Incorporating Data into EFSM Inference

This is the author accepted manuscript. The final version is available from Springer Verlag via the DOI in this record17th International Conference, SEFM 2019 Oslo, Norway, September 18–20, 2019Models are an important way of understanding software systems. If they do not already exist, then we need to infer them from system behaviour. Most current approaches infer classical FSM models that do not consider data, thus limiting applicability. EFSMs provide a way to concisely model systems with an internal state but existing inference techniques either do not infer models which allow outputs to be computed from inputs, or rely heavily on comprehensive white-box traces that reveal the internal program state, which are often unavailable. In this paper, we present an approach for inferring EFSM models, including functions that modify the internal state. Our technique uses black-box traces which only contain information visible to an external observer of the system. We implemented our approach as a prototype

Methods for economic evaluation of a factorial-design cluster randomised controlled trial of a nutrition supplement and an exercise programme among healthy older people living in Santiago, Chile: the CENEX study

BACKGROUND: In an effort to promote healthy ageing and preserve health and function, the government of Chile has formulated a package of actions into the Programme for Complementary Food in Older People (Programa de Alimentación Complementaria para el Adulto Mayor - PACAM). The CENEX study was designed to evaluate the impact, cost and cost-effectiveness of the PACAM and a specially designed exercise programme on pneumonia incidence, walking capacity and body mass index in healthy older people living in low- to medium-socio-economic status areas of Santiago. The purpose of this paper is to describe in detail the methods that will be used to estimate the incremental costs and cost-effectiveness of the interventions. METHODS AND DESIGN: The base-case analysis will adopt a societal perspective, including the direct medical and non-medical costs borne by the government and patients. The cost of the interventions will be calculated by the ingredients approach, in which the total quantities of goods and services actually employed in applying the interventions will be estimated, and multiplied by their respective unit prices. Relevant information on costs of interventions will be obtained mainly from administrative records. The costs borne by patients will be collected via exit and telephone interviews. An annual discount rate of 8% will be used, consistent with the rate recommended by the Government of Chile. All costs will be converted from Chilean Peso to US dollars with the 2007 average period exchange rate of US$1 = 522.37 Chilean Peso. To test the robustness of model results, we will vary the assumptions over a plausible range in sensitivity analyses. DISCUSSION: The protocol described here indicates our intent to conduct an economic evaluation alongside the CENEX study. It provides a detailed and transparent statement of planned data collection methods and analyses. TRIAL REGISTRATION: ISRCTN48153354

Evolution of predator dispersal in relation to spatio-temporal prey dynamics : how not to get stuck in the wrong place!

Peer reviewedPublisher PD

Meta-analysis of genome-wide association studies from the CHARGE consortium identifies common variants associated with carotid intima media thickness and plaque

Carotid intima media thickness (cIMT) and plaque determined by ultrasonography are established measures of subclinical atherosclerosis that each predicts future cardiovascular disease events. We conducted a meta-analysis of genome-wide association data in 31,211 participants of European ancestry from nine large studies in the setting of the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) Consortium. We then sought additional evidence to support our findings among 11,273 individuals using data from seven additional studies. In the combined meta-analysis, we identified three genomic regions associated with common carotid intima media thickness and two different regions associated with the presence of carotid plaque (P < 5 × 10 -8). The associated SNPs mapped in or near genes related to cellular signaling, lipid metabolism and blood pressure homeostasis, and two of the regions were associated with coronary artery disease (P < 0.006) in the Coronary Artery Disease Genome-Wide Replication and Meta-Analysis (CARDIoGRAM) consortium. Our findings may provide new insight into pathways leading to subclinical atherosclerosis and subsequent cardiovascular events

The yeast P5 type ATPase, Spf1, regulates manganese transport into the endoplasmic reticulum

The endoplasmic reticulum (ER) is a large, multifunctional and essential organelle. Despite intense research, the function of more than a third of ER proteins remains unknown even in the well-studied model organism Saccharomyces cerevisiae. One such protein is Spf1, which is a highly conserved, ER localized, putative P-type ATPase. Deletion of SPF1 causes a wide variety of phenotypes including severe ER stress suggesting that this protein is essential for the normal function of the ER. The closest homologue of Spf1 is the vacuolar P-type ATPase Ypk9 that influences Mn2+ homeostasis. However in vitro reconstitution assays with Spf1 have not yielded insight into its transport specificity. Here we took an in vivo approach to detect the direct and indirect effects of deleting SPF1. We found a specific reduction in the luminal concentration of Mn2+ in ∆spf1 cells and an increase following it’s overexpression. In agreement with the observed loss of luminal Mn2+ we could observe concurrent reduction in many Mn2+-related process in the ER lumen. Conversely, cytosolic Mn2+-dependent processes were increased. Together, these data support a role for Spf1p in Mn2+ transport in the cell. We also demonstrate that the human sequence homologue, ATP13A1, is a functionally conserved orthologue. Since ATP13A1 is highly expressed in developing neuronal tissues and in the brain, this should help in the study of Mn2+-dependent neurological disorders

Protective Immunity to Listeria Monocytogenes Infection Mediated by Recombinant Listeria innocua Harboring the VGC Locus

In this study we propose a novel bacterial vaccine strategy where non-pathogenic bacteria are complemented with traits desirable for the induction of protective immunity. To illustrate the proof of principle of this novel vaccination strategy, we use the model organism of intracellular immunity Listeria. We introduced a, low copy number BAC-plasmid harbouring the virulence gene cluster (vgc) of L. monocytogenes (Lm) into the non-pathogenic L. innocua (L.inn) strain and examined for its ability to induce protective cellular immunity. The resulting strain (L.inn::vgc) was attenuated for virulence in vivo and showed a strongly reduced host detrimental inflammatory response compared to Lm. Like Lm, L.inn::vgc induced the production of Type I Interferon's and protection was mediated by Listeria-specific CD8+ T cells. Rational vaccine design whereby avirulent strains are equipped with the capabilities to induce protection but lack detrimental inflammatory effects offer great promise towards future studies using non-pathogenic bacteria as vectors for vaccination

Automated Planar Patch-Clamp Recording of P2X Receptors

P2X receptors are a structurally and functionally distinctive family of ligand-gated ion channels that play important roles in mediating extracellular adenosine 5′-triphosphate (ATP) signaling in diverse physiological and pathophysiological processes. For several decades, the “manual” patch-clamp technique was regarded as the gold standard assay for investigating ion channel properties. More recently, breakthroughs in the development of automated patch-clamp technologies are enabling the study of ion channels, with much greater throughput capacities. These automated platforms, of which there are many, generate consistent, reliable, high-fidelity data. This chapter demonstrates the versatility of one of these technologies for ligand-gated ion channels, with a particular emphasis on protocols that address some of the issues of receptor desensitization that are commonly associated with P2X receptor-mediated currents

SIRT2 Ablation Has No Effect on Tubulin Acetylation in Brain, Cholesterol Biosynthesis or the Progression of Huntington's Disease Phenotypes In Vivo

Huntington's disease (HD) is a devastating neurodegenerative disorder for which there are no disease-modifying treatments. The molecular pathogenesis of HD is complex and many mechanisms and cellular processes have been proposed as potential sites of therapeutic intervention. However, prior to embarking on drug development initiatives, it is essential that therapeutic targets can be validated in mammalian models of HD. Previous studies in invertebrate and cell culture HD models have suggested that inhibition of SIRT2 could have beneficial consequences on disease progression. SIRT2 is a NAD[superscript +]-dependent deacetylase that has been proposed to deacetylate α-tubulin, histone H4 K16 and to regulate cholesterol biogenesis – a pathway which is dysregulated in HD patients and HD mouse models. We have utilized mice in which SIRT2 has been reduced or ablated to further explore the function of SIRT2 and to assess whether SIRT2 loss has a beneficial impact on disease progression in the R6/2 mouse model of HD. Surprisingly we found that reduction or loss of SIRT2 had no effect on the acetylation of α-tubulin or H4K16 or on cholesterol biosynthesis in the brains of wild type mice. Equally, genetic reduction or ablation of SIRT2 had no effect on HD progression as assessed by a battery of physiological and behavioural tests. Furthermore, we observed no change in aggregate load or levels of soluble mutant huntingtin transprotein. Intriguingly, neither the constitutive genetic loss nor acute pharmacological inhibition of SIRT2 affected the expression of cholesterol biosynthesis enzymes in the context of HD. Therefore, we conclude that SIRT2 inhibition does not modify disease progression in the R6/2 mouse model of HD and SIRT2 inhibition should not be prioritised as a therapeutic option for HD.American Parkinson Disease Association, Inc. (Fellowship)Johnson & Johnson. Pharmaceutical Research & Development (Fellowship

Soluble Rhesus Lymphocryptovirus gp350 Protects against Infection and Reduces Viral Loads in Animals that Become Infected with Virus after Challenge

Epstein-Barr virus (EBV) is a human lymphocryptovirus that is associated with several malignancies. Elevated EBV DNA in the blood is observed in transplant recipients prior to, and at the time of post-transplant lymphoproliferative disease; thus, a vaccine that either prevents EBV infection or lowers the viral load might reduce certain EBV malignancies. Two major approaches have been suggested for an EBV vaccine- immunization with either EBV glycoprotein 350 (gp350) or EBV latency proteins (e.g. EBV nuclear antigens [EBNAs]). No comparative trials, however, have been performed. Rhesus lymphocryptovirus (LCV) encodes a homolog for each gene in EBV and infection of monkeys reproduces the clinical, immunologic, and virologic features of both acute and latent EBV infection. We vaccinated rhesus monkeys at 0, 4 and 12 weeks with (a) soluble rhesus LCV gp350, (b) virus-like replicon particles (VRPs) expressing rhesus LCV gp350, (c) VRPs expressing rhesus LCV gp350, EBNA-3A, and EBNA-3B, or (d) PBS. Animals vaccinated with soluble gp350 produced higher levels of antibody to the glycoprotein than those vaccinated with VRPs expressing gp350. Animals vaccinated with VRPs expressing EBNA-3A and EBNA-3B developed LCV-specific CD4 and CD8 T cell immunity to these proteins, while VRPs expressing gp350 did not induce detectable T cell immunity to gp350. After challenge with rhesus LCV, animals vaccinated with soluble rhesus LCV gp350 had the best level of protection against infection based on seroconversion, viral DNA, and viral RNA in the blood after challenge. Surprisingly, animals vaccinated with gp350 that became infected had the lowest LCV DNA loads in the blood at 23 months after challenge. These studies indicate that gp350 is critical for both protection against infection with rhesus LCV and for reducing the viral load in animals that become infected after challenge. Our results suggest that additional trials with soluble EBV gp350 alone, or in combination with other EBV proteins, should be considered to reduce EBV infection or virus-associated malignancies in humans

The P2Y4 receptor forms homo-oligomeric complexes in several CNS and PNS neuronal cells

It is well established that several cell surface receptors interact with each other to form dimers and oligomers, which are essential for their activation. Since little is known about the quaternary structure of P2Y receptors, in the present work, we investigated the expression of the G-protein-coupled P2Y4 subunit as monomeric or higher-order complex protein. We examined both endogenously expressed P2Y4 subtype with the aid of specific anti-P2Y4 antiserum, and heterologously transfected P2Y4-tagged receptors with the use of antitag antibodies. In both cases, we found the P2Y4 receptor displaying molecular masses corresponding to monomeric, dimeric and oligomeric structures. Experiments performed in the absence of reducing agents demonstrated that there is a strict correlation among the multiple protein bands and that the multimeric forms are at least partially assembled by disulphide bonds. The direct demonstration of P2Y4 homodimerisation comes instead from co–transfection and differential co–immunoprecipitation experiments, with the use of differently tagged P2Y4 receptors and antitag antibodies. The structural propensity of the P2Y4 protein to form homo-oligomers may open the possibility of a novel regulatory mechanism of physiopathological functions for this and additional P2Y receptors