Transcriptome Kinetics of Circulating Neutrophils during Human Experimental Endotoxemia

Polymorphonuclear cells (neutrophils) play an important role in the systemic inflammatory response syndrome and the development of sepsis. These cells are essential for the defense against microorganisms, but may also cause tissue damage. Therefore, neutrophil numbers and activity are considered to be tightly regulated. Previous studies have investigated gene transcription during experimental endotoxemia in whole blood and peripheral blood mononuclear cells. However, the gene transcription response of the circulating pool of neutrophils to systemic inflammatory stimulation in vivo is currently unclear. We examined neutrophil gene transcription kinetics in healthy human subjects (n = 4) administered a single dose of endotoxin (LPS, 2 ng/kg iv). In addition, freshly isolated neutrophils were stimulated ex vivo with LPS, TNFα, G-CSF and GM-CSF to identify stimulus-specific gene transcription responses. Whole transcriptome microarray analysis of circulating neutrophils at 2, 4 and 6 hours after LPS infusion revealed activation of inflammatory networks which are involved in signaling of TNFα and IL-1α and IL-1β. The transcriptome profile of inflammatory activated neutrophils in vivo reflects extended survival and regulation of inflammatory responses. These changes in neutrophil transcriptome suggest a combination of early activation of circulating neutrophils by TNFα and G-CSF and a mobilization of young neutrophils from the bone marrow

High throughput mutagenesis for identification of residues regulating human prostacyclin (hIP) receptor

The human prostacyclin receptor (hIP receptor) is a seven-transmembrane G protein-coupled receptor (GPCR) that plays a critical role in vascular smooth muscle relaxation and platelet aggregation. hIP receptor dysfunction has been implicated in numerous cardiovascular abnormalities, including myocardial infarction, hypertension, thrombosis and atherosclerosis. Genomic sequencing has discovered several genetic variations in the PTGIR gene coding for hIP receptor, however, its structure-function relationship has not been sufficiently explored. Here we set out to investigate the applicability of high throughput random mutagenesis to study the structure-function relationship of hIP receptor. While chemical mutagenesis was not suitable to generate a mutagenesis library with sufficient coverage, our data demonstrate error-prone PCR (epPCR) mediated mutagenesis as a valuable method for the unbiased screening of residues regulating hIP receptor function and expression. Here we describe the generation and functional characterization of an epPCR derived mutagenesis library compromising >4000 mutants of the hIP receptor. We introduce next generation sequencing as a useful tool to validate the quality of mutagenesis libraries by providing information about the coverage, mutation rate and mutational bias. We identified 18 mutants of the hIP receptor that were expressed at the cell surface, but demonstrated impaired receptor function. A total of 38 non-synonymous mutations were identified within the coding region of the hIP receptor, mapping to 36 distinct residues, including several mutations previously reported to affect the signaling of the hIP receptor. Thus, our data demonstrates epPCR mediated random mutagenesis as a valuable and practical method to study the structurefunction relationship of GPCRs. © 2014 Bill et al

Phosphatidylinositol(4,5)bisphosphate coordinates actin-mediated mobilization and translocation of secretory vesicles to the plasma membrane of chromaffin cells

ORP5 and ORP8, members of the oxysterol-binding protein (OSBP)-related proteins (ORP) family, are endoplasmic reticulum membrane proteins implicated in lipid trafficking. ORP5 and ORP8 are reported to localize to endoplasmic reticulum-plasma membrane junctions via binding to phosphatidylinositol-4-phosphate (PtdIns(4)P), and act as a PtdIns(4)P/phosphatidylserine counter exchanger between the endoplasmic reticulum and plasma membrane. Here we provide evidence that the pleckstrin homology domain of ORP5/8 via PtdIns(4,5)P 2, and not PtdIns(4)P binding mediates the recruitment of ORP5/8 to endoplasmic reticulum-plasma membrane contact sites. The OSBP-related domain of ORP8 can extract and transport multiple phosphoinositides in vitro, and knocking down both ORP5 and ORP8 in cells increases the plasma membrane level of PtdIns(4,5)P 2 with little effect on PtdIns(4)P. Overall, our data show, for the first time, that phosphoinositides other than PtdIns(4)P can also serve as co-exchangers for the transport of cargo lipids by ORPs.ORP5/8 are endoplasmic reticulum (ER) membrane proteins implicated in lipid trafficking that localize to ER-plasma membrane (PM) contacts and maintain membrane homeostasis. Here the authors show that PtdIns(4,5)P 2 plays a critical role in the targeting and function of ORP5/8 at the PM

A Geospatial Modelling Approach Integrating Archaeobotany and Genetics to Trace the Origin and Dispersal of Domesticated Plants

Background: The study of the prehistoric origins and dispersal routes of domesticated plants is often based on the analysis of either archaeobotanical or genetic data. As more data become available, spatially explicit models of crop dispersal can be used to combine different types of evidence. Methodology/Principal Findings: We present a model in which a crop disperses through a landscape that is represented by a conductance matrix. From this matrix, we derive least-cost distances from the geographical origin of the crop and use these to predict the age of archaeological crop remains and the heterozygosity of crop populations. We use measures of the overlap and divergence of dispersal trajectories to predict genetic similarity between crop populations. The conductance matrix is constructed from environmental variables using a number of parameters. Model parameters are determined with multiple-criteria optimization, simultaneously fitting the archaeobotanical and genetic data. The consilience reached by the model is the extent to which it converges around solutions optimal for both archaeobotanical and genetic data. We apply the modelling approach to the dispersal of maize in the Americas. Conclusions/Significance: The approach makes possible the integrative inference of crop dispersal processes, whil

Reduced auditory steady state responses in autism spectrum disorder

Background Auditory steady state responses (ASSRs) are elicited by clicktrains or amplitude-modulated tones, which entrain auditory cortex at their specific modulation rate. Previous research has reported reductions in ASSRs at 40 Hz for autism spectrum disorder (ASD) participants and first-degree relatives of people diagnosed with ASD (Mol Autism. 2011;2:11, Biol Psychiatry. 2007;62:192–197). Methods Using a 1.5 s-long auditory clicktrain stimulus, designed to elicit an ASSR at 40 Hz, this study attempted to replicate and extend these findings. Magnetencephalography (MEG) data were collected from 18 adolescent ASD participants and 18 typically developing controls. Results The ASSR localised to bilateral primary auditory regions. Regions of interest were thus defined in left and right primary auditory cortex (A1). While the transient gamma-band response (tGBR) from 0-0.1 s following presentation of the clicktrain stimulus was not different between groups, for either left or right A1, the ASD group had reduced oscillatory power at 40 Hz from 0.5 to 1.5 s post-stimulus onset, for both left and right A1. Additionally, the ASD group had reduced inter-trial coherence (phase consistency over trials) at 40 Hz from 0.64-0.82 s for right A1 and 1.04-1.22 s for left A1. Limitations In this study, we did not conduct a clinical autism assessment (e.g. the ADOS), and therefore, it remains unclear whether ASSR power and/or ITC are associated with the clinical symptoms of ASD. Conclusion Overall, our results support a specific reduction in ASSR oscillatory power and inter-trial coherence in ASD, rather than a generalised deficit in gamma-band responses. We argue that this could reflect a developmentally relevant reduction in non-linear neural processing

Identification of recruitment and retention strategies for rehabilitation professionals in Ontario, Canada: results from expert panels

<p>Abstract</p> <p>Background</p> <p>Demand for rehabilitation services is expected to increase due to factors such as an aging population, workforce pressures, rise in chronic and complex multi-system disorders, advances in technology, and changes in interprofessional health service delivery models. However, health human resource (HHR) strategies for Canadian rehabilitation professionals are lagging behind other professional groups such as physicians and nurses. The objectives of this study were: 1) to identify recruitment and retention strategies of rehabilitation professionals including occupational therapists, physical therapists and speech language pathologists from the literature; and 2) to investigate both the importance and feasibility of the identified strategies using expert panels amongst HHR and education experts.</p> <p>Methods</p> <p>A review of the literature was conducted to identify recruitment and retention strategies for rehabilitation professionals. Two expert panels, one on <it>Recruitment and Retention </it>and the other on <it>Education </it>were convened to determine the importance and feasibility of the identified strategies. A modified-delphi process was used to gain consensus and to rate the identified strategies along these two dimensions.</p> <p>Results</p> <p>A total of 34 strategies were identified by the <it>Recruitment and Retention </it>and <it>Education </it>expert panels as being important and feasible for the development of a HHR plan for recruitment and retention of rehabilitation professionals. Seven were categorized under the <it>Quality of Worklife and Work Environment </it>theme, another seven in <it>Financial Incentives and Marketing</it>, two in <it>Workload and Skill Mix</it>, thirteen in <it>Professional Development </it>and five in <it>Education and Training</it>.</p> <p>Conclusion</p> <p>Based on the results from the expert panels, the three major areas of focus for HHR planning in the rehabilitation sector should include strategies addressing <it>Quality of Worklife and Work Environment</it>, <it>Financial Incentives and Marketing </it>and <it>Professional Development</it>.</p

A Functional and Structural Investigation of the Human Fronto-Basal Volitional Saccade Network

Almost all cortical areas are connected to the subcortical basal ganglia (BG) through parallel recurrent inhibitory and excitatory loops, exerting volitional control over automatic behavior. As this model is largely based on non-human primate research, we used high resolution functional MRI and diffusion tensor imaging (DTI) to investigate the functional and structural organization of the human (pre)frontal cortico-basal network controlling eye movements. Participants performed saccades in darkness, pro- and antisaccades and observed stimuli during fixation. We observed several bilateral functional subdivisions along the precentral sulcus around the human frontal eye fields (FEF): a medial and lateral zone activating for saccades in darkness, a more fronto-medial zone preferentially active for ipsilateral antisaccades, and a large anterior strip along the precentral sulcus activating for visual stimulus presentation during fixation. The supplementary eye fields (SEF) were identified along the medial wall containing all aforementioned functions. In the striatum, the BG area receiving almost all cortical input, all saccade related activation was observed in the putamen, previously considered a skeletomotor striatal subdivision. Activation elicited by the cue instructing pro or antisaccade trials was clearest in the medial FEF and right putamen. DTI fiber tracking revealed that the subdivisions of the human FEF complex are mainly connected to the putamen, in agreement with the fMRI findings. The present findings demonstrate that the human FEF has functional subdivisions somewhat comparable to non-human primates. However, the connections to and activation in the human striatum preferentially involve the putamen, not the caudate nucleus as is reported for monkeys. This could imply that fronto-striatal projections for the oculomotor system are fundamentally different between humans and monkeys. Alternatively, there could be a bias in published reports of monkey studies favoring the caudate nucleus over the putamen in the search for oculomotor functions

The autonomous city: towards a critical geography of occupation

This paper explores the recent resurgence of occupation-based practices across the globe, from the seizure of public space to the assembling of improvised protest camps. It re-examines the relationship between the figure of occupation and the affirmation of an alternative ‘right to the city’. The paper develops a critical understanding of occupation as a political process that prefigures and materializes the social order which it seeks to enact. The paper highlights the constituent role of occupation as an autonomous form of urban dwelling, as a radical politics of infrastructure and as a set of relations that produce common spaces for political action

Increased Risk of Fragility Fractures among HIV Infected Compared to Uninfected Male Veterans

BACKGROUND: HIV infection has been associated with an increased risk of fragility fracture. We explored whether or not this increased risk persisted in HIV infected and uninfected men when controlling for traditional fragility fracture risk factors. METHODOLOGY/PRINCIPAL FINDINGS: Cox regression models were used to assess the association of HIV infection with the risk for incident hip, vertebral, or upper arm fracture in male Veterans enrolled in the Veterans Aging Cohort Study Virtual Cohort (VACS-VC). We calculated adjusted hazard ratios comparing HIV status and controlling for demographics and other established risk factors. The sample consisted of 119,318 men, 33% of whom were HIV infected (34% aged 50 years or older at baseline, and 55% black or Hispanic). Median body mass index (BMI) was lower in HIV infected compared with uninfected men (25 vs. 28 kg/m²; p<0.0001). Unadjusted risk for fracture was higher among HIV infected compared with uninfected men [HR: 1.32 (95% CI: 1.20, 1.47)]. After adjusting for demographics, comorbid disease, smoking and alcohol abuse, HIV infection remained associated with an increased fracture risk [HR: 1.24 (95% CI: 1.11, 1.39)]. However, adjusting for BMI attenuated this association [HR: 1.10 (95% CI: 0.97, 1.25)]. The only HIV-specific factor associated with fragility fracture was current protease inhibitor use [HR: 1.41 (95% CI: 1.16, 1.70)]. CONCLUSIONS/SIGNIFICANCE: HIV infection is associated with fragility fracture risk. This risk is attenuated by BMI