Our findings indicate a new pathway for bradykinin formation in patients with HAE, in which FXII is cleaved and activated by plasmin. This should lead to the identification of new markers for diagnosis and targets for treatment.

OBJECTIVEPatients with type 2 diabetes have an increased risk of cardiovascular disease and show abnormalities in the coagulation cascade. We investigated whether increased synthesis of tissue factor (TF) by platelets could contribute to the hypercoagulant state.RESEARCH DESIGN AND METHODSPlatelets from type 2 diabetic patients and matched control subjects were adhered to different surface-coated proteins, and TF premRNA splicing, TF protein, and TF procoagulant activity were measured.RESULTSDifferent adhesive proteins induced different levels of TF synthesis. A mimetic of active clopidogrel metabolite (AR-C69931 MX) reduced TF synthesis by 56 ± 10%, an aspirin-like inhibitor (indomethacin) by 82 ± 9%, and the combination by 96 ± 2%, indicating that ADP release and thromboxane A2 production followed by activation of P2Y12 and thromboxane receptors mediate surface-induced TF synthesis. Interference with intracellular pathways revealed inhibition by agents that raise cAMP and interfere with phosphatidylinositol 3-kinase/protein kinase B. Insulin is known to raise cAMP in platelets and inhibited collagen III–induced TF premRNA splicing and reduced TF activity by 35 ± 5 and 47 ± 5% at 1 and 100 nmol/l. Inhibition by insulin was reduced in type 2 diabetes platelets resulting in an ∼1.6-fold higher TF synthesis than in matched control subjects.CONCLUSIONSWe characterized the extra- and intracellular mechanisms that couple surface activation to TF synthesis in adhering platelets. In healthy individuals, TF synthesis is inhibited by insulin, but in patients with type 2 diabetes inhibition is impaired. This leads to the novel finding that platelets from type 2 diabetic patients produce more TF than platelets from matched control subjects.

BackgroundThe endocannabinoid system has previously been implicated in the regulation of neurons and inflammatory cells. Additionally, it has been reported that endocannabinoid receptors are present on circulating platelets, but there has been conflicting evidence on their contribution to platelet function.ObjectivesOur aim was to examine the role of endocannabinoids in platelet function in vitro and in vivo.Methods and ResultsWe studied the effects of the well-characterized endogenous endocannabinoid anandamide on platelet aggregation in suspension, α-granule release, calcium mobilization, Syk phosphorylation, as well as platelet spreading and aggregate formation under flow. Anandamide inhibits platelet aggregation and α-granule release by collagen, collagen-derived peptide CRP-XL, ADP, arachidonic acid and thromboxane A2 analogue U46619. However, activation via thrombin receptor PAR-1 stays largely unaffected. Calcium mobilization is significantly impaired when platelets are stimulated with collagen or CRP-XL, but remains normal in the presence of the other agonists. In line with this finding, we found that anandamide prevents collagen-induced Syk phosphorylation. Furthermore, anandamide-treated platelets exhibit reduced spreading on immobilized fibrinogen, have a decreased capacity for binding fibrinogen in solution and show perturbed platelet aggregate formation under flow over collagen. Finally, we investigated the influence of Cannabis sativa consumption by human volunteers on platelet activation. Similar to our in vitro findings with anandamide, ex vivo collagen-induced platelet aggregation and aggregate formation on immobilized collagen under flow were impaired in whole blood of donors that had consumed Cannabis sativa.ConclusionsEndocannabinoid receptor agonists reduce platelet activation and aggregate formation both in vitro and ex vivo after Cannabis sativa consumption. Further elucidation of this novel regulatory mechanism for platelet function may prove beneficial in the search for new antithrombotic therapies.

Summary. Objectives: Platelets from healthy subjects are inhibited by insulin but type 2 diabetes mellitus (T2DM) platelets have become insulin-resistant, which might explain their hyperactivity. In the present study we investigated whether monocytes are responsive to insulin. Methods and results: LPSinduced tissue factor (TF) upregulation was measured in human monocytes and monocytic THP-1 cells in a factor Xa generation assay.) induced a dose-dependent inhibition in both cell types and in monocytes 100 nmol L )1 insulin inhibited cytosolic, membrane-bound and microparticle TF by 32 ± 2, 27 ± 3 and 52 ± 4% (n = 3). Insulin induced Tyr phosphorylation of the insulin receptor (INS-R) and formation of an INS-R -G i a 2 complex, suggesting interference with LPS-induced cAMP control. Indeed, insulin interfered with LPS-induced cAMP decrease and TF upregulation in a manner similar to an inhibitor of G i (pertussis toxin) and agents that raise cAMP (iloprost, forskolin, IBMX) reduced TF upregulation. Although LPS failed to raise cytosolic Ca 2+ , quenching of Ca 2+ increases (BAPTA-AM) reduced and induction of Ca 2+ entry (ionophore, P2X7 activation) enhanced upregulation of TF mRNA and procoagulant activity. Insulin interfered with MCP-1-induced Ca 2+ mobilization but not with ATP-induced Ca 2+ rises. Conclusions: Insulin inhibits TF expression in monocytes and monocyte-derived microparticles through interference with G i a 2 -mediated cAMP suppression, which attenuates Ca 2+ -mediated TF synthesis.