bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mouse Polyomavirus Enters Early Endosomes, Requires Their Acidic pH for Productive Infection, and Meets Transferrin Cargo in Rab11-Positive Endosomes
Authors: Liebl David , Difato Francesco , Horníková Lenka , Mannová Petra , Štokrová Jitka , Forstová Jitka 
Published: April 2006

ABSTRACT

Mouse polyomavirus (PyV) virions enter cells by internalization into smooth monopinocytic vesicles, which fuse under the cell membrane with larger endosomes. Caveolin-1 was detected on monopinocytic vesicles carrying PyV particles in mouse fibroblasts and epithelial cells (33). Here, we show that PyV can be efficiently internalized by Jurkat cells, which do not express caveolin-1 and lack caveolae, and that overexpression of a caveolin-1 dominant-negative mutant in mouse epithelial cells does not prevent their productive infection. Strong colocalization of VP1 with early endosome antigen 1 (EEA1) and of EEA1 with caveolin-1 in mouse fibroblasts and epithelial cells suggests that the monopinocytic vesicles carrying the virus (and vesicles containing caveolin-1) fuse with EEA1-positive early endosomes. In contrast to SV40, PyV infection is dependent on the acidic pH of endosomes. Bafilomycin A1 abolished PyV infection, and an increase in endosomal pH by NH
4
Cl markedly reduced its efficiency when drugs were applied during virion transport towards the cell nucleus. The block of acidification resulted in the retention of a fraction of virions in early endosomes. To monitor further trafficking of PyV, we used fluorescent resonance energy transfer (FRET) to determine mutual localization of PyV VP1 with transferrin and Rab11 GTPase at a 2- to 10-nm resolution. Positive FRET between PyV VP1 and transferrin cargo and between PyV VP1 and Rab11 suggests that during later times postinfection (1.5 to 3 h), the virus meets up with transferrin in the Rab11-positive recycling endosome. These results point to a convergence of the virus and the cargo internalized by different pathways in common transitional compartments.
Regulated Cleavages at the West Nile Virus NS4A-2K-NS4B Junctions Play a Major Role in Rearranging Cytoplasmic Membranes and Golgi Trafficking of the NS4A Protein
Authors: Roosendaal Jojanneke , Westaway Edwin G. , Khromykh Alexander , Mackenzie Jason M. 
Published: April 2006

ABSTRACT
A common feature associated with the replication of most RNA viruses is the formation of a unique membrane environment encapsulating the viral replication complex. For their part, flaviviruses are no exception, whereupon infection causes a dramatic rearrangement and induction of unique membrane structures within the cytoplasm of infected cells. These virus-induced membranes, termed paracrystalline arrays, convoluted membranes, and vesicle packets, all appear to have specific functions during replication and are derived from different organelles within the host cell. The aim of this study was to identify which protein(s) specified by the Australian strain of West Nile virus, Kunjin virus (KUNV), are responsible for the dramatic membrane alterations observed during infection. Thus, we have shown using immunolabeling of ultrathin cryosections of transfected cells that expression of the KUNV polyprotein intermediates NS4A-4B and NS2B-3-4A, as well as that of individual NS4A proteins with and without the C-terminal transmembrane domain 2K, resulted in different degrees of rearrangement of cytoplasmic membranes. The formation of the membrane structures characteristic for virus infection required coexpression of an NS4A-NS4B cassette with the viral protease NS2B-3pro which was shown to be essential for the release of the individual NS4A and NS4B proteins. Individual expression of NS4A protein retaining the C-terminal transmembrane domain 2K resulted in the induction of membrane rearrangements most resembling virus-induced structures, while removal of the 2K domain led to a less profound membrane rearrangement but resulted in the redistribution of the NS4A protein to the Golgi apparatus. The results show that cleavage of the KUNV polyprotein NS4A-4B by the viral protease is the key initiation event in the induction of membrane rearrangement and that the NS4A protein intermediate containing the uncleaved C-terminal transmembrane domain plays an essential role in these membrane rearrangements.
Generation of Infectious Hepatitis C Virus in Immortalized Human Hepatocytes
Authors: Kanda Tatsuo , Basu Arnab , Steele Robert , Wakita Takaji , Ryerse Jan S. , Ray Ranjit , Ray Ratna B. 
Published: April 2006

ABSTRACT

Progress in understanding hepatitis C virus (HCV) biology has remained a challenge due to the lack of an efficient cell culture system for virus growth. In this study, we examined HCV core protein-mediated immortalized human hepatocytes (IHH) for growth of HCV. In vitro-transcribed full-length RNA from HCV genotype 1a (clone H77) was introduced into IHH by electroporation. Reverse transcription-PCR of cellular RNA isolated from HCV genome-transfected IHH suggested that viral RNA replication occurred. IHH transfected with the full-length HCV genome also displayed viral protein expression by indirect immunofluorescence. In contrast, cells transfected with polymerase-defective HCV (H77/GND) RNA as a negative control did not exhibit expression of the viral genome. Immunogold labeling demonstrated localization of E1 protein in the rough endoplasmic reticulum of RNA-transfected IHH. Virus-like particles of ∼50 nm were observed in the cytoplasm. After being inoculated with culture media of cells transfected with the full-length HCV genome, naïve IHH displayed NS5a protein expression in a dilution-dependent manner, but expression of NS5a was inhibited by prior incubation of culture medium with HCV-infected patient sera. NS5a-positive immunofluorescence of cell culture media of IHH transfected with full-length H77 RNA yielded ∼4.5 × 10
4
to 1 × 10
5
focus-forming units/ml. A similar level of virus growth was observed upon transfection of RNA from HCV genotype 2a (JFH1) into IHH. Taken together, our results suggest that IHH support HCV genome replication and virus assembly.
DECISION TRAPS AND COMPETENCE DYNAMICS IN CHANGEABLE SPACES
Authors: YU P. L. , CHIANGLIN C. Y. 
Published: April 2006

There are many parameters in challenging decision problems, including the alternatives, the criteria, resources, the perception of decision problems, decision makers and their psychological states, information inputs from the environment, and self-suggestion, etc. At any moment of time, some of these parameters can catch our attention, called alerted parameters; some cannot, called unalerted parameters. Some parameters are visible, some are invisible. In addition, the parameters themselves can vary over certain ranges or domains. All of these make challenging decision problems very complex. We call this kind of problems as decision problems with changeable spaces (parameters). We may focus on certain parameters with certain assumed values to find an "optimal" solution, which may lead to solve wrong problem with bad solution. Quite often, our focus may be just a small part of what we know, or just a part of what we are most familiar with. We may often neglect what we are not familiar with, and pay no attention to what we do not know. As a consequence, we may see just a small part of the problem domain (including all parameters and their possible variations over time). The portion (of the problem domain) that we cannot see is our decision blind. Suppose our alerted domain (those parameters and their variations that are currently under our consideration) to be fixed in only a small part of the problem domains. Then very likely we could end up with a serious mistake. This situation is known as decision trap. In this article, we will introduce a systematic scheme, based on habitual domain theory, to help us reduce decision blinds and avoid decision traps so that we could make decision with good quality. Then we will also introduce the concept of competence set analysis to help us cope with challenging decision problems. This including: (i) how to effectively expand our competence (resources, skill, know-how, information, ideas, effort, etc.) as to solve a given problem effectively; and (ii) given a set of competence, how to maximize its value by solving a set of value added problems. Furthermore, we will introduce innovation dynamics which describe the dynamics of how to solve a set of problems with our existent or acquired competence (to relieve the pains or frustrations of "certain customers or decision makers" at certain situations) as to create value, and how to distribute this created value so that we can continuously expand out competence set to solve more challenging problems and create more value.
The influence of fibrin polymerization and platelet-mediated contractile forces on citrated whole blood thromboelastography profile
Authors: Chakroun Tahar , Gerotziafas Grigoris , Seghatchian Jerhad , Samama Meyer , Hatmi Mohamed , Elalamy Ismail 
Published: April 2006

SummaryThromboelastography analysis providing a global assessment of coagulation is gaining new interest in clinical practice. Minimal TF triggered whole blood thromboelastography provides a valuable tool for studying the kinetics of clot formation (expressed by the parameters R, K and α-angle) and the physical characteristics of the clot, such as its firmness and the elastic modulus shear (expressed by the parameters maximal amplitude MA and G). We studied the influence of fibrin polymerization and platelet functional status on each parameter of thromboelastographic trace obtained by minimal TF activation in WB by employing increasing concentrations of a fibrin polymerization inhibitors (the tetrapeptide Gly-Pro-Arg-Pro-OH. AcOH; Pefabloc-FG®) and an inhibitor of actin polymerization (Cytochalasin D). Pefabloc-FG® at concentrations higher than 5 mg/ml prolonged the R and K times and decreased the α-angle in a concentration-dependent manner but it did not modify MA and G parameters. At the concentration of 5 mg/ml, Pefabloc-FG® completely inhibited clot formation. Cytochalasin D had no effect on R time but decreased the α-angle, MA and G parameters by reachinga plateau at the concentration of 5 µM. The effect of cytochalasin D was more pronounced on MA and G than on the α-angle. A combination of both Pefabloc-FG® (0.5 mg/ml) and cytochalasin D (50 µM) significantly decreased α-angle compared to control as well as their single effect. However, G value was dramatically reduced in the presence of cytochalasin D exposure, without any additional effect when both inhibitors were combined. This study confirms the importance of fibrin polimerization on the kinetics of thrombus formation and demonstrates the close association between the quality of the thrombus and the functional status of platelets. Normal platelet contractile forces are of major importance for the maximum amplitude of TEG which is related to the strength and elastic modulus of the thrombus.
Platelets enhance CD4+ lymphocyte adhesion to extracellular matrix under flow conditions: Role of platelet aggregation, integrins, and non-integrin receptors
Authors: Vitkovsky Yuri , Brill Grigory , Koltakov Alexander , Farzam Nahid , Varon David , Bank Ilan , Solpov Alexey , Shenkman Boris , Savion Naphtali 
Published: April 2006

SummaryThe purpose of this study was to examine the role of platelets in CD4+ T lymphocyte adhesion to subendothelial extracellular matrix (ECM). Herpesvirus saimiri (HVS)-infected CD4+ T cells were incubated on ECM. An image analysis was used to evaluate T cell adhesion. Under static condition, T cell activation with 4-α-Phorbol 12-myristate 13-acetate (PMA) resulted in a 2.6-fold increase in cell adhesion. However, adhesion was not affected by platelets. In contrast, under flow (200s−1), platelets markedly enhanced both resting and PMA-activatedT cell adhesion (33- and 48-fold), forming lymphocyte-platelet co-aggregates that contain approximately 90% of the adherent T cells. Abrogation of platelet aggregation with tirofiban inhibited formation of platelet-T cell co-aggregates under flow and reduced T cell adhesion by 74%. Separate and combined blockade of CD40L and P-selectin glycoprotein-1 (PSGL-1) on PMA-activated lymphocytes reduced adhesion under flow in the presence of platelets by 28%, 33%, and 55%, respectively. Blockade of β1-integrins decreased adhesion under both static and flow conditions (by 35% and 44%, respectively), while blockade of β2-integrin reduced adhesion only under static condition (by 23%). A similar adhesion pattern was observed using CD4+ T cells isolated from normal donor peripheral blood. In conclusion, platelets support CD4+ lymphocyte adhesion to ECM under flow by formation of heterotypic platelet-lymphocyte co-aggregates involving αIIbβ3 integrin and β1-related integrins, as well as CD40L and PSGL-1.
Benefit/risk profile of high-dose antithrombin in patients with severe sepsis treated with and without concomitant heparin
Authors: Hoffmann Johannes , Wiedermann Christian , Juers Mathias , Ostermann Helmut , Kienast Joachim , Briegel Josef , Strauss Richard , Warren Brian , Opal Steven ,   for the KyberSept investigators
Published: April 2006

SummaryA randomised, prospective, placebo-controlled phase III multicentre clinical trial (KyberSept) has been performed to test the efficacy of high-dose antithrombin therapy in patients with severe sepsis. Concomitant low-dose heparin has been routinely given in two thirds of patients for deep vein thrombosis prophylaxis.This study analyses heparin – antithrombin interactions in terms of long-term mortality, adverse events, and thromboembolic events. From a total of 2,314 patients with severe sepsis (placebo: n=1,157; antithrombin: n=1,157) 1,616 patients (placebo: 811, antithrombin: 805) received heparin concomitantly with study drug (antithrombin 30,000 IU) over four days, whereas 698 patients (346 and 352, respectively) did not. In patients with no concomitant heparin, 28-day mortality was lower with antithrombin than with placebo (37.8% vs. 43.6%; absolute reduction: 5.8%; risk ratio: 0.860 [0.725–1.019]), which increased until day-90 (44.9% vs. 52.5%; absolute reduction: 7.6%; risk ratio: 0.851 [0.735–0.987]). In patients with concomitant heparin, no effect of antithrombin on mortality was seen (28-day mortality: 39.4% vs. 36.6%; absolute increase: 2.8%; risk ratio: 1.08 [0.96–1.22]). Frequency of use of concomitant heparin increased during conduct of the study. Increased bleeding incidences were reported with antithrombin plus concomitant heparin as compared to antithrombin alone. Rates of thromboembolic events were similar when antithrombin was given with or without concomitant heparin. In the treatment of severe sepsis, high-dose antithrombin may sufficiently protect against development of venous thromboembolism when no concomitant heparin is given. Combined administration of the two increases bleeding risk and probably abolishes efficacy of antithrombin.
Group B streptococcus isolates from septic patients and healthy carriers differentially activate platelet signaling cascades
Authors: Dornieden Catharina , Beyrich Claudia , Schinke Birgitta , Schubert-Unkmeir Alexandra , Abele-Horn Marianne , Speer Christian , Siauw Celine , Kobsar Anna , Eigenthaler Martin 
Published: April 2006

SummaryInfection with group B streptococcus (GBS) is the most common cause of early onset neonatal sepsis in many countries, leading to neonatal morbidity and mortality.There is much evidence fora direct involvement of platelets in the pathogenesis of inflammation and sepsis. Several bacteria are known to directly interact with platelets leading to activation and aggregation,a phenomenon also observed with GBS. Here, we demonstrate that GBS rapidly bound to platelets; however, only strains isolated from septic patients bound fibrinogen on their surface and induced platelet thromboxane synthesis, platelet aggregation, and P-selectin (CD62P) expression. In contrast, GBS strains isolated from healthy newborns or healthy pregnant women induced only shape change, but not platelet thromboxane synthesis, platelet aggregation, or CD62P expression. All GBS strains investigated were able to activate FcγRIIA receptor signaling pathways including phospholipase C gamma2 (PLCγ2), as well as calcium/calmodulin-dependent myosin kinase II (CaMKII) and phosphorylation of myosin light chain (MLC). In contrast, protein kinase C (PKC) was exclusively activated by GBS strains isolated from septic patients, and p38 mitogen activated protein kinase (p38 MAP kinase) was preferentially activated by septic GBS strains. Furthermore, stress signaling kinase SEK1/MKK4 and focal adhesion kinase (FAK) were activated by all tested GBS strains in a FcγRIIA-independent way.This study demonstrates that septic, but not colonizing, GBS strains bind fibrinogen on their surface, and that septic GBS strains influence platelet function not only via the FcγRIIA receptor, but also via pathways distinct from IgG-mediated signalling. These mechanisms lead to platelet aggregation and secretion, thereby possibly modulating the pathophysiologic course of GBS infections.
Successful silencing of plasminogen activator inhibitor-1 in human vascular endothelial cells using small interfering RNA
Authors: Hecke Anneke , Brooks Hilary , Meryet-Figuière Matthieu , Minne Stephanie , Konstantinides Stavros , Hasenfuss Gerd , Lebleu Bernard , Schäfer Katrin 
Published: April 2006

SummaryClinical as well as experimental evidence suggests that vascular overexpression of plasminogen activator inhibitor (PAI)-1, the primary physiological inhibitor of both urokinase and tissuetype plasminogen activator, may be involved in the pathophysiology of atherosclerosis and cardiovascular disease. We investigated the feasibility, efficacy and functional effects of PAI-1 gene silencing in human vascular endothelial cells using small interfering RNA. Double-stranded 21 bp-RNA molecules targeted at sequences within the human PAI-1 gene were constructed. Successful siRNA transfection of HUVEC was confirmed using fluorescence microscopy and flow cytometry. One of five candidate siRNA sequences reduced PAI-1 mRNA and protein in a concentrationand time-dependent manner. Suppression of PAI-1 mRNA was detected up to 72 hours after transfection. Moreover, siRNA treatment reduced the activity of PAI-1 released from HUVEC, and prevented the oxLDL- or LPS-induced upregulation of PAI-1 secretion. Importantly, siRNA treatment did not affect the expression of other endothelial-cell markers. Moreover, downregulation of PAI-1 significantly enhanced the ability of endothelial cells to adhere to vitronectin, and this effect could be reversed upon addition of recombinant PAI-1. SiRNAmediated reduction of PAI-1 expression may be a promising strategy for dissecting the effects of PAI-1 on vascular homeostasis.
Antithrombin reduces reperfusion-induced liver injury in mice by enhancing sensory neuron activation
Authors: Harada Naoaki , Uchiba Mitsuhiro , Kurihara Hiroki , Nakagata Naomi , Okajima Kenji 
Published: April 2006

SummaryWe recently demonstrated that antithrombin (AT) reduces ischemia/reperfusion (I/R)-induced liver injury in rats by increasing hepatic tissue levels of calcitonin gene-related peptide (CGRP),a neuropeptide released from the sensory nerve endings. In the present study, we examined the effect of AT on I/Rinduced liver injury in wild type mice (CGRP+/+) and congenitally αCGRP-deficient mice (CGRP−/−). We also investigated any effects of AT on CGRP release from dorsal root ganglion neurons (DRG) isolated from CGRP+/+. Based on results obtained in the present study, we attempted to determine if the anti-inflammatory activity of AT in vivo is dependent mainly on sensory neuron activation. AT enhanced ischemia/reperfusion-induced increases in hepatic tissue levels of CGRP and 6-keto-PGF1α
, a stable metabolite of PGI2, in CGRP+/+, but it did not enhance these increases in CGRP−/−. AT inhibited reperfusion-induced increases in serum alanine aminotransferase levels by increasing hepatic tissue blood flow and by attenuating increases in hepatic levels of tumor necrosis factor and myeloperoxidase in CGRP+/+,although it showed neither of these therapeutic effects in CGRP−/−. AT increased CGRP release from cultured DRGs only in the presence of anandamide, and AT-induced increase in CGRP release was not observed in the presence KT5720, an inhibitor of protein kinase A (PKA).AT markedly increased intracellular levels of cAMP in the presence of anandamide. These results strongly suggest that AT might reduce I/R-induced liver injury by enhancing activation of the sensory neurons through activation of PKA in sensory neurons.
Serum osteoprotegerin in young survivors of myocardial infarction
Authors: Brodin Ellen , Børvik Trond , Sveinbjørnsson Baldur , Hansen John-Bjarne , Vik Anders 
Published: April 2006

SummaryOsteoprotegerin (OPG) is a member of the tumour necrosis factor superfamily and is involved in the regulation of bone metabolism and vascular calcification. Increased serum OPG levels have been reported in patients with stable angina pectoris and survivors of myocardial infarction with heart failure. The purpose of the present study was to determine serum OPG levels in young survivors of acute myocardial infarction (MI), and the relationship between OPG, homocysteine, sCD40L and coagulation factors in blood. Fifty-eight patients with verified MI, 40–60 years of age, were recruited 1–4 years after the acute event into an age- and sex- matched case control study with controls recruited from the general population. Serum OPG levels were similar in cases (2.41 ng/ml, 2.11–2.77 ng/ml) (mean, 95% CI) and controls (2.43 ng/ml, 2.11–2.79 ng/ml) (p= 0.92). Significant correlation between OPG and homocysteine was found in patients (r=0.30, p=0.02) and controls (r=0.35, p=0.007). A significant negative correlation was found between OPG and sCD40L in patients (r=-0.51, p<0.001), but not in controls (r=0.001, p=0.96). No associations were found between serum OPG and markers of coagulation activation. The present study shows that serum OPG level was not increased in young survivors of uncomplicated myocardial infarction. Serum OPG levels were not associated with thrombin generation assessed by thrombin-antithrombin complexes (TAT), but a positive association between serum OPG and homocysteine was found.
The influence of sequence variations in factor VII, γ-glutamyl carboxylase and vitamin K epoxide reductase complex genes on warfarin dose requirement
Authors: Herman Darja , Peternel Polona , Stegnar Mojca , Breskvar Katja , Dolzan Vita 
Published: April 2006

SummaryThe degree of interpatient variability in the warfarin dose required to achieve the desired anticoagulant response can only partly be explained by polymorphisms in the CYP2C9 gene, suggesting that additional genetic factors such as polymorphisms in genes involved in blood coagulation may influence warfarin dose requirement. In total, 165 Caucasian outpatients on stable maintenance warfarin treatment previously genotyped for CYP2C9 were analysed for common polymorphisms in FVII, GGCX and VKORC1 genes. The -402G>A polymorphism and a variable number of repeats in intron 7 of FVII gene did not significantly influence warfarin dose.The mean warfarin doses increased with the number of (CAA) repeats in the GGCX gene, but the differences were significant only in the CYP2C9*1/*1 subgroup of patients (p=0.032). Common polymorphism (6484C>T) in intron 1 of the VKORC1 gene led to lower warfarin dose requirement; the means were 5.70 (95% C.I. 4.95 - 6.45), 3.49 (3.07 - 3.90) and 2.11 (1.80 - 2.42) mg/day for 6484 CC, CT and TT genotypes, respectively (p<0.001). In contrast, 9041G>A polymorphism in 3’UTR of theVKORC1 gene led to higher warfarin dose requirement; the means were 3.09 (2.58 - 3.60), 4.26 (3.69 - 4.82) and 5.86 (4.53 - 7.19) mg/day for 9041 GG, GA and AA genotypes, respectively (p<0.001).With a regression model we explained 60.0% of variability in warfarin dose, which was due to gene polymorphisms (CYP2C9,VKORC1), age and body-surfacearea. When aiming for individualised warfarin therapy, at least VKORC1 polymorphisms should be included in predictive genotyping besides CYP2C9.