Glycoprotein Ib activation by thrombin stimulates the energy metabolism in human platelets

<div><p>Thrombin-induced platelet activation requires substantial amounts of ATP. However, the specific contribution of each ATP-generating pathway <i>i</i>.<i>e</i>., oxidative phosphorylation (OxPhos) versus glycolysis and the biochemical mechanisms involved in the thrombin-induced activation of energy metabolism remain unclear. Here we report an integral analysis on the role of both energy pathways in human platelets activated by several agonists, and the signal transducing mechanisms associated with such activation. We found that thrombin, Trap-6, arachidonic acid, collagen, A23187, epinephrine and ADP significantly increased glycolytic flux (3–38 times <i>vs</i>. non-activated platelets) whereas ristocetin was ineffective. OxPhos (33 times) and mitochondrial transmembrane potential (88%) were increased only by thrombin. OxPhos was the main source of ATP in thrombin-activated platelets, whereas in platelets activated by any of the other agonists, glycolysis was the principal ATP supplier. In order to establish the biochemical mechanisms involved in the thrombin-induced OxPhos activation in platelets, several signaling pathways associated with mitochondrial activation were analyzed. Wortmannin and LY294002 (PI3K/Akt pathway inhibitors), ristocetin and heparin (GPIb inhibitors) as well as resveratrol, ATP (calcium-release inhibitors) and PP1 (Tyr-phosphorylation inhibitor) prevented the thrombin-induced platelet activation. These results suggest that thrombin activates OxPhos and glycolysis through GPIb-dependent signaling involving PI3K and Akt activation, calcium mobilization and protein phosphorylation.</p></div

Down selecting adjuvanted vaccine formulations: a comparative method for harmonized evaluation.

The need for rapid and accurate comparison of panels of adjuvanted vaccine formulations and subsequent rational down selection, presents several challenges for modern vaccine development. Here we describe a method which may enable vaccine and adjuvant developers to compare antigen/adjuvant combinations in a harmonized fashion. Three reference antigens: Plasmodium falciparum apical membrane antigen 1 (AMA1), hepatitis B virus surface antigen (HBsAg), and Mycobacterium tuberculosis antigen 85A (Ag85A), were selected as model antigens and were each formulated with three adjuvants: aluminium oxyhydroxide, squalene-in-water emulsion, and a liposome formulation mixed with the purified saponin fraction QS21. The nine antigen/adjuvant formulations were assessed for stability and immunogenicity in mice in order to provide benchmarks against which other formulations could be compared, in order to assist subsequent down selection of adjuvanted vaccines. Furthermore, mouse cellular immune responses were analyzed by measuring IFN-γ and IL-5 production in splenocytes by ELISPOT, and humoral responses were determined by antigen-specific ELISA, where levels of total IgG, IgG1, IgG2b and IgG2c in serum samples were determined. The reference antigens and adjuvants described in this study, which span a spectrum of immune responses, are of potential use as tools to act as points of reference in vaccine development studies. The harmonized methodology described herein may be used as a tool for adjuvant/antigen comparison studies

System-level determinants of immunization coverage disparities among health districts in Burkina Faso: a multiple case study

<p>Abstract</p> <p>Background</p> <p>Despite rapid and tangible progress in vaccine coverage and in premature mortality rates registered in sub-Saharan Africa, inequities to access remain firmly entrenched, large pockets of low vaccination coverage persist, and coverage often varies considerably across regions, districts, and health facilities' areas of responsibility. This paper focuses on system-related factors that can explain disparities in immunization coverage among districts in Burkina Faso.</p> <p>Methods</p> <p>A multiple-case study was conducted of six districts representative of different immunization trends and overall performance. A participative process that involved local experts and key actors led to a focus on key factors that could possibly determine the efficiency and efficacy of district vaccination services: occurrence of disease outbreaks and immunization days, overall district management performance, resources available for vaccination services, and institutional elements. The methodology, geared toward reconstructing the evolution of vaccine services performance from 2000 to 2006, is based on data from documents and from individual and group interviews in each of the six health districts. The process of interpreting results brought together the field personnel and the research team.</p> <p>Results</p> <p>The districts that perform best are those that assemble a set of favourable conditions. However, the leadership of the district medical officer (DMO) appears to be the main conduit and the rallying point for these conditions. Typically, strong leadership that is recognized by the field teams ensures smooth operation of the vaccination services, promotes the emergence of new initiatives and offers some protection against risks related to outbreaks of epidemics or supplementary activities that can hinder routine functioning. The same is true for the ability of nurse managers and their teams to cope with new situations (epidemics, shortages of certain stocks).</p> <p>Conclusion</p> <p>The discourse on factors that determine the performance or breakdown of local health care systems in lower and middle income countries remains largely concentrated on technocratic and financial considerations, targeting institutional reforms, availability of resources, or accessibility of health services. The leadership role of those responsible for the district, and more broadly, of those we label "the human factor", in the performance of local health care systems is mentioned only marginally. This study shows that strong and committed leadership promotes an effective mobilization of teams and creates the conditions for good performance in districts, even when they have only limited access to supports provided by external partners.</p> <p>Abstract in French</p> <p>See the full article online for a translation of this abstract in French.</p

Synergistic TLR2/6 and TLR9 Activation Protects Mice against Lethal Influenza Pneumonia

Lower respiratory tract infections caused by influenza A continue to exact unacceptable worldwide mortality, and recent epidemics have emphasized the importance of preventative and containment strategies. We have previously reported that induction of the lungs' intrinsic defenses by aerosolized treatments can protect mice against otherwise lethal challenges with influenza A virus. More recently, we identified a combination of Toll like receptor (TLR) agonists that can be aerosolized to protect mice against bacterial pneumonia. Here, we tested whether this combination of synthetic TLR agonists could enhance the survival of mice infected with influenza A/HK/8/68 (H3N2) or A/California/04/2009 (H1N1) influenza A viruses. We report that the TLR treatment enhanced survival whether given before or after the infectious challenge, and that protection tended to correlate with reductions in viral titer 4 d after infection. Surprisingly, protection was not associated with induction of interferon gene expression. Together, these studies suggest that synergistic TLR interactions can protect against influenza virus infections by mechanisms that may provide the basis for novel therapeutics

Protein 4.1B Contributes to the Organization of Peripheral Myelinated Axons

Neurons are characterized by extremely long axons. This exceptional cell shape is likely to depend on multiple factors including interactions between the cytoskeleton and membrane proteins. In many cell types, members of the protein 4.1 family play an important role in tethering the cortical actin-spectrin cytoskeleton to the plasma membrane. Protein 4.1B is localized in myelinated axons, enriched in paranodal and juxtaparanodal regions, and also all along the internodes, but not at nodes of Ranvier where are localized the voltage-dependent sodium channels responsible for action potential propagation. To shed light on the role of protein 4.1B in the general organization of myelinated peripheral axons, we studied 4.1B knockout mice. These mice displayed a mildly impaired gait and motility. Whereas nodes were unaffected, the distribution of Caspr/paranodin, which anchors 4.1B to the membrane, was disorganized in paranodal regions and its levels were decreased. In juxtaparanodes, the enrichment of Caspr2, which also interacts with 4.1B, and of the associated TAG-1 and Kv1.1, was absent in mutant mice, whereas their levels were unaltered. Ultrastructural abnormalities were observed both at paranodes and juxtaparanodes. Axon calibers were slightly diminished in phrenic nerves and preterminal motor axons were dysmorphic in skeletal muscle. βII spectrin enrichment was decreased along the axolemma. Electrophysiological recordings at 3 post-natal weeks showed the occurrence of spontaneous and evoked repetitive activity indicating neuronal hyperexcitability, without change in conduction velocity. Thus, our results show that in myelinated axons 4.1B contributes to the stabilization of membrane proteins at paranodes, to the clustering of juxtaparanodal proteins, and to the regulation of the internodal axon caliber

Lactate Produced by Glycogenolysis in Astrocytes Regulates Memory Processing

When administered either systemically or centrally, glucose is a potent enhancer of memory processes. Measures of glucose levels in extracellular fluid in the rat hippocampus during memory tests reveal that these levels are dynamic, decreasing in response to memory tasks and loads; exogenous glucose blocks these decreases and enhances memory. The present experiments test the hypothesis that glucose enhancement of memory is mediated by glycogen storage and then metabolism to lactate in astrocytes, which provide lactate to neurons as an energy substrate. Sensitive bioprobes were used to measure brain glucose and lactate levels in 1-sec samples. Extracellular glucose decreased and lactate increased while rats performed a spatial working memory task. Intrahippocampal infusions of lactate enhanced memory in this task. In addition, pharmacological inhibition of astrocytic glycogenolysis impaired memory and this impairment was reversed by administration of lactate or glucose, both of which can provide lactate to neurons in the absence of glycogenolysis. Pharmacological block of the monocarboxylate transporter responsible for lactate uptake into neurons also impaired memory and this impairment was not reversed by either glucose or lactate. These findings support the view that astrocytes regulate memory formation by controlling the provision of lactate to support neuronal functions

Metopic synostosis

Premature closure of the metopic suture results in a growth restriction of the frontal bones, which leads to a skull malformation known as trigonocephaly. Over the course of recent decades, its incidence has been rising, currently making it the second most common type of craniosynostosis. Treatment consists of a cranioplasty, usually preformed before the age of 1 year. Metopic synostosis is linked with an increased level of neurodevelopmental delays. Theories on the etiology of these delays range from a reduced volume of the anterior cranial fossa to intrinsic malformations of the brain. This paper aims to provide an overview of this entity by giving an update on the epidemiology, etiology, evolution of treatment, follow-up, and neurodevelopment of metopic synostosis