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)

We report functional differences in constitutive and agonist-mediated endothelial barrier function between cultured primary and Clonetics human umbilical vein endothelial cells (pHUVEC and cHUVEC) grown in soluble growth factors and heparin. Basal transendothelial resistance (TER) was much lower in pHUVEC than in cHUVEC grown in medium supplemented with growth factors, such as basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), and human epithelial growth factor (EGF), and heparin. On the basis of a numerical model of TER, the increased basal TER in cHUVEC was due to effects on cell-matrix adhesion and membrane capacitance. Heparin and bFGF increased constitutive TER in cultured pHUVEC, and heparin mediated additional increases in constitutive TER in pHUVEC supplemented with bFGF. EGF attenuated bFGF-mediated increases in TER. On the basis of the numerical model, in contrast to cHUVEC, heparin and bFGF augmented TER through effects on cell-cell adhesion and membrane capacitance in pHUVEC. Thrombin mediated quantitatively greater amplitude and a more sustained decline in TER in cultured cHUVEC than pHUVEC. Thrombin-mediated barrier dysfunction was attenuated in pHUVEC conditioned in EGF in the presence or absence of heparin. Thrombin-mediated barrier dysfunction was also attenuated when monolayers were exposed to low concentrations of heparin and further attenuated in the presence of bFGF. cAMP stimulation mediated differential attenuation of thrombin-mediated barrier dysfunction between pHUVEC and cHUVEC. VEGF displayed differential effects in TER in serum-free medium. Taken together, these data demonstrate marked differential regulation of constitutive and agonist-mediated endothelial barrier function in response to mitogens and heparin stimulation.
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PMID:Growth factor- and heparin-dependent regulation of constitutive and agonist-mediated human endothelial barrier function. 1667 98

1. Fibrin gel has been used as a carrier of angiogenic molecules to promote neovascularization in animal models of limb ischaemia. However, little is known about the effects of fibrin itself under such pathological conditions. Accordingly, the present study tested the efficacy of fibrin in a rabbit model of acute hindlimb ischaemia. 2. Unilateral ischaemia was induced by resection of the left femoral artery. Seven days after surgery, fibrin particles (FP), which were free of fibrinogen, thrombin and vascular endothelial growth factor, were injected directly into the ischaemic thigh muscles. Twenty-four rabbits were divided into four groups, namely a control group receiving phosphate-buffered saline and three FP-treated groups receiving 5, 10 or 20 mg FP. 3. Collateral vessel development and limb perfusion were assessed by angiography, measuring the calf blood pressure ratio (BPR), thermographic scanning and the histological determination of capillary density. 4. At day 35 post-surgery, the treatment with 5 mg FP produced an augmentation of collateral vessel development (P < 0.01), increased numbers of capillaries (P < 0.05) and improved perfusion manifested by a higher blood flow (P < 0.01) and calf BPR (P < 0.05) compared with controls. Treatment with 10 and 20 mg FP had similar effects to those observed with 5 mg FP. 5. The present study reveals that FP promotes angiogenesis in a rabbit model of hindlimb ischaemia, thus providing a feasible approach to therapeutic angiogenesis in ischaemic diseases.
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PMID:Therapeutic angiogenesis by intramuscular injection of fibrin particles into ischaemic hindlimbs. 1678 29

Evidence that platelets play a role in tumor metastasis includes the observation of circulating tumor cell-platelet aggregates and the anti-metastatic effect of thrombocytopenia and anti-platelet drugs. Platelets have recently been shown to contain vascular endothelial growth factor (VEGF) which is released during clotting. We therefore studied the effects of (1) tumor cell-platelet adherence and tumor cell TF activity on platelet VEGF release; and (2) the effects of GpIIb/IIIa blockade on tumor cell-induced platelet VEGF release, tumor cell-induced thrombocytopenia and experimental metastasis. Adherent A375 human melanoma cells (TF+) and KG1 myeloid leukemia (TF-) cells were cultured in RPMI containing 10% fetal bovine serum. Platelet-rich plasma was obtained from normal citrated whole blood and the presence of VEGF (34 and 44 kDa isoforms) confirmed by immunoblotting. Platelet-rich plasma with or without anti-GpIIb/IIIa (Abciximab) was added to A375 monolayers and supernatant VEGF measured by ELISA. Tumor cell-induced platelet activation and release were determined by CD62P expression and serotonin release respectively. In vitro, tumor cell-platelet adherence was evaluated by flow cytometry. In vivo, thrombocytopenia and lung seeding were assessed 30 min and 18 days, respectively, after i.v. injection of Lewis Lung carcinoma (LL2) cells into control or murine 7E3 F(ab')(2) (6 mg/ kg) athymic rats. Maximal in vitro platelet activation (72% serotonin release) occurred 30 min after adding platelets to tumor cells. At this time, 87% of the A375 cells had adhered to platelets. Abciximab significantly (P<0.05) reduced platelet adherence to tumor cells as evidenced by flow cytometry. Incubation of A375 cells with platelets induced VEGF release in a time-dependent manner. This release was significantly inhibited by Abciximab (81% at 30 min; P<0.05). In the presence of fibrinogen and FII, VEGF release induced by A375 (TF+) cells was significantly higher than that induced by KG1 (TF-) cells (105.5+/-24 vs. 42+/-7 pg/ml; P<0.001). Omitting fibrinogen or FII from the reaction mixture markedly decreased VEGF release. In vivo, GpIIb/IIIa blockade with murine 7E3 F(ab')(2) reduced LL2 tumor cell-induced thrombocytopenia by 90% (P<0.001) and lung seeding by 82% (P<0.05). We conclude that TF-bearing tumor cells can activate platelets largely via thrombin generation, and that such activation is associated with release of VEGF. This may enhance metastasis, possibly by increasing extravasation at points of adhesion to vascular endothelium.
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PMID:Blockade of GpIIb/IIIa inhibits the release of vascular endothelial growth factor (VEGF) from tumor cell-activated platelets and experimental metastasis. 1680 Nov 4

Data are emerging to support the idea that mediators of angiogenesis are found in the Alzheimer's disease (AD) brain. The objective of this study is to compare the expression of the angiogenic mediators vascular endothelial growth factor (VEGF), angiopoietin, and matrix metalloproteinases (MMPs) in the microcirculation of AD patients and age-matched controls. Our results indicate that angiopoietin-2 and VEGF are expressed by AD- but not control-derived microvessels. AD-derived microvessels also release higher levels of MMP-2 and MMP-9 compared to controls. The data show that despite high levels of MMP-9, assessed by western blot, MMP-9 activity is not detectable in AD microvessels. In this regard we find high levels of the tissue inhibitor of matrix metalloproteinases-1 (TIMP-1) in AD, but not control vessels. Furthermore, we explore the ability of thrombin, previously shown to be present in AD microvessels, to affect TIMP expression in cultured brain endothelial cells and find that thrombin causes up regulation of TIMP-1. These data show that angiogenic changes occur in the microcirculation of the AD brain and suggest that if these changes are contributory to disease pathogenesis, targeting the abnormal brain endothelial cell would provide a novel therapeutic approach for the treatment of this disease.
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PMID:Angiogenic proteins are expressed by brain blood vessels in Alzheimer's disease. 1698 87

Increased lung vascular permeability is an important contributor to respiratory failure in acute lung injury (ALI). We found that a function-blocking antibody against the integrin alphavbeta5 prevented development of lung vascular permeability in two different models of ALI: ischemia-reperfusion in rats (mediated by vascular endothelial growth factor [VEGF]) and ventilation-induced lung injury (VILI) in mice (mediated, at least in part, by transforming growth factor-beta [TGF-beta]). Knockout mice homozygous for a null mutation of the integrin beta5 subunit were also protected from lung vascular permeability in VILI. In pulmonary endothelial cells, both the genetic absence and blocking of alphavbeta5 prevented increases in monolayer permeability induced by VEGF, TGF-beta, and thrombin. Furthermore, actin stress fiber formation induced by each of these agonists was attenuated by blocking alphavbeta5, suggesting that alphavbeta5 regulates induced pulmonary endothelial permeability by facilitating interactions with the actin cytoskeleton. These results identify integrin alphavbeta5 as a central regulator of increased pulmonary vascular permeability and a potentially attractive therapeutic target in ALI.
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PMID:Integrin alphavbeta5 regulates lung vascular permeability and pulmonary endothelial barrier function. 1707 79

A methodology for the detection of protein biomarkers at picomolar concentrations that utilizes surface plasmon resonance imaging (SPRI) measurements of RNA aptamer microarrays is developed. The adsorption of proteins onto the RNA microarray is detected by the formation of a surface aptamer-protein-antibody complex. The SPRI response signal is then amplified using a localized precipitation reaction catalyzed by the enzyme horseradish peroxidase that is conjugated to the antibody. This enzymatically amplified SPRI methodology is first characterized by the detection of human thrombin at a concentration of 500 fM; the appropriate thrombin aptamer for the sandwich assay is identified from a microarray of three potential thrombin aptamer candidates. The SPRI method is then used to detect the protein vascular endothelial growth factor (VEGF) at a biologically relevant concentration of 1 pM. VEGF is a signaling protein that has been used as a serum biomarker for rheumatoid arthritis, breast cancer, lung cancer, and colorectal cancer and is also associated with age-related macular degeneration.
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PMID:Detection of protein biomarkers using RNA aptamer microarrays and enzymatically amplified surface plasmon resonance imaging. 1726 39

Preeclampsia (PE), intrauterine growth restriction (IUGR) and abruption with or without fetal loss are associated with reduced uteroplacental blood flow, decidual vasculopathy, endothelial cell dysfunction, thrombosis, inflammation and hemorrhage. Our hypothesis is that reduced uteroplacental blood flow causes focal decidual hypoxia that generates vascular endothelial growth factor (VEGF). The latter acts directly on decidual endothelial cells to induce aberrant expression of tissue factor (TF), the primary initiator of coagulation. This in turn generates thrombin that induces: i) further TF expression; and ii) inflammatory cytokines. Both VEGF and TF induce aberrant angiogenesis-vessel maintenance reflected by endothelial cell fenestrations and induction of a prothrombotic surface causing both the decidual hemorrhage (i.e. abruption) and thrombosis (i.e. uteroplacental vascular insufficiency) observed in these adverse pregnancy outcomes. This novel hypothesis is supported by our finding of TF expression in decidual endothelium of pregnancies complicated by IUGR and/or fetal loss. Moreover, treatment of cultured endometrial endothelial cells with VEGF or thrombin induces TF protein and mRNA expression. Quantitative RT-PCR analysis indicates that thrombin enhances (>10-fold) the output of diverse inflammatory cytokines in these cultures. The greatest effect (>2-log) was seen on macrophage inflammatory protein 3alpha (MIP3alpha). In vitro, thrombin results in endometrial endothelial cell aggregations and changes in the apoptotic pathway. Thus, we postulate that reductions in uteroplacental flow initiate a cascade of molecular effects leading to hypoxia, thrombosis, inflammation, and endothelial cell dysfunction resulting in untoward pregnancy outcomes.
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PMID:Thrombin activation of endometrial endothelial cells: a possible role in intrauterine growth restriction. 1726 54

This study was conducted to compare the effects of atorvastatin plus aspirin combined therapy on inflammatory responses, endothelial cell function, and blood coagulation system in patients undergoing coronary artery bypass grafting (CABG) to aspirin monotherapy. The patients were randomized into atorvastatin plus aspirin combined therapy group and aspirin monotherapy group. Reduced total cholesterol in the combined therapy group was found in a short term of medication for 14 days. On postoperative day (POD)-14, inhibitory effects of the combined therapy on whole blood aggregation as well as platelet activation assessed by flow cytometry were stronger than those of the monotherapy. Furthermore, cytokine, cytokine receptors, c-reactive protein and alpha1-acid glycoprotein in the combined therapy group were down-regulated on POD-14. At the same time, circulating levels of thromboxane A(2), vascular endothelial growth factor and thrombin-antithrombin III complex as well as P-selectin, L-selectin and intercellular adhesion molecule-1 were down-regulated, while E-selectin and transforming growth factor-beta1 was up-regulated. Atorvastatin plus aspirin combined therapy may improve inflammatory responses, accelerated platelet function, vascular endothelial cell function, blood coagulation system at the early stage such as 14th day after CABG. In conclusion, atorvastatin and aspirin combined therapy may bring beneficial effects to the patient after CABG.
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PMID:Effects of atorvastatin and aspirin combined therapy on inflammatory responses in patients undergoing coronary artery bypass grafting. 1730 Sep 51

Although fracture healing is a well-optimized biological process that leads to healing, approximately 10-20% of fractures result in impaired or delayed healing and these fractures may benefit from the use of biotechnologies to enhance skeletal repair. Peptide signaling molecules such as the bone morphogenetic proteins have been shown to stimulate the healing of fresh fractures, nonunions, and spinal fusions and side effects from their use appear to be minimal. Other growth factors currently being studied for local application include growth and differentiation factor-5 (GDF-5), vascular endothelial growth factor (VEGF), transforming growth factor beta (TGFbeta), and platelet-derived growth factor (PDGF). Molecules such as prostaglandin E receptor agonists and the thrombin-related peptide, TP508, have shown promise in animal models of fracture repair. Gene therapy using various growth factors or combinations of factors might also aid in fracture repair, particularly as new methods for delivery that do not require viral vectors are developed. Systemic therapy with agents such as parathyroid hormone (PTH), growth hormone (GH), and the HMG-CoA reductase inhibitors are also under investigation. As these and other technologies are shown to be safe and effective, their use will become a part of the standard of care in managing skeletal injuries.
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PMID:New technologies for the enhancement of skeletal repair. 1738 86

This study was purposed to investigate the effects of platelet-derived membrane microparticles (PMP) on the proliferation and apoptosis of human umbilical vein endothelial cells (HUVEC). Different concentrations of thrombin were adopted to activate the platelets so as to release PMPs. Flow cytometry (FCM) was adopted to evaluate the efficiencies of different concentrations of thrombin to release PMPs. By using the HUVEC cultivated in vitro as vector, the effects of PMPs on the proliferation and apoptosis of HUVEC were investigated by MTT and FCM. The results showed that the efficiencies releasing PMPs from platelets activated by 2.0, 1.5, 1.0, 0.5 U/ml thrombin were 28.7, 47.7, 50.1 and 43.9% respectively; PMPs induced proliferation of HUVEC in a dose dependent manner. At the concentration of 40 microg/ml PMPs, the proliferation rate of HUVEC was 1.8 +/- 0.3 times as much as blank control, the proliferation rate in group of vascular endothelial growth factor was 1.9 +/- 0.5 times of as much as blank control, there was no statistic difference (p > 0.05). The PMPs inhibited HUVEC apoptosis. Compared with the apoptosis rate of control (9.4 +/- 0.5)%, apoptosis rate in PMP group (40 microg/ml) was (3.9 +/- 0.4)% (p < 0.05). The addition of VEGF (10 microl/ml) did not successfully prevented apoptosis of HUVEC with apoptosis rate of (8.0 +/- 0.8)%. It is concluded that platelets activated by 1 U/ml thrombin gets the best efficiency of PMP release, which stimulates proliferation of HUVEC and inhibits its apoptosis.
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PMID:[Effects of platelet-derived membrane microparticles on the proliferation and apoptosis of human umbilical vein endothelial cells]. 1770 19


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