An optogenetic gene expression system with rapid activation and deactivation kinetics

Optogenetic gene expression systems can control transcription with spatial and temporal detail unequaled with traditional inducible promoter systems. However, current eukaryotic light-gated transcription systems are limited by toxicity, dynamic range, or slow activation/deactivation. Here we present an optogenetic gene expression system that addresses these shortcomings and demonstrate its broad utility. Our approach utilizes an engineered version of EL222, a bacterial Light-Oxygen-Voltage (LOV) protein that binds DNA when illuminated with blue light. The system has a large (\u3e100-fold) dynamic range of protein expression, rapid activation (\u3c 10 s) and deactivation kinetics (\u3c 50 s), and a highly linear response to light. With this system, we achieve light-gated transcription in several mammalian cell lines and intact zebrafish embryos with minimal basal gene activation and toxicity. Our approach provides a powerful new tool for optogenetic control of gene expression in space and time

Impact of Scotland’s comprehensive, smoke-free legislation on stroke

<p>Background: Previous studies have reported a reduction in acute coronary events following smoke-free legislation. Evidence is lacking on whether stroke is also reduced. The aim was to determine whether the incidence of stroke, overalland by sub-type, fell following introduction of smoke-free legislation across Scotland on 26 March 2006.</p> <p>Methods and Findings: A negative binomial regression model was used to determine whether the introduction of smokefree legislation resulted in a step and/or slope change in stroke incidence. The model was adjusted for age-group, sex, socioeconomic deprivation quintile, urban/rural residence and month. Interaction tests were also performed. Routine hospital administrative data and death certificates were used to identify all hospital admissions and pre-hospital deaths due to stroke (ICD10 codes I61, I63 and I64) in Scotland between 2000 and 2010 inclusive. Prior to the legislation, rates of all stroke, intracerebral haemorrhage and unspecified stroke were decreasing, whilst cerebral infarction was increasing at 0.97% per annum. Following the legislation, there was a dramatic fall in cerebral infarctions that persisted for around 20 months. No visible effect was observed for other types of stroke. The model confirmed an 8.90% (95% CI 4.85, 12.77, p,0.001) stepwise reduction in cerebral infarction at the time the legislation was implemented, after adjustment for potential cofounders.</p> <p>Conclusions: Following introduction of national, comprehensive smoke-free legislation there was a selective reduction in cerebral infarction that was not apparent in other types of stroke.</p&gt

Patterns of adherence to and compliance with the Portuguese smoke-free law in the leisure-hospitality sector

CIEC – Research Centre on Child Studies, UM (FCT R&D 317)Background: In 2008, the Portuguese smoke-free law came into effect including partial bans in the leisure-hospitality (LH) sector. The objective of the study is to assess the prevalence of smoking control policies (total ban, smoking permission and designated smoking areas) adopted by the LH sector in Portugal. The levels of noncompliance with each policy are investigated as well as the main factors associated with smoking permission and noncompliance with the law. Methods: Cross-sectional study conducted between January 2010 and May 2011. A random sample of venues was selected from the Portuguese LH sector database, proportionally stratified according to type, size and geographical area. All venues were assessed in loco by an observer. The independent effects of venues’ characteristics on smoking permission and the level of noncompliance with the law were explored using logistic regression. Results: Overall, 1.412 venues were included. Total ban policy was adopted by 75.9% of venues, while 8.4% had designated smoking areas. Smoking ban was more prevalent in restaurants (85.9%). Only 29.7% of discos/bars/pubs opted for complete ban. Full or partial smoking permission was higher in discos/bar/pubs (OR = 7.37; 95%CI 4.87 to 11.17). Noncompliance with the law was higher in venues allowing smoking and lower in places with complete ban (33.6% and 7.6% respectively, p, 0.001). Discos/bars/pubs with full smoking permission had the highest level of noncompliance (OR = 3.31; 95%CI 1.40 to 7.83). Conclusions: Our findings show a high adherence to smoking ban policy by the Portuguese LH sector. Nonetheless, one quarter of the venues is fully or partially permissive towards smoking, with the discos/bars/pubs considerably contributing to this situation. Venues with smoking permission policies were less compliant with the legislation. The implementation of a comprehensive smoke-free law, without any exceptions, is essential to effectively protect people from the second hand smoke.The work is part of a large Epidemiological Study on the Portuguese Tobacco Control Policy, developed by the Instituto de Medicina Preventiva da Faculdade de Medicina de Lisboa and supported, in its preliminary part, by the Direccao Geral da Saude (DGS) and, in the second part, by the national funding institution Fundacao para a Ciencia e Tecnologia (FCT). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Reduction in Phencyclidine Induced Sensorimotor Gating Deficits in the Rat Following Increased System Xc − Activity in the Medial Prefrontal Cortex

Rationale: Aspects of schizophrenia, including deficits in sensorimotor gating, have been linked to glutamate dysfunction and/or oxidative stress in the prefrontal cortex. System xc −, a cystine–glutamate antiporter, is a poorly understood mechanism that contributes to both cellular antioxidant capacity and glutamate homeostasis. Objectives: Our goal was to determine whether increased system xc − activity within the prefrontal cortex would normalize a rodent measure of sensorimotor gating. Methods: In situ hybridization was used to map messenger RNA (mRNA) expression of xCT, the active subunit of system xc −, in the prefrontal cortex. Prepulse inhibition was used to measure sensorimotor gating; deficits in prepulse inhibition were produced using phencyclidine (0.3–3 mg/kg, sc). N-Acetylcysteine (10–100 μM) and the system xc − inhibitor (S)-4-carboxyphenylglycine (CPG, 0.5 μM) were used to increase and decrease system xc − activity, respectively. The uptake of 14C-cystine into tissue punches obtained from the prefrontal cortex was used to assay system xc − activity. Results: The expression of xCT mRNA in the prefrontal cortex was most prominent in a lateral band spanning primarily the prelimbic cortex. Although phencyclidine did not alter the uptake of 14C-cystine in prefrontal cortical tissue punches, intraprefrontal cortical infusion of N-acetylcysteine (10–100 μM) significantly reduced phencyclidine- (1.5 mg/kg, sc) induced deficits in prepulse inhibition. N-Acetylcysteine was without effect when coinfused with CPG (0.5 μM), indicating an involvement of system xc −. Conclusions: These results indicate that phencyclidine disrupts sensorimotor gating through system xc − independent mechanisms, but that increasing cystine–glutamate exchange in the prefrontal cortex is sufficient to reduce behavioral deficits produced by phencyclidine

Pacific island regional preparedness for El Niño

The El Niño Southern Oscillation (ENSO) cycle is often blamed for disasters in Pacific island communities. From a disaster risk reduction (DRR) perspective, the challenges with the El Niño part of the ENSO cycle, in particular, are more related to inadequate vulnerability reduction within development than to ENSO-induced hazard influences. This paper analyses this situation, filling in a conceptual and geographic gap in El Niño-related research, by reviewing El Niño-related preparedness (the conceptual gap) for Pacific islands (the geographic gap). Through exploring El Niño impacts on Pacific island communities alongside their vulnerabilities, resiliences, and preparedness with respect to El Niño, El Niño is seen as a constructed discourse rather than as a damaging phenomenon, leading to suggestions for El Niño preparedness as DRR as part of development. Yet the attention which El Niño garners might bring resources to the Pacific region and its development needs, albeit in the short term while El Niño lasts. Conversely, the attention given to El Niño could shift blame from underlying causes of vulnerability to a hazard-centric viewpoint. Instead of focusing on one hazard-influencing phenomenon, opportunities should be created for the Pacific region to tackle wider DRR and development concerns

Intrahepatic cholestasis of pregnancy

Intrahepatic cholestasis of pregnancy (ICP) is a cholestatic disorder characterized by (i) pruritus with onset in the second or third trimester of pregnancy, (ii) elevated serum aminotransferases and bile acid levels, and (iii) spontaneous relief of signs and symptoms within two to three weeks after delivery. ICP is observed in 0.4–1% of pregnancies in most areas of Central and Western Europe and North America, while in Chile and Bolivia as well as Scandinavia and the Baltic states roughly 5–15% and 1–2%, respectively, of pregnancies are associated with ICP. Genetic and hormonal factors, but also environmental factors may contribute to the pathogenesis of ICP. Intrahepatic cholestasis of pregnancy increases the risk of preterm delivery (19–60%), meconium staining of amniotic fluid (27%), fetal bradycardia (14%), fetal distress (22–41%), and fetal loss (0.4–4.1%), particularly when associated with fasting serum bile acid levels > 40 μmol/L. The hydrophilic bile acid ursodeoxycholic acid (10–20 mg/kg/d) is today regarded as the first line treatment for intrahepatic cholestasis of pregnancy. Delivery has been recommended in the 38th week when lung maturity has been established

Safety and feasibility of switching from phenytoin to levetiracetam monotherapy for glioma-related seizure control following craniotomy: a randomized phase II pilot study

Seizures are common in patients with gliomas, and phenytoin (PHT) is frequently used to control tumor-related seizures. PHT, however, has many undesirable side effects (SEs) and drug interactions with glioma chemotherapy. Levetiracetam (LEV) is a newer antiepileptic drug (AED) with fewer SEs and essentially no drug interactions. We performed a pilot study testing the safety and feasibility of switching patients from PHT to LEV monotherapy for postoperative control of glioma-related seizures. Over a 13-month period, 29 patients were randomized in a 2:1 ratio to initiate LEV therapy within 24 h of surgery or to continue PHT therapy. 6 month follow-up data were available for 15 patients taking LEV and for 8 patients taking PHT. In the LEV group, 13 patients (87%) were seizure-free. In the PHT group, 6 patients (75%) were seizure-free. Reported SEs at 6 months was as follows (%LEV/%PHT group): dizziness (0/14), difficulty with coordination (0/29), depression (7/14) lack of energy or strength (20/43), insomnia (40/43), mood instability (7/0). The pilot data presented here suggest that it is safe to switch patients from PHT to LEV monotherapy following craniotomy for supratentorial glioma. A large-scale, double-blinded, randomized control trial of LEV versus PHT is required to determine seizure control equivalence and better assess differences in SEs

The spectral, spatial and contrast sensitivity of human polarization pattern perception

It is generally believed that humans perceive linear polarized light following its conversion into a luminance signal by diattenuating macular structures. Measures of polarization sensitivity may therefore allow a targeted assessment of macular function. Our aim here was to quantify psychophysical characteristics of human polarization perception using grating and optotype stimuli defined solely by their state of linear polarization. We show: (i) sensitivity to polarization patterns follows the spectral sensitivity of macular pigment; (ii) the change in sensitivity across the central field follows macular pigment density; (iii) polarization patterns are identifiable across a range of contrasts and scales, and can be resolved with an acuity of 15.4 cycles/degree (0.29 logMAR); and (iv) the human eye can discriminate between areas of linear polarization differing in electric field vector orientation by as little as 4.4°. These findings, which support the macular diattenuator model of polarization sensitivity, are unique for vertebrates and approach those of some invertebrates with a well-developed polarization sense. We conclude that this sensory modality extends beyond Haidinger's brushes to the recognition of quantifiable spatial polarization-modulated patterns. Furthermore, the macular origin and sensitivity of human polarization pattern perception makes it potentially suitable for the detection and quantification of macular dysfunction

Determination of Therapeutic Equivalence of Generic Products of Gentamicin in the Neutropenic Mouse Thigh Infection Model

Background: Drug regulatory agencies (DRA) support prescription of generic products of intravenous antibiotics assuming therapeutic equivalence from pharmaceutical equivalence. Recent reports of deaths associated with generic heparin and metoprolol have raised concerns about the efficacy and safety of DRA-approved drugs. Methodology/Principal Findings: To challenge the assumption that pharmaceutical equivalence predicts therapeutic equivalence, we determined in vitro and in vivo the efficacy of the innovator product and 20 pharmaceutically equivalent generics of gentamicin. The data showed that, while only 1 generic product failed in vitro (MIC = 45.3 vs. 0.7 mg/L, P,0.05), 10 products (including gentamicin reference powder) failed in vivo against E. coli due to significantly inferior efficacy (E max = 4.81 to 5.32 vs. 5.99 log 10 CFU/g, P#0.043). Although the design lacked power to detect differences in survival after thigh infection with P. aeruginosa, dissemination to vital organs was significantly higher in animals treated with generic gentamicin despite 4 days of maximally effective treatment. Conclusion: Pharmaceutical equivalence does not predict therapeutic equivalence of generic gentamicin. Stricter criteri