Defective glycosylation of coagulation factor XII underlies hereditary angioedema type III.

Hereditary angioedema type III (HAEIII) is a rare inherited swelling disorder that is associated with point mutations in the gene encoding the plasma protease factor XII (FXII). Here, we demonstrate that HAEIII-associated mutant FXII, derived either from HAEIII patients or recombinantly produced, is defective in mucin-type Thr309-linked glycosylation. Loss of glycosylation led to increased contact-mediated autoactivation of zymogen FXII, resulting in excessive activation of the bradykinin-forming kallikrein-kinin pathway. In contrast, both FXII-driven coagulation and the ability of C1-esterase inhibitor to bind and inhibit activated FXII were not affected by the mutation. Intravital laser-scanning microscopy revealed that, compared with control animals, both F12-/- mice reconstituted with recombinant mutant forms of FXII and humanized HAEIII mouse models with inducible liver-specific expression of Thr309Lys-mutated FXII exhibited increased contact-driven microvascular leakage. An FXII-neutralizing antibody abolished bradykinin generation in HAEIII patient plasma and blunted edema in HAEIII mice. Together, the results of this study characterize the mechanism of HAEIII and establish FXII inhibition as a potential therapeutic strategy to interfere with excessive vascular leakage in HAEIII and potentially alleviate edema due to other causes.

The plasma contact system 2.0.

The contact system is a protease cascade that is initiated by factor XII activation on cardiovascular cells. The system starts procoagulant and proinflammatory reactions, via the intrinsic pathway of coagulation and the kallikrein-kinin system, respectively. The biochemistry of the contact system in vitro is well understood. However, activators of the system in vivo and their contributions to disease states have remained enigmatic. Recent experimental and clinical data have identified misfolded proteins, collagens, and polyphosphates as the long-sought activators of the contact system in vivo. Here we present an overview about contact system activators and their contributions to health and pathology.

Mast cells increase vascular permeability by heparin-initiated bradykinin formation in vivo.

Activated mast cells trigger edema in allergic and inflammatory disease. We report a paracrine mechanism by which mast cell-released heparin increases vascular permeability in vivo. Heparin activated the protease factor XII, which initiates bradykinin formation in plasma. Targeting factor XII or kinin B2 receptors abolished heparin-triggered leukocyte-endothelium adhesion and interfered with a mast cell-driven drop in blood pressure in rodents. Intravital laser scanning microscopy and tracer measurements showed heparin-driven fluid extravasation in mouse skin microvessels. Ablation of factor XII or kinin B2 receptors abolished heparin-induced skin edema and protected mice from allergen-activated mast cell-driven leakage. In contrast, heparin and activated mast cells induced excessive edema in mice deficient in the major inhibitor of factor XII, C1 esterase inhibitor. Allergen exposure triggered edema attacks in hereditary angioedema patients, lacking C1 esterase inhibitor. The data indicate that heparin-initiated bradykinin formation plays a fundamental role in mast cell-mediated diseases.

Cytoskeleton assembly at endothelial cell-cell contacts is regulated by alphaII-spectrin-VASP complexes.

Directed cortical actin assembly is the driving force for intercellular adhesion. Regulated by phosphorylation, vasodilator-stimulated phosphoprotein (VASP) participates in actin fiber formation. We screened for endothelial proteins, which bind to VASP, dependent on its phosphorylation status. Differential proteomics identified alphaII-spectrin as such a VASP-interacting protein. alphaII-Spectrin binds to the VASP triple GP(5)-motif via its SH3 domain. cAMP-dependent protein kinase-mediated VASP phosphorylation at Ser157 inhibits alphaII-spectrin-VASP binding. VASP is dephosphorylated upon formation of cell-cell contacts and in confluent, but not in sparse cells, alphaII-spectrin colocalizes with nonphosphorylated VASP at cell-cell junctions. Ectopic expression of the alphaII-spectrin SH3 domain at cell-cell contacts translocates VASP, initiates cortical actin cytoskeleton formation, stabilizes cell-cell contacts, and decreases endothelial permeability. Conversely, the permeability of VASP-deficient endothelial cells (ECs) and microvessels of VASP-null mice increases. Reconstitution of VASP-deficient ECs rescues barrier function, whereas alphaII-spectrin binding-deficient VASP mutants fail to restore elevated permeability. We propose that alphaII-spectrin-VASP complexes regulate cortical actin cytoskeleton assembly with implications for vascular permeability.