Oxidative stress and proinflammatory cytokines contribute to demyelination and axonal damage in a cerebellar culture model of neuroinflammation

Background: Demyelination and axonal damage are critical processes in the pathogenesis of multiple sclerosis (MS). Oxidative stress and pro-inflammatory cytokines elicited by inflammation mediates tissue damage. Methods/Principal Findings: To monitor the demyelination and axonal injury associated with microglia activation we employed a model using cerebellar organotypic cultures stimulated with lipopolysaccharide (LPS). Microglia activated by LPS released pro-inflammatory cytokines (IL-1β, IL-6 and TNFα), and increased the expression of inducible nitric oxide synthase (iNOS) and production of reactive oxygen species (ROS). This activation was associated with demyelination and axonal damage in cerebellar cultures. Axonal damage, as revealed by the presence of non-phosphorylated neurofilaments, mitochondrial accumulation in axonal spheroids, and axonal transection, was associated with stronger iNOS expression and concomitant increases in ROS. Moreover, we analyzed the contribution of pro-inflammatory cytokines and oxidative stress in demyelination and axonal degeneration using the iNOS inhibitor ethyl pyruvate, a free-scavenger and xanthine oxidase inhibitor allopurinol, as well as via blockage of pro-inflammatory cytokines using a Fc-TNFR1 construct. We found that blocking microglia activation with ethyl pyruvate or allopurinol significantly decreased axonal damage, and to a lesser extent, demyelination. Blocking TNFα significantly decreased demyelination but did not prevented axonal damage. Moreover, the most common therapy for MS, interferon-beta, was used as an example of an immunomodulator compound that can be tested in this model. In vitro, interferon-beta treatment decreased oxidative stress (iNOS and ROS levels) and the release of pro-inflammatory cytokines after LPS stimulation, reducing axonal damage. Conclusion: The model of neuroinflammation using cerebellar culture stimulated with endotoxin mimicked myelin and axonal damage mediated by the combination of oxidative stress and pro-inflammatory cytokines. This model may both facilitate understanding of the events involved in neuroinflammation and aid in the development of neuroprotective therapies for the treatment of MS and other neurodegenerative diseases

SiRNA delivery of ENAC mediated by targeted nanocomplex: a therapeutic strategy for cystic fibrosis

The inhibition of ENaC may have therapeutic potential in CF airways by reducing sodium hyperabsorption, restoring lung epithelial surface fluid levels, airway hydration and mucociliary function. The challenge has been to deliver siRNA to the lung with sufficient efficacy for a sustained therapeutic effect. We have developed a self-assembling nanocomplex formulation for siRNA delivery to the airways that consists of a liposome (DOTMA/DOPE; L), an epithelial targeting peptide (P) and siRNA (R). LPR formulations were assessed for their ability to silence expression of the transcript of the gene encoding the α-subunit of the sodium channel ENaC in cell lines and primary epithelial cells, in submerged cultures or grown in air-liquid interface conditions. LPRs, containing 50 nM or 100 nM siRNA, showed high levels of silencing, particularly in primary airway epithelial cells. When nebulised these nanocomplexes still retained their biophysical properties and transfection efficiencies. The silencing ability was determined at protein level by confocal microscopy and western blotting. In vivo data demonstrated that these nanoparticles had the ability to silence expression of the α-ENaC subunit gene. In conclusion, these findings show that LPRs can modulate the activity of ENaC and this approach might be promising as co-adjuvant therapy for cystic fibrosis

A Long Baseline Neutrino Oscillation Experiment Using J-PARC Neutrino Beam and Hyper-Kamiokande

Document submitted to 18th J-PARC PAC meeting in May 2014. 50 pages, 41 figuresDocument submitted to 18th J-PARC PAC meeting in May 2014. 50 pages, 41 figuresDocument submitted to 18th J-PARC PAC meeting in May 2014. 50 pages, 41 figuresHyper-Kamiokande will be a next generation underground water Cherenkov detector with a total (fiducial) mass of 0.99 (0.56) million metric tons, approximately 20 (25) times larger than that of Super-Kamiokande. One of the main goals of Hyper-Kamiokande is the study of $CP$ asymmetry in the lepton sector using accelerator neutrino and anti-neutrino beams. In this document, the physics potential of a long baseline neutrino experiment using the Hyper-Kamiokande detector and a neutrino beam from the J-PARC proton synchrotron is presented. The analysis has been updated from the previous Letter of Intent [K. Abe et al., arXiv:1109.3262 [hep-ex]], based on the experience gained from the ongoing T2K experiment. With a total exposure of 7.5 MW $\times$ 10$^7$ sec integrated proton beam power (corresponding to $1.56\times10^{22}$ protons on target with a 30 GeV proton beam) to a $2.5$-degree off-axis neutrino beam produced by the J-PARC proton synchrotron, it is expected that the $CP$ phase $\delta_{CP}$ can be determined to better than 19 degrees for all possible values of $\delta_{CP}$, and $CP$ violation can be established with a statistical significance of more than $3\,\sigma$ ($5\,\sigma$) for $76$ ($58$) of the $\delta_{CP}$ parameter space

Predictors of children's secondhand smoke exposure at home: a systematic review and narrative synthesis of the evidence

BACKGROUND: Children's exposure to secondhand smoke (SHS) has been causally linked to a number of childhood morbidities and mortalities. Over 50% of UK children whose parents are smokers are regularly exposed to SHS at home. No previous review has identified the factors associated with children's SHS exposure in the home. AIM: To identify by systematic review, the factors which are associated with children's SHS exposure in the home, determined by parent or child reports and/or biochemically validated measures including cotinine, carbon monoxide or home air particulate matter. METHODS: Electronic searches of MEDLINE, EMBASE, PsychINFO, CINAHL and Web of Knowledge to July 2014, and hand searches of reference lists from publications included in the review were conducted. FINDINGS: Forty one studies were included in the review. Parental smoking, low socioeconomic status and being less educated were all frequently and consistently found to be independently associated with children's SHS exposure in the home. Children whose parents held more negative attitudes towards SHS were less likely to be exposed. Associations were strongest for parental cigarette smoking status; compared to children of non-smokers, those whose mothers or both parents smoked were between two and 13 times more likely to be exposed to SHS. CONCLUSION: Multiple factors are associated with child SHS exposure in the home; the best way to reduce child SHS exposure in the home is for smoking parents to quit. If parents are unable or unwilling to stop smoking, they should instigate smoke-free homes. Interventions targeted towards the socially disadvantaged parents aiming to change attitudes to smoking in the presence of children and providing practical support to help parents smoke outside the home may be beneficial

Global Assessment of Extinction Risk to Populations of Sockeye Salmon Oncorhynchus nerka

BACKGROUND: Concern about the decline of wild salmon has attracted the attention of the International Union for the Conservation of Nature (IUCN). The IUCN applies quantitative criteria to assess risk of extinction and publishes its results on the Red List of Threatened Species. However, the focus is on the species level and thus may fail to show the risk to populations. The IUCN has adapted their criteria to apply to populations but there exist few examples of this type of assessment. We assessed the status of sockeye salmon Oncorhynchus nerka as a model for application of the IUCN population-level assessments and to provide the first global assessment of the status of an anadromous Pacific salmon. METHODS/PRINCIPAL FINDINGS: We found from demographic data that the sockeye salmon species is not presently at risk of extinction. We identified 98 independent populations with varying levels of risk within the species' range. Of these, 5 (5%) are already extinct. We analyzed the risk for 62 out of 93 extant populations (67%) and found that 17 of these (27%) are at risk of extinction. The greatest number and concentration of extinct and threatened populations is in the southern part of the North American range, primarily due to overfishing, freshwater habitat loss, dams, hatcheries, and changing ocean conditions. CONCLUSIONS/SIGNIFICANCE: Although sockeye salmon are not at risk at the species-level, about one-third of the populations that we analyzed are at risk or already extinct. Without an understanding of risk to biodiversity at the level of populations, the biodiversity loss in salmon would be greatly underrepresented on the Red List. We urge government, conservation organizations, scientists and the public to recognize this limitation of the Red List. We also urge recognition that about one-third of sockeye salmon global population diversity is at risk of extinction or already extinct

Beyond BMI for self-estimates of body size and shape: A new method for developing stimuli correctly calibrated for body composition

Accurate self-assessment of body shape and size plays a key role in the prevention, diagnosis, and treatment of both obesity and eating disorders. These chronic conditions cause significant health problems, reduced quality of life, and represent a major problem for health services. Variation in body shape depends on two aspects of composition: adiposity and muscularity. However, most self-assessment tools are unidimensional. They depict variation in adiposity only, typically quantified by the body mass index. This can lead to substantial, and clinically meaningful, errors in estimates of body shape and size. To solve this problem, we detail a method of creating biometrically valid body stimuli. We obtained high-resolution 3D body shape scans and composition measures from 397 volunteers (aged 18–45 years) and produced a statistical mapping between the two. This allowed us to create 3D computer-generated models of bodies, correctly calibrated for body composition (i.e., muscularity and adiposity). We show how these stimuli, whose shape changes are based on change in composition in two dimensions, can be used to match the body size and shape participants believe themselves to have, to the stimulus they see. We also show how multivariate multiple regression can be used to model shape change predicted by these 2D outcomes, so that participants’ choices can be explained by their measured body composition together with other psychometric variables. Together, this approach should substantially improve the accuracy and precision with which self-assessments of body size and shape can be made in obese individuals and those suffering from eating disorders

EndoG Links Bnip3-Induced Mitochondrial Damage and Caspase-Independent DNA Fragmentation in Ischemic Cardiomyocytes

Mitochondrial dysfunction, caspase activation and caspase-dependent DNA fragmentation are involved in cell damage in many tissues. However, differentiated cardiomyocytes repress the expression of the canonical apoptotic pathway and their death during ischemia is caspase-independent. The atypical BH3-only protein Bnip3 is involved in the process leading to caspase-independent DNA fragmentation in cardiomyocytes. However, the pathway by which DNA degradation ensues following Bnip3 activation is not resolved. To identify the mechanism involved, we analyzed the interdependence of Bnip3, Nix and EndoG in mitochondrial damage and DNA fragmentation during experimental ischemia in neonatal rat ventricular cardiomyocytes. Our results show that the expression of EndoG and Bnip3 increases in the heart throughout development, while the caspase-dependent machinery is silenced. TUNEL-positive DNA damage, which depends on caspase activity in other cells, is caspase-independent in ischemic cardiomyocytes and ischemia-induced DNA high and low molecular weight fragmentation is blocked by repressing EndoG expression. Ischemia-induced EndoG translocation and DNA degradation are prevented by silencing the expression of Bnip3, but not Nix, or by overexpressing Bcl-xL. These data establish a link between Bnip3 and EndoG-dependent, TUNEL-positive, DNA fragmentation in ischemic cardiomyocytes in the absence of caspases, defining an alternative cell death pathway in postmitotic cells

Quality of Life After Sentinel Lymph Node Biopsy or Axillary Lymph Node Dissection in Stage I/II Breast Cancer Patients: A Prospective Longitudinal Study

Background:\ud Breast cancer patients’ quality of life (QoL) after surgery has been reported to improve significantly over time. Little is known about QoL recovery after sentinel lymph node biopsy (SLNB) in comparison to axillary lymph node dissection (ALND).\ud \ud Methods:\ud 175 of 195 stage I/II breast cancer patients completed the EORTC QLQ-C30: one day before surgery (T0) and after 6 (T1), 26 (T2), 52 (T3) and 104 (T4) weeks. Of these, 54 patients underwent SLNB, 56 SLNB+ALND and 65 ALND. General linear models and paired T-tests between T0–T4 and T1–T4 were computed. Complications, radiotherapy and systemic therapy were added to the model.\ud \ud Results:\ud Significant time effects were found on physical, role and emotional functioning. Physical and role functioning decreased between T0 and T1. At T4, SLNB patients’ functioning had increased to their T0 level; ALND (+/– SLNB) patients’ functioning had increased, but had not improved to T0 level. Emotional functioning increased linearly between T0 and T4. At T4, emotional functioning was significantly higher in all groups as compared with T0. No significant group or interaction (time × group) effects were found. Complications and chemotherapy had a significant negative effect on role, emotional and cognitive functioning. Complications had a significant effect on social functioning also. Effect sizes varied between 0.00 and 0.06.\ud \ud Conclusion:\ud Two years post surgery, breast cancer patients’ QoL is comparable to that shortly before surgery. Women rated their emotional functioning as even better. SLNB is not associated with a better QoL than ALND. However, undergoing systemic therapy and/or experiencing complications affects QoL negatively

Novel Prognostic and Therapeutic Targets for Oral Squamous Cell Carcinoma

In oral squamous cell carcinoma (OSCC), metastasis to lymph nodes is associated with a 50% reduction in 5-year survival. To identify a metastatic gene set based on DNA copy number abnormalities (CNAs) of differentially expressed genes, we compared DNA and RNA of OSCC cells laser-microdissected from non-metastatic primary tumors (n = 17) with those from lymph node metastases (n = 20), using Affymetrix 250K Nsp single-nucleotide polymorphism (SNP) arrays and U133 Plus 2.0 arrays, respectively. With a false discovery rate (FDR)<5%, 1988 transcripts were found to be differentially expressed between primary and metastatic OSCC. Of these, 114 were found to have a significant correlation between DNA copy number and gene expression (FDR<0.01). Among these 114 correlated transcripts, the corresponding genomic regions of each of 95 transcripts had CNAs differences between primary and metastatic OSCC (FDR<0.01). Using an independent dataset of 133 patients, multivariable analysis showed that the OSCC-specific and overall mortality hazards ratio (HR) for patients carrying the 95-transcript signature were 4.75 (95% CI: 2.03-11.11) and 3.45 (95% CI: 1.84-6.50), respectively. To determine the degree by which these genes impact cell survival, we compared the growth of five OSCC cell lines before and after knockdown of over-amplified transcripts via a high-throughput siRNA-mediated screen. The expression-knockdown of 18 of the 26 genes tested showed a growth suppression ≥ 30% in at least one cell line (P<0.01). In particular, cell lines derived from late-stage OSCC were more sensitive to the knockdown of G3BP1 than cell lines derived from early-stage OSCC, and the growth suppression was likely caused by increase in apoptosis. Further investigation is warranted to examine the biological role of these genes in OSCC progression and their therapeutic potentials

Designing antifilarial drug trials using clinical trial simulators

Lymphatic filariasis and onchocerciasis are neglected tropical diseases (NTDs) targeted for elimination by mass (antifilarial) drug administration. These drugs are predominantly active against the microfilarial progeny of adult worms. New drugs or combinations are needed to improve patient therapy and to enhance the effectiveness of interventions in persistent hotspots of transmission. Several therapies and regimens are currently in (pre-)clinical testing. Clinical trial simulators (CTSs) project patient outcomes to inform the design of clinical trials but have not been widely applied to NTDs, where their resource-saving payoffs could be highly beneficial. We demonstrate the utility of CTSs using our individual-based onchocerciasis transmission model (EPIONCHO-IBM) that projects trial outcomes of a hypothetical macrofilaricidal drug. We identify key design decisions that influence the power of clinical trials, including participant eligibility criteria and post-treatment follow-up times for measuring infection indicators. We discuss how CTSs help to inform target product profiles