Decreased endothelial glycocalyx thickness is an early predictor of mortality in sepsis.

Microcirculatory alterations play an important role in the early phase of sepsis. Shedding of the endothelial glycocalyx is regarded as a central pathophysiological mechanism causing microvascular dysfunction, contributing to multiple organ failure and death in sepsis. The objective of this study was to investigate whether endothelial glycocalyx thickness at an early stage in septic patients relates to clinical outcome.

Rational modulator design by exploitation of protein-protein complex structures.

The horizon of drug discovery is currently expanding to target and modulate protein-protein interactions (PPIs) in globular proteins and intrinsically disordered proteins that are involved in various diseases. To either interrupt or stabilize PPIs, the 3D structure of target protein-protein (or protein-peptide) complexes can be exploited to rationally design PPI modulators (inhibitors or stabilizers) through structure-based molecular design. In this review, we present an overview of experimental and computational methods that can be used to determine 3D structures of protein-protein complexes.

Targeting CD40-Induced TRAF6 Signaling in Macrophages Reduces Atherosclerosis.

Background: Disrupting the costimulatory CD40-CD40L dyad reduces atherosclerosis, but can result in immune suppression. The authors recently identified small molecule inhibitors that block the interaction between CD40 and tumor necrosis factor receptor-associated factor (TRAF) 6 (TRAF-STOPs), while leaving CD40-TRAF2/3/5 interactions intact, thereby preserving CD40-mediated immunity.

Objectives: This study evaluates the potential of TRAF-STOP treatment in atherosclerosis.

Chemokine interactome mapping enables tailored intervention in acute and chronic inflammation.

Chemokines orchestrate leukocyte trafficking and function in health and disease. Heterophilic interactions between chemokines in a given microenvironment may amplify, inhibit, or modulate their activity; however, a systematic evaluation of the chemokine interactome has not been performed. We used immunoligand blotting and surface plasmon resonance to obtain a comprehensive map of chemokine-chemokine interactions and to confirm their specificity.

Presence of Cytotoxic Extracellular Histones in Machine Perfusate of Donation After Circulatory Death Kidneys.

Background: Extracellular histones are cytotoxic molecules that are related to cell stress and death. They have been shown to play a crucial role in multiple pathophysiologic processes like sepsis, inflammation, vascular dysfunction, and thrombosis. Their role in organ donation and graft function and survival is still unknown.

Structure-Based Design of Peptidic Inhibitors of the Interaction between CC Chemokine Ligand 5 (CCL5) and Human Neutrophil Peptides 1 (HNP1).

Protein-protein interactions (PPIs) are receiving increasing interest, much sparked by the realization that they represent druggable targets. Recently, we successfully developed a peptidic inhibitor, RRYGTSKYQ ("SKY" peptide), that shows high potential in vitro and in vivo to interrupt a PPI between the platelet-borne chemokine CCL5 and the neutrophil-derived granule protein HNP1. This PPI plays a vital role in monocyte adhesion, representing a key mechanism in acute and chronic inflammatory diseases.

Recruitment of classical monocytes can be inhibited by disturbing heteromers of neutrophil HNP1 and platelet CCL5.

In acute and chronic inflammation, neutrophils and platelets, both of which promote monocyte recruitment, are often activated simultaneously. We investigated how secretory products of neutrophils and platelets synergize to enhance the recruitment of monocytes. We found that neutrophil-borne human neutrophil peptide 1 (HNP1, α-defensin) and platelet-derived CCL5 form heteromers.

Pharmacological Targeting of AMP-Activated Protein Kinase and Opportunities for Computer-Aided Drug Design.

As a central regulator of metabolism, the AMP-activated protein kinase (AMPK) is an established therapeutic target for metabolic diseases. Beyond the metabolic area, the number of medical fields that involve AMPK grows continuously, expanding the potential applications for AMPK modulators. Even though indirect AMPK activators are used in the clinics for their beneficial metabolic outcome, the few described direct agonists all failed to reach the market to date, which leaves options open for novel targeting methods.

iPPI-DB: an online database of modulators of protein-protein interactions.

In order to boost the identification of low-molecular-weight drugs on protein-protein interactions (PPI), it is essential to properly collect and annotate experimental data about successful examples. This provides the scientific community with the necessary information to derive trends about privileged physicochemical properties and chemotypes that maximize the likelihood of promoting a given chemical probe to the most advanced stages of development. To this end we have developed iPPI-DB (freely accessible at http://www.

Extracellular histone H3 levels are inversely correlated with antithrombin levels and platelet counts and are associated with mortality in sepsis patients.

Objective: Sepsis is a leading cause of death worldwide. Extracellular histones are cytotoxic compounds mediating death in murine sepsis and circulating nucleosome levels predict mortality in human inflammation and sepsis. Whether or not circulating extracellular histone H3 correlates with other plasma parameters and/or ICU scoring systems has not been completely established, nor if levels of circulating extracellular histones can be used as predictive markers for clinical outcome in sepsis.

Identification of small-molecule inhibitors against SecA by structure-based virtual ligand screening.

The rapid rise of antibiotic-resistant bacteria is one of the major concerns in modern medicine. Therefore, to treat bacterial infections, there is an urgent need for new antibacterials-preferably directed against alternative bacterial targets. One such potential target is the preprotein translocation motor SecA.

Targeting the adaptive immune system: new strategies in the treatment of atherosclerosis.

Atherosclerosis is a lipid-driven chronic inflammatory disease of the arterial wall. Current treatment of atherosclerosis is focused on limiting its risk factors, such as hyperlipidemia or hypertension. However, treatments that target the inflammatory nature of atherosclerosis are still under development.

Discovery of small molecule CD40-TRAF6 inhibitors.

The CD154-CD40 receptor complex plays a pivotal role in several inflammatory pathways. Attempts to inhibit the formation of this complex have resulted in systemic side effects. Downstream inhibition of the CD40 signaling pathway therefore seems a better way to ameliorate inflammatory disease.

The structure function of the death domain of human IRAK-M.

Background: IRAK-M is an inhibitor of Toll-like receptor signaling that acts by re-directing IRAK-4 activity to TAK1 independent NF-κB activation and by inhibition of IRAK-1/IRAK-2 activity. IRAK-M is expressed in monocytes/macrophages and lung epithelial cells. Lack of IRAK-M in mice greatly improves the resistance to nosocomial pneumonia and lung tumors, which entices IRAK-M as a potential therapeutic target.

Molecular simulation studies of human coagulation factor VIII C domain-mediated membrane binding.

The C-terminal C domains of activated coagulation factor VIII (FVIIIa) are essential to membrane binding of this crucial coagulation cofactor protein. To provide an overall membrane binding mechanism for FVIII, we performed simulations of membrane binding through coarse-grained molecular dynamics simulations of the C1 and C2 domain, and the combined C-domains (C1+C2). We found that the C1 and C2 domain have different membrane binding properties.

Understanding the functional difference between growth arrest-specific protein 6 and protein S: an evolutionary approach.

Although protein S (PROS1) and growth arrest-specific protein 6 (GAS6) proteins are homologous with a high degree of structural similarity, they are functionally different. The objectives of this study were to identify the evolutionary origins from which these functional differences arose. Bioinformatics methods were used to estimate the evolutionary divergence time and to detect the amino acid residues under functional divergence between GAS6 and PROS1.

Effects of exogenous recombinant APC in mouse models of ischemia reperfusion injury and of atherosclerosis.

Activated protein C (APC) is a serine protease that has both anticoagulant and cytoprotective properties. The cytoprotective effects are protease activated receptor 1 (PAR-1) and endothelial protein C receptor (EPCR) dependent and likely underlie protective effects of APC in animal models of sepsis, myocardial infarction and ischemic stroke. S360A-(A)PC, a variant (A)PC that has no catalytic activity, binds EPCR and shifts pro-inflammatory signaling of the thrombin-PAR-1 complex to anti-inflammatory signaling.

Identification of novel small molecule inhibitors of activated protein C.

Introduction: Activated protein C (APC) is the central enzyme of the anticoagulant protein C pathway. Low concentrations of APC circulate in plasma and are believed to contribute to the maintenance of a normal haemostatic balance.

Materials And Methods: We have used a structure-based virtual screening approach to discover small drug-like molecules that inhibit the interaction between APC and its substrate FVa through inhibition of a predominant APC exosite, known to be involved in FVa substrate binding.

Nonanticoagulant heparin prevents histone-mediated cytotoxicity in vitro and improves survival in sepsis.

Extracellular histones are considered to be major mediators of death in sepsis. Although sepsis is a condition that may benefit from low-dose heparin administration, medical doctors need to take into consideration the potential bleeding risk in sepsis patients who are already at increased risk of bleeding due to a consumption coagulopathy. Here, we show that mechanisms that are independent of the anticoagulant properties of heparin may contribute to the observed beneficial effects of heparin in the treatment of sepsis patients.

Rational design of small molecules targeting the C2 domain of coagulation factor VIII.

The C domains of coagulation factors V (FV) and VIII (FVIII) are structurally conserved domains and share a common and essential function in membrane binding. In vivo regulation of thrombin formation strongly depends on the expression and regulation of the cofactor activities of FVIII and FV. With this study, we explored the possibility of inhibition of thrombin formation in full blood with small druglike molecules.

Optimization of compound ranking for structure-based virtual ligand screening using an established FRED-Surflex consensus approach.

The use of multiple target conformers has been applied successfully in virtual screening campaigns; however, a study on how to best combine scores for multiple targets in a hierarchic method that combines rigid and flexible docking is not available. In this study, we used a data set of 59 479 compounds to screen multiple conformers of four distinct protein targets to obtain an adapted and optimized combination of an established hierarchic method that employs the programs FRED and Surflex. Our study was extended and verified by application of our protocol to ten different data sets from the directory of useful decoys (DUD).

Factor Xa activation of factor V is of paramount importance in initiating the coagulation system: lessons from a tick salivary protein.

Background: Generation of active procoagulant cofactor factor Va (FVa) and its subsequent association with the enzyme activated factor X (FXa) to form the prothrombinase complex is a pivotal initial event in blood coagulation and has been the subject of investigative effort, speculation, and controversy. The current paradigm assumes that FV activation is initiated by limited proteolysis by traces of (meizo) thrombin.

Methods And Results: Recombinant tick salivary protein TIX-5 was produced and anticoagulant properties were studied with the use of plasma, whole blood, and purified systems.

Comparative analysis of pharmacophore screening tools.

The pharmacophore concept is of central importance in computer-aided drug design (CADD) mainly because of its successful application in medicinal chemistry and, in particular, high-throughput virtual screening (HTVS). The simplicity of the pharmacophore definition enables the complexity of molecular interactions between ligand and receptor to be reduced to a handful set of features. With many pharmacophore screening softwares available, it is of the utmost interest to explore the behavior of these tools when applied to different biological systems.

Goodpasture antigen-binding protein/ceramide transporter binds to human serum amyloid P-component and is present in brain amyloid plaques.

Serum amyloid P component (SAP) is a non-fibrillar glycoprotein belonging to the pentraxin family of the innate immune system. SAP is present in plasma, basement membranes, and amyloid deposits. This study demonstrates, for the first time, that the Goodpasture antigen-binding protein (GPBP) binds to human SAP.

The structure-function relationship of activated protein C. Lessons from natural and engineered mutations.

Protein C is the central enzyme of the natural anticoagulant pathway and its activated form APC (activated protein C) is able to proteolyse non-active as well as active coagulation factors V and VIII. Proteolysis renders these cofactors inactive, resulting in an attenuation of thrombin formation and overall down-regulation of coagulation. Presences of the APC cofactor, protein S, thrombomodulin, endothelial protein C receptor and a phospholipid surface are important for the expression of anticoagulant APC activity.