"Waiting for DAAs": A retrospective chart review of patients with untreated hepatitis C in Rwanda.
Access to treatment for hepatitis C virus (HCV) in sub-Saharan Africa is extremely limited. With the advent of direct acting antivirals (DAAs), highly effective and easy-to-deliver oral regimens are now available on the global market. This study was conducted to understand the background and characteristics of a national cohort of patients with HCV infection enrolled in care and awaiting therapy with DAAs. We conducted a retrospective chart review of all adult patients with confirmed HCV infection who were currently enrolled in care and treatment at the four existing hepatitis referral centers in Rwanda. Patient charts at these centers were reviewed, and routinely collected data were recorded and analyzed. Overall, 253 patients were identified; median age was 56 years (IQR: 43, 65), and 149 (58.9%) were female. Median viral load was 688,736 IU/ml and 96.7% were HCV genotype 4. As classified by FIB-4 score, 64.8% of the patients had moderate to severe fibrosis. Fibrosis stage was associated with age (OR 1.12, CI 1.09-1.17), but not with time since diagnosis, gender, treatment center, or type of insurance. There was a low frequency of documented co-morbid conditions, including hypertension, diabetes, HIV, and hepatitis B virus. Compared to an estimated 55,000 patients eligible for HCV treatment in Rwanda, this study identified only 253 patients currently diagnosed and engaged in care, highlighting an immense treatment gap in HCV, likely due to the lack of accessible and affordable screening, diagnostic, and treatment modalities. The patients that were enrolled in care had a disproportionately advanced fibrosis stage, possibly indicating late presentation to care or lack of treatment options. In the context of newly available and effective treatment options, this study supports the overall need to accelerate access to HCV screening, diagnostics, and care and treatment services in resource-limited settings in sub-Saharan Africa.


Neuroimaging manifestations in children with SARS-CoV-2 infection: a multinational, multicentre collaborative study.
The CNS manifestations of COVID-19 in children have primarily been described in case reports, which limit the ability to appreciate the full spectrum of the disease in paediatric patients. We aimed to identify enough cases that could be evaluated in aggregate to better understand the neuroimaging manifestations of COVID-19 in the paediatric population. An international call for cases of children with encephalopathy related to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and abnormal neuroimaging findings was made. Clinical history and associated plasma and cerebrospinal fluid data were requested. These data were reviewed by a central neuroradiology panel, a child neurologist, and a paediatric infectious diseases expert. The children were categorised on the basis of their time of probable exposure to SARS-CoV-2. In addition, cases were excluded when a direct link to SARS-CoV-2 infection could not be established or an established alternate diagnostic cause could be hypothesised. The accepted referral centre imaging data, from ten countries, were remotely reviewed by a central panel of five paediatric neuroradiologists and a consensus opinion obtained on the imaging findings. 38 children with neurological disease related to SARS-CoV-2 infection were identified from France (n=13), the UK (n=8), the USA (n=5), Brazil (n=4), Argentina (n=4), India (n=2), Peru (n=1), and Saudi Arabia (n=1). Recurring patterns of disease were identified, with neuroimaging abnormalities ranging from mild to severe. The most common imaging patterns were postinfectious immune-mediated acute disseminated encephalomyelitis-like changes of the brain (16 patients), myelitis (eight patients), and neural enhancement (13 patients). Cranial nerve enhancement could occur in the absence of corresponding neurological symptoms. Splenial lesions (seven patients) and myositis (four patients) were predominantly observed in children with multisystem inflammatory syndrome. Cerebrovascular complications in children were less common than in adults. Significant pre-existing conditions were absent and most children had favourable outcomes. However, fatal atypical CNS co-infections developed in four previously healthy children infected with SARS-CoV-2. Acute-phase and delayed-phase SARS-CoV-2-related CNS abnormalities are seen in children. Recurring patterns of disease and atypical neuroimaging manifestations can be found and should be recognised being as potentially due to SARS-CoV-2 infection as an underlying aetiological factor. Studies of paediatric specific cohorts are needed to better understand the effects of SARS-CoV-2 infection on the CNS at presentation and on long-term follow-up in children. American Society of Pediatric Neuroradiology, University of Manchester (Manchester, UK). VIDEO ABSTRACT.


In Vitro-In Vivo Extrapolation Scaling Factors for Intestinal P-Glycoprotein and Breast Cancer Resistance Protein: Part I: A Cross-Laboratory Comparison of Transporter-Protein Abundances and Relative Expression Factors in Human Intestine and Caco-2 Cells.
Over the last 5 years the quantification of transporter-protein absolute abundances has dramatically increased in parallel to the expanded use of in vitro-in vivo extrapolation (IVIVE) and physiologically based pharmacokinetics (PBPK)-linked models, for decision-making in pharmaceutical company drug development pipelines and regulatory submissions. Although several research groups have developed laboratory-specific proteomic workflows, it is unclear if the large range of reported variability is founded on true interindividual variability or experimental variability resulting from sample preparation or the proteomic methodology used. To assess the potential for methodological bias on end-point abundance quantification, two independent laboratories, the University of Manchester (UoM) and Bertin Pharma (BPh), employing different proteomic workflows, quantified the absolute abundances of Na/K-ATPase, P-gp, and breast cancer resistance protein (BCRP) in the same set of biologic samples from human intestinal and Caco-2 cell membranes. Across all samples, P-gp abundances were significantly correlated (P = 0.04, Rs = 0.72) with a 2.4-fold higher abundance (P = 0.001) generated at UoM compared with BPh. There was a systematically higher BCRP abundance in Caco-2 cell samples quantified by BPh compared with UoM, but not in human intestinal samples. Consequently, a similar intestinal relative expression factor (REF), derived from distal jejunum and Caco-2 monolayer samples, between laboratories was found for P-gp. However, a 2-fold higher intestinal REF was generated by UoM (2.22) versus BPh (1.11). We demonstrate that differences in absolute protein abundance are evident between laboratories and they probably result from laboratory-specific methodologies relating to peptide choice.


Activity of erythrocyte sodium-hydrogen exchange in normal pregnancy.
1. Pregnancy is associated with a 30-50% rise in cardiac output and a 50% increase in blood volume. The contribution of changes in the activity of primary and secondary active transporters to these haemodynamic adaptations remains unknown. For the first time, we measured sodium-hydrogen exchange activity over the course of normal pregnancy. 2. Eighteen healthy pregnant women were studied at 14, 24 and 33 weeks of gestation and compared with 18 non-pregnant healthy women. None of the pregnancies was complicated by hypertension. At each antenatal visit, body weight and blood pressure were recorded, blood and 24 h-urine samples were taken to control renal function and metabolic equilibrium, maternal glucose tolerance was evaluated by oral glucose test and glycated haemoglobin testing, and erythrocyte sodium-hydrogen antiport was also measured. 3. Erythrocyte antiport activity values were 10.0 +/- 3.0, 9.6 +/- 2.9 and 8.4 +/- 3.5 mmol h-1 (litre of cells)-1 in the three gestational trimesters respectively, significantly higher at each trimester than in control women [6.8 +/- 2.5 mmol h-1 (litre of cells)-1]. The clearances of urea and creatinine were constantly elevated in pregnant women; at each trimester their serum concentrations were lower than in non-pregnant women. Serum potassium significantly decreased during pregnancy. Serum total cholesterol and triacylglycerol levels, already above the normal range from the first trimester, further increased until the third trimester. The area under the glycaemic curve became larger during pregnancy, and the area under the insulinaemic curve increased to a lesser extent. There was a significant association between antiport activity and serum triacylglycerol levels. 4. The observed hyperactivity of the transporter, peaking at the fourteenth week of gestation, may be a contributing factor to the haemodynamic adjustments attending upon normal pregnancy.


Role of the alpha-, beta-, and gamma-subunits of epithelial sodium channel in a model of polygenic hypertension.
The pathophysiological basis of Liddle's syndrome, a rare autosomal dominant form of arterial hypertension, has been found to rest on missense mutations or truncations of the beta- and gamma-subunits of the epithelial sodium channel. The hypothesis has been advanced that molecular variants of these genes might also contribute to the common polygenic forms of hypertension. We tested this hypothesis by performing a cosegregation study in a reciprocal cross between the stroke-prone spontaneously hypertensive rat (SHRSPHD) and a Wistar-Kyoto rat (WKY-1HD) reference strain. We carried out genetic mapping and chromosomal assignment of the alpha-, beta-, and gamma-subunits of the epithelial sodium channel using both linkage analysis and fluorescent in situ hybridization techniques. We demonstrate that in the rat, the beta- and gamma-subunits, as in humans, are in close linkage; they map to rat chromosome 1 and cosegregate with systolic pressure after dietary NaCl (logarithm of the odds [LOD] score, 3.7), although the peak LOD score of 5.0 for this quantitative trait locus was detected 4.4 cM away from the beta-/gamma-subunit locus. The alpha-subunit was mapped to chromosome 4 and exhibited no linkage to blood pressure phenotype. Comparative analysis of the complete coding sequences of all three subunits in the SHRSPHD and WKY-1HD strains revealed no biologically relevant mutations. Furthermore, Northern blot comparison of mRNA levels for all three subunits in the kidney showed no differences between SHRSPHD and WKY-1HD. Our results fail to support a material contribution of the epithelial sodium channel genes to blood pressure regulation in this model of polygenic hypertension.


Flies Exploit Predictable Perspectives and Backgrounds to Enhance Iridescent Signal Salience and Mating Success.
Communication requires both the encoding of information and its effective transmission, but little is known about display traits that primarily serve to enhance efficacy. Here we examined the visual courtships of Lispe cana, a cursorial fly that lives and mates in heterogeneous foreshores, and tested the prediction that males should seek to enhance signal salience and consequent fitness through the flexible choice of display locations. We show that courting males access the field of view of females by straddling them and holding their wings closed before moving ahead to present their structurally colored faces in ritualized dances. Males preferentially present these UV-white signals against darker backgrounds and the magnitude of contrast predicts female attention, which in turn predicts mating success. Our results demonstrate a striking interplay between the physical and attentional manipulation of receivers and reveal novel routes to the enhancement of signal efficacy in noisy environments.


Blood Glucose Levels of Subelite Athletes During 6 Days of Free Living.
Continuous glucose monitoring (CGM) devices, with their 1-5 min measurement interval, allow blood glucose (BG) concentration dynamics to be captured more frequently and less invasively than traditional BG measures. One cohort CGM could provide insight is athletes. This study investigates what impact their heightened energy expenditure and dietary intake may have on their ability to achieve optimal BG. Ten subelite athletes (resting HR<60 bpm, training>6 hrs per week) were recruited. Two Ipro2 CGM devices (Medtronic Minimed, Northridge, CA) were inserted into the abdomen and remained in place for ~6 days. Time in band was calculated as the percentage of CGM BG measurements with in the 4.0-6.0 mmol/L. Fasting glucose was calculated using CGM calibration BG measurements and postprandial glucose response was also calculated using the CGM values. 4/10 athletes studied spent more than 70% of the total monitoring time above 6.0 mmol/L even with the 2-hour period after meals is excluded. Fasting BG was also in the ADA defined prediabetes range for 3/10 athletes. Only 1 participant spent substantial time below 4.0 mmol/L which was largely due to significantly lower energy intake compared to recommendations. Contrary to expectations high BG appears to be more of a concern for athletes then low BG even in those with the highest energy expenditure and consuming below the recommended carbohydrate intake. This study warrants further investigation on the recommended diets and the BG of athletes to better determine the causes and impact of this hyperglycemia on overall athlete health.


A successful implementation of e-epidemiology: the Danish pregnancy planning study 'Snart-Gravid'.
The attraction of being able to use the internet for the recruitment of an epidemiologic cohort stems mainly from cost efficiency and convenience. The pregnancy planning study ('Snart-Gravid')-a prospective cohort study of Danish women planning a pregnancy-was conducted to evaluate the feasibility and cost efficiency of using internet-based recruitment and follow-up. Feasibility was assessed by examining patient accrual data over time, questionnaire-specific response rates and losses to follow-up. The relative cost efficiency was examined by comparing the study costs with those of an alternative non internet-based study approach. The target recruitment of 2,500 participants over 6 months was achieved using advertisements on a health-related website, supported by a coordinated media strategy at study initiation. Questionnaire cycle-specific response rates ranged from 87 to 90% over the 12-month follow-up. At 6 months, 87% of women had a known outcome or were still under follow-up; at 12 months the figure was 82%. The study cost of $400,000 ($160 per enrolled subject) compared favorably with the estimated cost to conduct the same study using a conventional non-internet based approach ($322 per subject). The gain in efficiency with the internet-based approach appeared to be even more substantial with longer follow-up and larger study sizes. The successful conduct of this pilot study suggests that the internet may be a useful tool to recruit and follow subjects in prospective cohort studies.


Sleep, diurnal preference, health, and psychological well-being: a prospective single-allelic-variation study.
Individual differences in sleep and diurnal preference associate with physical and mental health characteristics, but few genetic determinants of these differences have been identified. A variable number tandem repeat (VNTR) polymorphism in the PERIOD3 (PER3) gene (rs57875989) has been reported to associate with diurnal preference, i.e., preferred timing of waking and sleep. Here, the authors investigate in a prospective single-candidate genetic variant study whether allelic variation for this polymorphism associates also with reported actual sleep timing and sleep duration, as well as psychological and health measures. Six hundred and seventy-five subjects, aged 20 to 35 yrs, completed questionnaires to assess sleep and psychological and health characteristics and were genotyped for the PER3 VNTR. Homozygosity for the longer allele (PER3(5/5)) of the VNTR was associated with increased morning preference, earlier wake time and bedtime, and reduced daytime sleepiness. Separate analyses of work and rest days demonstrated that the increase in time in bed during rest days was greatest in PER3(5/5) homozygotes. PER3 genotype modified the effects of sleep timing and duration on fluid intelligence and body mass index. Genotype was not associated with physical or psychological characteristics as assessed by the SF-36 Health Questionnaire, the General Health Questionnaire, the Big Five Inventory, the Behavioral Inhibition System-Behavioral Activation System scales, and the Positive and Negative Affect Scale, even though these measures varied significantly with diurnal preference as assessed by the Morningness-Eveningness Questionnaire. Whereas diurnal preference also predicts mental health and psychological characteristics, as well as sleep timing, the PER3 VNTR specifically affects measures of sleep timing and may also modify the effects of sleep on health outcome measures.


Peroxisome proliferator-activated receptor gamma agonists promote TRAIL-induced apoptosis by reducing survivin levels via cyclin D3 repression and cell cycle arrest.
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising cancer therapy that preferentially induces apoptosis in cancer cells. However, many neoplasms are resistant to TRAIL by mechanisms that are poorly understood. Here we demonstrate that human breast cancer cells, but not normal mammary epithelial cells, are dramatically sensitized to TRAIL-induced apoptosis and caspase activation by peroxisome proliferator-activated receptor gamma (PPARgamma) agonists of the thiazolidinedione (TZD) class. Although TZDs do not significantly alter the expression of components of the TRAIL signaling pathway, they profoundly reduce protein levels of cyclin D3, but not other D-type cyclins, by decreasing cyclin D3 mRNA levels and by inducing its proteasomal degradation. Importantly, both TRAIL sensitization and reduction in cyclin D3 protein levels induced by TZDs are likely PPARgamma-independent because a dominant negative mutant of PPARgamma did not antagonize these effects of TZDs, nor were they affected by the expression levels of PPARgamma. TZDs also inhibit G(1) to S cell cycle progression. Furthermore, silencing cyclin D3 by RNA interference inhibits S phase entry and sensitizes breast cancer cells to TRAIL, indicating a key role for cyclin D3 repression in these events. G(1) cell cycle arrest sensitizes breast cancer cells to TRAIL at least in part by reducing levels of the anti-apoptotic protein survivin: ectopic expression of survivin partially suppresses apoptosis induced by TRAIL and TZDs. We also demonstrate for the first time that TZDs promote TRAIL-induced apoptosis of breast cancer in vivo, suggesting that this combination may be an effective therapy for cancer.


Changes in the pressure and the dynamic stiffness index of medical elastic compression stockings after having been worn for eight hours: a pilot study.
There is no data available at present on the changes in the exerted pressure together with the dynamic stiffness index (DSI) of medical elastic compression stockings (MECS). The objective of this pilot study was to measure the pressure and calculate the DSI of 12 different brands of MECS before and after having been worn for eight hours. In all, 12 different commercially available brands of MECS that were divided into two categories (class I round-knitted and class II flat-knitted MECS) were tested. The pressure was measured, and the DSI of the MECS was first calculated at the B1 level before wearing in the morning and again eight hours after they had been worn. All laboratory measurements were performed using a newly developed dynamic leg-segment model. The pressure at the B1 level dropped significantly in all 12 brands of MECS after having been worn for eight hours, whereas the DSI remained unchanged. The DSI of MECS reflects an important and particularly consistent therapeutic effect. As the pressure drops during the day, the pressure amplitude or pulsations remain the same. The pressure drop may be due to fatigue of the elastic material. The DSI would therefore form a valuable indicator for prescribing the most effective MECS for the patient.


Squeeze-film hydrogel deposition and dry micropatterning.
In this technical note, we demonstrate a squeeze-film based spacer-free method for creating controllable submicrometer hydrogel films on planar substrates that can be used to photolithographically fabricate hydrogel microstructures. This new technique improves the photolithographic resolution and yield by providing a uniform and low-defect hydrogel film. The optimum polymerization initiation time for achieving such a layer was determined to be around 1 min. For patterning, the dried hydrogel film was coated with a parylene-C masking layer. Subsequent etching in oxygen plasma was used to transfer selected patterns of hydrogel to the substrate in a batch scale.


Simultaneous simulations of uptake in plants and leaching to groundwater of cadmium and lead for arable land amended with compost or farmyard manure.
The water budget of soil, the uptake in plants and the leaching to groundwater of cadmium (Cd) and lead (Pb) were simulated simultaneously using a physiological plant uptake model and a tipping buckets water and solute transport model for soil. Simulations were compared to results from a ten-year experimental field study, where four organic amendments were applied every second year. Predicted concentrations slightly decreased (Cd) or stagnated (Pb) in control soils, but increased in amended soils by about 10% (Cd) and 6% to 18% (Pb). Estimated plant uptake was lower in amended plots, due to an increase of K(d) (dry soil to water partition coefficient). Predicted concentrations in plants were close to measured levels in plant residues (straw), but higher than measured concentrations in grains. Initially, Pb was mainly predicted to deposit from air into plants (82% in 1998); the next years, uptake from soil became dominating (30% from air in 2006), because of decreasing levels in air. For Cd, predicted uptake from air into plants was negligible (1-5%).


Complex 2D photonic crystals with analogue local symmetry as 12-fold quasicrystals.
We construct fourteen complex periodic two-dimensional (2D) photonic structures with different structural symmetries by arranging the small portions of a 12-fold quasicrystal on square or hexagonal lattices. The corresponding reciprocal lattices confirm that all of them demonstrate the 12-fold-like characteristics due to the analogue short-range arrangements. We then investigate their photonic bandgap properties at different dielectric contrast levels (dielectric rods in air background). Our results suggest that all structures possess analogue transverse magnetic (TM) gaps in both Si and glass photonic crystals due to the similarity of their local geometries. However, the arrangements of the basic elements, total symmetries, and the coupling between the local and the lattice symmetries have greater impact on the glass photonic crystals, which show much larger deviation of gap sizes from different structures. Furthermore, we find that the minimal dielectric contrast to achieve the TM gap in the complex lattices (dielectric-in-air) can be as low as epsilon = 1.44, whereas the inverse structures may open a 2D complete gap in silicon nitride (epsilon = 4.1).


Co-culture with NK-92MI cells enhanced the anti-cancer effect of bee venom on NSCLC cells by inactivation of NF-κB.
In the present study we experimented on a multimodal therapeutic approach, such as combining chemotherapy agent (Bee venom) with cellular (NK-92MI) immunotherapy. Previously bee venom has been found to show anti-cancer effect in various cancer cell lines. In lung cancer cells bee venom showed an IC(50) value of 3 μg/ml in both cell lines. The co-culture of NK-92MI cell lines with lung cancer cells also show a decrease in viability upto 50 % at 48 h time point. Hence we used bee venom treated NK-92MI cells to co-culture with NSCLC cells and found that there is a further decrease in cell viability upto 70 and 75 % in A549 and NCI-H460 cell lines respectively. We further investigated the expression of various apoptotic and anti-apoptotic proteins and found that Bax, cleaved caspase-3 and -8 were increasing where as Bcl-2 and cIAP-2 was decreasing. The expression of various death receptor proteins like DR3, DR6 and Fas was also increasing. Concomitantly the expression of various death receptor ligands (TNFalpha, Apo3L and FasL) was also increasing of NK-92MI cells after co-culture. Further the DNA binding activity and luciferase activity of NF-κB was also inhibited after co-culture with bee venom treated NK-92MI cell lines. The knock down of death receptors with si-RNA has reversed the decrease in cell viability and NF-κB activity after co-culture with bee venom treated NK-92MI cells. Thus this new approach can enhance the anti-cancer effect of bee venom at a much lower concentration.


Bumblebees are not deterred by ecologically relevant concentrations of nectar toxins.
Bees visit flowers to collect nectar and pollen that contain nutrients and simultaneously facilitate plant sexual reproduction. Paradoxically, nectar produced to attract pollinators often contains deterrent or toxic plant compounds associated with herbivore defence. The functional significance of these nectar toxins is not fully understood, but they may have a negative impact on pollinator behaviour and health, and, ultimately, plant pollination. This study investigates whether a generalist bumblebee, Bombus terrestris, can detect naturally occurring concentrations of nectar toxins. Using paired-choice experiments, we identified deterrence thresholds for five compounds found in the nectar of bee-pollinated plants: quinine, caffeine, nicotine, amygdalin and grayanotoxin. The deterrence threshold was determined when bumblebees significantly preferred a sucrose solution over a sucrose solution containing the compound. Bumblebees had the lowest deterrence threshold for the alkaloid quinine (0.01 mmol l(-1)); all other compounds had higher deterrence thresholds, above the natural concentration range in floral nectar. Our data, combined with previous work using honeybees, suggest that generalist bee species have poor acuity for the detection of nectar toxins. The fact that bees do not avoid nectar-relevant concentrations of these compounds likely indicates that it is difficult for them to learn to associate floral traits with the presence of toxins, thus maintaining this trait in plant populations.


Evidence based management guidelines for the detection and treatment of congenital CMV.
CMV is the most common congenital infection in newborns worldwide. Congenital CMV causes sensorineural hearing loss in a significant proportion of infected newborns, while the majority of newborns are asymptomatic. In the last three years there have been significant advances in the diagnosis and treatment of congenital CMV. We have developed practical evidence based guidelines for the management of congenital CMV.


Psychoeducation for relapse prevention in bipolar disorder: a systematic review of efficacy in randomized controlled trials.
Previous reviews have concluded that interventions including psychoeducation are effective in preventing relapse in bipolar disorder, but the efficacy of psychoeducation itself has not been systematically reviewed. Our aim was to evaluate the efficacy of psychoeducation for bipolar disorder in preventing relapse and other outcomes, and to identify factors that relate to clinical outcomes. We employed the systematic review of randomized controlled trials of psychoeducation in participants with bipolar disorder not in an acute illness episode, compared with treatment-as-usual, and placebo or active interventions. Pooled odds ratios (ORs) for non-relapse into any episode, mania/hypomania, and depression were calculated using an intent-to-treat (ITT) analysis, assigning dropouts to relapse, with a sensitivity analysis in which dropouts were assigned to non-relapse (optimistic ITT). Sixteen studies were included, eight of which provided data on relapse. Although heterogeneity in the data warrants caution, psychoeducation appeared to be effective in preventing any relapse [n = 7; OR: 1.98-2.75; number needed to treat (NNT): 5-7, depending on the method of analysis] and manic/hypomanic relapse (n = 8; OR: 1.68-2.52; NNT: 6-8), but not depressive relapse. Group, but not individually, delivered interventions were effective against both poles of relapse; the duration of follow-up and hours of therapy explained some of the heterogeneity. Psychoeducation improved medication adherence and short-term knowledge about medication. No consistent effects on mood symptoms, quality of life, or functioning were found. Group psychoeducation appears to be effective in preventing relapse in bipolar disorder, with less evidence for individually delivered interventions. Better understanding of mediating mechanisms is needed to optimize efficacy and personalize treatment.


Saffron: Its Phytochemistry, Developmental Processes, and Biotechnological Prospects.
The present state of knowledge concerning developmental processes and the secondary metabolism of saffron, Crocus sativus L. (Iridaceae), along with the genes involved in these processes so far known, is reviewed. Flowers and corms constitute the most valuable parts of saffron. Corm and flower development are two key aspects to be studied in saffron to increase the yield and quality of the spice, to raise its reproductive rate, and to implement new production systems. Important knowledge about the physiology of flowering and vegetative growth has been acquired in recent years, but there is still only limited information on molecular mechanisms controlling these processes. Although some genes involved in flower formation and meristem transition in other species have been isolated in saffron, the role of these genes in this species awaits further progress. Also, genes related with the synthesis pathway of abscisic acid and strigolactones, growth regulators related with bud endodormancy and apical dominance (paradormancy), have been isolated. However, the in-depth understanding of these processes as well as of corm development is far from being achieved. By contrast, saffron phytochemicals have been widely studied. The different flower tissues and the corm have been proved to be an important source of phytochemicals with pharmacological properties. The biotechnological prospects for saffron are here reviewed on the basis of the discovery of the enzymes involved in key aspects of saffron secondary metabolism, and we also analyze the possibility of transferring current knowledge about flowering and vegetative propagation in model species to the Crocus genus.


Online sparse Gaussian process regression and its applications.
We present a new Gaussian process (GP) inference algorithm, called online sparse matrix Gaussian processes (OSMGP), and demonstrate its merits by applying it to the problems of head pose estimation and visual tracking. The OSMGP is based upon the observation that for kernels with local support, the Gram matrix is typically sparse. Maintaining and updating the sparse Cholesky factor of the Gram matrix can be done efficiently using Givens rotations. This leads to an exact, online algorithm whose update time scales linearly with the size of the Gram matrix. Further, we provide a method for constant time operation of the OSMGP using matrix downdates. The downdates maintain the Cholesky factor at a constant size by removing certain rows and columns corresponding to discarded training examples. We demonstrate that, using these matrix downdates, online hyperparameter estimation can be included at cost linear in the number of total training examples. We describe a robust appearance-based head pose estimation system based upon the OSMGP. Numerous experiments and comparisons with existing methods using a large dataset system demonstrate the efficiency and accuracy of our system. Further, to showcase the applicability of OSMGP to a wide variety of problems, we also describe a regression-based visual tracking method. Experiments show that our OSMGP algorithm generalizes well using online learning.


Dissecting the fungal biology of Bipolaris papendorfii: from phylogenetic to comparative genomic analysis.
Bipolaris papendorfii has been reported as a fungal plant pathogen that rarely causes opportunistic infection in humans. Secondary metabolites isolated from this fungus possess medicinal and anticancer properties. However, its genetic fundamental and basic biology are largely unknown. In this study, we report the first draft genome sequence of B. papendorfii UM 226 isolated from the skin scraping of a patient. The assembled 33.4 Mb genome encodes 11,015 putative coding DNA sequences, of which, 2.49% are predicted transposable elements. Multilocus phylogenetic and phylogenomic analyses showed B. papendorfii UM 226 clustering with Curvularia species, apart from other plant pathogenic Bipolaris species. Its genomic features suggest that it is a heterothallic fungus with a putative unique gene encoding the LysM-containing protein which might be involved in fungal virulence on host plants, as well as a wide array of enzymes involved in carbohydrate metabolism, degradation of polysaccharides and lignin in the plant cell wall, secondary metabolite biosynthesis (including dimethylallyl tryptophan synthase, non-ribosomal peptide synthetase, polyketide synthase), the terpenoid pathway and the caffeine metabolism. This first genomic characterization of B. papendorfii provides the basis for further studies on its biology, pathogenicity and medicinal potential.


Effects of long-term supplementation with moderate pharmacologic doses of vitamin E are saturable and reversible in patients with type 1 diabetes.
Vitamin E supplementation has been proposed as adjunctive therapy to counteract the increased LDL oxidation in diabetes and thus prevent or delay cardiovascular complications. The objective of this study was to investigate the effect of a moderate pharmacologic dose of vitamin E for </=1 y in patients with type 1 diabetes. The study was double blind and the subjects were randomly assigned to 2 groups: the supplemented group (group S; n = 22) received 250 IU (168 mg) RRR-alpha-tocopherol 3 times/d for 1 y and the placebo group (group P; n = 22) received a placebo for 6 mo followed by 250 IU (168 mg) RRR-alpha-tocopherol 3 times/d for an additional 6 mo. Serum vitamin E doubled after 3 mo of supplementation, from a mean (+/-SD) of 36.9 +/- 10.9 to 66.4 +/- 18.3 micromol/L (P: < 0.0005). Although lipid profiles, glycated hemoglobin, and blood biochemistry values did not change significantly, copper-induced in vitro peroxidizability of LDL and VLDL decreased after 3 mo of supplementation: the production of thiobarbituric acid-reactive substances decreased by 30-60% (P: < 0. 005) and the lag time for the appearance of fluorescent products increased from 107 +/- 25 to 123 +/- 30 min in group S (P: = 0.002 compared with group P). Vitamin E supplementation for an additional 3-9 mo resulted in no further changes in serum vitamin E and lipoprotein peroxidizability. Values returned to baseline after supplementation ended. Because the improvement in lipoprotein peroxidizability is saturable and reversible, life-long supplementation with vitamin E should be considered in patients with type 1 diabetes.


Clay and soil organic matter drive wood multi-elemental composition of a tropical tree species: Implications for timber tracing.
Forensic methods to independently trace timber origin are essential to combat illegal timber trade. Tracing product origin by analysing their multi-element composition has been successfully applied in several commodities, but its potential for timber is not yet known. To evaluate this potential the drivers of wood multi-elemental composition need to be studied. Here we report on the first study relating wood multi-elemental composition of forest trees to soil chemical and physical properties. We studied the reactive soil element pools and the multi-elemental composition in sapwood and heartwood for 37 Azobé (Lophira alata) trees at two forest sites in Cameroon. A total of 46 elements were measured using ICP-MS. We also measured three potential drivers of soil and wood elemental composition: clay content, soil organic matter and pH. We tested associations between soil and wood using multiple regressions and multivariate analyses (Mantel test, db-RDA). Finally, we performed a Random Forest analysis of heartwood elemental composition to check site assignment accuracy. We found elemental compositions of soil, sapwood and heartwood to be significantly associated. Soil clay content and organic matter positively influenced individual element concentrations (for 13 and 9 elements out of 46 respectively) as well as the multi-elemental composition in wood. However, associations between wood and topsoil elemental concentrations were only significant for one element. We found close associations between element concentrations and composition in sapwood and heartwood. Lastly, the Random Forest assignment success was 97.3 %. Our findings indicate that wood elemental composition is associated with that in the topsoil and its variation is related to soil clay and organic matter content. These associations suggests that the multi-elemental composition of wood can yield chemical fingerprints obtained from sites that differ in soil properties. This finding in addition to the high assignment accuracy shows potential of multi-element analysis for tracing wood origin.


Laying hens can convert high doses of folic acid added to the feed into natural folates in eggs providing a novel source of food folate.
There are few good sources of natural food folates apart from green leafy vegetables and these may have a limited potential to increase folate status because of substantial losses that can occur during cooking. Fortified foods can overcome this but are controversial because of safety concerns regarding chronic exposure to high-dose folic acid (FA; the synthetic form). The aim of the present study was to develop eggs with an enriched natural folate content and minimal unmetabolised FA. Forty-eight, 30-week-old laying hens were randomised to receive the basal feed (formulated to provide 1 mg folate/kg feed) to which had been added one of the following FA levels (0, 2, 4, 8, 16, 32 mg/kg feed). Total folate was measured in eggs collected throughout the 12-week study period and the FA content estimated at 12 weeks. Results showed that the maximal egg folate content was achieved by adding 16 mg FA/kg feed. At this optimal dose, the total folate content per egg was 75 microg (compared with 32 microg in a regular egg) of which FA represented at most 10%, a level which would probably be converted into natural folates by humans after ingestion. The results demonstrate that it is possible to use synthetic FA at high doses to produce novel animal foods enriched with natural folates in a cost-efficient process. Such foods may be particularly relevant to European populations without access to FA fortification and therefore dependent on natural food folate sources for the primary prevention of folate-related disease.


DNA-Based Nanodevices Controlled by Purely Entropic Linker Domains.
We demonstrate here the rational design of purely entropic domains as a versatile approach to achieve control of the input/output response of synthetic molecular receptors. To do so and to highlight the versatility and generality of this approach, we have rationally re-engineered two model DNA-based receptors: a clamp-like DNA-based switch that recognizes a specific DNA sequence and an ATP-binding aptamer. We show that, by varying the length of the linker domain that connects the two recognition portions of these receptors, it is possible to finely control their affinity for their specific ligand. Through mathematical modeling and thermodynamic characterization, we also demonstrate for both systems that entropy changes associated with changes in linker length are responsible for affinity modulation and that the linker we have designed behaves as a disordered random-coil polymer. The approach also allows us to regulate the ligand concentration range at which the receptors respond and show optimal specificity. Given these attributes, the use of purely entropic domains appears as a versatile and general approach to finely control the activity of synthetic receptors in a highly predictable and controlled fashion.


Ultra slow light achievement in photonic crystals by merging coupled cavities with waveguides.
We explore slow light behavior of a specially designed optical waveguide by carrying out structural dispersion using numerical techniques. The structure proposed is composed of square-lattice photonic crystal waveguide integrated with side-coupled cavities. We report three orders of magnitude reduction in group velocity at around υ(g) ≅ 0.0008c with strongly suppressed group velocity dispersion. The analysis is performed by using both plane-wave expansion and finite-difference time-domain methods. For the first time, we succeeded to show such a low group velocity in photonic structures. Slow light pulse propagation accompanied by light tunneling between each cavity is observed. These achievements show the feasibility of photonic devices to generate extremely large group index which in turn will eventually pave the way to new frontiers in nonlinear optics, optical buffers and low threshold lasers.


Identification of zyklopen, a new member of the vertebrate multicopper ferroxidase family, and characterization in rodents and human cells.
We previously detected a membrane-bound, copper-containing oxidase that may be involved in iron efflux in BeWo cells, a human placental cell line. We have now identified a gene encoding a predicted multicopper ferroxidase (MCF) with a putative C-terminal membrane-spanning sequence and high sequence identity to hephaestin (Heph) and ceruloplasmin (Cp), the other known vertebrate MCF. Molecular modeling revealed conservation of all type I, II, and III copper-binding sites as well as a putative iron-binding site. Protein expression was observed in multiple diverse mouse tissues, including placenta and mammary gland, and the expression pattern was distinct from that of Cp and Heph. The protein possessed ferroxidase activity, and protein levels decreased in cellular copper deficiency. Knockdown with small interfering RNA in BeWo cells indicates that this gene represents the previously detected oxidase. We propose calling this new member of the MCF family "zyklopen."


Chronic morphine treatment increases cyclic AMP-dependent protein kinase activity in the rat locus coeruleus.
We have studied a possible role for cyclic AMP-dependent protein kinase in mediating opiate addiction in the central nervous system by focusing on the rat locus coeruleus. This brain region is well suited for these studies because it is relatively homogeneous and because a wealth of electrophysiological and behavioral data indicate that it plays an important role in mediating the chronic effects of opiates in animals, including humans. It was found that chronic, but not acute, in vivo treatment of rats with morphine increased cyclic AMP-dependent protein kinase activity in the locus coeruleus with a time course that closely paralleled the time course by which locus coeruleus neurons become tolerant to and dependent on opiates, based on electrophysiological studies. Concomitant administration of the opiate receptor antagonist naltrexone was found to block the effect of chronic morphine treatment on protein kinase activity, indicating that the effect is mediated via specific activation of opiate receptors. In contrast, chronic morphine treatment did not alter protein kinase activity in several other brain regions studied, including the neostriatum, frontal cortex, and dorsal raphe. We propose that the observed increase in cyclic AMP-dependent protein kinase activity in the locus coeruleus contributes to the biochemical basis of opiate addiction.


The impact of iodine supplementation and bread fortification on urinary iodine concentrations in a mildly iodine deficient population of pregnant women in South Australia.
Mild iodine deficiency during pregnancy can have significant effects on fetal development and future cognitive function. The purpose of this study was to characterise the iodine status of South Australian women during pregnancy and relate it to the use of iodine-containing multivitamins. The impact of fortification of bread with iodized salt was also assessed. Women (n = 196) were recruited prospectively at the beginning of pregnancy and urine collected at 12, 18, 30, 36 weeks gestation and 6 months postpartum. The use of a multivitamin supplement was recorded at each visit. Spot urinary iodine concentrations (UIC) were assessed. Median UICs were within the mildly deficient range in women not taking supplements (<90 μg/L). Among the women taking iodine-containing multivitamins UICs were within WHO recommendations (150-249 μg/L) for sufficiency and showed an increasing trend through gestation. The fortification of bread with iodized salt increased the median UIC from 68 μg/L to 84 μg/L (p = .011) which was still in the deficient range. Pregnant women in this region of Australia were unlikely to reach recommended iodine levels without an iodine supplement, even after the mandatory iodine supplementation of bread was instituted in October 2009.


Molecular Analysis of a Plum pox virus W Isolate in Plum Germplasm Hand Carried into the USA from the Ukraine Shows a Close Relationship to a Latvian Isolate.
Four of 19 Prunus germplasm accessions hand carried from the Ukraine into the United States without authorization were found to be infected with Plum pox virus (PPV). Of the three isolates characterized, isolates UKR 44189 and UKR 44191 were confirmed to be isolates of PPV strain W, and UKR 44188 was confirmed to be an isolate of PPV strain D. UKR 44189 and UKR 44191 are very closely related to the PPV strain W isolate LV-145bt (HQ670748) from Latvia. Nucleotide and amino acid sequence identities between these three isolates were greater than 99%. This indicates that the isolates are very closely related and likely originated from a common source. The high genetic diversity among PPV-W strain isolates allowed the identification of potential recombination events between PPV isolates. It appears also that GF 305 peach and Prunus tomentosa are not hosts for the PPV isolate UKR 44189.


A hybrid controller method with genetic algorithm optimization to measure position and angular for mobile robot motion control.
Due to the complexity of autonomous mobile robot's requirement and drastic technological changes, the safe and efficient path tracking development is becoming complex and requires intensive knowledge and information, thus the demand for advanced algorithm has rapidly increased. Analyzing unstructured gain data has been a growing interest among researchers, resulting in valuable information in many fields such as path planning and motion control. Among those, motion control is a vital part of a fast, secure operation. Yet, current approaches face problems in managing unstructured gain data and producing accurate local planning due to the lack of formulation in the knowledge on the gain optimization. Therefore, this research aims to design a new gain optimization approach to assist researcher in identifying the value of the gain's product with a qualitative comparative study of the up-to-date controllers. Gains optimization in this context is to classify the near perfect value of the gain's product and processes. For this, a domain controller will be developed based on the attributes of the Fuzzy-PID parameters. The development of the Fuzzy Logic Controller requires information on the PID controller parameters that will be fuzzified and defuzzied based on the resulting 49 fuzzy rules. Furthermore, this fuzzy inference will be optimized for its usability by a genetic algorithm (GA). It is expected that the domain controller will give a positive impact to the path planning position and angular PID controller algorithm that meet the autonomous demand.


Identifying determinants of cullin binding specificity among the three functionally different Drosophila melanogaster Roc proteins via domain swapping.
Cullin-dependent E3 ubiquitin ligases (CDL) are key regulators of protein destruction that participate in a wide range of cell biological processes. The Roc subunit of CDL contains an evolutionarily conserved RING domain that binds ubiquitin charged E2 and is essential for ubiquitylation. Drosophila melanogaster contains three highly related Roc proteins: Roc1a and Roc2, which are conserved in vertebrates, and Roc1b, which is specific to Drosophila. Our previous genetic data analyzing Roc1a and Roc1b mutants suggested that Roc proteins are functionally distinct, but the molecular basis for this distinction is not known. Using co-immunoprecipitation studies we show that Drosophila Roc proteins bind specific Cullins: Roc1a binds Cul1-4, Roc1b binds Cul3, and Roc2 binds Cul5. Through domain swapping experiments, we demonstrate that Cullin binding specificity is strongly influenced by the Roc NH(2)-terminal domain, which forms an inter-molecular beta sheet with the Cullin. Substitution of the Roc1a RING domain with that of Roc1b results in a protein with similar Cullin binding properties to Roc1a that is active as an E3 ligase but cannot complement Roc1a mutant lethality, indicating that the identity of the RING domain can be an important determinant of CDL function. In contrast, the converse chimeric protein with a substitution of the Roc1b RING domain with that of Roc1a can rescue the male sterility of Roc1b mutants, but only when expressed from the endogenous Roc1b promoter. We also identified mutations of Roc2 and Cul5 and show that they cause no overt developmental phenotype, consistent with our finding that Roc2 and Cul5 proteins are exclusive binding partners, which others have observed in human cells as well. The Drosophila Roc proteins are highly similar, but have diverged during evolution to bind a distinct set of Cullins and to utilize RING domains that have overlapping, but not identical, function in vivo.


Supported employment: evidence for an evidence-based practice.
Supported employment for people with severe mental illnesses is an evidence-based practice, based on converging findings from 4 studies of the conversion of day treatment to supported employment and 9 randomized controlled trials comparing supported employment to a variety of alternative approaches. These two lines of research suggest that between 40% and 60% of consumers enrolled in supported employment obtain competitive employment while less than 20% of similar consumers do so when not enrolled in supported employment. Consumers who hold competitive jobs for a sustained period of time show benefits such as improved self-esteem and better symptom control, although by itself, enrollment in supported employment has no systematic impact on nonvocational outcomes, either on undesirable outcomes, such as rehospitalization, or on valued outcomes, such as improved quality of life. The psychiatric rehabilitation field has achieved consensus on a core set of principles of supported employment, although efforts continue to develop enhancements. A review of the evidence suggests strong support for 4 of 7 principles of supported employment, while the evidence for the remaining 3 is relatively weak. Continued innovation and research on principles is recommended.


Effect of different forms of information produced for cancer patients on their use of the information, social support, and anxiety: randomised trial.
To explore the hypothesis that different methods of selecting and printing information for cancer patients could improve emotional support by affecting interaction with others, and so lead to improved psychological wellbeing. Randomised trial with eight groups (three factors, 2x2x2). Data collected at recruitment and three month follow-up. 400 patients starting radiotherapy, of whom 325 with breast or prostate cancer and complete anxiety and depression data were included in the analysis. Printed booklets: half had only general information from CancerBACUP about each patient's cancer and half had personalised information from the patient's medical record plus selected general information; half were composed of information chosen interactively by the patient and half were produced automatically with a larger volume of material; and half had additional advice on anxiety management and half did not. Patients' views of the information, use of their booklets with others; change in reported social support; change in anxiety and depression. The larger booklets produced automatically were more likely to be found useful and to tell patients something new and less likely to be seen as too limited than the booklets produced interactively, but they were also more likely to overwhelm some patients. Personalised booklets were more likely than general booklets to tell patients something new. There was no difference in patients' perceived understanding of their cancer by any of the intervention factors. Patients with personalised information were more likely to show their booklets to others and to think it helped in discussing their cancer or its treatment. There were no major differences in social support, anxiety, or depression by any intervention factors. Patients were more likely to show personalised information to their confidants than general information. Further research is needed into the effects of sharing information on patients' social support and anxiety. Trial registration US Government Clinical Trials Database NCT00127465.


A Novel Method of Determining the Functional Effects of a Minor Genetic Modification of a Protein.
Contraction of muscles results from the ATP-coupled cyclic interactions of the myosin cross-bridges with actin filaments. Macroscopic parameters of contraction, such as maximum tension, speed of shortening, or ATPase activity, are unlikely to reveal differences between the wild-type and mutated (MUT) proteins when the level of transgenic protein expression is low. This is because macroscopic measurements are made on whole organs containing trillions of actin and myosin molecules. An average of the information collected from such a large assembly is bound to conceal any differences imposed by a small fraction of MUT molecules. To circumvent the averaging problem, the measurements were done on isolated ventricular myofibril (MF) in which thin filaments were sparsely labeled with a fluorescent dye. We isolated a single MF from a ventricle, oriented it vertically (to be able measure the orientation), and labeled 1 in 100,000 actin monomers with a fluorescent dye. We observed the fluorescence from a small confocal volume containing approximately three actin molecules. During the contraction of a ventricle actin constantly changes orientation (i.e., the transition moment of rigidly attached fluorophore fluctuates in time) because it is repetitively being "kicked" by myosin cross-bridges. An autocorrelation functions (ACFs) of these fluctuations are remarkably sensitive to the mutation of myosin. We examined the effects of Alanine to Threonine (A13T) mutation in the myosin regulatory light chain shown by population studies to cause hypertrophic cardiomyopathy. This is an appropriate example, because mutation is expressed at only 10% in the ventricles of transgenic mice. ACFs were either "Standard" (Std) (decaying monotonically in time) or "Non-standard" (NStd) (decaying irregularly). The sparse labeling of actin also allowed the measurement of the spatial distribution of actin molecules. Such distribution reflects the interaction of actin with myosin cross-bridges and is also remarkably sensitive to myosin mutation. The result showed that the A13T mutation caused 9% ACFs and 9% of spatial distributions of actin to be NStd, while the remaining 91% were Std, suggesting that the NStd performances were executed by the MUT myosin heads and that the Std performances were executed by non-MUT myosin heads. We conclude that the method explored in this study is a sensitive and valid test of the properties of low prevalence mutations in sarcomeric proteins.


Tumors in von Hippel-Lindau Syndrome: From Head to Toe-Comprehensive State-of-the-Art Review.
Von Hippel-Lindau syndrome (VHL) is an autosomal-dominant hereditary tumor disease that arises owing to germline mutations in the VHL gene, located on the short arm of chromosome 3. Patients with VHL may develop multiple benign and malignant tumors involving various organ systems, including retinal hemangioblastomas (HBs), central nervous system (CNS) HBs, endolymphatic sac tumors, pancreatic neuroendocrine tumors, pancreatic cystadenomas, pancreatic cysts, clear cell renal cell carcinomas, renal cysts, pheochromocytomas, paragangliomas, and epididymal and broad ligament cystadenomas. The VHL/hypoxia-inducible factor pathway is believed to play a key role in the pathogenesis of VHL-related tumors. The diagnosis of VHL can be made clinically when the characteristic clinical history and findings have manifested, such as the presence of two or more CNS HBs. Genetic testing for heterozygous germline VHL mutation may also be used to confirm the diagnosis of VHL. Imaging plays an important role in the diagnosis and surveillance of patients with VHL. Familiarity with the clinical and imaging manifestations of the various VHL-related tumors is important for early detection and guiding appropriate management. The purpose of this article is to discuss the molecular cytogenetics and clinical manifestations of VHL, review the characteristic multimodality imaging features of the various VHL-related tumors affecting multiple organ systems, and discuss the latest advances in management of VHL, including current recommendations for surveillance and screening. ©RSNA, 2018 An earlier incorrect version of this article appeared online. This article was corrected on April 9, 2018.


The structure and evolution of centromeric transition regions within the human genome.
An understanding of how centromeric transition regions are organized is a critical aspect of chromosome structure and function; however, the sequence context of these regions has been difficult to resolve on the basis of the draft genome sequence. We present a detailed analysis of the structure and assembly of all human pericentromeric regions (5 megabases). Most chromosome arms (35 out of 43) show a gradient of dwindling transcriptional diversity accompanied by an increasing number of interchromosomal duplications in proximity to the centromere. At least 30% of the centromeric transition region structure originates from euchromatic gene-containing segments of DNA that were duplicatively transposed towards pericentromeric regions at a rate of six-seven events per million years during primate evolution. This process has led to the formation of a minimum of 28 new transcripts by exon exaptation and exon shuffling, many of which are primarily expressed in the testis. The distribution of these duplicated segments is nonrandom among pericentromeric regions, suggesting that some regions have served as preferential acceptors of euchromatic DNA.


Hemodynamic changes during otolaryngological office-based flexible endoscopic procedures.
A preponderance of literature supports the safety of office-based flexible endoscopic procedures of the upper aerodigestive tract; however, until recently there were no data regarding hemodynamic stability during these procedures. A recent study showed intraprocedure changes in patients' hemodynamic parameters, raising the concern that perhaps patients should be monitored during these procedures. The aim of our study was to determine whether physiologically significant alterations in vital signs occur during office-based flexible endoscopic procedures. We performed a retrospective review of 100 consecutive patients who underwent office-based flexible endoscopic procedures of the upper aerodigestive tract from July 2010 to October 2011. Baseline values and the maximal changes in systolic blood pressure, diastolic blood pressure, heart rate, and oxygen saturation were recorded and compared. One hundred consecutive patients were included in the study. Twenty-one patients (21%) had severe hypertension and 40 patients (40%) had tachycardia during the procedure. The mean change overall in systolic blood pressure was 26.2 mm Hg (p < 0.001), the mean change in diastolic blood pressure was 13.9 mm Hg (p < 0.001), the mean change in heart rate was 16.6 beats per minute (p < 0.001), and the mean change in oxygen saturation was 1.6% (p < 0.001). These changes were significant. On further breakdown into groups, patients over 50 years of age and patients who were undergoing esophageal or laser procedures had significant elevations in heart rate (p = 0.01 and p = 0.04, respectively). An elevation in diastolic blood pressure was also significant in patients who were undergoing esophageal or laser procedures (p = 0.04 for both). These data concur with those of the previous report that found potentially significant hemodynamic changes during office-based procedures. Although preliminary, our findings suggest that it may be wise to monitor vital signs in patients over 50 years of age and patients who are undergoing an esophageal or laser procedure who are at risk for complications that could arise from tachycardia and hypertension.


Attenuated cold defense responses in orexin neuron-ablated rats.
Recent reports of the use of transgenic mice targeting orexin neurons show that the ablation of orexin neurons in the hypothalamus causes hypothermia during cold exposure. This suggests the importance of orexin neurons for cold-induced autonomic and physiological defense responses, including brown adipose tissue (BAT) thermogenesis and vasoconstriction in thermoregulatory cutaneous vascular bed. The present study investigated whether the ablation of orexin neurons attenuated cold-elicited BAT thermogenesis and cutaneous vasoconstriction. The study took advantage of our established conscious rat experimental model of direct measurement of BAT and body temperature and tail cutaneous blood flow. The study used transgenic orexin neurons-ablated (ORX-AB) rats and wild type (WT) rats. BAT temperature and tail artery blood flow with pre-implanted probes were measured, as well as behavioral locomotor activity under conscious free-moving condition. Gradually, the ambient temperature was decreased to below 5°C. ORX-AB rats showed an attenuated cold-induced BAT thermogenesis and behavioral activity, and delayed tail vasoconstriction. An ambient temperature that initiated BAT thermogenesis and established full cutaneous vasoconstriction was 14.1 ± 1.9 °C, which was significantly lower than 20.5 ± 1.9 °C, the corresponding value in WT rats (n = 10, P < 0.01). The results from this study suggest that the integrity of orexin-synthesising neurons in thermoregulatory networks is important for full expression of the cold defense responses.


Simultaneous determination of the HIV-protease inhibitors indinavir, amprenavir, ritonavir, saquinavir and nelfinavir in human plasma by reversed-phase high-performance liquid chromatography.
A rapid, simple and sensitive high-performance liquid chromatographic (HPLC) assay has been developed for the simultaneous quantification of the HIV-protease inhibitors indinavir, amprenavir, ritonavir, saquinavir and nelfinavir in human plasma. The method involved the solid-phase extraction of the five drugs and the internal standard (I.S., verapamil) from 400 microl of human plasma. The HPLC analysis used a reversed-phase C18 analytical column and a mobile phase consisting of a gradient with 15 mM phosphate buffer (pH 5.75)-acetonitrile and UV monitoring. The method was linear over the therapeutic concentration range for the five HIV-protease inhibitors. The accuracy of the method ranged from 98.2 to 106.7% and the precision values ranged from 1.4 to 8.1% for intra-day precision and from 3.1 to 6.4% for the inter-day values.


Argyrophilic nucleolar organizer regions (AgNORs) in relation to p53 and bcl-2 protein expression in leukemia patients.
The aim of this study was to assess the possible relationship between the silver stained nucleolar organizer regions (AgNOR) and immunocytochemically detected p53 and bcl-2 proteins in ALL, AML, B-CLL and CML patients (adults and children) at the initial presentation. AgNORs are loops of DNA, correlated with proliferative potential of cells. Alteration in p53 and bcl-2 proteins expression may characterize the malignant potential of leukemic cells. The patients were subdivided according to the p53 positivity and negativity. The frequency of p53-positive patients was relatively low in T-ALL (29%) and B-CLL (16%). B-ALL, AML and CML patients revealed higher frequency of p53 protein (46%, 47% and 88%, respectively). The overall frequency of positive cytoplasmic staining for bcl-2 protein was demonstrated in the majority of patients. No significant differences in the percentage of p53-positive cells among leukemia subtypes were seen. The proportion of bcl-2 protein positive cells did not differ significantly among various leukemia subtypes, except for significant differences between p53-positive and p53-negative peripheral blood (p=0.0073) and bone marrow (p=0.0175) cells of B-CLL patients. The samples from healthy subjects used as controls exhibited relatively low numbers of AgNOR dots in both, peripheral blood and bone marrow cells. Highly significant differences in AgNOR quantities between healthy donors and p53 protein positive peripheral blood as well as bone marrow cells of distinct leukemia subtypes (except for bone marrow cells in B-CLL patients, p=0.1727) were observed. Significant differences in AgNOR count between p53 protein positive and p53 protein negative samples of peripheral blood cells of B-ALL (p=0.0099) as well as B-CLL (p=0.0117) cases were found. No significant differences (except for B-CLL, p=0.0558) were encountered in bone marrow cells. P53 protein positivity or negativity did not influence the amount of AgNOR proteins in cells of our T-ALL and AML cases. Mutual comparing the number of AgNOR dots among different leukemias showed that for both peripheral blood and bone marrow cells the differences between ALL and AML (p=0.0383 and p=0.0033, respectively) as well as for peripheral blood of AML and CML (p=0.0302) were statistically significant. The bcl-2 protein positivity did not affect significantly the AgNOR distribution either in p53 protein positive or p53-negative cases of our leukemia patients. However, an association between the lowest AgNOR quantity and highest bcl-2 protein expression in p53-negative B-CLL patients was seen for both peripheral blood and bone marrow cells. The correlation between relatively high AgNOR numbers and relatively increased percentage of bcl-2 protein in the p53-positive cases of CML patients was found in some cases. Regarding the age and sex, the AgNOR distribution in p53-positive and p53-negative leukemia cases in children and adults showed neither relationship nor dependence. The WBC count differed evidently among distinct leukemia subtypes, with enormous heterogeneity in range as well. Larger studies are needed in order to consolidate these preliminary results and characterize the possible prognostic value of AgNOR in association with p53 and bcl-2 proteins expression.


Integration of palliative care into COVID-19 pandemic planning.
The COVID-19 pandemic is expected to surpass the healthcare system's capacity to provide intensive care to all patients who deteriorate as a result of the disease. This poses a unique challenge to healthcare teams of rationing care during pandemic when resources are scarce. Healthcare providers will need to acquire new skills in care decision making and effective symptom control for patients who do not receive life-saving measures. In this review, we describe some of the important palliative care considerations that need to be incorporated into COVID-19 pandemic planning. The main aspects to be considered include decision algorithms for rationing care, training on effective symptoms management, alternative delivery methods of palliative care services such as telemedicine and finally death and bereavement support for surviving family members who are likely to be isolated from their loved one at the moment of death.


Constitutive activity of the melanocortin-4 receptor is maintained by its N-terminal domain and plays a role in energy homeostasis in humans.
The melanocortin-4 receptor (MC4R), a centrally expressed G protein-coupled receptor (GPCR), is essential for the maintenance of long-term energy balance in humans. Mutations in MC4R are the most common genetic cause of obesity. Since activation of this receptor leads to a decrease in food intake, MC4R is also a major therapeutic target for the treatment of obesity. Control of MC4R activity in vivo is modulated by the opposing effects of the anorexigenic agonist alpha-melanocyte-stimulating hormone (alpha-MSH) and the orexigenic antagonist agouti-related protein (AGRP). In addition, experiments in vitro have demonstrated that the human MC4R has an intrinsic constitutive activity on which AGRP also acts as an inverse agonist. The physiological role of this constitutive activity in the control of energy balance as well as the domain of the protein implicated in its maintenance are unknown. By systematically studying functional defects in naturally occurring MC4R mutations from obese patients, we defined a cluster of N-terminal mutations that selectively impair the constitutive activity of the receptor. Further characterization of this domain demonstrated that it functions as a tethered intramolecular ligand that maintains the constitutive activity of MC4R and may provide novel avenues for the design of drugs targeting this receptor. Our results also suggest that the tonic satiety signal provided by the constitutive activity of MC4R may be required for maintaining long-term energy homeostasis in humans.


Hyperbaric oxygen therapy restored traumatic stress-induced dysregulation of fear memory and related neurochemical abnormalities.
Individuals with posttraumatic stress disorder (PTSD) are characterized by fear memory problems and hypocortisolemia of which traumatic stress-induced monoaminergic disruption over infralimbic (IL) cortex is considered the key mechanism. Hyperbaric oxygen therapy (HBOT) has recently proven its utility in treating several mental disorders but remains unexplored for PTSD. The present study aimed to examine the effects of 5-day HBO paradigm on traumatic stress (single prolonged stress, SPS, an animal model of PTSD)-induced dysregulation of fear memory/anxiety profiles and related abnormalities in IL monoamines and plasma corticosterone. Rats were randomly assigned to four groups (CON-sham, CON-HBOT, SPS-sham, and SPS-HBOT) and received Pavlovian fear conditioning test or elevated-T maze (ETM). The extracellular and tissue levels of monoamines over the IL cortex and the activity of the hypothalamus-pituitary-adrenal axis (i.e., the plasma corticosterone level and expression of the glucocorticoid receptor (GR) in the IL, hippocampus, amygdala, and hypothalamus) were measured. The results demonstrated that HBOT restored behaviorally the SPS-impaired fear extinction retrieval ability and SPS-induced conditioned anxiety, and neurochemically the SPS-reduced IL monoamines efflux level, and the corticosterone profiles. The present study shows some positive effects of HBOT in both behavioral and neurochemical profiles of PTSD outcomes.


Spatiotemporal imaging of complexity.
What are the functional neuroimaging measurements required for more fully characterizing the events and locations of neocortical activity? A prime assumption has been that modulation of cortical activity will inevitably be reflected in changes in energy utilization (for the most part) changes of glucose and oxygen consumption. Are such a measures complete and sufficient? More direct measures of cortical electrophysiological activity show event or task-related modulation of amplitude or band-limited oscillatory power. Using magnetoencephalography (MEG), these measures have been shown to correlate well with energy utilization sensitive BOLD fMRI. In this paper, we explore the existence of state changes in electrophysiological cortical activity that can occur independently of changes in averaged amplitude, source power or indices of metabolic rates. In addition, we demonstrate that such state changes can be described by applying a new measure of complexity, rank vector entropy (RVE), to source waveform estimates from beamformer-processed MEG. RVE is a non-parametric symbolic dynamic informational entropy measure that accommodates the wide dynamic range of measured brain signals while resolving its temporal variations. By representing the measurements by their rank values, RVE overcomes the problem of defining embedding space partitions without resorting to signal compression. This renders RVE-independent of absolute signal amplitude. In addition, this approach is robust, being relatively free of tunable parameters. We present examples of task-free and task-dependent MEG demonstrating that RVE provides new information by uncovering hidden dynamical structure in the apparent turbulent (or chaotic) dynamics of spontaneous cortical activity.


Tree of motility - A proposed history of motility systems in the tree of life.
Motility often plays a decisive role in the survival of species. Five systems of motility have been studied in depth: those propelled by bacterial flagella, eukaryotic actin polymerization and the eukaryotic motor proteins myosin, kinesin and dynein. However, many organisms exhibit surprisingly diverse motilities, and advances in genomics, molecular biology and imaging have showed that those motilities have inherently independent mechanisms. This makes defining the breadth of motility nontrivial, because novel motilities may be driven by unknown mechanisms. Here, we classify the known motilities based on the unique classes of movement-producing protein architectures. Based on this criterion, the current total of independent motility systems stands at 18 types. In this perspective, we discuss these modes of motility relative to the latest phylogenetic Tree of Life and propose a history of motility. During the ~4 billion years since the emergence of life, motility arose in Bacteria with flagella and pili, and in Archaea with archaella. Newer modes of motility became possible in Eukarya with changes to the cell envelope. Presence or absence of a peptidoglycan layer, the acquisition of robust membrane dynamics, the enlargement of cells and environmental opportunities likely provided the context for the (co)evolution of novel types of motility.


A comprehensive molecular study on Coffin-Siris and Nicolaides-Baraitser syndromes identifies a broad molecular and clinical spectrum converging on altered chromatin remodeling.
Chromatin remodeling complexes are known to modify chemical marks on histones or to induce conformational changes in the chromatin in order to regulate transcription. De novo dominant mutations in different members of the SWI/SNF chromatin remodeling complex have recently been described in individuals with Coffin-Siris (CSS) and Nicolaides-Baraitser (NCBRS) syndromes. Using a combination of whole-exome sequencing, NGS-based sequencing of 23 SWI/SNF complex genes, and molecular karyotyping in 46 previously undescribed individuals with CSS and NCBRS, we identified a de novo 1-bp deletion (c.677delG, p.Gly226Glufs*53) and a de novo missense mutation (c.914G>T, p.Cys305Phe) in PHF6 in two individuals diagnosed with CSS. PHF6 interacts with the nucleosome remodeling and deacetylation (NuRD) complex implicating dysfunction of a second chromatin remodeling complex in the pathogenesis of CSS-like phenotypes. Altogether, we identified mutations in 60% of the studied individuals (28/46), located in the genes ARID1A, ARID1B, SMARCB1, SMARCE1, SMARCA2, and PHF6. We show that mutations in ARID1B are the main cause of CSS, accounting for 76% of identified mutations. ARID1B and SMARCB1 mutations were also found in individuals with the initial diagnosis of NCBRS. These individuals apparently belong to a small subset who display an intermediate CSS/NCBRS phenotype. Our proposed genotype-phenotype correlations are important for molecular screening strategies.


Distinguishing hyperglycemic changes by Set7 in vascular endothelial cells.
Epigenetic changes are implicated in the persisting vascular effects of hyperglycemia. The precise mechanism whereby chromatin structure and subsequent gene expression are regulated by glucose in vascular endothelial cells remain to be fully defined. We have studied the molecular and functional mechanism whereby the Set7 methyltransferase associates with chromatin formation and histone methylation in vascular cells in response to current and previous exposure to glucose. To characterize the molecular and functional identity of the Set7 protein, we used vascular cells overexpressing or lacking Set7. Chromatin fractionation for mono-methylation of lysine 4 on histone H3 identified methyltransferase activity. Immunofluorescence experiments strongly suggest that Set7 protein accumulates in the nucleus in response to hyperglycemia. Moreover, activation of proinflammatory genes by high glucose is dependent on Set7 but distinguished by H3K4m1 gene patterns. We show that transient hyperglycemia regulates the expression of proinflammatory genes in vascular endothelial cells in vitro and the persistent increase in glucose-induced gene expression in the aorta of nondiabetic mice. This study uncovers that the response to hyperglycemia in vascular endothelial cells involves the H3K4 methyltransferase, Set7. This enzyme appears to regulate glucose-induced chromatin changes and gene expression not only by H3K4m1-dependent but also H3K4m1-independent pathways. Furthermore, Set7 appears to be responsible for sustained vascular gene expression in response to prior hyperglycemia and is a potential molecular mechanism for the phenomenon of hyperglycemic memory.


Poly (ADP-ribose) polymerase activation and circulatory shock.
Sepsis is associated with increased production of reactive oxidant species. Oxidative and nitrosative stress can lead to activation of the nuclear enzyme poly (ADP-ribose) polymerase (PARP), with subsequent loss of cellular functions. Activation of PARP may dramatically lower the intracellular concentration of its substrate, NAD thus slowing the rate of glycolysis, electron transport and subsequently ATP formation. This process can result in cell dysfunction and cell death. In addition, PARP enhances the expression of various pro-inflammatory mediators, via activation of NF-kappaB, MAP kinase and AP-1 and other signal transduction pathways. Preclinical studies in various rodent and large animal models demonstrate that PARP inhibition or PAR deficiency exerts beneficial effects on the haemodynamic and metabolic alterations associated with septic and haemorrhagic shock. Recent human data also support the role of PARP in septic shock: In a retrospective study in 25 septic patients, an increase in plasma troponin level was related to increased mortality risk. In patients who died, significant myocardial damage was detected, and histological analysis of heart showed inflammatory infiltration, increased collagen deposition, and derangement of mitochondrial criptae. Immunohistochemical staining for poly(ADP-ribose) (PAR), the product of activated PARP was demonstrated in septic hearts. There was a positive correlation between PAR staining and troponin I; and a correlation of PAR staining and LVSSW. Thus, there is significant PARP activation in animal models subjected to circulatory shock, as well as in the hearts of septic patients. Based on the interventional studies in animals and the correlations observed in patients we propose that PARP activation may be, in part responsible for the cardiac depression and haemodynamic failure seen in humans with severe sepsis. Interestingly, recent studies reveal that the protective effects of PARP inhibitors are predominant in male animals, and are not apparent in female animals. Oestrogen, by providing a baseline inhibitory effect on PARP activation, may be partially responsible for this gender difference.


The Impact of Emerging Bioconjugation Chemistries on Radiopharmaceuticals.
The use of radiolabeled antibodies, immunoglobulin fragments, and other proteins are an increasingly important sector of research for diagnostic imaging and targeted radiotherapy in nuclear medicine. As with all radiopharmaceuticals, efficient radiochemistry is a prerequisite to clinical translation. For proteins, variations in the primary amino acid sequence, the secondary structures, and tertiary folds, as well as differences in the size, charge, polarity, lipophilicity, and the presence of posttranslational modifications, add complexity to the system. The choice of radionuclide or chelate, and its impact on the thermodynamic, kinetic, and metabolic stability of a radiotracer, has attracted much attention but the chemistry by which the radionuclide is conjugated to the protein scaffold is of equal importance. Recently, a wealth of creative advances in protein ligation methods based on chemical, photochemical, and enzyme-mediated processes has emerged. As radiochemists explore alternative bioconjugation strategies, this article considers their potential impact on radiotracer design.


At issue: The future of cognitive rehabilitation of schizophrenia.
Cognitive rehabilitation is becoming an increasingly popular intervention in treatment programs for people with schizophrenia. Despite this increased acceptance, however, the evidence base on its effectiveness is not impressive. Moreover, given the evidence of cognitive recovery in treated patients who do not receive cognitive rehabilitation--from newer medications and from other evidence-based psychosocial interventions--it is not clear whether cognitive rehabilitation is worth its expense in time and resources. We believe that the slow progress in the field of cognitive rehabilitation of schizophrenia is related to failure to address several critical issues: (1) the importance of manipulating stimulus and context structure in rehabilitative interventions; (2) the need to base a cognitive rehabilitation of schizophrenia on cognitive neuroscience as opposed to neuropsychology; (3) the importance of systematically addressing motivation, self-esteem, and affective factors when designing cognition-enhancing interventions; (4) the need to move beyond one-size-fits-all interventions and develop individualized treatments; and (5) the need to address abnormalities in the experience of the self when designing interventions to optimize cognitive and behavioral performance. Suggestions for addressing these issues are discussed.


Neurologic melioidosis.
Melioidosis is an important cause of morbidity and mortality in northern Australia and Southeast Asia. Diagnosis is best made by isolation of Burkholderia pseudomallei from clinical specimens. A variety of clinical presentations are described, including neurologic disease. The aim of this study was to review admissions with confirmed neurologic melioidosis to a regional hospital in a region to which melioidosis is endemic during 1995-2011. There were 12 culture-confirmed cases of neurologic melioidosis, of which two were detected by analysis of cerebrospinal fluid. Four of these cases were in children. Significant clinical features were fever, headache, and ataxia. Common changes on magnetic resonance imaging T2-weighted scans included ring-enhancing lesions and leptomeningeal enhancement. There were four deaths and an additional four patients had significant long-term neurologic sequelae. When considering the etiology of undifferentiated neurologic disease, an awareness of the possibility of neurologic melioidosis is important in disease-endemic regions.


OARSI Clinical Trials Recommendations: Hand imaging in clinical trials in osteoarthritis.
Tremendous advances have occurred in our understanding of the pathogenesis of hand osteoarthritis (OA) and these are beginning to be applied to trials targeted at modification of the disease course. The purpose of this expert opinion, consensus driven exercise is to provide detail on how one might use and apply hand imaging assessments in disease modifying clinical trials. It includes information on acquisition methods/techniques (including guidance on positioning for radiography, sequence/protocol recommendations/hardware for MRI); commonly encountered problems (including positioning, hardware and coil failures, sequences artifacts); quality assurance/control procedures; measurement methods; measurement performance (reliability, responsiveness, validity); recommendations for trials; and research recommendations.


Quality of evidence-based guidelines for platelet transfusion and use: A systematic review.
Guidelines for platelet (PLT) transfusion are an important source of information for clinicians. Although guidelines intend to increase consistency and quality of care, variation in methodology and recommendations may exist that could impact the value of a guideline. We aimed to determine the quality of existing PLT transfusion guidelines using the Appraisal of Guidelines for Research and Evaluation (AGREE II) instrument and to describe the inconsistencies in recommendations. A systematic search was undertaken for evidence-based guidelines from January 1, 2013, to January 25, 2019. Citations were reviewed in duplicate for inclusion and descriptive data extracted. Four physicians appraised the guideline using the AGREE II instrument and the scaled score for each item evaluated was calculated. The protocol was registered in PROSPERO. Of 6744 citations, 6740 records were screened. Seven of 28 full-text studies met the inclusion criteria. The median scaled score (and the interquartile range of the scaled score) for the following items were as follows: scope and purpose, 94% (8%); stakeholder involvement, 63% (18%); rigor of development, 83% (14%); clarity of presentation, 94% (6%); applicability, 58% (20%); and editorial independence, 77% (4%). Overall quality ranged from 4 to 7 (7 is the maximum score). Inconsistent recommendations were on prophylactic PLT transfusion in hypoproliferative thrombocytopenia in the presence of risk factors and dose recommendations. Inconsistencies between guidelines and variable quality highlight areas for future guideline writers to address. Areas of specific attention include issues of stakeholder involvement and applicability.


Supercontinuum assisted trapped electron accumulation in titanium oxide gel by femtosecond laser pulses.
Long-lived trapped electrons are generated in titanium oxide gels in a convergent part of a loosely focused beam from a millijoule, femtosecond Ti:Sapphire laser. The appearance of trapped electrons is accompanied by the generation of the white supercontinuum. Considering the relevant physical mechanisms of trapped electron accumulation, we show that two-photon interband transitions with participation of photons at the fundamental frequency and from the blue part of the supercontinuum explain the experimental findings.


Exploring the link between MORF4L1 and risk of breast cancer.
Proteins encoded by Fanconi anemia (FA) and/or breast cancer (BrCa) susceptibility genes cooperate in a common DNA damage repair signaling pathway. To gain deeper insight into this pathway and its influence on cancer risk, we searched for novel components through protein physical interaction screens. Protein physical interactions were screened using the yeast two-hybrid system. Co-affinity purifications and endogenous co-immunoprecipitation assays were performed to corroborate interactions. Biochemical and functional assays in human, mouse and Caenorhabditis elegans models were carried out to characterize pathway components. Thirteen FANCD2-monoubiquitinylation-positive FA cell lines excluded for genetic defects in the downstream pathway components and 300 familial BrCa patients negative for BRCA1/2 mutations were analyzed for genetic mutations. Common genetic variants were genotyped in 9,573 BRCA1/2 mutation carriers for associations with BrCa risk. A previously identified co-purifying protein with PALB2 was identified, MRG15 (MORF4L1 gene). Results in human, mouse and C. elegans models delineate molecular and functional relationships with BRCA2, PALB2, RAD51 and RPA1 that suggest a role for MRG15 in the repair of DNA double-strand breaks. Mrg15-deficient murine embryonic fibroblasts showed moderate sensitivity to γ-irradiation relative to controls and reduced formation of Rad51 nuclear foci. Examination of mutants of MRG15 and BRCA2 C. elegans orthologs revealed phenocopy by accumulation of RPA-1 (human RPA1) nuclear foci and aberrant chromosomal compactions in meiotic cells. However, no alterations or mutations were identified for MRG15/MORF4L1 in unclassified FA patients and BrCa familial cases. Finally, no significant associations between common MORF4L1 variants and BrCa risk for BRCA1 or BRCA2 mutation carriers were identified: rs7164529, Ptrend = 0.45 and 0.05, P2df = 0.51 and 0.14, respectively; and rs10519219, Ptrend = 0.92 and 0.72, P2df = 0.76 and 0.07, respectively. While the present study expands on the role of MRG15 in the control of genomic stability, weak associations cannot be ruled out for potential low-penetrance variants at MORF4L1 and BrCa risk among BRCA2 mutation carriers.


DNA Transformer for Visualizing Endogenous RNA Dynamics in Live Cells.
The functions of RNA are tightly regulated via diverse intracellular mechanisms. However, probing the complex dynamics of endogenous RNA in live cells is a challenging task. In the present study, a DNA transformer is designed for visualizing the abundance, distribution, and mobility of endogenous mRNAs in live human cells. The transformable tetrahedral DNA (T-TED) probe has a flexible hinge structure and is programmed to conform into a 3D tetrahedron upon binding with the target mRNA. By incorporating Förster resonance energy transfer (FRET) imaging, super-resolution localization, and single particle tracking, the T-TED biosensor is applied for investigating the dynamics of Delta-like ligand 4 (Dll4) mRNA, which encodes a transmembrane protein, in human pulmonary microvascular endothelial cells. The data reveal unprecedented subpopulations of Dll4 mRNA with distinct mobility organized spatially in association with the endoplasmic reticulum and microtubule networks. The ability to monitor the dynamics of endogenous RNA in live human cells will provide a useful tool for studying the functions and regulation of RNA.


Volatile Gas Production by Methyl Halide Transferase: An In Situ Reporter Of Microbial Gene Expression In Soil.
Traditional visual reporters of gene expression have only very limited use in soils because their outputs are challenging to detect through the soil matrix. This severely restricts our ability to study time-dependent microbial gene expression in one of the Earth's largest, most complex habitats. Here we describe an approach to report on dynamic gene expression within a microbial population in a soil under natural water levels (at and below water holding capacity) via production of methyl halides using a methyl halide transferase. As a proof-of-concept application, we couple the expression of this gas reporter to the conjugative transfer of a bacterial plasmid in a soil matrix and show that gas released from the matrix displays a strong correlation with the number of transconjugant bacteria that formed. Gas reporting of gene expression will make possible dynamic studies of natural and engineered microbes within many hard-to-image environmental matrices (soils, sediments, sludge, and biomass) at sample scales exceeding those used for traditional visual reporting.


IL-18 Gene Promoter Region 607C/A Polymorphism in HIV-1 Infected North Indian Population.
Several host genetic factors play an important role in susceptibility to human immunodeficiency virus type 1 (HIV-1) infection and in its progression to acquired immune deficiency syndrome (AIDS). The interleukin-18 (IL-18) is a multifunctional proinflammatory cytokine that regulates immune responses and plays a pathogenic role in HIV-1 infection by enhancing viral replication. Single nucleotide polymorphisms (SNPs) in the IL-18 gene promoter region may lead to altered transcriptional activity and IL-18 production, and may account for variation in the risk of HIV-1 infection. We have investigated the association between IL-18 promoter polymorphism -607C>A and HIV-1 infection through a case-control study of 500 patients with HIV-1/AIDS and an equal number of age and sex matched controls in a north Indian population. Genotyping using sequence specific primer-polymerase chain reaction (SSP-PCR) showed a statistically significant reduced risk of HIV-1 infection for the A>A genotype [odds ratio (OR) = 0.57, 95% confidence interval (95% CI) = 0.33-0.98, p = 0.040], but not for the C>A genotype (OR = 0.87, 95% CI = 0.66-1.14, p = 0.321). We concluded that the -607A allele of the IL-18 gene promoter polymorphism may play a protective role against the progression of HIV-1 infection in this population.


The influence of AHI1 variants on the diagnosis and treatment outcome in schizophrenia.
The present study aimed to explore whether four single nucleotide polymorphisms (SNPs) within the AHI1 gene could be associated with schizophrenia (SCZ) and whether they could predict the clinical outcomes in SCZ patients treated with antipsychotics. Four hundred twenty-six (426) in-patients with SCZ and 345 controls were genotyped for four AHI1 SNPs (rs11154801, rs7750586, rs9647635 and rs9321501). Baseline and clinical measures for SCZ patients were assessed through the Positive and Negative Syndrome Scale (PANSS). Allelic and genotypic frequencies in SCZ subjects were compared with those of controls using the χ2 statistics. The repeated-measure ANOVA was used for the assessment of treatment outcomes measured by PANSS changes. The case-control analysis did not show any difference in the genotypic distribution of the SNPs, while in the allelic analysis, a weak association was found between the rs9647635 A allele and SCZ. Furthermore, in the haplotype analysis, three haplotypes resulted in being associated with SCZ. On the other hand, two SNPs (rs7750586 and rs9647635) were associated with clinical improvement of negative symptoms in the allelic analysis, although in the genotypic analysis, only trends of association were found for the same SNPs. Our findings suggest a possible influence of AHI1 variants on SCZ susceptibility and antipsychotic response, particularly concerning negative symptomatology. Subsequent well-designed studies would be mandatory to confirm our results due to the methodological shortcomings of the present study.


Correlation of intrinsic in vitro and in vivo clearance for drugs metabolized by hepatic UDP-glucuronosyltransferases in rats.
A method for quantitatively predicting the hepatic clearance of drugs by UDP-glucuronosyltransferases (UGTs) from in vitro data has not yet been established. We examined the relationship between in vitro and in vivo intrinsic clearance by rat hepatic UGTs using 10 drugs. For these 10 drugs, the in vitro intrinsic clearance by UGTs (CL(int, in vitro)) measured using alamethicin-activated rat liver microsomes was in the range 0.10-4500 ml/min/kg. Microsomal binding (f(u, mic)) was determined to be in the range 0.29-0.95 and the unbound intrinsic clearance (CL(uint, in vitro)) to be in the range 0.11-9600 ml/min/kg. The contribution of rat hepatic glucuronidation to drug elimination was 12.0%-76.6% and in vivo intrinsic clearance by UGTs was 5.7-9000 ml/min/kg. To evaluate the discrepancy between the in vitro and in vivo values, a scaling factor was calculated (CL(int, in vivo)/CL(int, in vitro)); the values were found to be in the range 0.89-110. The average fold error of the scaling factor values incorporating f(u, mic) was closer to unity than that without f(u, mic). The scaling factor values incorporating f(u, mic) were <10 in 8/10 drugs and <2 in 6/10 drugs, indicating a small discrepancy between in vitro and in vivo values. Thus, using alamethicin-activated liver microsomes, incorporating f(u, mic) into CL(int, in vitro), and considering the contribution of glucuronidation may enable us to quantitatively predict in vivo hepatic glucuronidation from in vitro data.


Sex differences in adolescent methylphenidate sensitization: effects on glial cell-derived neurotrophic factor and brain-derived neurotrophic factor.
This study analyzed sex differences in methylphenidate (MPH) sensitization and corresponding changes in glial cell-derived neurotrophic factor (GDNF) and brain-derived neurotprhic factor protein (BDNF) in adolescent male and female rats. After habituation to a locomotor arena, animals were sensitized to MPH (5mg/kg) or saline from postnatal day (P) 33-49, tested every second day. On P50, one group of animals were injected with saline and behavior assessed for conditioned hyperactivity. Brain tissue was harvested on P51 and analyzed for GDNF protein. A second group of animals was also sensitized to MPH from P33 to 49, and expression of behavioral sensitization was analyzed on a challenge given at P60, and BDNF protein analyzed at P61. Females demonstrated more robust sensitization to MPH than males, but only females given MPH during sensitization demonstrated conditioned hyperactivity. Interestingly, MPH resulted in a significant increase in striatal and accumbal GDNF with no sex differences revealed. Results of the challenge revealed that females sensitized and challenged with MPH demonstrated increased activity compared to all other groups. Regarding BDNF, only males given MPH demonstrated an increase in dorsal striatum, whereas MPH increased accumbal BDNF with no sex differences revealed. A hierarchical regression analysis revealed that behavioral sensitization and the conditioned hyperactivity test were reliable predictors of striatal and accumbal GDNF, whereas sensitization and activity on the challenge were reliable predictors of accumbal BDNF, but had no relationship to striatal BDNF. These data have implications for the role of MPH in addiction and dopamine system plasticity.