Mechanism of rapid desensitization of muscarinic K+ current in adult rat and guinea pig atrial cells.
Acetylcholine (Ach) activates the muscarinic K+ current in atrial cells via the inhibitory GTP binding protein. After activation, the whole-cell K+ current decreases rapidly (rapid desensitization) to approximately half of the initial current within approximately 20 seconds. The mechanism of this rapid desensitization was investigated in adult rat and guinea pig atrial cells. Whole-cell voltage-clamp and patch-clamp techniques were used to study the K+ current. In voltage-clamped whole cells, ACh activated a K+ current that desensitized rapidly during the initial approximately 20 seconds followed by a slower decrease over several minutes. The rapid K+ current desensitization (a rapid decrease in channel open probability) was also observed at the single-channel level in cell-attached patches and was associated with a progressive shortening of the channel open time and prolongation of the closed time. These changes in channel current and kinetics were abolished by removal of the cytoplasm (by forming inside-out patches) and were partially inhibited by phosphatase inhibitors, suggesting an involvement of cytosolic phosphatase(s) in K+ current desensitization. In inside-out patches with ACh in the pipette and GTP in the bath, the open time of muscarinic K+ channels and channel open probability were increased by 1 mM Mg(2+)-ATP (but not by the nonhydrolyzable analogue, adenylylimidodiphosphate) and decreased by alkaline phosphatase. These results suggest that the rapid K+ current desensitization in adult rat or guinea pig atrial cells is produced by changes in the gating kinetics of the K+ channel, possibly mediated via membrane-associated protein kinase and cytosolic phosphatase(s).


Novel Bat Influenza Virus NS1 Proteins Bind Double-Stranded RNA and Antagonize Host Innate Immunity.
We demonstrate that novel bat HL17NL10 and HL18NL11 influenza virus NS1 proteins are effective interferon antagonists but do not block general host gene expression. Solving the RNA-binding domain structures revealed the canonical NS1 symmetrical homodimer, and RNA binding required conserved basic residues in this domain. Interferon antagonism was strictly dependent on RNA binding, and chimeric bat influenza viruses expressing NS1s defective in this activity were highly attenuated in interferon-competent cells but not in cells unable to establish antiviral immunity.


Repurposing of Guanabenz acetate by encapsulation into long-circulating nanopolymersomes for treatment of triple-negative breast cancer.
Poor patient response and limited treatment modalities are the major challenges against combating triple-negative breast cancer (TNBC). The high related mortality urges for novel cancer therapeutics. Guanabenz acetate (GA) is an orphan antihypertensive drug with a short half-life. Re-purposing (GA) by developing a polymersome (PS)-based cancer nanomedicine is an innovative approach in treating TNBC. Formulation and optimization of GA-loaded PEGylated Polycaprolactone PS through different process variables (solvent selection, the order of addition, pH of the aqueous phase, and drug to polymer ratio) were achieved by the nanoprecipitation method. The in vitro cellular uptake, anti-cancer, and anti-metastatic activity of GA and GA-loaded PS were tested in MDA-MB 231(TNBC cell line) and MCF-7 cell line. Western blot analysis was performed to elucidate the molecular anti-cancer mechanism. The in vivo biodistribution study and antitumor activity were investigated in the TNBC-xenograft model implanted in mice. Under optimized formulation conditions, GA-loaded PS had a nanosize of 90.5 nm with PDI < 0.2, a zeta potential -9.11 mV, drug encapsulation efficiency of 92.11% and sustained drug release for 6-days. GA-loaded PS exhibited enhanced cellular uptake and achieved a significantly lower IC50 in both breast cancer cell lines compared to free GA. Treatment with GA-loaded PS (60 µM) showed a significant reduction of 60.5 and 78.1% in cancer migration and metastasis in the case of MDA-MB 231 and MCF-7, respectively. Besides, drug-loaded PS increased phosphorylation of translational regulator eIF2α and decreased expression of Rac1 which were essential for decreasing cancer cell survival and metastasis. In vivo biodistribution study of GA-loaded PS showed long-circulating PS with high passively targeted tumor accumulation. Treatment with GA-loaded PS resulted in a significant decrease in tumor size and weight compared to free GA. In conclusion, GA-loaded PS is a new promising cancer therapeutics for the treatment of TNBC.


Phospholemman: a novel cardiac stress protein.
Phospholemman (PLM), a member of the FXYD family of regulators of ion transport, is a major sarcolemmal substrate for protein kinases A and C in cardiac and skeletal muscle. In the heart, PLM co-localizes and co-immunoprecipitates with Na(+)-K(+)-ATPase, Na(+)/Ca(2+) exchanger, and L-type Ca(2+) channel. Functionally, when phosphorylated at serine(68), PLM stimulates Na(+)-K(+)-ATPase but inhibits Na(+)/Ca(2+) exchanger in cardiac myocytes. In heterologous expression systems, PLM modulates the gating of cardiac L-type Ca(2+) channel. Therefore, PLM occupies a key modulatory role in intracellular Na(+) and Ca(2+) homeostasis and is intimately involved in regulation of excitation-contraction (EC) coupling. Genetic ablation of PLM results in a slight increase in baseline cardiac contractility and prolongation of action potential duration. When hearts are subjected to catecholamine stress, PLM minimizes the risks of arrhythmogenesis by reducing Na(+) overload and simultaneously preserves inotropy by inhibiting Na(+)/Ca(2+) exchanger. In heart failure, both expression and phosphorylation state of PLM are altered and may partly account for abnormalities in EC coupling. The unique role of PLM in regulation of Na(+)-K(+)-ATPase, Na(+)/Ca(2+) exchanger, and potentially L-type Ca(2+) channel in the heart, together with the changes in its expression and phosphorylation in heart failure, make PLM a rational and novel target for development of drugs in our armamentarium against heart failure. Clin Trans Sci 2010; Volume 3: 189-196.


Characterisation of the putative effector interaction site of the regulatory HbpR protein from Pseudomonas azelaica by site-directed mutagenesis.
Bacterial transcription activators of the XylR/DmpR subfamily exert their expression control via σ(54)-dependent RNA polymerase upon stimulation by a chemical effector, typically an aromatic compound. Where the chemical effector interacts with the transcription regulator protein to achieve activation is still largely unknown. Here we focus on the HbpR protein from Pseudomonas azelaica, which is a member of the XylR/DmpR subfamily and responds to biaromatic effectors such as 2-hydroxybiphenyl. We use protein structure modeling to predict folding of the effector recognition domain of HbpR and molecular docking to identify the region where 2-hydroxybiphenyl may interact with HbpR. A large number of site-directed HbpR mutants of residues in- and outside the predicted interaction area was created and their potential to induce reporter gene expression in Escherichia coli from the cognate P(C) promoter upon activation with 2-hydroxybiphenyl was studied. Mutant proteins were purified to study their conformation. Critical residues for effector stimulation indeed grouped near the predicted area, some of which are conserved among XylR/DmpR subfamily members in spite of displaying different effector specificities. This suggests that they are important for the process of effector activation, but not necessarily for effector specificity recognition.


Evidence for stabilization of aquaporin-2 folding mutants by N-linked glycosylation in endoplasmic reticulum.
Aquaporin-2 (AQP2) is the vasopressin-sensitive water channel that regulates water reabsorption in the distal nephron collecting duct. Inherited AQP2 mutations that disrupt folding lead to nephrogenic diabetes insipidus (NDI) by targeting newly synthesized protein for degradation in the endoplasmic reticulum (ER). During synthesis, a subset of wild-type (WT) AQP2 is covalently modified by N-linked glycosylation at residue Asn123. To investigate the affect of glycosylation, we expressed WT AQP2 and four NDI-related mutants in Xenopus laevis oocytes and compared stability of glycosylated and nonglycosylated isoforms. In all constructs, approximately 15-20% of newly synthesized AQP2 was covalently modified by N-linked glycosylation. At steady state, however, core glycosylated WT protein was nearly undetectable, whereas all mutants were found predominantly in the glycosylated form (60-70%). Pulse-chase metabolic labeling studies revealed that glycosylated isoforms of mutant AQP2 were significantly more stable than their nonglycosylated counterparts. For nonglycosylated isoforms, the half-life of WT AQP2 was significantly greater (>48 h) than that of mutant AQP2 (T126M 4.1 +/- 1.0 h, A147T 4.2 +/- 0.60 h, C181W 4.5 +/- 0.50 h, R187C 6.8 +/- 1.2 h). This is consistent with rapid turnover in the ER as previously reported. In contrast, the half-lives of mutant proteins containing N-linked glycans were similar to WT (approximately 25 h), indicating that differences in steady-state glycosylation profiles are caused by increased stability of glycosylated mutant proteins. These results suggest that addition of a single N-linked oligosaccharide moiety can partially compensate for ER folding defects induced by disease-related mutations.


A pilot randomized controlled trial evaluating motivationally matched pedometer feedback to increase physical activity behavior in older adults.
Increasing physical activity (PA) levels in older adults represents an important public health challenge. The purpose of this study was to evaluate the feasibility of combining individualized motivational messaging with pedometer walking step targets to increase PA in previously inactive and insufficiently active older adults. In this 12-week intervention study older adults were randomized to 1 of 4 study arms: Group 1--control; Group 2--pedometer 10,000 step goal; Group 3--pedometer step goal plus individualized motivational feedback; or Group 4--everything in Group 3 augmented with biweekly telephone feedback. 81 participants were randomized into the study, 61 participants completed the study with an average age of 63.8 ± 6.0 years. Group 1 did not differ in accumulated steps/day following the 12-week intervention compared with participants in Group 2. Participants in Groups 3 and 4 took on average 2159 (P < .001) and 2488 (P < .001) more steps/day, respectively, than those in Group 1 after the 12-week intervention. In this 12-week pilot randomized control trial, a pedometer feedback intervention partnered with individually matched motivational messaging was an effective intervention strategy to significantly increase PA behavior in previously inactive and insufficiently active older adults.


The histone deacetylase inhibitor panobinostat is a potent antitumor agent in canine diffuse large B-cell lymphoma.
Non-Hodgkin lymphoma (NHL) is one of the most common causes of cancer-related death in the United States and Europe. Although the outcome of NHL patients has improved over the last years with current therapies, the rate of mortality is still high. A plethora of new drugs is entering clinical development for NHL treatment; however, the approval of new treatments remains low due in part to the paucity of clinically relevant models for validation. Canine lymphoma shares remarkable similarities with its human counterpart, making the dog an excellent animal model to explore novel therapeutic molecules and approaches. Histone deacetylase inhibitors (HDACis) have emerged as a powerful new class of anti-cancer drugs for human therapy. To investigate HDACi antitumor properties on canine diffuse large B-cell lymphoma, a panel of seven HDACi compounds (CI-994, panobinostat, SBHA, SAHA, scriptaid, trichostatin A and tubacin) was screened on CLBL-1 canine B-cell lymphoma cell line. Our results demonstrated that all HDACis tested exhibited dose-dependent inhibitory effects on proliferation of CLBL-1 cells, while promoting increased H3 histone acetylation. Amongst all HDACis studied, panobinostat proved to be the most promising compound and was selected for further in vitro and in vivo evaluation. Panobinostat cytotoxicity was linked to H3 histone and α-tubulin acetylation, and to apoptosis induction. Importantly, panobinostat efficiently inhibited CLBL-1 xenograft tumor growth, and strongly induced acetylation of H3 histone and apoptosis in vivo. In conclusion, these results provide new data validating HDACis and, especially, panobinostat as a novel anti-cancer therapy for veterinary applications, while contributing to comparative oncology.


Stretch induced hyperexcitability of mice callosal pathway.
Memory and learning are thought to result from changes in synaptic strength. Previous studies on synaptic physiology in brain slices have traditionally been focused on biochemical processes. Here, we demonstrate with experiments on mouse brain slices that central nervous system plasticity is also sensitive to mechanical stretch. This is important, given the host of clinical conditions involving changes in mechanical tension on the brain, and the normal role that mechanical tension plays in brain development. A novel platform is developed to investigate neural responses to mechanical stretching. Flavoprotein autofluoresence (FA) imaging was employed for measuring neural activity. We observed that synaptic excitability substantially increases after a small (2.5%) stretch was held for 10 min and released. The increase is accumulative, i.e., multiple stretch cycles further increase the excitability. We also developed analytical tools to quantify the spatial spread and response strength. Results show that the spatial spread is less stable in slices undergoing the stretch-unstretch cycle. FA amplitude and activation rate decrease as excitability increases in stretch cases but not in electrically enhanced cases. These results collectively demonstrate that a small stretch in physiological range can modulate neural activities significantly, suggesting that mechanical events can be employed as a novel tool for the modulation of neural plasticity.


Experience with high frequency oscillation ventilation during the 2009 H1N1 influenza pandemic in Australia and New Zealand.
During the 2009 H1N1 pandemic, large numbers of patients had severe respiratory failure. High frequency oscillation ventilation was used as a salvage technique for profound hypoxaemia. Our aim was to compare this experience with high frequency oscillation ventilation during the 2009 H1N1 pandemic with the same period in 2008 by performing a three-month period prevalence study in Australian and New Zealand intensive care units. The main study end-points were clinical demographics, care delivery and survival. Nine intensive care units contributed data. During 2009 there were 22 H1N1 patients (17 adults, five children) and 10 non-H1N1 patients (five adults, five children), while in 2008, 18 patients (two adults, 16 children) received high frequency oscillation ventilation. The principal non-H1N1 high frequency oscillation ventilation indication was bacterial or viral pneumonia (56%). For H1N1 patients, the median duration of high frequency oscillation ventilation was 3.7 days (interquartile range 1.8 to 5) with concomitant therapies including recruitment manoeuvres (22%), prone ventilation (41%), inhaled prostacyclins (18%) and inhaled nitric oxide (36%). Seven patients received extracorporeal membrane oxygenation, six having H1N1. Three patients had extracorporeal membrane oxygenation concurrently, two as salvage therapy following the commencement of high frequency oscillation ventilation. In 2008, no high frequency oscillation ventilation patient received extracorporeal membrane oxygenation. Overall hospital survival was 77% in H1N1 patients, while survival in patients having adjunctive extracorporeal membrane oxygenation was similar to those receiving high frequency oscillation ventilation alone (65% compared to 71%, P = 1.00). Survival rates were comparable to published extracorporeal membrane oxygenation outcomes. High frequency oscillation ventilation was used successfully as a rescue therapy for severe respiratory failure. High frequency oscillation ventilation was only available in a limited number of intensive care units during the H1N1 pandemic.


Circadian Expression of Migratory Factors Establishes Lineage-Specific Signatures that Guide the Homing of Leukocyte Subsets to Tissues.
The number of leukocytes present in circulation varies throughout the day, reflecting bone marrow output and emigration from blood into tissues. Using an organism-wide circadian screening approach, we detected oscillations in pro-migratory factors that were distinct for specific vascular beds and individual leukocyte subsets. This rhythmic molecular signature governed time-of-day-dependent homing behavior of leukocyte subsets to specific organs. Ablation of BMAL1, a transcription factor central to circadian clock function, in endothelial cells or leukocyte subsets demonstrated that rhythmic recruitment is dependent on both microenvironmental and cell-autonomous oscillations. These oscillatory patterns defined leukocyte trafficking in both homeostasis and inflammation and determined detectable tumor burden in blood cancer models. Rhythms in the expression of pro-migratory factors and migration capacities were preserved in human primary leukocytes. The definition of spatial and temporal expression profiles of pro-migratory factors guiding leukocyte migration patterns to organs provides a resource for the further study of the impact of circadian rhythms in immunity.


Impact of long-term diesel contamination on soil microbial community structure.
Microbial community composition and diversity at a diesel-contaminated railway site were investigated by pyrosequencing of bacterial and archaeal 16S rRNA gene fragments to understand the interrelationships among microbial community composition, pollution level, and soil geochemical and physical properties. To this end, 26 soil samples from four matrix types with various geochemical characteristics and contaminant concentrations were investigated. The presence of diesel contamination significantly impacted microbial community composition and diversity, regardless of the soil matrix type. Clean samples showed higher diversity than contaminated samples (P < 0.001). Bacterial phyla with high relative abundances in all samples included Proteobacteria, Firmicutes, Actinobacteria, Acidobacteria, and Chloroflexi. High relative abundances of Archaea, specifically of the phylum Euryarchaeota, were observed in contaminated samples. Redundancy analysis indicated that increased relative abundances of the phyla Chloroflexi, Firmicutes, and Euryarchaeota correlated with the presence of contamination. Shifts in the chemical composition of diesel constituents across the site and the abundance of specific operational taxonomic units (OTUs; defined using a 97% sequence identity threshold) in contaminated samples together suggest that natural attenuation of contamination has occurred. OTUs with sequence similarity to strictly anaerobic Anaerolineae within the Chloroflexi, as well as to Methanosaeta of the phylum Euryarchaeota, were detected. Anaerolineae and Methanosaeta are known to be associated with anaerobic degradation of oil-related compounds; therefore, their presence suggests that natural attenuation has occurred under anoxic conditions. This research underscores the usefulness of next-generation sequencing techniques both to understand the ecological impact of contamination and to identify potential molecular proxies for detection of natural attenuation.


Rhizobium common nod genes are required for biofilm formation.
In legume nitrogen-fixing symbioses, rhizobial nod genes are obligatory for initiating infection thread formation and root nodule development. Here we show that the common nod genes, nodD1ABC, whose products synthesize core Nod factor, a chitin-like oligomer, are also required for the establishment of the three-dimensional architecture of the biofilm of Sinorhizobium meliloti. Common nod gene mutants form a biofilm that is a monolayer. Moreover, adding Nod Factor antibody to S. meliloti cells inhibits biofilm formation, while chitinase treatment disrupts pre-formed biofilms. These results attest to the involvement of core Nod factor in rhizobial biofilm establishment. However, luteolin, the plant-derived inducer of S. meliloti's nod genes, is not required for mature biofilm formation, although biofilm establishment is enhanced in the presence of this flavonoid inducer. Because biofilm formation is plant-inducer-independent and because all nodulating rhizobia, both alpha- and beta-proteobacteria have common nod genes, the role of core Nod factor in biofilm formation is likely to be an ancestral and evolutionarily conserved function of these genes.


Role of periostin, FENO, IL-13, lebrikzumab, other IL-13 antagonist and dual IL-4/IL-13 antagonist in asthma.
Asthma markedly diminishes quality of life due to limited activity, absences from work or school and hospitalizations. Patients with severe asthma which are not controlled despite taking effective therapy are most in need of new treatment approaches. IL-13 was demonstrated as 'central mediator of allergic asthma'. IL-13 has been implicated in the pathogenesis of asthma, idiopathic pulmonary fibrosis and COPD. IL-13 levels in the sputum and bronchial biopsy samples remain elevated in severe asthma despite the use of inhaled and systemic corticosteroids. Thus, IL-13 is a mediator involved in corticosteroid resistance. Periostin enhances profibrotic TGF-β signaling in subepithelial fibrosis associated with asthma. IL-13 induces bronchial epithelial cells to secrete periostin. Periostin may be a biomarker for Th2 induced airway inflammation. Lebrikizumab is a monoclonal antibody against IL-13. Lebrikizumab improved lung function in asthmatics who were symptomatic despite treatment with long acting beta agonist and inhaled corticosteroids and provided benefit in the treatment of severe uncontrolled asthma. Lebrikizumab block IL-13 signaling through the IL-13Rα1/IL-4Rα receptor. There was a larger reduction in FENO in the high periostin subgroup than in the low periostin subgroup (34.4 vs 4.3%). Serum CCL17, CCL13 and total IgE levels decreased in the lebrikizumab group.


Development of a liposomal nanoparticle formulation of 5-fluorouracil for parenteral administration: formulation design, pharmacokinetics and efficacy.
5-Fluorouracil (5-FU) is a small, very membrane permeable drug that is poorly retained within the aqueous compartment of liposomal nanoparticles (LNP). To address this problem a novel method relying on formation of a ternary complex comprising copper, low molecular weight polyethylenimine (PEI) and 5-FU has been developed. More specifically, in the presence of entrapped copper and PEI, externally added 5-FU can be efficiently encapsulated (>95%) in DSPC/Chol (1,2-Distearoyl-sn-Glycero-3-Phosphocholine/cholesterol; 55:45 mol%) liposomes (130-170 nm) to achieve drug-to-lipid ratios of 0.1 (mol:mol). Drug release studies completed using this LNP formulation of 5-FU demonstrated significant improvements in drug retention in vitro and in vivo. Plasma concentrations of 5-FU were 7- to 23-fold higher when the drug was administered intravenously to mice as the LNP 5-FU formulation compared to free 5-FU. Further, the therapeutic effects of the LNP 5-FU formulation, as determined in a HT-29 subcutaneous colorectal cancer model where treatment was given QDx5, was greater than that which could be achieved with free 5-FU when compared at equivalent doses. This is the first time an active loading method has been described for 5-FU. The use of ternary metal complexation strategy to encapsulate therapeutic agents may define a unique platform for preparation of LNP drug formulations.


Motor learning and metaplasticity in striatal neurons: relevance for Parkinson's disease.
Nigro-striatal dopamine transmission is central to a wide range of neuronal functions, including skill learning, which is disrupted in several pathologies such as Parkinson's disease. The synaptic plasticity mechanisms, by which initial motor learning is stored for long time periods in striatal neurons, to then be gradually optimized upon subsequent training, remain unexplored. Addressing this issue is crucial to identify the synaptic and molecular mechanisms involved in striatal-dependent learning impairment in Parkinson's disease. In this study, we took advantage of interindividual differences between outbred rodents in reaching plateau performance in the rotarod incremental motor learning protocol, to study striatal synaptic plasticity ex vivo. We then assessed how this process is modulated by dopamine receptors and the dopamine active transporter, and whether it is impaired by overexpression of human α-synuclein in the mesencephalon; the latter is a progressive animal model of Parkinson's disease. We found that the initial acquisition of motor learning induced a dopamine active transporter and D1 receptors mediated long-term potentiation, under a protocol of long-term depression in striatal medium spiny neurons. This effect disappeared in animals reaching performance plateau. Overexpression of human α-synuclein reduced striatal dopamine active transporter levels, impaired motor learning, and prevented the learning-induced long-term potentiation, before the appearance of dopamine neuronal loss. Our findings provide evidence of a reorganization of cellular plasticity within the dorsolateral striatum that is mediated by dopamine receptors and dopamine active transporter during the acquisition of a skill. This newly identified mechanism of cellular memory is a form of metaplasticity that is disrupted in the early stage of synucleinopathies, such as Parkinson's disease, and that might be relevant for other striatal pathologies, such as drug abuse.


Reducing overestimation in reported mobile phone use associated with epidemiological studies.
Case-control studies of mobile phones are commonly based on retrospective, self-reported exposure information, which are often characterized as involving substantial uncertainty concerning data validity. We assessed the validity of self-reported mobile phone use and developed a statistical model to account for the over-reporting of exposure. We collected information on mobile phone use from 70 volunteers using two sources of data: self-report in an interview and network operator records. We used regression models to obtain bias-corrected estimates of exposure. A correlation coefficient of 0.71 was obtained between the self-reported and the network operators' data on average calling time (log-transformed minutes per month). A simple linear regression model, where the duration of calls acquired from network operators is explained with the self-reported duration fitted the data reasonably well (adjusted R(2) 0.51). The constant term was 2.71 and the regression coefficient 0.49 (logarithmic scale). No significant improvement in the model fit was achieved by including potential predictors of accuracy in self-reported exposure estimates, such as the pattern of mobile phone use, the modality of response to the questionnaire or demographic characteristics. Overestimation in self-reported intensity of mobile phone use can be accounted for by the use of regression calibration. The estimates obtained in our study may not be applicable in other contexts, but similar methods could be used to reduce bias in other studies.


A Parkinson's disease measurement system using laser lines and a CMOS image sensor.
This paper presents a non-invasive, non-contact system for the measurement of the arterial dorsum manus vibration waveforms of Parkinson disease patients. The laser line method is applied to detect the dorsum manus vibration in rest and postural situations. The proposed measurement system mainly consists of a laser diode and a low cost complementary metal-oxide semiconductor (CMOS) image sensor. Laser line and centroid methods are combined with the Fast Fourier Transform (FFT) in this study. The shape and frequency and relative frequency of the dorsum manus vibration waveforms can be detected rapidly using our Parkinson's disease measurement system. A laser line near the wrist joint is used as the testing line. The experimental results show an obvious increase in the amplitude and frequency of dorsum manus variation in the measured region in patients suffering from Parkinson's disease, indicating the obvious effects of the disease. Both in postural and rest state measurements, as the patient disease age increases the vibration frequency increases. The measurement system is well suited for evaluating and pre-diagnosing early stage Parkinson's disease.


Performance of an adapted household food insecurity access scale in measuring seasonality in household food insecurity in rural Ethiopia: a cohort analysis.
Seasonality poses a considerable food security challenge in Ethiopia. Yet, measuring seasonal variations in food insecurity, particularly the dimension of food access, lacks an adequately validated tool. We therefore evaluated the performance of an adapted Household Food Insecurity Access Scale (HFIAS) to estimate seasonal variations in food insecurity (FI) among subsistence villagers in Ethiopia. We employed a cohort study design using a panel of four repeated measurements taken in June, September, and December in the year 2017, and in March 2018. The study recruited 473 villagers from the drought-affected Wolaita area in southwest Ethiopia. The performance of the HFIAS was evaluated via internal consistency (Chronbach's alpha values) and criterion validation techniques. The set of criteria include: parallelism between affirmative responses to FI questions and wealth strata; dose-response relationship between FI and dietary intake; and also FI severity and household wealth status. This study revealed that the HFIAS had satisfactory performance in four repeated measurements. The likelihood of affirmative responses to questions about FI decreased with ascending wealth quintiles. We observed an inverse dose-response relationship between FI and wealth status, and between FI and household dietary diversity. The HFIAS showed an acceptable potential for measuring seasonal variations in FI in the study area. Our findings complement efforts to evaluate the scale's applicability in various settings, in order to promote cross-culture monitoring and comparisons. However, it required a careful adaption for contextual and cultural sensitivities.


Heterogeneous thermal-lens immunoassay for small organic compounds: determination of 4-aminophenol.
4-Aminophenol was selected as a model hapten for thermal-lens detection in a heterogeneous indirect immunoassay for small organic compounds. The assay is based on the competition of the free hapten and 4-aminophenol labeled with tetramethyl rhodamine isothiocyanate (TRITC) for rabbit anti-4-aminophenol antibodies (4-AP-GA-BSA) immobilized on a transparent poly(ethylene terephthalate) plate. The amount of the colored analyte was directly measured by thermal lensing at the plate surface. The developed method is specific for 4-aminophenol. The limit of detection is 2 x 10(-7) M, which corresponds to the absolute amount of 3 x 10(-11) mol of 4-aminophenol. The sensitivities of the developed thermal-lens and fluorescence-based measurements of the immunoassay under the same conditions are in good agreement with the expected and discussed theoretical sensitivities of photothermal and fluorescence measurements.


Domestic homicide cases related to schizophrenic offenders.
The goal of this study was identification of highly specific patterns of schizophrenia related domestic homicides by comparing schizophrenic homicide offenders with related domestic homicide group of offenders diagnosed with other psychiatric disorders. This study was based on the comparison of schizophrenic homicide group and other homicide group of offenders on the basis of differences in psychosocial and sociodemographic patterns and the modality of crime. The survey was conducted on mentally insane domestic homicide offenders diagnosed with schizophrenia (n=44) and second group of mentally insane offenders diagnosed with other psychiatric diagnosis (n=43). All offenders were admitted to Department of Forensic Psychiatry (Neuropsychiatric Hospital "Dr. Ivan Barbot", Popovaca, Croatia) for psychiatric evaluation. They have undergone psychological testing and psychiatric evaluation in order to make forensic expert analyses of each case particularly. This study showed some specific characteristics in the cases of schizophrenic offenders; they are more often commit parricides and siblicides, the victims are often males with their own physical strength. Furthermore, schizophrenic offenders were indifferent upon killing their victim; they were less often provoked by a victim itself and were sober tempore criminis. Moreover, in the same homicide group we found young, single offenders with high school education, average intelligence and with positive psychiatric heredity. Finally, in the same group of offenders we have found no history of military serving, less social developmental disruptions, less history of drug and alcohol abuse during adolescence and adult age.


sn-1,2-Diacylglycerols and phorbol diesters stimulate thromboxane synthesis by de novo synthesis of prostaglandin H synthase in human promyelocytic leukemia cells.
We studied the regulation of thromboxane (TX) synthesis in promyelocytic leukemia cells during macrophage differentiation. Cells treated with 12-O-tetradecanoylphorbol-13-acetate (TPA) showed rates of TXB2 synthesis from exogenous arachidonic acid that exceeded that of control cells by a factor of up to 81. Cells treated with sn-1,2-dioctanoylglycerol (diC8) showed similarly high TXB2 synthesis rates when diC8 was added concomitantly with a subthreshold concentration of TPA or when given in multiple doses. These activities depended on de novo synthesis of prostaglandin H (PGH) synthase because: microsomal PGH synthase activity showed large increases in Vmax values, and mass measurements of PGH synthase revealed the presence of PGH synthase in differentiating cells whereas the enzyme was undetectable in control cells. These results indicate that macrophage differentiation is associated with stimulation of TXB2 synthesis that requires both activation of protein kinase C and de novo synthesis of PGH synthase.


A conjugated carbon-dot-tyrosinase bioprobe for highly selective and sensitive detection of dopamine.
In this work, a bioprobe for the detection of dopamine was designed and fabricated through covalently linking fluorescent carbon dots (CDs) and tyrosinase (TYR). The bioprobe (named CDs-TYR) can catalyze oxidation of dopamine and produce dopaquinone, and consequently the fluorescence of the CDs was quenched due to an efficient electron transfer mechanism from excited CDs to dopaquinone. The fluorescence intensity of CDs decreased in a dopamine-concentration-dependent manner, which built the foundation of dopamine quantification. The bioprobe provided a wide linear range from 0.1 to 6.0 μM for dopamine sensing. Additionally, excellent selectivity of the bioprobe to dopamine was achieved because of the specific catalytic character of the conjugated TYR. Furthermore, the bioprobe was successfully employed for the detection of dopamine in spiked human serum. To the best of our knowledge, this is the first example of the construction of a bioprobe through conjugating CDs and an enzyme. This work would open new opportunities to develop CD-based photoinduced electron transfer bioprobes for other analytes via linking typical enzymes onto CDs.


Surface deformation during an action potential in pearled cells.
Electric pulses in biological cells (action potentials) have been reported to be accompanied by a propagating cell-surface deformation with a nanoscale amplitude. Typically, this cell surface is covered by external layers of polymer material (extracellular matrix, cell wall material, etc.). It was recently demonstrated in excitable plant cells (Chara braunii) that the rigid external layer (cell wall) hinders the underlying deformation. When the cell membrane was separated from the cell wall by osmosis, a mechanical deformation, in the micrometer range, was observed upon excitation of the cell. The underlying mechanism of this mechanical pulse has, to date, remained elusive. Herein we report that Chara cells can undergo a pearling instability, and when the pearled fragments were excited even larger and more regular cell shape changes were observed (∼10-100μm in amplitude). These transient cellular deformations were captured by a curvature model that is based on three parameters: surface tension, bending rigidity, and pressure difference across the surface. In this paper these parameters are extracted by curve-fitting to the experimental cellular shapes at rest and during excitation. This is a necessary step to identify the mechanical parameters that change during an action potential.


Wnt, Ptk7, and FGFRL expression gradients control trunk positional identity in planarian regeneration.
Mechanisms enabling positional identity re-establishment are likely critical for tissue regeneration. Planarians use Wnt/beta-catenin signaling to polarize the termini of their anteroposterior axis, but little is known about how regeneration signaling restores regionalization along body or organ axes. We identify three genes expressed constitutively in overlapping body-wide transcriptional gradients that control trunk-tail positional identity in regeneration. ptk7 encodes a trunk-expressed kinase-dead Wnt co-receptor, wntP-2 encodes a posterior-expressed Wnt ligand, and ndl-3 encodes an anterior-expressed homolog of conserved FGFRL/nou-darake decoy receptors. ptk7 and wntP-2 maintain and allow appropriate regeneration of trunk tissue position independently of canonical Wnt signaling and with suppression of ndl-3 expression in the posterior. These results suggest that restoration of regional identity in regeneration involves the interpretation and re-establishment of axis-wide transcriptional gradients of signaling molecules.


Processing speed differences between 70- and 83-year-olds matched on childhood IQ.
Processing speed is an important human cognitive capability that might underlie differences in other cognitive skills and their aging. We aimed to test aging-related processing speed differences using a novel cross-sectional design that adjusted for cognitive ability tested in youth. We examined aging differences on three different ways of assessing processing speed: psychometric, experimental, and psychophysical. We compared large narrow-age cohorts of 70- and 83-year-old people who were matched for cognitive ability in childhood. There were decrements of substantial effect size in all processing speed assessments in the older group that were not accounted for by prior cognitive ability, health, or fitness differences, though these factors also contributed to processing speed differences. These findings confirm age-related cognitive slowing using an unusual research design, and provide evidence against recent theories characterizing aging-related cognitive decline as a myth.


Modelling spatio-temporal patterns of disease for spatially misaligned data: An application on measles incidence data in Namibia from 2005-2014.
Making inferences about measles distribution patterns at small area level is vital for more focal targeted intervention. However, in statistical literature, the analysis of originally collected data on one resolution with the purpose to make inferences on a different level of spatial resolution is referred to as the misalignment problem. In Namibia the measles data were available in aggregated format at regional level for the period 2005 to 2014. This leads to a spatial misalignment problem if the purpose is to make decisions at constituency level. Moreover, although data on risk covariates of measles could be obtained at constituency level, they were not available each year between 2005 and 2014. Thus, assuming that covariates were constant through the study period would induce measurement errors which might have effects on the analysis results. This paper presents a spatio-temporal model through a multi-step approach in order to deal with misalignment and measurement error. For the period 2005-2014, measles data from the Ministry of Health and Social Services (MoHSS) were analysed in two steps. First, a multi-step approach was applied to correct spatial misalignment in the data. Second, a classical measurement error model was fitted to account for measurement errors. The time effects were specified using a nonparametric formulation for the linear trend through first order random walk. An interaction between area and time was modelled through type I and type II interaction structures. The study showed that there was high variation in measles risk across constituencies and as well as over the study period (2005-2014). Furthermore, the risk of measles was found to be associated with (i) the number of people aged between 0 and 24 years, (ii) the percentages of women aged 15-49 with an educational level more than secondary, (iii) the percentages of children age 12-23 months who received measles vaccine, (iv) the percentages of malnourished children under 5 years, and (vi) the measles cases for each previous year. The study showed some of the determinants of measles risk and revealed areas at high risk through disease mapping. Additionally, the study showed a non-linear temporal distribution of measles risk over the study period. Finally, it was shown that ignoring the measurement errors may yield misleading results. It was recommended that group and geographically targeted intervention, prevention and control strategies can be tailored on the basis these findings.


Diagnostic accuracy of somatosensory evoked potential monitoring during scoliosis fusion.
The goal of this review was to ascertain the diagnostic accuracy of intraoperative somatosensory evoked potential (SSEP) changes to predict perioperative neurological outcome in patients undergoing spinal deformity surgery to correct adolescent idiopathic scoliosis (AIS). The authors searched PubMed/MEDLINE and World Science databases to retrieve reports and/or experiments from January 1950 through January 2014 for studies on SSEP use during AIS surgery. All motor and sensory deficits were noted in the neurological examination administered after the procedure which was used to determine the effectiveness of SSEP as an intraoperative monitoring technique. Fifteen studies identified a total of 4763 procedures on idiopathic patients. The observed incidence of neurological deficits was 1.11% (53/4763) of the sample population. Of the patients with new postoperative neurological deficits 75.5% (40/53) showed significant SSEP changes, and 24.5% (13/53) did not show significant change. Pooled analysis using the bivariate model showed SSEP change with pooled sensitivity (average 84%, 95% confidence interval 59-95%) and specificity (average 98%, 95% confidence interval 97-99%). The diagnostic odds ratio of a patient who had a new neurological deficit with SSEP changes was a diagnostic odds ratio of 340 (95% confidence interval 125-926). Overall, detection of SSEP changes had excellent discriminant ability with an area under the curve of 0.99. Our meta-analysis covering 4763 operations on idiopathic patients showed that it is a highly sensitive and specific test and that iatrogenic spinal cord injury resulting in new neurological deficits was 340 times more likely to have changes in SSEP compared to those without any new deficits.


Self-assembly of actin scaffolds on lipid microbubbles.
Microbubbles offer unique properties as combined carriers of therapeutic payloads and diagnostic agents. Here we report on the development of novel microbubble architectures that in addition to the usual lipid shell have an actin cytoskeletal cortex assembled on their exterior. We show, using atomic force microscopy that this biomimetic coating creates a thin mesh that allows tuning of the mechanical properties of microbubbles and that the nature of actin assembly is determined by the fluidity of the lipid layer. Further, we show that it is possible to attach payloads and targeting-ligands to the actin scaffold. Resistance to gas permeation showed that the additional actin layer reduces gas diffusion across the shell and thus increases bubble lifetime. This study demonstrates a one step method to creating more complex microbubble architectures, which would be capable of further modification and tuning through the inclusion of actin binding proteins.


Beta-adrenergic stimulation of adenine nucleotide catabolism and purine release in human adipocytes.
The effects of beta-adrenergic agonists on ATP utilization and adenine nucleotide breakdown in human adipocytes were examined. The catecholamine-induced increase in cAMP was associated with an enhancement of adenine nucleotide catabolism resulting in an increase in release of inosine and hypoxanthine which can not be reutilized for adenine nucleotide synthesis. Therefore, one-third of total cellular adenine nucleotides were irreversibly lost in the presence of 1 mumol/liter isoproterenol. The catecholamine-induced increase in purine release could be blocked by phosphodiesterase inhibitors, suggesting that cAMP is the main precursor of purines in the presence of beta-adrenergic agonists. However, epinephrine (in the simultaneous presence of the alpha 2-adrenergic blocking agent, yohimbine) and isoproterenol were 10 times more potent in stimulating purine release than in elevating cAMP. In addition, purine release ceased when cAMP was still markedly increased, suggesting a compartmentation of the cyclic nucleotide and/or involvement of the hormone-sensitive, low Km cAMP phosphodiesterase. The results document that white fat cells have an enormous potential for dissipating energy, and demonstrate that the pathway involving cAMP formation and hydrolysis constitutes the principle route of adenine nucleotide catabolism in the presence of beta-adrenergic agonists.


Monitoring neural progenitor fate through multiple rounds of division in an intact vertebrate brain.
The behaviour of neural progenitors in the intact vertebrate brain and spinal cord is poorly understood, chiefly because of the inaccessibility and poor optical qualities inherent in many model systems. To overcome these problems we have studied the optically superior brain of the zebrafish embryo and have monitored the in vivo behaviour of fluorescently labelled neural progenitors and their daughter cells throughout a substantial period of hindbrain development. We find the majority (84%) of hindbrain neurons are born from progenitor divisions that generate two neurons and 68% of reconstructed lineage trees contained no asymmetric stem cell-like divisions. No progenitors divided in the manner expected of a classic stem cell; i.e. one that repeatedly self-renews and generates a differentiated cell type by asymmetric division. We also analysed the orientation of progenitor divisions relative to the plane of the ventricular zone (VZ) and find that this does not correlate with the fate of the daughter cells. Our results suggest that in this vertebrate system the molecular determinants that control whether a cell will become a neuron are usually not linked to a mechanism that generates asymmetric divisions.


An NCI perspective on creating sustainable biospecimen resources.
High-quality biospecimens with appropriate clinical annotation are critical in the era of personalized medicine. It is now widely recognized that biospecimen resources need to be developed and operated under established scientific, technical, business, and ethical/legal standards. To date, such standards have not been widely practiced, resulting in variable biospecimen quality that may compromise research efforts. The National Cancer Institute (NCI) Office of Biorepositories and Biospecimen Research (OBBR) was established in 2005 to coordinate NCI's biospecimen resource activities and address those issues that affect access to the high-quality specimens and data necessary for its research enterprises as well as the broader translational research field. OBBR and the NCI Biorepository Coordinating Committee developed NCI's "Best Practices for Biospecimen Resources" after consultation with a broad array of experts. A Biospecimen Research Network was established to fund research to develop additional evidence-based practices. Although these initiatives will improve the overall availability of high-quality specimens and data for cancer research, OBBR has been authorized to implement a national biobanking effort, cancer HUman Biobank (caHUB). caHUB will address systematically the gaps in knowledge needed to improve the state-of-the-science and strengthen the standards for human biobanking. This commentary outlines the progressive efforts by NCI in technical, governance, and economic considerations that will be important as the new caHUB enterprise is undertaken.


Oxidation of acid-volatile sulfide in surface sediments increases the release and toxicity of copper to the benthic amphipod Melita plumulosa.
Acid-volatile sulfides (AVS) are an important metal-binding phase in sediments. For sediments that contain an excess of AVS over simultaneously extracted metal (SEM) concentrations, acute or chronic effects should not result from the metals Cd, Cu, Ni, Pb and Zn. While AVS phases may exist in surface sediments, the exposure to dissolved oxygen may oxidize the AVS and release metals to more bioavailable forms. We investigated the role of oxidation of AVS, and specifically copper sulfide phases, in surface sediments, in the toxicity to juveniles of the epibenthic amphipod, Melita plumulosa. Sediments containing known amounts of copper sulfide were prepared either in situ by reacting dissolved copper with AVS that had formed in field sediments or created in sediments within the laboratory, or by addition of synthesised CuS to sediments. Regardless of the form of the copper sulfide, considerable oxidation of AVS occurred during the 10-d tests. Sediments that had a molar excess of AVS compared to SEM at the start of the tests, did not always have an excess at the end of the tests. Consistent with the AVS-SEM model, no toxicity was observed for sediments with an excess of AVS throughout the tests. However, the study highlights the need to carefully consider the changes in AVS concentrations during tests, and that measurements of AVS and SEM concentrations should carefully target the materials to which the organisms are being exposed throughout tests, which in the case of juvenile M. plumulosa is the top few mm of the sediments.


Homeopathic Rhus toxicodendron treatment increased the expression of cyclooxygenase-2 in primary cultured mouse chondrocytes.
Rhus toxicodendron (Rhus tox) is a homeopathic remedy with anti-inflammatory activities used for arthritis pain. We studied the effects of 4×, 30×, 30c and 200c homeopathic dilutions of Rhus tox in primary cultured mouse chondrocytes. We examined the expression of collagen type II, a marker protein of chondrocytes, and cyclooxygenase-2 (COX-2), which is responsible for the biosynthesis of prostaglandin E2 (PGE2) and the regulation of the inflammatory response. We assessed the expression of collagen type II and COX-2 using biochemical and immunological methods, such as reverse transcription polymerase chain reaction (RT-PCR), quantitative (or real-time) RT-PCR (qRT-PCR) and immunoblot assays. Stimulation with different concentrations of Rhus tox increased the mRNA expression of COX-2, and stimulation with 30× Rhus tox showed the most prominent mRNA expression in both RT-PCR and qRT-PCR analyses. We also observed that homeopathic dilutions of 4×, 30× and 30c Rhus tox inhibited collagen type II expression, suggesting that Rhus tox induced the dedifferentiation of chondrocytes. In addition, treatment with 30× Rhus tox significantly increased PGE2 release compared with other homeopathic dilutions of Rhus tox. Taken together, these results suggest that homeopathic treatment with Rhus tox induced chondrocyte dedifferentiation and inflammatory responses, such as COX-2 expression and PGE2 production, in primary cultured chondrocytes.


Receiving a hug is associated with the attenuation of negative mood that occurs on days with interpersonal conflict.
Interpersonal touch is emerging as an important topic in the study of adult relationships, with recent research showing that such behaviors can promote better relationship functioning and individual well-being. This investigation considers whether being hugged is associated with reduced conflict-related decreases in positive affect and increases in negative affect as well as whether these associations differ between women and men. A sample of 404 adults were interviewed every night for 14 consecutive days about their conflicts, hug receipt, and positive and negative affect. Results indicated that there was an interaction between hug receipt and conflict exposure such that receiving a hug was associated with a smaller conflict-related decrease in positive affect and a smaller conflict-related increase in negative affect when assessed concurrently. Hug receipt was also prospectively associated with a smaller conflict-related increase in next day negative affect but was not associated with next day positive affect. Associations between hug receipt and conflict-related changes in affect did not differ between women and men, between individuals who were married or in a marital-like relationship and those who were not, or as a function of individual differences in baseline perceived social support. While correlational, these results are consistent with the hypothesis that hugs buffer against deleterious changes in affect associated with experiencing interpersonal conflict. Possible mechanisms through which hugs facilitate positive adaptation to conflict are discussed.


Elevated expression of TDP-43 in the forebrain of mice is sufficient to cause neurological and pathological phenotypes mimicking FTLD-U.
TDP-43 is a multifunctional DNA/RNA-binding factor that has been implicated in the regulation of neuronal plasticity. TDP-43 has also been identified as the major constituent of the neuronal cytoplasmic inclusions (NCIs) that are characteristic of a range of neurodegenerative diseases, including the frontotemporal lobar degeneration with ubiquitin(+) inclusions (FTLD-U) and amyotrophic lateral sclerosis (ALS). We have generated a FTLD-U mouse model (CaMKII-TDP-43 Tg) in which TDP-43 is transgenically overexpressed in the forebrain resulting in phenotypic characteristics mimicking those of FTLD-U. In particular, the transgenic (Tg) mice exhibit impaired learning/memory, progressive motor dysfunction, and hippocampal atrophy. The cognitive and motor impairments are accompanied by reduced levels of the neuronal regulators phospho-extracellular signal-regulated kinase and phosphorylated cAMP response element-binding protein and increased levels of gliosis in the brains of the Tg mice. Moreover, cells with TDP-43(+), ubiquitin(+) NCIs and TDP-43-deleted nuclei appear in the Tg mouse brains in an age-dependent manner. Our data provide direct evidence that increased levels of TDP-43 protein in the forebrain is sufficient to lead to the formation of TDP-43(+), ubiquitin(+) NCIs and neurodegeneration. This FTLD-U mouse model should be valuable for the mechanistic analysis of the role of TDP-43 in the pathogenesis of FTLD-U and for the design of effective therapeutic approaches of the disease.


Non-occupational physical activity levels of shift workers compared with non-shift workers.
Lack of physical activity (PA) has been hypothesised as an underlying mechanism in the adverse health effects of shift work. Therefore, our aim was to compare non-occupational PA levels between shift workers and non-shift workers. Furthermore, exposure-response relationships for frequency of night shifts and years of shift work regarding non-occupational PA levels were studied. Data of 5980 non-shift workers and 532 shift workers from the European Prospective Investigation into Cancer and Nutrition-Netherlands (EPIC-NL) were used in these cross-sectional analyses. Time spent (hours/week) in different PA types (walking/cycling/exercise/chores) and intensities (moderate/vigorous) were calculated based on self-reported PA. Furthermore, sports were operationalised as: playing sports (no/yes), individual versus non-individual sports, and non-vigorous-intensity versus vigorous-intensity sports. PA levels were compared between shift workers and non-shift workers using Generalized Estimating Equations and logistic regression. Shift workers reported spending more time walking than non-shift workers (B=2.3 (95% CI 1.2 to 3.4)), but shift work was not associated with other PA types and any of the sports activities. Shift workers who worked 1-4 night shifts/month (B=2.4 (95% CI 0.6 to 4.3)) and ≥5 night shifts/month (B=3.7 (95% CI 1.8 to 5.6)) spent more time walking than non-shift workers. No exposure-response relationships were found between years of shift work and PA levels. Shift workers spent more time walking than non-shift workers, but we observed no differences in other non-occupational PA levels. To better understand if and how PA plays a role in the negative health consequences of shift work, our findings need to be confirmed in future studies.


Engineering enzyme specificity by "substrate-assisted catalysis".
A novel approach to engineering enzyme specificity is presented in which a catalytic group from an enzyme is first removed by site-directed mutagenesis causing inactivation. Activity is then partially restored by substrates containing the missing catalytic functional group. Replacement of the catalytic His with Ala in the Bacillus amyloliquefaciens subtilisin gene (the mutant is designated His64Ala) by site-directed mutagenesis reduces the catalytic efficiency (kcat/Km) by a factor of a million when assayed with N-succinyl-L-Phe-L-Ala-L-Ala-L-Phe-p-nitroanilide (sFAAF-pNA). Model building studies showed that a His side chain at the P2 position of a substrate bound at the active site of subtilisin could be virtually superimposed on the catalytic His side chain of this serine protease. Accordingly, the His64Ala mutant hydrolyzes a His P2 substrate (sFAHF-pNA) up to 400 times faster than a homologous Ala P2 or Gln P2 substrate (sFAAF-pNA or sFAQF-pNA) at pH 8.0. In contrast, the wild-type enzyme hydrolyzes these three substrates with similar catalytic efficiencies. Additional data from substrate-dependent pH profiles and hydrolysis of large polypeptides indicate that the His64Ala mutant enzyme can recover partially the function of the lost catalytic histidine from a His P2 side chain on the substrate. Such "substrate-assisted catalysis" provides a new basis for engineering enzymes with very narrow and potentially useful substrate specificities. These studies also suggest a possible functional intermediate in the evolution of the catalytic triad of serine proteases.


Ergosterol attenuates cigarette smoke extract-induced COPD by modulating inflammation, oxidative stress and apoptosis in vitro and in vivo.
Cigarette smoke (CS) is the major cause of chronic obstructive pulmonary disease (COPD). CS heightens inflammation, oxidative stress and apoptosis. Ergosterol is the main bioactive ingredient in Cordyceps sinensis (C. sinensis), a traditional medicinal herb for various diseases. The objective of this work was to investigate the effects of ergosterol on anti-inflammatory and antioxidative stress as well as anti-apoptosis in a cigarette smoke extract (CSE)-induced COPD model both in vitro and in vivo Our results demonstrate that CSE induced inflammatory and oxidative stress and apoptosis with the involvement of the Bcl-2 family proteins via the nuclear factor kappa B (NF-κB)/p65 pathway in both 16HBE cells and Balb/c mice. CSE induced epithelial cell death and increased the expression of nitric oxide (NO), interleukin-6 (IL-6), tumor necrosis factor α (TNF-α), malondialdehyde (MAD) and the apoptosis-related proteins cleaved caspase 3/7/9 and cleaved-poly-(ADP)-ribose polymerase (PARP) both in vitro and in vivo, whereas decreased the levels of superoxide dismutase (SOD) and catalase (CAT). Treatment of 16HBE cells and Balb/c mice with ergosterol inhibited CSE-induced inflammatory and oxidative stress and apoptosis by inhibiting the activation of NF-κB/p65. Ergosterol suppressed apoptosis by inhibiting the expression of the apoptosis-related proteins both in vitro and in vivo Moreover, the usage of QNZ (an inhibitor of NF-κB) also partly demonstrated that NF-κB/p65 pathway was involved in the ergosterol protective progress. These results show that ergosterol suppressed COPD inflammatory and oxidative stress and apoptosis through the NF-κB/p65 pathway, suggesting that ergosterol may be partially responsible for the therapeutic effects of cultured C. sinensis on COPD patients.


Effects of financing smoking cessation outpatient services in Taiwan.
This study assesses the effects of a 2005 increase in funding for smoking cessation services on provider participation, patient utilisation of smoking cessation services and cessation outcome at a six-month follow-up. Analyses are based on existing databases and on a follow-up study among smokers participating in the smoking cessation service. The effect of the policy is evaluated by comparing year 2004 (old policy) with year 2005 (new policy). The generalised estimating equations (GEE) method was conducted to examine the effects of increasing funding for smoking cessation services on monthly smoking cessation services provided per physician and yearly consultations received per patient. Logistic regression was used to examine the effects of increasing funding on smoking cessation outcome. The study found the increased reimbursement rates and medication subsidies for smoking cessation to be positively related to the number of physicians enrolling in the programme (1841 in 2004 vs 3466 in 2005), the number of cessation consultations per month per physician (5.1 vs 14.6) and the number of cessation visits per year per patient (2.0 vs 2.5). Male providers and providers belonging to the private sector were found to offer more cessation consultations. The number of subjects receiving this counselling increased from 22 167 in 2004 to 109 508 in 2005. After adjusting for consumer and provider factors the likelihood of successful quitting among those counselled did not change. Overall, smokers who were older, had attempted to quit in the past year, had lower nicotine dependence, had gone to more smoking cessation service visits, had received consultations in the public sector and were seen by physicians delivering fewer consultations were more likely to have quit smoking at the six-month follow-up. Based on increases in physician enrollment and consultations and the increase in number of subjects receiving counselling and number of visits, the policy of increasing provider incentives and medication subsidies appears to have successfully promoted smoking cessation services.


Agrobacterium tumefaciens mutants affected in crown gall tumorigenesis and octopine catabolism.
Mutants of Agrobacterium tumefaciens which affect virulence or the ability to catabolize octopine were isolated after Tn5-induced mutagenesis. Of 8,900 colonies tested, 7 mutants with Tn5 insertions in a specific region of other Ti plasmid unable to catabolize octopine were isolated. Thirty-seven mutants affected in tumorigenesis resulted from insertions in the Ti plasmid and the Agrobacterium chromosome. Of these mutations, 12 were chromosomal and 25 mapped on the plasmid. Twenty-three mapped within a 20-megadalton region, which is distinct from the Ti plasmid sequences found stably integrated into the plant cell genome T-deoxyribonucleic acid). Included in these were mutants that were either a virulent or produced tumors with unusual morphologies. Three mutants contained insertions in the T-deoxyribonucleic acid. These three mutants incited tumors which synthesized octopine but had an altered morphology due to either extensive proliferation of shoots or roots from the tumor callus. Three additional mutants not caused by Tn5 contained mutations in the Ti plasmid.


Contact force and scanning velocity during active roughness perception.
Haptic perception is bidirectionally related to exploratory movements, which means that exploration influences perception, but perception also influences exploration. We can optimize or change exploratory movements according to the perception and/or the task, consciously or unconsciously. This paper presents a psychophysical experiment on active roughness perception to investigate movement changes as the haptic task changes. Exerted normal force and scanning velocity are measured in different perceptual tasks (discrimination or identification) using rough and smooth stimuli. The results show that humans use a greater variation in contact force for the smooth stimuli than for the rough stimuli. Moreover, they use higher scanning velocities and shorter break times between stimuli in the discrimination task than in the identification task. Thus, in roughness perception humans spontaneously use different strategies that seem effective for the perceptual task and the stimuli. A control task, in which the participants just explore the stimuli without any perceptual objective, shows that humans use a smaller contact force and a lower scanning velocity for the rough stimuli than for the smooth stimuli. Possibly, these strategies are related to aversiveness while exploring stimuli.


Ageing and the telomere connection: An intimate relationship with inflammation.
Telomeres are the heterochromatic repeat regions at the ends of eukaryotic chromosomes, whose length is considered to be a determinant of biological ageing. Normal ageing itself is associated with telomere shortening. Here, critically short telomeres trigger senescence and eventually cell death. This shortening rate may be further increased by inflammation and oxidative stress and thus affect the ageing process. Apart from shortened or dysfunctional telomeres, cells undergoing senescence are also associated with hyperactivity of the transcription factor NF-κB and overexpression of inflammatory cytokines such as TNF-α, IL-6, and IFN-γ in circulating macrophages. Interestingly, telomerase, a reverse transcriptase that elongates telomeres, is involved in modulating NF-κB activity. Furthermore, inflammation and oxidative stress are implicated as pre-disease mechanisms for chronic diseases of ageing such as neurodegenerative diseases, cardiovascular disease, and cancer. To date, inflammation and telomere shortening have mostly been studied individually in terms of ageing and the associated disease phenotype. However, the interdependent nature of the two demands a more synergistic approach in understanding the ageing process itself and for developing new therapeutic approaches. In this review, we aim to summarize the intricate association between the various inflammatory molecules and telomeres that together contribute to the ageing process and related diseases.


Thermal lensing in an end-pumped Yb:KGW slab laser with high power single emitter diodes.
We demonstrate that it is possible to pump an Ng-cut Yb:KGW crystal slab with up to 18W by a single emitter laser diode and still achieve a nearly diffraction limited output beam with a M(2) value below M(2) < 1.1 and optical to optical efficiencies of more than 44%. Furthermore, we have measured the focal length of thermally induced lenses in an Yb:KGW crystal slab which was end-pumped by 12 W and 18 W single-emitter diodes. We have observed that the focal lengths not only depend on the pump power and beam waists, but are also dependent on the specific diode beam profile, as well as the collimation optics. We have been able to correlate this behavior with specific patterns of the beam profile.


Cerebral blood flow and cardiovascular risk effects on resting brain regional homogeneity.
Regional homogeneity (ReHo) is a measure of local functional brain connectivity that has been reported to be altered in a wide range of neuropsychiatric disorders. Computed from brain resting-state functional MRI time series, ReHo is also sensitive to fluctuations in cerebral blood flow (CBF) that in turn may be influenced by cerebrovascular health. We accessed cerebrovascular health with Framingham cardiovascular risk score (FCVRS). We hypothesize that ReHo signal may be influenced by regional CBF; and that these associations can be summarized as FCVRS→CBF→ReHo. We used three independent samples to test this hypothesis. A test-retest sample of N = 30 healthy volunteers was used for test-retest evaluation of CBF effects on ReHo. Amish Connectome Project (ACP) sample (N = 204, healthy individuals) was used to evaluate association between FCVRS and ReHo and testing if the association diminishes given CBF. The UKBB sample (N = 6,285, healthy participants) was used to replicate the effects of FCVRS on ReHo. We observed strong CBF→ReHo links (p<2.5 × 10-3) using a three-point longitudinal sample. In ACP sample, marginal and partial correlations analyses demonstrated that both CBF and FCVRS were significantly correlated with the whole-brain average (p<10-6) and regional ReHo values, with the strongest correlations observed in frontal, parietal, and temporal areas. Yet, the association between ReHo and FCVRS became insignificant once the effect of CBF was accounted for. In contrast, CBF→ReHo remained significantly linked after adjusting for FCVRS and demographic covariates (p<10-6). Analysis in N = 6,285 replicated the FCVRS→ReHo effect (p = 2.7 × 10-27). In summary, ReHo alterations in health and neuropsychiatric illnesses may be partially driven by region-specific variability in CBF, which is, in turn, influenced by cardiovascular factors.


Deletion of Braun lipoprotein and plasminogen-activating protease-encoding genes attenuates Yersinia pestis in mouse models of bubonic and pneumonic plague.
Currently, there is no FDA-approved vaccine against Yersinia pestis, the causative agent of bubonic and pneumonic plague. Since both humoral immunity and cell-mediated immunity are essential in providing the host with protection against plague, we developed a live-attenuated vaccine strain by deleting the Braun lipoprotein (lpp) and plasminogen-activating protease (pla) genes from Y. pestis CO92. The Δlpp Δpla double isogenic mutant was highly attenuated in evoking both bubonic and pneumonic plague in a mouse model. Further, animals immunized with the mutant by either the intranasal or the subcutaneous route were significantly protected from developing subsequent pneumonic plague. In mice, the mutant poorly disseminated to peripheral organs and the production of proinflammatory cytokines concurrently decreased. Histopathologically, reduced damage to the lungs and livers of mice infected with the Δlpp Δpla double mutant compared to the level of damage in wild-type (WT) CO92-challenged animals was observed. The Δlpp Δpla mutant-immunized mice elicited a humoral immune response to the WT bacterium, as well as to CO92-specific antigens. Moreover, T cells from mutant-immunized animals exhibited significantly higher proliferative responses, when stimulated ex vivo with heat-killed WT CO92 antigens, than mice immunized with the same sublethal dose of WT CO92. Likewise, T cells from the mutant-immunized mice produced more gamma interferon (IFN-γ) and interleukin-4. These animals had an increasing number of tumor necrosis factor alpha (TNF-α)-producing CD4(+) and CD8(+) T cells than WT CO92-infected mice. These data emphasize the role of TNF-α and IFN-γ in protecting mice against pneumonic plague. Overall, our studies provide evidence that deletion of the lpp and pla genes acts synergistically in protecting animals against pneumonic plague, and we have demonstrated an immunological basis for this protection.


Kupffer cell-dependent TNF-alpha signaling mediates injury in the arterialized small-for-size liver transplantation in the mouse.
Implantation of small liver grafts causes liver injury and defective regeneration leading to graft failure. We investigated whether Kupffer cell-dependent TNF-alpha signaling contributes to this poor outcome. Partial 30% liver transplantation was performed in C57BL/6 wild-type mice (control group), and in three groups with down-regulation of the TNF-alpha pathway: (i) TNF receptor 1 knockout [TNFR-1(-/-)] mice, and mice pretreated with (ii) gadolinium chloride or (iii) pentoxifylline (PTX). Fifty-percent partial liver transplantation, a model associated with full recovery, and transplantation in IL-6 knockout [IL-6(-/-)] mice were performed in some experiments. Graft injury, regeneration, portal flow, liver microcirculation, leukocyte adhesion, and animal survival were assessed. Animal survival rates were 14% in the control group vs. 43% in the gadolinium chloride group, 57% for the TNFR-1(-/-) group, and 86% in the PTX group (P < 0.001). Markers of liver injury were reduced in all treated groups when compared with controls. Each treated group disclosed better portal flow and sinusoid perfusion, decreased leukocyte adherence, particularly in the PTX group. Liver regeneration occurred only in the treated groups. IL-6 and IL-10 levels were dramatically up-regulated (50x) in the PTX group, and at lower levels in other experimental groups. The protective effect of PTX was lost in IL-6(-/-) mice and protection was restored by a single dose of r-IL-6. In conclusion, interruption of TNF-alpha signaling or depletion of Kupffer cells improves survival after 30% liver transplantation, reduces liver injury, and enhances regeneration. The superior effects of PTX are mediated by IL-6.


Myocardin in biology and disease.
Myocardin (MYOCD) is a potent transcriptional coactivator that functions primarily in cardiac muscle and smooth muscle through direct contacts with serum response factor (SRF) over cis elements known as CArG boxes found near a number of genes encoding for contractile, ion channel, cytoskeletal, and calcium handling proteins. Since its discovery more than 10 years ago, new insights have been obtained regarding the diverse isoforms of MYOCD expressed in cells as well as the regulation of MYOCD expression and activity through transcriptional, post-transcriptional, and post-translational processes. Curiously, there are a number of functions associated with MYOCD that appear to be independent of contractile gene expression and the CArG-SRF nucleoprotein complex. Further, perturbations in MYOCD gene expression are associated with an increasing number of diseases including heart failure, cancer, acute vessel disease, and diabetes. This review summarizes the various biological and pathological processes associated with MYOCD and offers perspectives to several challenges and future directions for further study of this formidable transcriptional coactivator.


Occupational exposure and new-onset asthma in a population-based study in Northern Europe (RHINE).
In a large population-based study among adults in northern Europe the relation between occupational exposure and new-onset asthma was studied. The study comprised 13 284 subjects born between 1945 and 1973, who answered a questionnaire 1989-1992 and again 1999-2001. Asthma was defined as 'Asthma diagnosed by a physician' with reported year of diagnose. Hazard ratios (HR), for new-onset adult asthma during 1980-2000, were calculated using a modified job-exposure matrix as well as high-risk occupations in Cox regression models. The analyses were made separately for men and women and were also stratified for atopy. During the observation period there were 429 subjects with new-onset asthma with an asthma incidence of 1.3 cases per 1000 person-years for men and 2.4 for women. A significant increase in new-onset asthma was seen for men exposed to plant-associated antigens (HR = 3.6; 95% CI [confidence interval] = 1.4-9.0), epoxy (HR = 2.4; 95% CI = 1.3-4.5), diisocyanates (HR = 2.1; 95% CI = 1.2-3.7) and accidental peak exposures to irritants (HR = 2.4; 95% CI = 1.3-4.7). Both men and women exposed to cleaning agents had an increased asthma risk. When stratifying for atopy an increased asthma risk were seen in non-atopic men exposed to acrylates (HR = 3.3; 95% CI = 1.4-7.5), epoxy compounds (HR = 3.6; 95% CI = 1.6-7.9), diisocyanates and accidental peak exposures to irritants (HR = 3.0; 95% CI = 1.2-7.2). Population attributable risk for occupational asthma was 14% for men and 7% for women. This population-based study showed that men exposed to epoxy, diisocyanates and acrylates had an increased risk of new-onset asthma. Non-atopics seemed to be at higher risk than atopics, except for exposure to high molecular weight agents. Increased asthma risks among cleaners, spray painters, plumbers, and hairdressers were confirmed.


Disruption of IRS-2 causes type 2 diabetes in mice.
Human type 2 diabetes is characterized by defects in both insulin action and insulin secretion. It has been difficult to identify a single molecular abnormality underlying these features. Insulin-receptor substrates (IRS proteins) may be involved in type 2 diabetes: they mediate pleiotropic signals initiated by receptors for insulin and other cytokines. Disruption of IRS-1 in mice retards growth, but diabetes does not develop because insulin secretion increases to compensate for the mild resistance to insulin. Here we show that disruption of IRS-2 impairs both peripheral insulin signalling and pancreatic beta-cell function. IRS-2-deficient mice show progressive deterioration of glucose homeostasis because of insulin resistance in the liver and skeletal muscle and a lack of beta-cell compensation for this insulin resistance. Our results indicate that dysfunction of IRS-2 may contribute to the pathophysiology of human type 2 diabetes.


Short Sprints Accumulated at School Modulate Postprandial Metabolism in Boys.
This study examined the efficacy of maximal sprint running accumulated during a typical school day to modulate postprandial metabolism in adolescent boys. Nineteen adolescent boys completed three 2-d experimental conditions: a standard-practice control (CON), an accumulated in-school sprint running (ACC), and a single block of afterschool sprint running (BLO). On day 1, a fasting capillary blood sample was taken at 0735 h in the school. Three subsequent postprandial blood samples were taken at predetermined times after consumption of standardized breakfast and lunch. During ACC, participants accumulated four sets of 10 × 30-m maximal-intensity sprint runs across natural breaks in lessons. During BLO, participants performed the same number of sprints (40) in a single after-school exercise session. The blood samples from day 1 were replicated on the day after exercise (day 2). On day 1, no significant differences in total area under the plasma triacylglycerol concentration versus time curve (TAUC-TAG) were observed between conditions (P = 0.126). However, TAUC-insulin was lower in ACC compared with BLO (-26%, effect size [ES] = 0.86, P = 0.001) and CON (-22%, ES = 0.72, P = 0.010). On day 2, TAUC-TAG was 12% lower after ACC (ES = 0.49; P = 0.002) and 10% lower after BLO (ES = 0.37; P = 0.019) compared with CON. No significant differences were observed between conditions on day 2 for postprandial insulin or glucose (P ≥ 0.738). Four sets of 10 × 30-m sprints, accumulated in four separate bouts (<5 min) during the school day, reduced postprandial triacylglycerol and insulin concentrations in adolescent boys and may represent an effective in-school exercise strategy to promote metabolic health.


Molecular phylogenetic analysis of the Amiota taurusata species group within the Chinese species, with descriptions of two new species.
The relationships among six species of the Amiota taurusata Takada, Beppu, & Toda (Diptera: Drosophilidae) species group were investigated based on DNA sequence data of the mitochondrial NADH dehydrogenase subunit 2 ( ND2) gene, using three species of the genus Amiota as outgroups. A mitochondrial gene, cytochrome c oxidase I ( COI), can be used to discriminate between species of the taurusata group. Two new species are described from South China: A. protuberantis Shao et Chen, sp. nov. and A. shennongi Shao et Chen, sp. nov. A key to all the species of the taurusata group based on morphological characters is provided.


KRAS and guanine nucleotide-binding protein mutations in pancreatic juice collected from the duodenum of patients at high risk for neoplasia undergoing endoscopic ultrasound.
Pancreatic imaging can identify neoplastic cysts but not microscopic neoplasms. Mutation analysis of pancreatic fluid after secretin stimulation might identify microscopic neoplasias in the pancreatic duct system. We determined the prevalence of mutations in KRAS and guanine nucleotide-binding protein α-stimulating genes in pancreatic juice from subjects undergoing endoscopic ultrasound for suspected pancreatic intraepithelial neoplasia, intraductal papillary mucinous neoplasms, or pancreatic adenocarcinoma. Secretin-stimulated juice samples were collected from the duodenum of 272 subjects enrolled in Cancer of the Pancreas Screening studies; 194 subjects were screened because of a family history of, or genetic predisposition to, pancreatic cancer, and 78 subjects were evaluated for pancreatic cancer (n = 30) or other disorders (controls: pancreatic cysts, pancreatitis, or normal pancreata, n = 48). Mutations were detected by digital high-resolution melt-curve analysis and pyrosequencing. The number of replicates containing a mutation determined the mutation score. KRAS mutations were detected in pancreatic juice from larger percentages of subjects with pancreatic cancer (73%) or undergoing cancer screening (50%) than controls (19%) (P = .0005). A greater proportion of patients with pancreatic cancer had at least 1 KRAS mutation detected 3 or more times (47%) than screened subjects (21%) or controls (6%, P = .002). Among screened subjects, mutations in KRAS (but not guanine nucleotide-binding protein α-stimulating) were found in similar percentages of patients with or without pancreatic cysts. However, a greater proportion of patients older than age 50 years had KRAS mutations (54.6%) than younger patients (36.3%) (P = .032); the older subjects also had more mutations in KRAS (P = .02). Mutations in KRAS are detected in pancreatic juice from the duodenum of 73% of patients with pancreatic cancer, and 50% of asymptomatic individuals with a high risk for pancreatic cancer. However, KRAS mutations were detected in pancreatic juice from 19% of controls. Mutations detected in individuals without pancreatic abnormalities, based on imaging analyses, likely arise from small pancreatic intraepithelial neoplasia lesions. ClinicalTrials.gov no: NCT00438906 and NCT00714701.


Sustained decreases in risk exposure and youth problem behaviors after installation of the Communities That Care prevention system in a randomized trial.
To test whether the Communities That Care (CTC) prevention system reduced levels of risk and adolescent problem behaviors community-wide 6 years after installation of CTC and 1 year after study-provided resources ended. A community randomized trial. Twenty-four small towns in 7 states, matched within state, randomly assigned to control or intervention condition in 2003. A panel of 4407 fifth-grade students was surveyed annually through 10th grade from 2004 to 2009. A coalition of community stakeholders received training and technical assistance to install CTC, used epidemiologic data to identify elevated risk factors and depressed protective factors in the community, and implemented programs to address their community's elevated risks from a menu of tested and effective programs for youths aged 10 to 14 years, their families, and schools. Levels of risk and incidence and prevalence of tobacco, alcohol, and other drug use; delinquency; and violent behavior by grade 10. Mean levels of targeted risks increased less rapidly between grades 5 and 10 in CTC than in control communities and were significantly lower in CTC than control communities in grade 10. The incidence of delinquent behavior, alcohol use, and cigarette use and the prevalence of current cigarette use and past-year delinquent and violent behavior were significantly lower in CTC than in control communities in grade 10. Using the CTC system can produce enduring reductions in community-wide levels of risk factors and problem behaviors among adolescents beyond the years of supported implementation, potentially contributing to long-term public health benefits. clinicaltrials.gov Identifier: NCT01088542.


Protection of rat skeletal muscle fibers by either L-carnitine or coenzyme Q10 against statins toxicity mediated by mitochondrial reactive oxygen generation.
Mitochondrial redox imbalance has been implicated in mechanisms of aging, various degenerative diseases and drug-induced toxicity. Statins are safe and well-tolerated therapeutic drugs that occasionally induce myotoxicity such as myopathy and rhabdomyolysis. Previous studies indicate that myotoxicity caused by statins may be linked to impairment of mitochondrial functions. Here, we report that 1-h incubation of permeabilized rat soleus muscle fiber biopsies with increasing concentrations of simvastatin (1-40 μM) slowed the rates of ADP-or FCCP-stimulated respiration supported by glutamate/malate in a dose-dependent manner, but caused no changes in resting respiration rates. Simvastatin (1 μM) also inhibited the ADP-stimulated mitochondrial respiration supported by succinate by 24% but not by TMPD/ascorbate. Compatible with inhibition of respiration, 1 μM simvastatin stimulated lactate release from soleus muscle samples by 26%. Co-incubation of muscle samples with 1 mM L-carnitine, 100 μM mevalonate or 10 μM coenzyme Q10 (Co-Q10) abolished simvastatin effects on both mitochondrial glutamate/malate-supported respiration and lactate release. Simvastatin (1 μM) also caused a 2-fold increase in the rate of hydrogen peroxide generation and a decrease in Co-Q10 content by 44%. Mevalonate, Co-Q10 or L-carnitine protected against stimulation of hydrogen peroxide generation but only mevalonate prevented the decrease in Co-Q10 content. Thus, independently of Co-Q10 levels, L-carnitine prevented the toxic effects of simvastatin. This suggests that mitochondrial respiratory dysfunction induced by simvastatin, is associated with increased generation of superoxide, at the levels of complexes-I and II of the respiratory chain. In all cases the damage to these complexes, presumably at the level of 4Fe-4S clusters, is prevented by L-carnitine.


Regulating cellular cyclic adenosine monophosphate: "Sources," "sinks," and now, "tunable valves".
A number of hormones and growth factors stimulate target cells via the second messenger pathways, which in turn regulate cellular phenotypes. Cyclic adenosine monophosphate (cAMP) is a ubiquitous second messenger that facilitates numerous signal transduction pathways; its production in cells is tightly balanced by ligand-stimulated receptors that activate adenylate cyclases (ACs), that is, "source" and by phosphodiesterases (PDEs) that hydrolyze it, that is, "sinks." Because it regulates various cellular functions, including cell growth and differentiation, gene transcription and protein expression, the cAMP signaling pathway has been exploited for the treatment of numerous human diseases. Reduction in cAMP is achieved by blocking "sources"; however, elevation in cAMP is achieved by either stimulating "source" or blocking "sinks." Here we discuss an alternative paradigm for the regulation of cellular cAMP via GIV/Girdin, the prototypical member of a family of modulators of trimeric GTPases, Guanine nucleotide Exchange Modulators (GEMs). Cells upregulate or downregulate cellular levels of GIV-GEM, which modulates cellular cAMP via spatiotemporal mechanisms distinct from the two most often targeted classes of cAMP modulators, "sources" and "sinks." A network-based compartmental model for the paradigm of GEM-facilitated cAMP signaling has recently revealed that GEMs such as GIV serve much like a "tunable valve" that cells may employ to finetune cellular levels of cAMP. Because dysregulated signaling via GIV and other GEMs has been implicated in multiple disease states, GEMs constitute a hitherto untapped class of targets that could be exploited for modulating aberrant cAMP signaling in disease states. This article is categorized under: Models of Systems Properties and Processes > Mechanistic Models Biological Mechanisms > Cell Signaling.


Occupational Therapy Education Research: Results of a National Survey.
Despite a strong tradition of education scholarship in occupational therapy, no systematic evaluation of the amount and nature of research in this arena, or its associated challenges, has been conducted. This study examined the scope and range of education-focused research conducted in Canada and identified perceived supports and barriers for this area of inquiry. An online survey was sent to faculty of all 14 Canadian occupational therapy programs. Descriptive statistics were used to summarize areas of study focus, and qualitative thematic analysis captured respondents' perceptions of personal and environmental factors influencing their engagement in education research. Respondents (n = 47) reported engagement in education scholarship spanning more than 20 focus areas, often with limited resources. Strategies are proposed to promote the viability of education research in occupational therapy and to improve resources and other supports for this type of research.


The transcriptional regulator gene phrR in Sinorhizobium meliloti WSM419 is regulated by low pH and other stresses.
The phrR gene in Sinorhizobium meliloti (previously known as Rhizobium meliloti) WSM419, directly downstream from actA, is induced by low pH or certain stresses (e.g. high concentrations of Zn2+, Cu2+, H2O2 or ethanol), but not in stationary phase or by other stresses (e.g. phosphate limitation, elevated temperature, high concentrations of sucrose or iron). A DNA fragment containing the wild-type phrR gene could not be cloned and inverse PCR was therefore used to amplify a 3.5 kb BamHI fragment containing phrR from the mutant S. meliloti TG2-6 (actA::Tn5). DNA fragments from a BamHI/SalI digest of the amplified product were cloned into pUK21 and sequenced. The phrR open reading frame contiguous to actA appears to code for a 15.2 kDa protein showing significant identity with the proteins encoded by y4wC and y4aM in Rhizobium sp. NGR234. All three proteins resemble transcriptional regulators in containing a DNA-binding helix-turn-helix motif similar to that reported for URF4 in Rhodospirillum rubrum and repressors in coliphage.


Podocyte effacement closely links to suPAR levels at time of posttransplantation focal segmental glomerulosclerosis occurrence and improves with therapy.
Focal segmental glomerulosclerosis (FSGS) recurs after kidney transplantation in more than 30% of cases and can lead to allograft loss. Serum soluble urokinase-type plasminogen activator receptor (suPAR) is implicated in the pathogenesis of native and recurrent FSGS. We conducted a retrospective study of 25 adults with posttransplantation FSGS. We investigated the relationship between suPAR levels and podocyte changes and the impact of therapy on podocyte structure. We assessed response to therapy by improvement in proteinuria, allograft function, and resolution of histologic changes. A median (interquartile range) of 15 (10-23) plasmapheresis sessions was administered; 13 of the subjects also received rituximab. Median pretreatment suPAR levels were higher among those with severe (≥75%) versus those with mild (≤25%) podocyte foot process effacement (13,030 vs. 4806 pg/mL; P=0.02). Overall, mean±SD of proteinuria improved from 5.1±3.8 to 2.1±2.8 mg/dL (P=0.003), mean podocyte effacement decreased from 57%±33% to 22%±22% (P=0.0001), estimated glomerular filtration rates increased from median (interquartile range) of 32.9 (20.6-44.2) to 39.3 (28.8-63.4; P<0.0001), and suPAR levels decreased from a median of 6.781 to 4.129 pg/mL (P=0.02) with therapy. Podocyte effacement is the first pathologic manifestation of FSGS after transplantation. The degree of podocyte effacement correlates with suPAR levels at time of diagnosis. Response to therapy results in significant reduction of suPAR levels and complete or significant improvement of podocyte effacement.


Characterization of new regulatory mutants of bacteriophage T4. II. New class of mutants.
New mutants of bacteriophage T4 that overproduce the enzyme dihydrofolate reductase were investigated. Unlike previously described overproducers of this enzyme (Johnson and Hall, 1974), these mutants did not overproduce deoxycytidylate deaminase. Overproduction of dihydrofolate reductase by the new mutants occurred because enzymatic activity continued to increase for a longer period of time in cells infected by the mutants than in cells infected by wild-type phage. This continued increase occurred even in the presence of rifampin, indicating that the overproduction is probably due to a post-transcriptional event. Both these new overproducers and the previously described overproducers were studied by using polyacrylamide gel electrophoresis. The two types of overproducers appeared to be very different. The previously described overproducers showed a delay and/or reduction in the synthesis of several proteins that normally started to be made 4 to 6 min after infection. Several proteins could be seen to be overproduced on the gels. The new overproducers did not show the delay in the synthesis of some proteins and only overproduced a few proteins. The new gene defined by the new overproducers is between the gene coding for thymidine kinase and the gene coding for lysozyme.


Salivary cortisol is related to atherosclerosis of carotid arteries.
Dysregulation of the hypothalamic-pituitary-adrenal axis has been suggested as an independent risk factor for ischemic heart disease. The aim of our study was to evaluate whether two markers of the hypothalamic-pituitary-adrenal axis activity, the level of salivary cortisol and the diurnal salivary cortisol pattern, are associated with atherosclerosis of the carotid arteries in an elderly population. A total of 1866 participants of the Rotterdam Study, a population-based cohort study in the elderly, provided four salivary cortisol samples throughout 1 d, and underwent ultrasonography to examine the presence of plaques in the common, internal, and bifurcation sites of both carotid arteries. Two summary measures of the separate cortisol values were computed: area under the curve (AUC), which is a measure of total cortisol exposure while awake; and the slope, which is a measure of diurnal cortisol decline. Total cortisol exposure while awake (AUC) was associated with higher plaque scores (beta = 0.08 per sd of AUC, 95% confidence interval 0.00-0.16; P = 0.04) in a fully adjusted linear regression model. Persons with an AUC in the highest tertile had a higher number of plaques of carotid arteries compared with those in the lowest tertile (3.08 vs. 2.80, 95% confidence interval of difference 0.09-0.48; P = 0.005). There was no relation between diurnal cortisol decline and plaque score. Our results support the hypothesis that increased total cortisol exposure is independently associated with atherosclerosis of the carotid arteries.


Mutations in Either TUBB or MAPRE2 Cause Circumferential Skin Creases Kunze Type.
Circumferential skin creases Kunze type (CSC-KT) is a specific congenital entity with an unknown genetic cause. The disease phenotype comprises characteristic circumferential skin creases accompanied by intellectual disability, a cleft palate, short stature, and dysmorphic features. Here, we report that mutations in either MAPRE2 or TUBB underlie the genetic origin of this syndrome. MAPRE2 encodes a member of the microtubule end-binding family of proteins that bind to the guanosine triphosphate cap at growing microtubule plus ends, and TUBB encodes a β-tubulin isotype that is expressed abundantly in the developing brain. Functional analyses of the TUBB mutants show multiple defects in the chaperone-dependent tubulin heterodimer folding and assembly pathway that leads to a compromised yield of native heterodimers. The TUBB mutations also have an impact on microtubule dynamics. For MAPRE2, we show that the mutations result in enhanced MAPRE2 binding to microtubules, implying an increased dwell time at microtubule plus ends. Further, in vivo analysis of MAPRE2 mutations in a zebrafish model of craniofacial development shows that the variants most likely perturb the patterning of branchial arches, either through excessive activity (under a recessive paradigm) or through haploinsufficiency (dominant de novo paradigm). Taken together, our data add CSC-KT to the growing list of tubulinopathies and highlight how multiple inheritance paradigms can affect dosage-sensitive biological systems so as to result in the same clinical defect.