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OBJECTIVEHigh-mobility group box-1 (HMGB1) protein is a nuclear DNA-binding protein released from necrotic cells, inducing inflammatory responses and promoting tissue repair and angiogenesis. Diabetic human and mouse tissues contain lower levels of HMGB1 than their normoglycemic counterparts. Deficient angiogenesis after ischemia contributes to worse outcomes of peripheral arterial disease in patients with diabetes. To test the hypothesis that HMGB1 enhances ischemia-induced angiogenesis in diabetes, we administered HMGB1 protein in a mouse hind limb ischemia model using diabetic mice.RESEARCH DESIGN AND METHODSAfter the induction of diabetes by streptozotocin, we studied ischemia-induced neovascularization in the ischemic hind limb of normoglycemic, diabetic, and HMGB1-treated diabetic mice.RESULTSWe found that the perfusion recovery was significantly attenuated in diabetic mice compared with normoglycemic control mice. Interestingly, HMGB1 protein expression was lower in the ischemic tissue of diabetic mice than in normoglycemic mice. Furthermore, we observed that HMGB1 administration restored the blood flow recovery and capillary density in the ischemic muscle of diabetic mice, that this process was associated with the increased expression of vascular endothelial growth factor (VEGF), and that HMGB1-induced angiogenesis was significantly reduced by inhibiting VEGF activity.CONCLUSIONSThe results of this study show that endogenous HMGB1 is crucial for ischemia-induced angiogenesis in diabetic mice and that HMGB1 protein administration enhances collateral blood flow in the ischemic hind limbs of diabetic mice through a VEGF-dependent mechanism.

Prothrombin (factor II [FII]) deficiency is a rare inherited coagulation disorder, having a prevalence of approximately 1 in 2,000,000. Two phenotypes can be distinguished: (1) true hypoprothrombinemia (type I deficiency), characterized by concomitantly low levels of the zymogen antigen; and (2) dysprothrombinemia (type II deficiency), characterized by the normal or near-normal synthesis of a dysfunctional protein. In the latter case, recent studies showed that particular mutations in the catalytic domain of active thrombin can even impair the enzyme interaction with antithrombin, favoring thromboembolic diseases. In some cases, hypoprothrombinemia associated with dysprothrombinemia was also described in compound heterozygous defects. Prothrombin is essential for the development of mammalian organisms. No living patient with undetectable plasma prothrombin has been reported to date. Prothrombin is encoded by a ≈21 kb gene located on chromosome 11 and containing 14 exons. Thirty-nine different mutations have been identified and characterized in prothrombin deficiency. Many of these are present in the catalytic site, whereas some involve regulatory domains, such as the anion-binding exosite I, the Na+-binding loop, and the light A-chain. Most hypoprothrombinemia-associated mutations are missense, but nonsense mutations leading to stop codons and one single nucleotide deletion have also been identified. Finally, recent developments in the therapy of congenital prothrombin deficiency are presented and discussed.

Inflammatory lung disease is a primary cause of morbidity and mortality in cystic fibrosis (CF). Mechanisms of unresolved acute inflammation in CF are not completely known, although the involvement of cystic fibrosis transmembrane conductance regulator (CFTR) in nonrespiratory cells is emerging. Here we examined CFTR expression and function in human platelets (PLTs) and found that they express a biologically active CFTR. CFTR blockade gave an ∼50% reduction in lipoxin A(4) (LXA(4)) formation during PLT/polymorphonuclear leukocytes (PMN) coincubations by inhibiting the lipoxin synthase activity of PLT 12-lipoxygenase. PLTs from CF patients generated ∼40% less LXA(4) compared to healthy subject PLTs. CFTR inhibition increased PLT-dependent PMN viability (33.0±5.7 vs. 61.2±8.2%; P=0.033), suppressed nitric oxide generation (0.23±0.04 vs. 0.11±0.002 pmol/10(8) PLTs; P=0.004), while reducing AKT (1.02±0.12 vs. 0.71±0.007 U; P=0.04), and increasing p38 MAPK phosphorylation (0.650±0.09 vs. 1.04±0.24 U; P=0.03). Taken together, these findings indicate that PLTs from CF patients are affected by the molecular defect of CFTR. Moreover, this CF PLT abnormality may explain the failure of resolution in CF.

quantitative modifications of von Willebrand factor in patients with essential thrombocythemia and controlled platelet count. J Thromb Haemost 2015; 13: 1226-37.

Inflammation and coagulation constantly influence each other and are constantly in balance. Emerging evidence supports this statement in acute inflammatory diseases, such as sepsis, but it also seems to be very important in chronic inflammatory settings, such as inflammatory bowel disease (IBD). Patients with Crohn's disease and ulcerative colitis have an increased risk of thromboembolic events, and several abnormalities concerning coagulation components occur in the endothelial cells of intestinal vessels, where most severe inflammatory abnormalities occur. The aims of this review are to update and classify the type of coagulation system abnormalities in IBD, and analyze the strict and delicate balance between coagulation and inflammation at the mucosal level. Recent studies on possible therapeutic applications arising from investigations on coagulation abnormalities associated with IBD pathogenesis will also be briefly presented and critically reviewed.

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The expression of the elongated fibrinogen ␥ chain, termed ␥, derives from alternative splicing of mRNA and causes an insertion sequence of 20 amino acids. This insertion domain interacts with the anion-binding exosite (ABE)-II of thrombin. This study investigated whether and how ␥ chain binding to ABE-II affects thrombin interaction with its platelet receptors, i.e. glycoprotein Ib␣ (GpIb␣), protease-activated receptor (PAR) 1, and PAR4. Both synthetic ␥ peptide and fibrinogen fragment D*, containing the elongated ␥ chain, inhibited thrombin-induced platelet aggregation up to 70%, with IC 50 values of 42 ؎ 3.5 and 0.47 ؎ 0.03 M, respectively. Solid-phase binding and spectrofluorimetric assays showed that both fragment D* and the synthetic ␥ peptide specifically bind to thrombin ABE-II and competitively inhibit the thrombin binding to GpIb␣ with a mean K i ≈ 0.5 and ≈35 M, respectively. Both these ␥ chain-containing ligands allosterically inhibited thrombin cleavage of a synthetic PAR1 peptide, of native PAR1 molecules on intact platelets, and of the synthetic chromogenic peptide D-Phe-pipecolyl-Arg-p-nitroanilide. PAR4 cleavage was unaffected. In summary, fibrinogen ␥ chain binds with high affinity to thrombin and inhibits with combined mechanisms the platelet response to thrombin. Thus, its variations in vivo may affect the hemostatic balance in arterial circulation.Fibrinogen is a key molecule in both primary and secondary hemostasis, because of its role in forming the platelet plug by connecting activated platelets and in forming plasma fibrin clot upon thrombin cleavage. Fibrinogen consists of two symmetric half-molecules, each containing a set of three different polypeptide chains termed A␣, B␤, and ␥. The latter contains several sites that interact with different ligands such as other fibrin(ogen) molecules, coagulation enzymes, growth factors, and integrins (1). The product of thrombin digestion of fibrinogen, i.e. fibrin, binds with a considerable specificity thrombin, so that in the early studies fibrin was termed antithrombin I (2). Thrombin has a divalent interaction with two classes of binding sites on fibrin, one of low affinity in the E domain and the other of high affinity in the D domain of fibrin(ogen) molecules (3). Binding of thrombin to fibrinogen involves sequences of both A␣ and B␤ chain, which contain recognition sites in the fibrinogen E domain. These recognition sites are still able to interact with thrombin after cleavage of fibrinopeptide A and B and form the low affinity binding site for the enzyme. The D domains contain a ␥ chain variant, termed ␥Ј, arising from an alternative mRNA splicing (4), resulting in an elongated chain composed of 427 instead of 411 residues. The inserted region at the C terminus is composed of 20 amino acids ( 408 VRPEH-PAETEYDSLYPEDDL 427 ), rich of acidic side chains and two sulfate anions linked to Tyr 418 and Tyr 422 (5). The elongated ␥ chain, termed ␥Ј, mainly heterodimerizes in the fibrinogen molecule with the more abundant ␥A chain, thus generating ...