired von Willebrand syndrome (AVWS), characterized by a loss of high-molecular-weight multimers of von Willebrand factor (VWF) leading to an increased bleeding risk. Onset of AVWS occurs within hours, persists during the whole period of mechanical support, and subsides rapidly after explantation. Patients with the older HeartMate II exhibit more severe AVWS than those with the newer HeartMate III, thanks to lower shear stress in the latter. All ECMO and VAD patients exhibit thrombocytopathia and often thrombocytopenia which further increases the bleeding risk. Etiological models for AVWS are increased cleavage by the metalloproteinase ADAMSTS13, mechanical destruction of VWF, and shear-induced VWF binding to platelets. Platelet secretion defects may be caused by transient platelet activation leading to degranulation. AVWS can be diagnosed by detection of VWF multimers using gel-electrophoresis and functional assays of varying sensitivity (VWF ristocetin cofactor activity, VWF activity, VWF collagen binding). Platelet dysfunction is monitored using light transmission aggregometry and secretion defects are detectable using flow cytometry. Modest use of anticoagulants and a target-controlled therapy based on VWF parameters and other coagulation and platelet parameters are shown to be beneficial in this patient group. Persistent hemorrhages may be controlled with tranexamic acid and platelet concentrates. Prompt weaning from the device, when indicated, is the best therapeutic option to prevent recurrent bleeding.