bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effect of factor VIII on tissue factor-initiated spatial clot growth
Authors: Ovanesov Mikhail , Lopatina Elena , Ananyeva Natalya , Ul’yanova Ljudmila , Plyushch Olga , Butilin Andrey , Ataullakhanov Fazly , Saenko Evgueni 
Published: March 2018

SummaryUsing time-lapse videomicroscopy, we studied the role of coagulation factor VIII (fVIII) in tissue factor-initiated spatial clot growth on fibroblast monolayers in a thin layer of non-stirred recalcified plasma from healthy donors or patients with severe Haemophilia A. Analysis of temporal evolution of light-scattering profiles from a growing clot revealed existence of two phases in the clot growth-initiation phase in a narrow (0.2 mm) zone adjacent to activator surface and elongation phase in plasma volume. While the initiation phase did not differ in normal and haemophilic plasmas, the rate of clot growth in the elongation phase in haemophilic plasma constituted only 30% of that in normal plasma. Supplementation of haemophilic plasma with 0.05 U/ml fVIII restored the normal clot growth rate (44.9 ± 2.5 μm/min) at high but not at low fibroblast density. Our results indicate that the functioning of the intrinsic tenase complex is critical for normal spatial clot growth.
Deficiency of tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) impairs nutritionally induced obesity in mice
Authors: Demeulemeester Diego , Van Hoef Berthe , Collen Désiré , Maquoi Erik , Roger Lijnen H. 
Published: March 2018

SummaryTissue inhibitor of matrix metalloproteinase-1 deficient (TIMP-1–/–) mice and wild-type (TIMP-1+/+) controls were kept on a standard (SFD) or a high fat diet (HFD) for 15 weeks. At the time of sacrifice, TIMP-1–/– mice on HFD had a significantly lower body weight (29 ± 1.5 versus 41 ± 1.8 g, p <0.005), and significantly less subcutaneous (0.81 ± 0.19 versus 1.78 ± 0.21 g, p <0.05) and gonadal (0.87 ± 0.17 versus 1.85 ± 0.18 g, p <0.005) fat mass. These differences were much less pronounced for mice on SFD. On HFD but not on SFD, adipocyte diameters were significantly lower in the adipose tissue of TIMP-1–/– mice. Plasma leptin levels in TIMP-1–/– mice on HFD were significantly lower as compared to TIMP-1+/– mice, and strongly correlated with adipose tissue mass for both genotypes. Staining with an endothelial cell specific lectin revealed a significantly higher blood vessel density, larger stained area and vessel size in adipose tissue of TIMP-1–/– mice on HFD. This difference disappeared after normalization to the adipocyte number, suggesting that it does not represent a true enhancement of angiogenesis. Thus, in a murine model of nutritionally induced obesity, TIMP-1 promotes adipose tissue development.
In vivo pig models of venous thrombosis mimicking human disease
Authors: Kang Chantal , Bonneau Michel , Brouland Jean-Philippe , dit Sollier Claire Bal , Drouet Ludovic 
Published: March 2018

SummaryMost animal models of venous thrombosis involve acute thrombosis with hypercoagulability in small rodents. To better replicate human disease, we developed two models in the pig, a species similar to humans in size and in vascular and coagulation reactivity. One model involves de-endothelialisation with 50% or 80% stenosis and the other replacement of a venous segment by a Gore-Tex™ vascular prosthesis. Both models were tested with and without acute induced hypercoagulability (thromboplastin infusion). Thrombi obtained without thromboplastin infusion were composed of a multilayered platelet and a fibrin meshwork structure similar to that usually found in humans. With thromboplastin infusion, the thrombi were homogeneous fibrin structures imprisoning red blood cells. The high incidence of thrombosis obtained with the 80% stenosis model would be useful for studying anticoagulant treatments, whereas the low incidence with 50% stenosis would be useful for evaluating procoagulant effects of conditions or treatments. These new models shed further light on the development of venous thrombi under conditions similar to those seen in humans and may prove useful for investigating anticoagulant and procoagulant effects.
Differential regulation of P-selectin expression by protein kinase A and protein kinase G in thrombin-stimulated human platelets
Authors: Libersan Danielle , Rousseau Guy , Merhi Yahye 
Published: March 2018

SummaryP-selectin is rapidly translocated from platelet α-granules following activation. Intracellular cyclic AMP (cAMP) is a potent inhibitory pathway that results in global downregulation of platelet activation. While cAMP-dependent protein kinase (PKA) has long been considered as the main mediator of cAMP-dependent effects, no study has yet evaluated its effect on P-selectin expression in human platelets. Pretreatment of thrombin-stimulated platelets with forskolin resulted in a concentration-dependent inhibition of P-selectin expression that correlated with adenylyl cyclase activity. Inhibition of PKA with H-89 reversed cAMP-induced inhibition of P-selectin while cGMP-dependent protein kinase (PKG) inhibition with KT5823 significantly potentiated cAMP-dependent inhibition of P-selectin. Similar results were also observed in a platelet/neutrophil binding assay. In conclusion, cAMP-induced inhibition of P-selectin expression is, in large part, mediated through activation of PKA. PKG appears to be sollicited for P-selectin expression when cAMP levels are elevated which suggest a cAMP/PKG-dependent pathway of platelet activation.
A critical role of lipid rafts in the organization of a key FcγRIIa-mediated signaling pathway in human platelets
Authors: Bodin Stéphane , Viala Cécile , Ragab Ashraf , Payrastre Bernard 
Published: March 2018

SummaryThe involvement of platelet FcγRIIa in heparin-associated thrombocytopenia (HIT) is now well established. However, the precise sequence of molecular events initiated by FcγRIIa cross-linking in platelets remains partly characterized. We investigated here the role of lipid rafts in the spatio-temporal organization of the FcγRIIa-dependent signaling events. Upon cross-linking, FcγRIIa relocated in rafts where the kinase Lyn and the adapter LAT were among the major phosphotyrosyl proteins. Upon stimulation by HIT sera, the second messenger phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P3) accumulated in rafts in a P2Y12 adenosine diphosphate (ADP) recep- tor-dependent manner. PtdIns(3,4,5)P3 was then essential to specifically recruit phospholipase Cγ2 (PLCγ2) to these membrane microdomains. Controlled disruption of rafts by methyl γ-cyclodextrin reversibly abolished PtdIns(3,4,5)P3 production, PLC activation and platelet responses induced by FcγRIIa cross-linking without affecting the tyrosine phosphorylation events. This work demonstrates that platelet rafts are essential for the integration of a key signaling complex leading to the rapid production of PtdIns(3,4,5)P3 and in turn PLCγ2 activation during HIT.
Activation-dependent surface expression of gC1qR/p33 on human blood platelets
Authors: Murphy Tara , Ghebrehiwet Berhane , Peerschke Ellinor 
Published: March 2018

SummaryGC1qR/p33 (gC1qR) is expressed by a variety of somatic and cultured cells, including blood platelets. It interacts with several cellular, viral, bacterial, and plasma proteins, suggesting a potential role in thrombosis, inflammation, and infection. Considerable controversy has surrounded the surface membrane localization of gC1qR, however, since its cDNA sequence does not predict a traditional membrane-anchoring domain, and bears a typical mitochondrial targeting sequence. The present study examined gC1qR expression on resting and activated human blood platelets using flow cytometry and confocal microscopy with two monoclonal antibodies, 74.5.2 and 60.11, directed against gC1qR C-terminal amino acids 204-218, and N-terminal amino acids 76-93, respectively. Unstimulated platelets reacted minimally with either antibody. In contrast, platelet activation with TRAP, epinephrine, or ADP produced markedly increased gC1qR expression as reflected by 74.5.2 binding but not 60.11 binding. Platelet activation was verified using PAC-1 and anti CD 62 antibodies. Whereas PAC-1 binding to activated platelets could be reversed following platelet incubation with PGE1, 74.5.2 binding remained unchanged, suggesting the sustained expression of gC1qR following platelet stimulation. The data further demonstrate that detection of cell surface gC1qR may be dependent on antibody specificity. The ability of gC1qR to bind proteins involved in complement, coagulation, and kinin systems, as well as viral and bacterial pathogens including S. aureus protein A, supports the hypothesis that gC1qR expressed on activated platelets may contribute directly to thrombosis, inflammation, and endovascular infections.
Aging of stromal-derived human breast fibroblasts might contribute to breast cancer progression
Authors: Sieuwerts Anieta , Bolt-de Vries Joan , Bosma Peter , Swiggers Susan , Klijn Jan , Foekens John , Martens John 
Published: March 2018

SummaryAge is an important factor in the development and spread of breast cancer. Stromal cells also contribute to breast cancer growth and metastasis through the production of extracellular matrix (ECM) modifiers such as urokinase type plasminogen activator (uPA), its receptor (uPAR), its inhibitors (PAI-1 and PAI-2), matrix metalloproteinases (MMPs), and growth factors, including the fibroblast and insulin-like growth factors (FGF’s and IGF’s). In the present study we have investigated whether breast fibroblasts aged in vitro through passage in culture display altered levels of the plasminogen activator system and growth factors that are known to modulate that system.With real-time RT-PCR we found that during passage human breast fibroblasts, whether derived from the tumour burden or from matched adjacent normal breast tissue, exhibited a consistent increase in PAI-1 and FGF-1 and a decrease in MMP-2 mRNA expression. In addition, in 5 out of 7 fibroblast strains we observed an induction of uPA expression in combination with a reduced IGF-1 expression. Interestingly, while during aging MMP-2 protein increased in all tumour-derived fibroblast strains, these protein levels were reduced in all normal-tissue-derived fibroblasts. No other clear-cut age-dependent alterations were found in the all-together 25 factors investigated. We furthermore demonstrate in one tumour-derived fibroblast strain that the increases in uPA and PAI-1 mRNA and MMP-2 protein production are inversely related to the telomere length. Artificially increasing telomere length in this fibroblast strain by expressing human telomerase reverse transcriptase (hTERT) prevented senescence and resulted in late passage cultures with early passage uPA, PAI-1 and MMP-2 levels.Our results show that aging accompanied by telomere loss induces PAI-1 and FGF-1 mRNA expression in all breast fibroblast strains, increases uPA and decreases IGF-1 mRNA expression in a subset, and increases MMP-2 protein expression only in tumour-derived breast fibroblasts. These age-induced levels of PAI-1, FGF-1, uPA and MMP-2 in stromal breast fibroblast could contribute to breast cancer progression.
Suppression of argatroban-induced endogenous thrombolysis by PKSI-527, and antibodies to TPA and UPA, evaluated in a rat arterial thrombolysis model
Authors: Oiwa Kazuhiro , Matsuo Osamu , Ueshima Shigeru , Okada Kiyotaka , Okada Yoshio , Okamoto Shosuke , Giddings John , Yamamoto Junichiro , Hashimoto Masaru 
Published: March 2018

SummaryWe have previously confirmed, using a rat mesenteric arteriole thrombolysis model, that thrombin inhibition induces endogenous thrombolysis in vivo. In addition, we have shown that thrombin-activatable fibrinolysis inhibitor (TAFI) plays a role in the down regulation of endogenous thrombolysis. However, the mechanism of endogenous thrombolysis or spontaneous plasmin generation in vivo remains unclear.It has been shown in an in vitro system that plasma kallikrein activates pro-urokinase (pro uPA) and/or plasminogen, resulting in plasmin generation. These findings suggest that spontaneous fibrinolysis might be mediated by tPA and plasma kallikrein-dependent uPA. The aim of the present study was to examine whether these mechanisms play a dominant role in endogenous thrombolysis in vivo, using our rat mesenteric arterial thrombolysis model.Argatroban infusion enhanced endogenous thrombolysis. PKSI-527, anti uPA and anti tPA IgGs suppressed argatroban-induced thrombolysis. Also, the antibody IgG preparations suppressed endogenous thrombolysis in the absence of argatroban. In the presence of PKSI-527, anti tPA IgG was more effective than anti uPA IgG in suppressing argatroban-induced thrombolysis. The results suggested that both tPA and plasma kallikrein-mediated uPA activation and tPA release contribute to endogenous fibrinolytic or thrombolytic mechanisms.
Inhibition of PMN-elastase activity by semisynthetic glucan sulfates
Authors: Becker Markus , Franz Gerhard , Alban Susanna 
Published: March 2018

SummaryProteolysis of connective tissue by enzymes such as PMN-elastase (PMNE) is a crucial step during inflammation and metastasis. Semisynthetic sulfated carbohydrates (SC) were shown to exhibit potent antiinflammatory and antimetastatic activity in vivo. The aim of the present study was to examine whether interferences with PMN-elastase may contribute to these effects. Therefore, the interactions of these compounds with PMNE were evaluated in various test systems. Besides semisynthetic α-1,4/1,6- and β-1,3-glucan sulfates, UFH, a LMWH and pentosan polysulfate (PPS) were included in the study. The inhibitory activity of SC improves not only with increasing molecular weight (MW 10 – 250 kDa: 37 – 54% inhibition at 0.25 µg/ml) and degree of sulfation (DS 0.25 - 2.0: 16 - 50% inhibition at 0.25 µg/ml), but depends also on their genuine polysaccharide structure (IC50 β-1,3-glucan sulfate 0.18 / α-1,4/1,6-glucan sulfate 0.25 / UFH 0.5 µg/ml). Using physiological substrate assays (collagen, elastin), β-1,3- and α-1,4/1,6-glucan sulfates are more active than UFH (inhibition at 1.5 µg/ml: 41 / 32 / 12%). According to enzyme-inhibitor binding studies, SC exhibit structure dependent affinity to the enzyme (Kd for PMNE: β-1,3 < α-1,4/1,6 < UFH). Finally, SC were shown to inhibit cancer cell-mediated elastinolysis.
Rapid genotyping of haemostatic gene polymorphisms using the 5´ nuclease assay
Authors: Jood Katarina , Ladenvall Per , Tjärnlund Anna , Andersson Maria , Jern Christina 
Published: March 2018

SummaryHemostatic gene polymorphisms have been shown to be associated with arterial and venous thrombotic disease. To date these polymorphisms have mainly been detected by labor-intensive conventional gel based methods. Aim of the present study was to design and optimize high throughput 5 nuclease assays for the detection of a set of 10 single-nucleotide polymorphisms (SNP) in genes of importance for hemostasis: plasminogen activator inhibitor type 1 -675 4G>5G, thrombin activatable fibrinolysis inhibitor Ala147Thr and 1,542C>G, β-fibrinogen -455G>A, von Willebrand factor -1,051A>G, factor VII Arg353Gln, factor XIII Val34Leu, prothrombin 20,210G>A, tissue factor pathway inhibitor -287T>C, and methylenetetrahydrofolate reductase 1,298A>C. Specificity of each genotyping assay was confirmed by sequence-based typing and reproducibility was evaluated by repeated genotyping. The genotyping protocols presented here may serve as a valuable tool for clinical researchers interested in exploring associations between these SNPs and thrombotic disease.
Accelerated thrombolysis in a rabbit model of carotid artery thrombosis with liposome-encapsulated and microencapsulated streptokinase
Authors: Leach J. Kent , Patterson Eugene , Miao Yiwei , Johnson Arthur , O’Rear Edgar 
Published: March 2018

SummaryThe present study compares the efficacy of two formulations of encapsulated streptokinase to streptokinase in a rabbit model of carotid artery thrombosis. Arterial thrombosis followed the injection of thrombin mixed with autologous whole blood into a carotid artery of New Zealand white rabbits. Thirty minutes after the confirmation of an occlusive thrombus, one of four streptokinase formulations was infused at a dosage of 6,000 IU/kg into the jugular vein. Free streptokinase (FREE SK) was compared to identical dosages of streptokinase encapsulated in a liposome (LESK), streptokinase entrapped in a water-soluble polymer (MESK), and free streptokinase admixed with blank microparticles (FREE SK + BLANK). Carotid arterial blood flow was determined by pulsed Doppler flowmetry to confirm clot formation and reperfusion. Two hours after drug infusion, the rabbits were killed and the residual thrombus mass was determined.Compared to FREE SK (74.5 ± 16.9 min; mean ± SEM), LESK demonstrated significantly reduced reperfusion times (19.3 ± 4.6 min) while MESK exhibited even greater improvement (7.3 ± 1.6 min). FREE SK + BLANK showed no statistical improvement versus FREE SK. LESK and MESK also resulted in reduced residual clot mass and greater return of arterial blood flow. These studies suggest that encapsulation of streptokinase offers a potential method of improved fibrinolytic treatment for patients with clot-based disorders. MESK performed slightly better than LESK with improved production and storage characteristics.
Impact of pre-treatment INR level on the effect of intravenous low dose vitamin K in patients with excessive anticoagulation
Authors: Shasha Yigal , Olchovsky David , Loebstein Ronen , Halkin Hillel , Ezra David , Lubetsky Aharon 
Published: March 2018

SummaryAdministration of vitamin K is the common mode of treatment in excessively anticoagulated patients. However, patient’s response to vitamin K varies, depending on the vitamin K dose and the route of administration. Another potential source of variation is the pre-treatment INR which has not been accounted for in most previous studies. In the present study the effect of baseline INR on the response to a single dose of intravenous vitamin K (0.5 mg) was studied in 95 episodes of excessively anticoagulated patients (n = 76).In 67 episodes of moderately excessive baseline INR (6-10) mean INR declined from 8.0 ± 1.2 to 2.6 ± 0.9 at 24 hours, 59/67 (88%) responding within the first 12 hours and not requiring a second dose. In contrast, in 28 episodes with highly excessive baseline INR (> 10) response was slower; mean INR declining from 13.6 ± 2.7 to 4.0 ± 2.1 at 24 hours. In 14/28 of these episodes, patients failed to respond to vitamin K in the first 12 hours and required a second vitamin K dose.We conclude that INR at presentation affects the response to vitamin K and that this INR value should be considered in determining appropriate vitamin K doses.
Fibrinolytic Proteins and Progression of Coronary Artery Disease in Relation to Gemfibrozil Therapy
Authors: Syvänne Mikko , Silveira Angela , Luong Le-Anh , Nieminen Markku , Humphries Steve , Frick M. , Taskinen Marja-Riitta , Hamsten Anders 
Published: March 2018

SummaryImpaired fibrinolytic function, mainly due to increased plasma plasminogen activator inhibitor-1 (PAI-1) activity, is common in patients with manifest coronary artery disease (CAD) and a predictor of recurrent cardiovascular events. We investigated the relationships of plasma tissue-type plasminogen activator (tPA) and PAI-1 antigen levels, plasma PAI-1 activity and PAI 4/5-guanosine (4G/5G) genotype to CAD progression in 203 middle-aged men participating in the Lopid Coronary Angiography Trial (LOCAT).A higher tPA antigen concentration, whether baseline or on-trial, was associated with a more severe global angiographic response (p < 0.05), an association mainly accounted for by progression of diffuse lesions in graft-affected segments (change in per-patient means of average diameters of segments haemodynamically related to bypass grafts). Plasma PAI-1 activity and mass concentration and 4G/5G PAI-1 genotype were unrelated to angiographic outcome measurements. tPA and PAI-1 antigen increased significantly in the gemfibrozil group (+11.3% and + 16.4%, respectively, p < 0.001), whereas there was no treatment effect on PAI-1 activity (median change 0.0%).It is concluded that fibrinolytic function does not substantially influence progression of CAD as assessed by angiography in middle-aged men. Furthermore, pronounced long-term lowering of serum triglycerides by gemfibrozil treatment does not significantly affect the plasma PAI-1 activity level but increases the plasma tPA and PAI-1 antigen concentrations.
Prospective Analysis after Coronary-artery Bypass Grafting: Platelet GP IIIa Polymorphism (HPA-1b /PlA2) Is a Risk Factor for Bypass Occlusion, Myocardial Infarction, and Death
Authors: Klein Michael , Dauben Hans , Moser Christiane , Gams Emmeran , Scharf Rüdiger , Zotz Rainer 
Published: March 2018

SummaryRecently, we have demonstrated that human platelet antigen 1b (HPA-1b or PlA2) is a hereditary risk factor for platelet thrombogenicity leading to premature myocardial infarction in preexisting coronary artery disease. However, HPA-1b does not represent a risk factor for coronary artery disease itself. The aim of our present study was to evaluate the role of HPA-1b on the outcome in patients after coronaryartery bypass surgery. We prospectively determined the HPA-1 genotype in 261 consecutive patients prior to saphenous-vein coronaryartery bypass grafting. The patients were followed for one year. Among patients with bypass occlusion, myocardial infarction, or death more than 30 days after surgery, the prevalence of HPA-1b was significantly higher than among patients without postoperative complications (60 percent, 6/10, vs. 24 percent, 58/241, p <0.05, odds ratio 4.7). Using a stepwise logistic regression analysis with the variables HPA1b, age, sex, body mass index, smoking (pack-years), hypertension, diabetes, cholesterol and triglyceride concentration, only HPA-1b had a significant association with bypass occlusion, myocardial infarction, or death after bypass surgery (p = 0.019, odds ratio 4.7). This study shows that HPA-1b is a hereditary risk factor for bypass occlusion, myocardial infarction, or death in patients after coronary-artery bypass surgery.
A Novel Fibrinogen-clotting Enzyme, TL-BJ, from the Venom of the Snake
Bothrops jararaca:
Purification and Characterization
Authors: Sampaio Claudio , Mentele Reinhard , Camargo Antonio , Fink Edwin , Serrano Solange 
Published: March 2018

SummaryThree chromatographically distinct forms of a novel fibrinogenclotting serine endopeptidase, TL-BJ 1, 2 and 3, were purified from the venom of Bothrops jararaca by a combination of ammonium sulfate precipitation and chromatographic steps. The three forms of TL-BJ have similar amidolytic and plasma coagulating activities. TL-BJ 1, TL-BJ 2 and TL-BJ 3 cause the specific clotting of fibrinogen with release of fibrinopeptide A, the specific activities are 16.8 NIH U/mg (TL-BJ 1), 16.7 NIH U/mg (TL-BJ 2) and 20.8 NIH U/mg (TL-BJ 3). The most sensitive chromogenic substrates for measuring the amidolytic activity of TL-BJ 3 were D-Pro-Phe-Arg-pNA, D-Phe-pipecolyl-ArgpNA and Z-D-Arg-Gly-Arg-pNA. The amidolytic and coagulant activities of TL-BJ were inhibited by phenylmethylsulfonyl fluoride but not by hirudin. Benzamidine derivatives, which are competitive inhibitors of trypsin-like serine endopeptidases, also inhibited the amidolytic activity of TL-BJ. In SDS/PAGE the main bands of TL-BJ 1, 2 and 3 showed molecular masses of 30 kDa, 31 kDa and 32 kDa. Upon incubation with N-glycosidase F only TL-BJ 3 remained unchanged, whereas TL-BJ 1 and TL-BJ 2 showed products with molecular masses around 23 kDa. Thus, TL-BJ 3 does not seem to be N-glycosylated. The N-terminal amino acid sequences of TL-BJ 2 and TL-BJ 3 are identical while TL-BJ 1 has five substitutions.