Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.5 (thrombin)
33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Recombinant immunoglobulin (Ig) fusion proteins of cell surface and intracellular proteins have wide applications. For example, fusion proteins have been used in the isolation, identification and study of ligands and the effects of binding or blocking a receptor-ligand pair, either in vivo or in vitro. For some applications, removal of the immunoglobulin Fc region is advantageous. We have developed two vectors for the expression of Ig fusion proteins that contain recognition sequences for protease cleavage using thrombin. In one vector, the sequence encoding the thrombin cleavage site is located at the junction of the DNA fragment encoding the protein or protein fragment to be studied and the hinge and constant regions of the immunoglobulin, allowing the generation of a monomeric form of the protein of interest. In the second vector, the sequence encoding the thrombin cleavage site is located between the sequences encoding the hinge and constant regions of the immunoglobulin, allowing for the generation of covalent dimers of the recombinant protein without the constant Fc domains. We have used these vectors to produce the constructs encoding two forms of the extracellular domain of CD40, CD40ThrIg and CD40HinThrIg, allowing production of a monomeric and dimeric form of recombinant CD40. Cleavage is efficient and complete. Following cleavage, there was no detectable binding of the monomeric form of CD40 to a soluble form of gp39, the ligand of CD40, while the dimeric form was able to bind. These vectors have been constructed to allow facile substitution with other sequences to generate cleavable forms of other proteins of interest.
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PMID:Cleavable CD40Ig fusion proteins and the binding to sgp39. 855 Oct 28

Recently, we have demonstrated that human platelets carry preformed CD40 ligand (CD154) molecules, which rapidly appear on the platelet surface following stimulation by thrombin. Once on the surface, platelet CD154 induces an inflammatory reaction of CD40-bearing endothelial cells. This study shows that strong platelet agonists other than thrombin also lead to the expression of CD154 on the platelet surface. At the same time, several lines of evidence are presented that together indicate that thrombotic events in the vasculature are generally accompanied by activation of the inflammatory potential of platelet CD154. This study also reports the constitutive expression of CD40, the receptor for CD154, on platelets. The binding of CD154 to coexpressed CD40 in the platelet aggregate leads within minutes to hours to the cleavage of membrane-bound surface CD154 and the release of an 18-kd soluble form of the molecule. Soluble CD154 (sCD154), in contrast to transmembrane CD154, can no longer induce an inflammatory reaction of endothelial cells. These findings indicate that the interaction of platelet CD154 with CD40 on neighboring cells is temporally limited to prevent an uncontrolled inflammation at the site of thrombus formation. Thus, similar to the very tight regulation of the CD154-CD40 interaction in the immune system, an effective mechanism controls the inflammatory potential of platelet CD154 in the vascular system. (Blood. 2001;98:1047-1054)
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PMID:The inflammatory action of CD40 ligand (CD154) expressed on activated human platelets is temporally limited by coexpressed CD40. 1149 50

We report here that soluble CD40 ligand (sCD40L) is released from human platelets when activated with collagen or thrombin. The sCD40L was detectable in the culture supernatants of platelets within 30 min after stimulation in vitro, and reached maximal levels in 3 h. The release was blocked by the metalloproteinase inhibitor, KB8301, indicating that the soluble CD40L is made by cleaving the membrane bound CD40L expressed on activated platelets. The sCD40L was undetectable in the supernatant of the activated platelets obtained from patients with X-linked hyper IgM syndrome (XHIM), who have defects in CD40L gene. Since sCD40L has been shown to have biologic function on the activation of vascular endothelial cells and B cells, these findings suggest that platelets play some roles in both inflammation and humoral immune response by releasing soluble CD40L.
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PMID:Characterization of soluble CD40 ligand released from human activated platelets. 1216 Feb 39

Platelets are known for their role in haemostasis where they help prevent blood loss at sites of vascular injury. To do this, they adhere, aggregate and form a procoagulant surface leading to thrombin generation and fibrin formation. Platelets also release substances that promote tissue repair and influence the reactivity of vascular and other blood cells in angiogenesis and inflammation. They contain storage pools of growth factors including PDGF, TGF-beta?and VEGF as well as cytokines including proteins such as PF4 and CD40L. Chemokines and newly synthesised active metabolites are also released. The fact that platelets secrete growth factors and active metabolites means that their applied use can have a positive influence in clinical situations requiring rapid healing and tissue regeneration. Their administration in fibrin clot or fibrin glue provides an adhesive support that can confine secretion to a chosen site. Additionally,the presentation of growth factors attached to platelets and/or fibrin may result in enhanced activity over recombinant proteins. Dental implant surgery with guided bone regeneration is one situation where an autologous platelet-rich clot clearly accelerates ossification after tooth extraction and/or around titanium implants. The end result is both marked reductions in the time required for implant stabilisation and an improved success rate. Orthopaedic surgery, muscle and/or tendon repair, reversal of skin ulcers, hole repair in eye surgery and cosmetic surgery are other situations where autologous plate-lets accelerate healing. Our aim is to review these advances and discuss the ways in which platelets may provide such unexpected beneficial therapeutic effects.
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PMID:Autologous platelets as a source of proteins for healing and tissue regeneration. 1469 63

There is evidence of activation of both blood coagulation and platelets in sickle cell disease. For example, plasma samples obtained in the steady state and during painful crisis demonstrate high levels of thrombin generation, depletion of anticoagulant proteins, and abnormal activation of the fibrinolytic system. Similarly, exposure of surface markers such as CD62P and CD40L, along with increased circulating levels of thrombospondin, signal platelet activation. In addition to its effects on the cleavage of fibrinogen and its ability to activate platelets, the increase in circulating thrombin levels, with its wide-ranging effects on endothelial cells and blood vessels, may be important in the pathophysiology of sickle cell disease. Therefore, treatments that could decrease thrombin generation or platelet activation may be beneficial in both the treatment of sickle cell disease and the prevention of complications that characterize this genetic disorder. This review discusses hypercoagulability in the various forms of sickle cell disease, including homozygous sickle cell anemia, hemoglobin SC disease, hemoglobin SD disease, and sickle cell-beta-thalassemia.
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PMID:Hypercoagulability in sickle cell disease: a curious paradox. 1469 25

The pro-inflammatory CD40/CD40L dyad participates in atherogenesis. Plasma levels of the soluble ligand (sCD40L) predict cardiovascular events and are elevated in diabetic patients. This study compared CD40/CD40L surface expression on platelets and T lymphocytes of diabetic and control subjects, and tested whether glucose and advanced glycation end products (AGEs) stimulate sCD40L release. Constitutive and inducible surface expression of CD40/CD40L on platelets or T lymphocytes did not differ between diabetic patients (n = 9) and controls (n = 13). Platelets from diabetic patients contained higher intracellular CD40L than controls (p < 0.05) and thrombin stimulated greater platelet sCD40L release in diabetic patients (15.11 +/- 16.77 ng/ml) compared to controls (3.64 +/- 2.03 ng/ml; p < 0.05). Glucose and AGEs induced platelet sCD40L release and CD40L expression in mouse megakaryocytes. This study demonstrates elevated CD40L content and inducible release from platelets of diabetic patients, and identifies glucose and AGEs as potential triggers of expression and release accounting for the elevated sCD40L plasma levels in these patients.
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PMID:Elevated release of sCD40L from platelets of diabetic patients by thrombin, glucose and advanced glycation end products. 1630 11

Peroxisome proliferator-activated receptor gamma (PPARgamma) is an important transcription factor for lipid and glucose metabolism. Currently, the PPARgamma ligands rosiglitazone and pioglitazone are used for the treatment of type 2 diabetes mellitus because they are potent insulin sensitizers. Recently, PPARgamma has emerged as an important anti-inflammatory factor. Platelets, anucleate cells involved in hemostasis, have also been implicated as key contributors to inflammation, because they produce many pro-inflammatory and pro-atherogenic mediators when activated. Surprisingly, it was discovered recently that platelets contain PPARgamma and that PPARgamma ligands, both natural and synthetic, inhibit platelet activation and release of bioactive mediators. In particular, release of soluble CD40 ligand (sCD40L) and thromboxane (TXA(2)) was inhibited by PPARgamma ligands in thrombin-activated platelets. CD40L signaling induces pro-inflammatory processes in many cell types, and increased blood levels of sCD40L are closely associated with inflammation, diabetes, and cardiovascular disease. Targeting platelet PPARgamma will, therefore, be an important treatment strategy for the attenuation of chronic inflammatory processes and prevention of thrombus formation.
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PMID:Platelets as a novel target for PPARgamma ligands : implications for inflammation, diabetes, and cardiovascular disease. 1683 Oct 22

The tissue factor (TF) pathway is the primary mechanism for initiation of blood coagulation. Circulating blood contains TF, which originates mainly from monocytes and is thrombogenic. The presence of somatostatin (SMS) receptors on monocytes suggests the possibility that SMS may regulate TF synthesis and/or release. Circulating TF procoagulant activity (TF-PCA), factor VIIa activity (FVIIa; clotting assays), TF antigen (TF-Ag; ELISA), prothrombin fragment 1.2 (F1.2), thrombin-antithrombin complexes (ELISAs), CD40 ligand expression on platelets, and monocyte-platelet aggregates (flow cytometry) were determined in blood from normal volunteers undergoing 24 h of basal glucose/basal insulin (BG/BI) clamps and high-glucose/high-insulin (HG/HI) clamps with and without SMS. Infusions of SMS under basal conditions (BG/BI) raised TF-PCA 1.8-fold (P < 0.03), TF-Ag 2.3-fold (P < 0.001), and TF expression on monocytes by 36% (P < 0.001) and decreased plasma levels of FVIIa by 30% (P < 0.001). Infusion of SMS reduced the 8.6-fold HG/HI-induced increase in TF-Ag by 26% and the 8.6-fold increase in TF-PCA by 100%. SMS also prevented the 60% increase in TF expression on monocytes, the 2.2-fold increase in F1.2, the 40% increase in CD40L expression on platelets, and the 17% increase in monocyte-platelet aggregates seen during HG/HI. We conclude that SMS completely prevented HG/HI-induced TF activation in normal volunteers and may be of use to reduce the procoagulant state and acute vascular events in hyperinsulinemic insulin-resistant patients with type 2 diabetes.
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PMID:Differential effects of somatostatin on circulating tissue factor procoagulant activity and protein. 1721 71

Tissue factor (TF) is the primary initiator of blood coagulation. Circulating TF procoagulant activity (TF-PCA) is associated with blood cells and microparticles and is elevated in patients with type 2 diabetes mellitus. Combined hyperinsulinemia and hyperglycemia and to a lesser degree selective hyperinsulinemia for 24 hours in healthy volunteers increased circulating TF-PCA, monocyte TF surface expression and mRNA, plasma thrombin generation, and coagulation factors VII and VIII activities, suggesting that the coagulation system had been activated. In addition, platelet CD40L and platelet-monocyte aggregates increased, indicating platelet activation. Somatostatin abolished these changes. We conclude that hyperinsulinemia, but particularly the combination of hyperinsulinemia and hyperglycemia, creates a prothrombotic state and may, in addition, be proinflammatory and proatherogenic by virtue of the actions of CD40L and TF.
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PMID:Effects of hyperglycemia and hyperinsulinemia on the tissue factor pathway of blood coagulation. 1754 39

We hypothesized that restenosis after coronary stenting is predicted by elevated levels of markers of thrombus formation and inflammation. Plasma levels of representative markers of inflammation, the thrombin and plasmin activation systems and adhesion molecules were measured in 59 patients with stable angina pectoris before, immediately after and 6 hours (h), 12 h, 24 h, one month and six months after elective stent implantation (radioactive phosphorus-32 stents/RSs/ n = 16, bare-metal stents/BMSs/ n = 43). All patients underwent clinical and angiographic follow-up (FUP) six months after stenting. RSs had significantly higher angiographic severity of restenosis than BMSs (47.1 +/- 20.1% vs. 27.6 +/- 22.0%, p = 0.003). Repeated measures ANOVA revealed significant differences between the BMS and RS groups as regards the increases in plasma levels of vascular cell adhesion molecule-1 (VCAM-1, p = 0.022), plasminogen activator inhibitor-1 (PAI-1, p = 0.047), tissue-type plasminogen activator (tPA, p = 0.047) and CD40 ligand (CD40L, p = 0.038). tPA levels tended to increase immediately after stenting in both groups, whereas the PAI-1 level one month after stenting was elevated significantly only in the RS group. In the RS group, the plasma levels of CD40L were increased at 24 h and six months after stenting, and the VCAM-1 level rose immediately after stenting and remained high during the FUP. Multivariate analysis on pooled laboratory data of both groups revealed elevated levels of VCAM-1 at 12 h and at six months as significant predictors of the severity of stent restenosis. In conclusion, the process of inflammation and thrombosis occurring after coronary interventions seems to be prolonged and enhanced in patients with high-grade restenosis at the follow up.
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PMID:Time course of prothrombotic and proinflammatory substance release after intracoronary stent implantation. 1839 32


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