AgNPs Change Microbial Community Structures of Wastewater.
Due to their strong antimicrobial activity, silver nanoparticles (AgNPs) are massively produced, applied, consumed and, as a negative consequence, released into wastewater treatment plants. Most AgNPs are assumed to be bound by sludge, and thus bear potential risk for microbial performance and stability. In this lab-scale study, flow cytometry as a high-throughput method and 16S rRNA gene amplicon Illumina MiSeq sequencing were used to track microbial community structure changes when being exposed to AgNPs. Both methods allowed deeper investigation of the toxic impact of chemicals on microbial communities than classical EC50 determination. In addition, ecological metrics were used to quantify microbial community variations depending on AgNP types (10 and 30 nm) and concentrations. Only low changes in α- and intra-community β-diversity values were found both in successive negative and positive control batches and batches that were run with AgNPs below the EC50 value. Instead, AgNPs at EC50 concentrations caused upcoming of certain and disappearance of formerly dominant subcommunities. Flavobacteriia were among those that almost disappeared, while phylotypes affiliated with Gammaproteobacteria (3.6-fold) and Bacilli (8.4-fold) increased in cell abundance in comparison to the negative control. Thus, silver amounts at the EC50 value affected community structure suggesting a potential negative impact on functions in wastewater treatment systems.


The endothelial safety of using a gentian violet dry-ink "S" stamp for precut corneal tissue.
To determine the endothelium damage resulting from the application of "dry" gentian violet (GV) stromal markings on corneas precut for Descemet stripping automated endothelial keratoplasty (DSAEK) using a Moria "S" Stamp. Five precut corneas had the stromal graft bed marked with GV using the Moria "S" stamp and 6 precut corneas were left unmarked as controls. After applying the ink to the stamp, care was taken to allow the alcohol carrier to dry for 10 seconds before applying the dry dye to the stromal surface. Tissue was then trephinated and stained with calcein AM to assess endothelial viability. Grafts were photographed and digital pixel planometry, using an established analysis technique, was used to compare the damage between the control and experimental groups. The mean percent cell damage of corneas treated with GV "S" stamp (n = 5) was 8.6% (range 4.4-12.9), and it was 8.1% (range 3.9-15.1) in the DSAEK control set (n = 6). Median percent cell damage was 6.7% among GV-treated corneas and 7.4% among control corneas. The distributions were not significantly different between groups (Mann-Whitney U test = 15.0, two-tailed P = 1.0). Moreover, no "S" pattern of damage was seen in any study eye. There were no significant differences in endothelial damage between the 2 groups. GV stromal markings may be applied without undue damage to the endothelium using the dry-ink technique described.


Association between preoperative diuretic use and in-hospital outcomes after cardiac surgery.
There is a paucity of evidence on the association between preoperative diuretics use and outcomes following cardiac surgery. We hypothesized that diuretic use prior to cardiac surgery will be associated with adverse in-hospital clinical outcomes. We evaluated patients undergoing cardiac surgery at a single institution between January 1, 2000, and December 31, 2011. Patients were grouped as follows: isolated coronary artery bypass grafting (CABG) (n = 8759), CABG plus valve surgery (n = 1188), and valve surgery only (n = 2646). A fourth group "All cardiac surgery" is comprised of patients from all three groups. Preoperative diuretic use was defined as patient on any diuretic till the day of surgery. Primary outcome was the incidence of major adverse events (MAEs) defined as the composite of mortality, postoperative renal dysfunction, myocardial infarction, stroke, and new-onset atrial fibrillation (AF). Logistic regression analysis and propensity score matching were performed. We included 12,593 patients [3546 on diuretic (28%)]. After logistic regression analyses, preoperative diuretic use was associated with an increased risk of the following: (1) MAE among all groups except the concomitant CABG and valve surgery group, (2) AF in "All cardiac surgery" and isolated CABG groups, (3) postoperative renal dysfunction in all groups. After propensity score matching (n = 3050 in each group), preoperative diuretic use was significantly associated with MAE (48% vs. 43%; P < 0.0001), postoperative renal dysfunction (19% vs. 14%; P < 0.0001), and AF (34% vs. 32%; P = 0.03) in the "All cardiac surgery" group. Preoperative diuretics use is associated with an increased incidence of MAEs after cardiac surgery.


Unusual distribution of mitochondrial large subunit rRNA in the cytosol during conjugation in Tetrahymena thermophila.
The distribution of mitochondria during conjugation of the ciliated protozoan Tetrahymena thermophila was surveyed using a mitochondrial stain and fluorescence in situ hybridization (FISH). When the mitochondria-specific stain, Mito-Tracker, was used, the majority of mitochondria were detected in the cortex; their distribution was not changed during conjugation. On the other hand, FISH using mitochondrial large subunit (LSU) rRNA as a probe showed an unusual distribution of signals during conjugation. Unexpectedly, the signals were detected throughout the cytoplasm of conjugating cells. These signals were not observed in pre-mating cells and in exconjugants. The cytosolic localization of mitochondrial rRNA was supported by northern blot analysis using post-mitochondrial RNA fraction at the later stages of conjugation. These observations suggest selective mitochondrial breakdown or transport of LSU rRNA into cytosol. The biological significance of the conjugation-specific appearance of the cytosolic mitochondrial rRNA is discussed.


Quality-score refinement of SSU rRNA gene pyrosequencing differs across gene region for environmental samples.
Due to potential sequencing errors in pyrosequencing data, species richness and diversity indices of microbial systems can be miscalculated. The "traditional" sequence refinement method is not sufficient to account for overestimations (e.g., length, primer errors, ambiguous nucleotides). Recent in silico and single-organism studies have revealed the importance of sequence quality scores in the estimation of ecological indices; however, this is the first study to compare quality-score stringencies across four regions of the SSU rRNA gene sequence (V1V2, V3, V4, and V6) with actual environmental samples compared directly to corresponding clone libraries produced from the same primer sets. The nucleic acid sequences determined via pyrosequencing were subjected to varying quality-score cutoffs that ranged from 25 to 32, and at each quality-score cutoff, either 10 or 15 % of the nucleotides were allowed to be below the cutoff. When species richness estimates were compared for the tested samples, the cutoff values of Q27(15%), Q30(10%), and Q32(15%) for V1V2, V4, and V6, respectively, estimated similar values as obtained with clone libraries and Sanger sequencing. The most stringent Q tested (Q32(10%)) was not enough to account for species richness inflation of the V3 region pyrosequence data. Results indicated that quality-score assessment greatly improved estimates of ecological indices for environmental samples (species richness and α-diversity) and that the effect of quality-score filtering was region-dependent.


Melatonin induces neuritogenesis at early stages in N1E-115 cells through actin rearrangements via activation of protein kinase C and Rho-associated kinase.
Melatonin increases neurite formation in N1E-115 cells through microtubule enlargement elicited by calmodulin antagonism and vimentin intermediate filament reorganization caused by protein kinase C (PKC) activation. Microfilament rearrangement is also a necessary process in growth cone formation during neurite outgrowth. In this work, we studied the effect of melatonin on microfilament rearrangements present at early stages of neurite formation and the possible participation of PKC and the Rho-associated kinase (ROCK), which is a downstream kinase in the PKC signaling pathway. The results showed that 1 nm melatonin increased both the number of cells with filopodia and with long neurites. Similar results were obtained with the PKC activator phorbol 12-myristate 13-acetate (PMA). Both melatonin and PMA increased the quantity of filamentous actin. In contrast, the PKC inhibitor bisindolylmaleimide abolished microfilament organization elicited by either melatonin or PMA, while the Rho inhibitor C3, or the ROCK inhibitor Y27632, abolished the bipolar neurite morphology of N1E-115 cells. Instead, these inhibitors prompted neurite ramification. ROCK activity measured in whole cell extracts and in N1E-115 cells was increased in the presence of melatonin and PMA. The results indicate that melatonin increases the number of cells with immature neurites and suggest that these neurites can be susceptible to differentiation by incoming extracellular signals. Data also indicate that PKC and ROCK are involved at initial stages of neurite formation in the mechanism by which melatonin recruits cells for later differentiation.


ABT-431: the diacetyl prodrug of A-86929, a potent and selective dopamine D1 receptor agonist: in vitro characterization and effects in animal models of Parkinson's disease.
(-)-Trans 9,10-hydroxy-2-propyl-4,5,5a,6,7,11b-hexahydro-3-thia-5- azacyclopent-1-ena[c]phenanthrene hydrochloride (A-86929) is a potent and selective full agonist at the dopamine (DA) D1-like receptor. Judging by its binding affinities to the D1 and D2 classes of receptors, the compound is approximately 20-fold D1 receptor-selective, whereas relative potencies based on functional in vitro assays indicate that A-86929 is greater than 400-fold D1-selective. A-86929 has moderate to weak (Ki > 1 microM) affinity at other monoaminergic and peptidergic receptors, at ion channels and at monoamine uptake sites. The catechol of A-86929 was bis-acetylated to produce the prodrug, (-)-trans 9,10-acetoxy-2-propyl-4,5,5a,6,7,11-b-hexahydro-3-thia- 5-azacyclopent-1-ena[c]phenanthrene hydrochloride (ABT-431), which is more chemically stable yet is rapidly converted to the parent compound with a half-life of less than 1 min in plasma. Both A-86929 and ABT-431 produced contralateral rotation in rats bearing unilateral 6-hydroxydopamine lesions, with ED50 values of 0.24 mumol/kg s.c. and 0.54 mumol/kg s.c., respectively. A-86929 and ABT-431 improved behavioral disability scores and increased locomotor activity in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-lesioned marmoset model of Parkinson's disease in a dose-dependent manner (the minimum effective dose was 0.10 mumol/kg s.c.). When administered three times daily for 30 consecutive days to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-lesioned marmosets, A-86929 significantly improved disability scores throughout the duration of the study. Current Parkinson's disease therapy includes L-dopa, which stimulates both classes of DA receptors by virtue of its conversion to DA in vivo, and direct-acting D2-selective agonists. Stimulation of the D2 receptor, which is associated with all current DA agonist-based therapies, may contribute to their dose-limiting side effects. An agent such as A-86929 (or its prodrug ABT-431), which selectively stimulates the D1 receptor, may represent a novel mechanism for Parkinson's disease therapy with the potential for an improved side-effect profile and, consequently, improved patient compliance.


A search for cytomegalovirus and herpes viral antigen in brains of schizophrenic patients.
The peroxidase-antiperoxidase method was employed to search for evidence of cytomegalovirus (CMV) or herpes simplex virus (HSV) antigen in the brains of 25 patients with a diagnosis of schizophrenia, 25 nonschizophrenic neuropsychiatric patients, and 16 nonpsychiatric control subjects. Brain specimens from patients with acute CMV and herpes encephalitis served as positive controls. Although early results with low-titer CMV antisera suggested immunoreactivity in specific brain regions of a small number of schizophrenic and control cases, the present studies with high-titer anti-CMV IgG did not give a positive immunoperoxidase reaction in sections from the basal forebrain, hypothalamus, or midbrain. Scattered neurons in the lateral vestibular nucleus and hippocampus showed questionable staining with CMV IgG in one schizophrenic patient and none in control subjects. No schizophrenic or control cases demonstrated an immune reaction to HSV antisera.


Extending life for people with a terminal illness: a moral right and an expensive death? Exploring societal perspectives.
Many publicly-funded health systems apply cost-benefit frameworks in response to the moral dilemma of how best to allocate scarce healthcare resources. However, implementation of recommendations based on costs and benefit calculations and subsequent challenges have led to 'special cases' with certain types of health benefits considered more valuable than others. Recent debate and research has focused on the relative value of life extensions for people with terminal illnesses. This research investigates societal perspectives in relation to this issue, in the UK. Q methodology was used to elicit societal perspectives from a purposively selected sample of data-rich respondents. Participants ranked 49 statements of opinion (developed for this study), onto a grid, according to level of agreement. These 'Q sorts' were followed by brief interviews. Factor analysis was used to identify shared points of view (patterns of similarity between individuals' Q sorts). Analysis produced a three factor solution. These rich, shared accounts can be broadly summarised as: i) 'A population perspective - value for money, no special cases', ii) 'Life is precious - valuing life-extension and patient choice', iii) 'Valuing wider benefits and opportunity cost - the quality of life and death'. From the factor descriptions it is clear that the main philosophical positions that have long dominated debates on the just allocation of resources have a basis in public opinion. The existence of certain moral positions in the views of society does not ethically imply, and pragmatically cannot mean, that all are translated into policy. Our findings highlight normative tensions and the importance of critically engaging with these normative issues (in addition to the current focus on a procedural justice approach to health policy). Future research should focus on i) the extent to which these perspectives are supported in society, ii) how respondents' perspectives relate to specific resource allocation questions, and iii) the characteristics of respondents associated with each perspective.


A Quantitative Tri-fluorescent Yeast Two-hybrid System: From Flow Cytometry to In cellula Affinities.
We present a technological advancement for the estimation of the affinities of Protein-Protein Interactions (PPIs) in living cells. A novel set of vectors is introduced that enables a quantitative yeast two-hybrid system based on fluorescent fusion proteins. The vectors allow simultaneous quantification of the reaction partners (Bait and Prey) and the reporter at the single-cell level by flow cytometry. We validate the applicability of this system on a small but diverse set of PPIs (eleven protein families from six organisms) with different affinities; the dissociation constants range from 117 pm to 17 μm After only two hours of reaction, expression of the reporter can be detected even for the weakest PPI. Through a simple gating analysis, it is possible to select only cells with identical expression levels of the reaction partners. As a result of this standardization of expression levels, the mean reporter levels directly reflect the affinities of the studied PPIs. With a set of PPIs with known affinities, it is straightforward to construct an affinity ladder that permits rapid classification of PPIs with thus far unknown affinities. Conventional software can be used for this analysis. To permit automated analysis, we provide a graphical user interface for the Python-based FlowCytometryTools package.


Opposing actions of Stat1 and Stat6 on IL-13-induced up-regulation of early growth response-1 and platelet-derived growth factor ligands in pulmonary fibroblasts.
IL-13 is a key cytokine involved in airway remodeling in asthma. We previously reported that IL-13 stimulated the mitogenesis of lung fibroblasts via platelet-derived growth factor (PDGF)-AA. In this report, we show that IL-13 increases PDGF-A and PDGF-C mRNA levels through a dual intracellular cascade that requires coactivation of Stat6 and Stat1 to impact transcriptional regulation of the early growth response (Egr)-1 gene, which then drives PDGF expression. Increased levels of PDGF-AA and PDGF-CC protein were observed in vivo in the airways of IL-13 transgenic mice. IL-13 up-regulated PDGF-A and PDGF-C mRNA levels in lung fibroblasts isolated from three different background strains of mice. However, IL-13-induced PDGF-A and PDGF-C mRNA levels were significantly reduced in Stat6-deficient (Stat6(-/-)) fibroblasts as compared with wild-type Stat6(+/+) fibroblasts. In contrast, IL-13-induced PDGF-A and PDGF-C mRNAs were enhanced in Stat1(-/-) fibroblasts as compared with Stat1(+/+) fibroblasts. IL-13 did not up-regulate PDGF-A or PDGF-C mRNA levels in Egr-1(-/-) fibroblasts. Moreover, IL-13 did not increase Egr-1 mRNA and protein levels in Stat6(-/-) fibroblasts and yet enhanced Egr-1 mRNA and protein levels in Stat1(-/-) fibroblasts. Our findings support the hypothesis that Stat6 and Stat1 exert stimulatory and inhibitory effects on Egr-1 and PDGF ligand mRNA transcription, respectively. This novel mechanism could aid in identifying molecular targets for the treatment of chronic airway remodeling and fibrosis in asthma.


Stressful life events predict eating disorder relapse following remission: six-year prospective outcomes.
To examine prospectively the natural course of bulimia nervosa (BN) and eating disorder not-otherwise-specified (EDNOS) and test for the effects of stressful life events (SLE) on relapse after remission from these eating disorders. 117 female patients with BN (N = 35) or EDNOS (N = 82) were prospectively followed for 72 months using structured interviews performed at baseline, 6- and 12-months, and then yearly thereafter. ED were assessed with the structured clinical interview for DSM-IV, and monitored over time with the longitudinal interval follow-up evaluation. Personality disorders were assessed with the diagnostic interview for DSM-IV-personality-disorders, and monitored over time with the follow-along-version. The occurrence and specific timing of SLE were assessed with the life events assessment interview. Cox proportional-hazard-regression-analyses tested associations between time-varying levels of SLE and ED relapse, controlling for comorbid psychiatric disorders, ED duration, and time-varying personality-disorder status. ED relapse probability was 43%; BN and EDNOS did not differ in time to relapse. Negative SLE significantly predicted ED relapse; elevated work and social stressors were significant predictors. Psychiatric comorbidity, ED duration, and time-varying personality-disorder status were not significant predictors. Higher work and social stress represent significant warning signs for triggering relapse for women with remitted BN and EDNOS.


Multisensory processing in event-based prospective memory.
Failures in prospective memory (PM) - that is, the failure to remember intended future actions - can have adverse consequences. It is therefore important to study those processes that may help to minimize such cognitive failures. Although multisensory integration has been shown to enhance a wide variety of behaviors, including perception, learning, and memory, its effect on prospective memory, in particular, is largely unknown. In the present study, we investigated the effects of multisensory processing on two simultaneously-performed memory tasks: An ongoing 2- or 3-back working memory (WM) task (20% target ratio), and a PM task in which the participants had to respond to a rare predefined letter (8% target ratio). For PM trials, multisensory enhancement was observed for congruent multisensory signals; however, this effect did not generalize to the ongoing WM task. Participants were less likely to make errors for PM than for WM trials, thus suggesting that they may have biased their attention toward the PM task. Multisensory advantages on memory tasks, such as PM and WM, may be dependent on how attention resources are allocated across dual tasks.


Neuroserpin Is Strongly Expressed in the Developing and Adult Mouse Neocortex but Its Absence Does Not Perturb Cortical Lamination and Synaptic Proteome.
Neuroserpin is a serine protease inhibitor that regulates the activity of tissue-type plasminogen activator (tPA) in the nervous system. Neuroserpin is strongly expressed during nervous system development as well as during adulthood, when it is predominantly found in regions eliciting synaptic plasticity. In the hippocampus, neuroserpin regulates developmental neurogenesis, synaptic maturation and in adult mice it modulates synaptic plasticity and controls cognitive and social behavior. High expression levels of neuroserpin in the neocortex starting from prenatal stage and persisting during adulthood suggest an important role for the serpin in the formation of this brain region and in the maintenance of cortical functions. In order to uncover neuroserpin function in the murine neocortex, in this work we performed a comprehensive investigation of its expression pattern during development and in the adulthood. Moreover, we assessed the role of neuroserpin in cortex formation by comparing cortical lamination and neuronal maturation between neuroserpin-deficient and control mice. Finally, we evaluated a possible regulatory role of neuroserpin at cortical synapses in neuroserpin-deficient mice. We observed that neuroserpin is expressed starting from the beginning of corticogenesis until adulthood throughout the neocortex in several classes of glutamatergic projection neurons and GABA-ergic interneurons. However, in the absence of neuroserpin we did not detect any alteration either in cortical layer formation, or in neuronal soma size and dendritic length. Furthermore, no significant quantitative changes were observed in the proteome of cortical synapses upon neuroserpin deficiency. We conclude that, although strongly expressed in the neocortex, absence of neuroserpin does not lead to gross developmental abnormalities, and does not perturb the composition of the cortical synaptic proteome.


Radiofrequency coblation decreases blood loss during endoscopic sinonasal and skull base tumor removal.
Minimizing bleeding during transnasal resections of sinonasal tumors is imperative for optimizing visualization and decreasing complications. The purpose of the present study was to determine whether radiofrequency coblation decreases blood loss during endoscopic tumor removal. Sinonasal/skull base tumors treated in 2008 with endoscopic techniques were reviewed. The data collected included demographics, histopathology, technique, duration, complications and estimated blood loss (EBL). Full operative videoendoscopy was available in all cases and scored by the authors using the 11-point Wormald surgical field grading scale. Twenty-three patients (average age: 46 years) with sinonasal or skull base tumors treated with transnasal endoscopic techniques were identified. Coblation was used in 10 cases. The sinus/skull base tumors included were esthesioneuroblastoma (n = 6), melanoma (n = 3), squamous cell carcinoma (n = 3), inverted papilloma (n = 3), adenocarcinoma (n = 2), intracranial dermoid cyst (n = 2), adenoid cystic carcinoma (n = 1), craniopharyngioma (n = 1), fibromyxosarcoma (n = 1) and undifferentiated carcinoma (n = 1). The use of the coblation device was associated with a significant decrease in all categories including EBL (350 vs. 1,000 ml; p = 0.0001), EBL per operative time (66 vs. 166 ml/h; p = 0.0001) and Wormald grade (3.3 vs. 6.4; p = 0.0001). Radiofrequency coblation significantly decreased blood loss during endoscopic tumor removal and is a useful tool in the armamentarium of the endoscopic skull base surgeon.


[Quality of life in women with pelvic floor dysfunction].
Pelvic floor dysfunction is a frequent problem affecting more than 50% of women in peri- and postmenopause. Considering that ageing and menopause befall in the significant factors causing this issue, as well as the expected longevity of women in the world and in our country, pelvic floor dysfunction prevelence is foreseen to be even higher. The aim of the study was to evaluate impact of the symptoms of pelvic dysfunction on quality of life and examine body image satisfaction in adult women with pelvic organ prolapse presenting to tertiary care clinic for surgical treatment. This prospective case-control study included 50 patients who presented to tertiary care gynecology clinic for surgical treatment and 50 controls with normal pelvic floor support and without urinary incontinence who presented tertiary care gynecology clinic for other reasons. Both, patients and controls, completed two quastionnaires recommended for the evaluation of symptoms (Pelvic floor distress inventory - short forms) and quality of life impact (Pelvic floor impact questionnaire - short form) of pelvic organ prolapse, and Body Image Scale. The patients scored significantly worse on the prolapse, urinary, colorectal scales and overall score of Pelvic floor distress inventory--20 than controls subjects (134.91 vs 78.08; p < 0.01). The patients also measured significant decrease in condition-specific quality of life (89.23 vs 3.1; p < 0.01). They were more likely to feel self-conscious (78% vs 42%; p < 0.01), less likely to feel physically attractive (78% vs 22%; p < 0.01), more likely to have difficulty looking at themselves naked (70% vs 42%; p < 0.01), less likely to feel sexually attractive (64% vs 32%; p < 0.01), and less likely to feel feminine (56% vs 16%; p < 0.05), than controls. There were no differencies in their feeling of dissatisfaction with appearance when dressed, avoiding people because of appereance and overall dissatisfaction with their body. There was a positive correlation between decreased quality of life and body image in women with pelvic dysfunction. Women with pelvic floor dysfunction have decreased quality of life and body image.


Commitment of cell fate in the early zebrafish embryo.
When do single cells in the early zebrafish embryo become irreversibly committed to a specific fate? Work with lineage tracing and fate mapping has shown that the marginal cells of the blastoderm give rise to hypoblast-derived fates (mesoderm and endoderm). However, experiments described here show that these marginal blastoderm cells remain pluripotent and uncommitted throughout the late blastula and early gastrula stages. Embryonic cells become committed to a hypoblast-derived fate at mid-gastrulation. Time-lapse photographic analysis reveals that committed cells, when transplanted heterotopically and heterochronically, can migrate along atypical pathways to reposition themselves within a more correct environment.


Social representation of palliative care in the Spanish printed media: A qualitative analysis.
Lack of social awareness is a major barrier to the development of palliative care. Mass media influences public opinion, and frequently deal with palliative care contributing to its image and public understanding. To analyse how palliative care is portrayed in Spanish newspapers, as well as the contribution made by the press to its social representation. Based on criteria of scope and editorial plurality, four print newspapers were selected. Using the newspaper archive MyNews (www.mynews.es), articles published between 2009 and 2014 containing the words "palliative care" or "palliative medicine" were identified. Sociological discourse analysis was performed on the identified texts on two levels: a) contextual analysis, focusing on the message as a statement; b) interpretative analysis, considering the discourse as a social product. We examined 262 articles. Politician and healthcare professionals were the main representatives transmitting messages on palliative care. The discourses identified were characterised by: strong ideological and moral content focusing on social debate, strong ties linking palliative care and death and, to a lesser degree, as a healthcare service. The messages transmitted by representatives with direct experience in palliative care (professionals, patients and families) contributed the most to building a positive image of this healthcare practice. Overall, media reflect different interests in framing public understanding about palliative care. The knowledge generated about how palliative care is reflected in the printed media may help to understand better one of the main barriers to its development not only in Spain, but also in other contexts.


Total and added sugars consumption in Argentina: Their contribution to daily energy intake. Results from Latin American Study of Nutrition and Health (ELANS).
The aim of the present study is to determine the intake of total sugars (TS) and added sugars (AS) in Argentina based on the local data of the Latin American Study of Nutrition and Health (ELANS). This is a cross-sectional study of a representative sample of the urban Argentine population (n = 1266). The sample was stratified by age group (15-65 years), gender, geographic region and socioeconomic level (SEL). TS and AS intake were obtained by two 24-hour recalls (R24) and analysed using the Nutrition Data System for Research Software 2013. On average, TS consumption in Argentina was 114.3 g/day, accounting for 39.8% of the total carbohydrate intake and 20.6% total energy (TE) intake. Overall, 77.2% of the TS intake consisted of AS (90.4 g/day), contributing to 30.4% of total carbohydrate intake and 15.9%TE. Men consume more TS and AS (in g/day), with no difference in the AS %TE between men and women. The consumption of sugars decreased with age, with adolescents consuming more AS and older adults more intrinsic sugars. The intake of AS was higher in low SEL. In Argentina, the intake of AS was 50% above the recommendations. Younger and socially vulnerable people are at higher risk of excessive intake.


Mass-Spectrometry-Based Identification of Cross-Links in Proteins Exposed to Photo-Oxidation and Peroxyl Radicals Using 18O Labeling and Optimized Tandem Mass Spectrometry Fragmentation.
Protein cross-links are formed in regulated biochemical processes in many biological systems, but they are also generated inadvertently via the reactions of exogenous or endogenous oxidants. Site-specific identification and characterization of such cross-links is challenging, and the goal was, therefore, to develop mass-spectrometry-based approaches tailored for proteins subjected to oxidative challenges that also are applicable for the analysis of complex samples. Using trypsin-mediated 18O isotopic labeling, different types of data acquisition workflows, and designated database software tools, we successfully identified tyrosine-tyrosine, tyrosine-tryptophan, tyrosine-lysine, and histidine-lysine cross-links in proteins subjected to sensitizer-mediated photo-oxidation with rose bengal or chemical oxidation with peroxyl radicals generated from the water-soluble compound 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH). Subsequently, AAPH was also applied to a protein extract from the Gram-positive bacterium Lactococcus lactis, demonstrating the feasibility to identify tyrosine-tyrosine, tyrosine-tryptophan, and tryptophan-tryptophan cross-linked peptides in a complex system. Different fragmentation techniques were evaluated, and it was observed that higher-energy collisional dissociation (HCD) resulted in a higher number of identified cross-link peptides, while electron-transfer dissociation supplemented with HCD (EThcD) generally provides higher fragment ion coverage of the cross-linked peptides.


Long term effects of olfactory training in patients with post-infectious olfactory loss.
There is evidence of the effectiveness of repeated exposure to odours on short-term olfactory function. The aim of this study was to assess the long-term effects of olfactory training. We conducted a prospective study of 111 patients with post-infectious olfactory dysfunction. Two groups of patients performed olfactory training for 16 and 56 weeks, respectively, and were compared with a control group. The training was performed twice daily using four odours (phenyl ethyl alcohol, eucalyptol, citronellal, eugenol). Olfactory testing was performed by means of the Sniffin Sticks test as a baseline assessment and then every 8 weeks for 56 weeks. Subjective ratings were performed using a visual analogue scale (0-100). Both training groups presented significantly higher scores than the controls. The long-term group had better results than the short-term group. Short-term training patients sustained their improvement within the follow-up period. Subsets analysis showed that training patients mainly increased identification and discrimination. Subjective ratings were in accordance with the olfactory test results. Long-term olfactory training seems to be associated with better results in patients with post-infectious olfactory loss than a short-term scheme. Short-term training provides sustainable results at 56 weeks follow-up assessment.


Prolonged activation of mitogen-activated protein kinases during NSAID-induced apoptosis in HT-29 colon cancer cells.
The mechanisms of the antineoplastic effect of nonsteroidal anti-inflammatory drugs (NSAIDs) still are unknown, but the induction of apoptosis is one of the possible mechanisms. We attempted to demonstrate the role of mitogen-activated protein (MAP) kinases, generally considered to be important mediators of proliferative and apoptotic signals, in NSAID-induced colon cancer cell apoptosis. Apoptosis was detected by demonstration of DNA fragmentation in agarose gel electrophoresis. Cell death was assessed by trypan blue dye exclusion method. MAP kinase activation was assessed by Western blot using phosphospecific antibodies to MAP kinases. Kinase assay using activating transcription factor-2 (ATF-2) fusion protein as a substrate was also performed for measuring p38 MAP kinase activity. For the inhibition of p38 MAP kinase, pyridinylimidazole compound (SB203580) was utilized. Caspase-3 activity was measured using the tetrapeptide fluorogenic substrate Ac-DEVD-AMC. Treatment of HT-29 cells with NSAIDs results in time- and dose-dependent induction of apoptosis, accompanied by sustained activation of all three MAP kinase subfamilies. The SB203580, a p38 MAP kinase inhibitor, reduced indomethacin-induced cell death by 43%, while PD098059, a MAPK/ERK kinase (MEK)1 inhibitor, did not affect cell death. p38 MAP kinase and caspase-3 activation were not significantly interlinked in indomethacin-induced apoptosis. From these results, we conclude that NSAIDs can induce prolonged activation of MAP kinases in colon cancer cells and that, of these, p38 MAP kinase may play a partial but significant role in indomethacin-induced apoptosis.


ISG15 modification of ubiquitin E2 Ubc13 disrupts its ability to form thioester bond with ubiquitin.
ISG15 was the first ubiquitin-like modifier to be identified. However, the function of ISG15 modification has been an enigma for many years. At present, no data are available about the function of ISGylation for any target. In this paper, we report the identification of Ubc13, which forms a unique ubiquitin-conjugating enzyme (Ubc) complex with ubiquitin enzyme variant Mms2 and generates atypical Lys63-linked ubiquitin conjugates, as one of the targets of ISG15 modification. Furthermore, we identify Lys92 as the only ISG15 modification site in Ubc13, which is the first report about the ISG15 modification site. Using the "covalent affinity" purification assay, we found that unmodified Ubc13 can bind to the ubiquitin-agarose, whereas ISGylated Ubc13 cannot. This result indicates that ISGylation of Ubc13 disrupts its ability to form thioester bond with ubiquitin.


The biological effectiveness of targeted radionuclide therapy based on a whole-body pharmacokinetic model.
Biologically effective dose (BED) may be more of a relevant quantity than absorbed dose for establishing tumour response relationships. By taking into account the dose rate and tissue-specific parameters such as repair and radiosensitivity, it is possible to compare the relative biological effects of different targeted radionuclide therapy (TRT) agents. The aim of this work was to develop an analytical tumour BED calculation for TRT that could predict a relative biological effect based on normal body and tumour pharmacokinetics. This work represents a step in the direction of establishing relative pharmacokinetic criteria of when the BED formalism is more applicable than absorbed dose for TRT. A previously established pharmacokinetic (PK) model for TRT was used and adapted into the BED formalism. An analytical equation for the protraction factor, which incorporates dose rate and repair rate, was derived. Dose rates within the normal body and tumour were related to the slopes of their time-activity curves which were determined by the ratios of their respective PK parameters. The relationships between the tumour influx-to-efflux ratio (k(34):k(43)), central compartment efflux-to-influx ratio (k(12):k(21)), central elimination (k(el)), and tumour repair rate (μ), and tumour BED were investigated. As the k(34):k(43) ratio increases and the k(12):k(21) ratio decreases, the difference between tumour BED and D increases. In contrast, as the k(34):k(43) ratios decrease and the k(12):k(21) ratios increase, the tumour BED approaches D. At large k(34):k(43) ratios, the difference between tumour BED and D increases to a maximum as k(el) increases. At small k(34):k(43) ratios, the tumour BED approaches D at very small k(el). At small μ and small k(34):k(43) ratios, the tumour BED approaches D. For large k(34):k(43) ratios, large μ values cause tumour BED to approach D. This work represents a step in the direction of establishing relative PK criteria of when the BED formalism is more applicable than absorbed dose for TRT. It also provides a framework by which the biological effects of different TRT agents can be compared in order to predict efficacy.


Ca2+ and segment length dependence of isometric force kinetics in intact ferret cardiac muscle.
The influence of Ca2+ and sarcomere length on myocardial crossbridge kinetics was studied in ferret papillary muscle by measuring the rate of force redevelopment following a rapid length step that dropped the force to zero. Tetanic stimulation with 5 mumol/L ryanodine was used to obtain a steady-state contraction, and segment length was measured and controlled using a sense-coil technique that measures changes in the cross-sectional area of the central region of the muscle. The rate constant for the recovery of force (ktr) following a rapid length release was obtained by fitting the data with a single exponential function. Contrary to results from skinned skeletal fibers in which ktr increases almost 10-fold from low to maximal activation levels, ktr was found not to increase at higher activation levels in this study. Similarly, although force increased with segment length under all conditions, ktr never increased with length. Data presented here are consistent with a model of myocardial Ca2+ activation in which Ca2+ modulates the number of crossbridges interacting with the thin filament and are inconsistent with a model in which Ca2+ modulates the kinetics of transitions to force producing states within the actomyosin cycle. Differences in the activation dependence of the force redevelopment rate between cardiac and skeletal muscle suggest that there are fundamental differences in the mechanism of Ca2+ activation between these two muscle types.


Modalities of fatigue in multiple sclerosis: correlation with clinical and biological factors.
Although different factors are probably involved in the etiology of fatigue in multiple sclerosis patients, no definite mechanism has been proposed. We have proposed that fatigue is a complex symptom that includes three clinical different entities (asthenia, fatigability and worsening of symptoms with effort). The goal of this study is to demonstrate if there is a peculiar mechanism for each of the different varieties of fatigue. A control sample of 155 patients (105 women, 50 men) with clinically definite MS was studied. Fatigue was measured using the Fatigue Descriptive Scale (FDS) and the Fatigue Severity Scale (FSS). Treatment, depression, anxiety, sleep and cellular immune status were studied too. Fatigue was a symptom in 118 patients (76.13%); 26 patients (22.03%) described it as asthenia (fatigue at rest); 85 patients (72.03%) as fatigability (fatigue with exercise), and seven patients (5.9%) as worsening of symptoms. The severity of pyramidal involvement was significantly more severe in patients suffering from fatigue; some immunological parameters were associated with fatigue as well. The discriminant analysis of the data shows that some of the immunoactivation parameters are associated with asthenia (F=21.5, P<0.001), and pyramidal tract involvement is associated with fatigability (F=10.5, P<0.001). Sleep disorders, anxiety and depression were linked with fatigue in a few patients. No relationship with treatment was proven. In conclusion, fatigue in MS seems to be a heterogeneous entity. Asthenia and fatigability may be different clinical entities. Certain immunoactivation parameters correlate with the presence of asthenia while pyramidal involvement is associated with fatigability.


De novo point mutations in patients diagnosed with ataxic cerebral palsy.
Cerebral palsy is a sporadic disorder with multiple likely aetiologies, but frequently considered to be caused by birth asphyxia. Genetic investigations are rarely performed in patients with cerebral palsy and there is little proven evidence of genetic causes. As part of a large project investigating children with ataxia, we identified four patients in our cohort with a diagnosis of ataxic cerebral palsy. They were investigated using either targeted next generation sequencing or trio-based exome sequencing and were found to have mutations in three different genes, KCNC3, ITPR1 and SPTBN2. All the mutations were de novo and associated with increased paternal age. The mutations were shown to be pathogenic using a combination of bioinformatics analysis and in vitro model systems. This work is the first to report that the ataxic subtype of cerebral palsy can be caused by de novo dominant point mutations, which explains the sporadic nature of these cases. We conclude that at least some subtypes of cerebral palsy may be caused by de novo genetic mutations and patients with a clinical diagnosis of cerebral palsy should be genetically investigated before causation is ascribed to perinatal asphyxia or other aetiologies.


The effect of tapered toothbrush filaments compared to end-rounded filaments on dental plaque, gingivitis and gingival abrasion: a systematic review and meta-analysis.
This systematic review was performed to establish the effect of a manual toothbrush with tapered toothbrush filaments (TFTBs) compared to a manual toothbrush with end-rounded toothbrush filaments (ERTB) on clinical parameters of dental plaque, gingivitis and gingival abrasion. MEDLINE-PubMed and Cochrane-CENTRAL databases were searched. The inclusion criteria were (randomized) controlled clinical trials, participants ≥18 years and papers evaluating the effect of a TFTB compared to an ERTB. Data were extracted for dental plaque index (PI), bleeding scores (BS), gingival index scores (GI) and gingival abrasion scores (GA). A descriptive analysis and a meta-analysis were performed when appropriate. An independent screening of 33 unique papers resulted in seven eligible publications, which included eight comparisons. Meta-analysis did not show a significant difference between TFTB and ERTB with respect to PI scores. The meta-analysis of the GI scores showed a significant mean difference in favour of the TFTB (DiffM=-0.12 [95% CI: -0.17; -0.07]). Of the three comparisons evaluating GA, no differences were found. With respect to plaque removal, evidence that supports the recommendation for usage of a TFTB over an ERTB is lacking. Regarding GI, there is minimal evidence favouring a TFTB over an ERTB and the clinical relevance of this difference is probably negligible. Therefore, based on the collective evidence emerging from this systematic review, the strength and direction of the recommendation, there appears to be no firm evidence for a dental healthcare professional to advise the use of a TFTB over the use of an ERTB.


Using relative distributions to investigate the body mass index in England and Canada.
In this paper we use relative distributions to examine changes in the distribution of the body mass index (BMI) in England and Canada during the period 1994/5-2000/1. The use of relative distributions allows us to describe changes in the whole distribution of the BMI in a non-parametric fashion. While statistics analogous to the Gini index can be constructed based on the relative distribution, important characteristics of changes in the distribution of the BMI such as changes in the proportions overweight and obese are more naturally handled using measures of relative polarization. Our results show that while BMI has increased in both countries, BMI in England has increased at a much faster rate than in Canada. Both groups show polarization over time towards both tails of the weight distribution, with the English polarizing towards the upper tail at a faster rate than Canadians.


Ozone-initiated particle formation, particle aging, and precursors in a laser printer.
An increasing number of researchers have hypothesized that ozone may be involved in the particle formation processes that occur during printing, however no studies have investigated this further. In the current study, this hypothesis was tested in a chamber study by adding supplemental ozone to the chamber after a print job without measurable ozone emissions. Subsequent particle number concentration and size distribution measurements showed that new particles were formed minutes after the addition of ozone. The results demonstrated that ozone did react with printer-generated volatile organic compounds (VOCs) to form secondary organic aerosols (SOAs). The hypothesis was further confirmed by the observation of correlations among VOCs, ozone, and particles concentrations during a print job with measurable ozone emissions. The potential particle precursors were identified by a number of furnace tests, which suggested that squalene and styrene were the most likely SOA precursors with respect to ozone. Overall, this study significantly improved scientific understanding of the formation mechanisms of printer-generated particles, and highlighted the possible SOA formation potential of unsaturated nonterpene organic compounds by ozone-initiated reactions in the indoor environment.


Enhancement of anti-DIII antibodies by the C3d derivative P28 results in lower viral titers and augments protection in mice.
Antibodies generated against West Nile virus (WNV) during infection are essential for controlling dissemination. Recent studies have demonstrated that epitopes in all three domains of the flavivirus envelope protein (E) are targets for neutralizing antibodies, with determinants in domain III (DIII) eliciting antibodies with strong inhibitory properties. In order to increase the magnitude and quality of the antibody response against the WNV E protein, DNA vaccines with derivatives of the WNV E gene (full length E, truncated E, or DIII region, some in the context of the pre-membrane [prM] gene) were conjugated to the molecular adjuvant P28. The P28 region of the complement protein C3d is the minimum CR2-binding domain necessary for the adjuvant activity of C3d. Delivery of DNA-based vaccines by gene gun and intramuscular routes stimulated production of IgG antibodies against the WNV DIII region of the E protein. With the exception of the vaccine expressing prM/E given intramuscularly, only mice that received DNA vaccines by gene gun produced protective neutralizing antibody titers (FRNT80 titer >1/40). Correspondingly, mice vaccinated by the gene gun route were protected to a greater level from lethal WNV challenge. In general, mice vaccinated with P28-adjuvated vaccines produced higher IgG titers than mice vaccinated with non-adjuvanted vaccines.


Multivesicular endosomes containing internalized EGF-EGF receptor complexes mature and then fuse directly with lysosomes.
We have followed the transfer of EGF-EGF receptor (EGFR) complexes from endosomal vacuoles that contain transferrin receptors (TfR) to lysosome vacuoles identified by their content of HRP loaded as a 15-min pulse 4 h previously. We show that the HRP-loaded lysosomes are lysosomal-associated membrane protein-1 (LAMP-1) positive, mannose-6-phosphate receptor (M6PR) negative. and contain active acid hydrolase. EGF-EGFR complexes are delivered to these lysosomes intact and are then rapidly degraded. Preactivating the HRP contained within the preloaded lysosomes inhibits the delivery of EGFR and degradation of EGF, and results in the accumulation of EGFR-containing multivesicular bodies (MVB). With time these accumulating MVB undergo a series of maturation changes that include the loss of TfR, the continued recruitment of EGFR, and the accumulation of internal vesicles, but they remain LAMP-1 and M6PR negative. The mature MVB are often seen to make direct contact with lysosomes containing preactivated HRP, but their perimeter membranes remain intact. Together our observations suggest that the transfer of EGF-EGFR complexes from the TfR-containing endosome compartment to the lysosomes that degrade them employs a single vacuolar intermediate, the maturing MVB, and can be achieved by a single heterotypic fusion step.


Phosphoinositide hydrolysis in mitogen-stimulated human peripheral-blood T lymphocytes.
Both phytohaemagglutinin and antibodies to the CD3 molecule induced proliferation and phosphoinositide hydrolysis in human peripheral-blood T lymphocytes, but the magnitude of the inositol phosphate response was small and the rate of accumulation slow [significant increases in Ins(1,4,5)P3 were observed only after 10 min]. Hence this response differs from the well-characterized Ins(1,4,5)P3 responses of many other systems. This slow response, its abrogation in Ca2+-depleted medium, the slow and maintained increase in Ca2+ as measured by Quin-2, and the ability of the Ca2+ ionophore A23187 to stimulate Ins(1,4,5)P3 accumulation all suggest that the increase in Ins(1,4,5)P3 occurs, at least in part, as a result of receptor-mediated Ca2+ influx in mitogen-stimulated T lymphocytes.


Effect of amyloid on memory and non-memory decline from preclinical to clinical Alzheimer's disease.
High amyloid has been associated with substantial episodic memory decline over 18 and 36 months in healthy older adults and individuals with mild cognitive impairment. However, the nature and magnitude of amyloid-related memory and non-memory change from the preclinical to the clinical stages of Alzheimer's disease has not been evaluated over the same time interval. Healthy older adults (n = 320), individuals with mild cognitive impairment (n = 57) and individuals with Alzheimer's disease (n = 36) enrolled in the Australian Imaging, Biomarkers and Lifestyle study underwent at least one positron emission tomography neuroimaging scan for amyloid. Cognitive assessments were conducted at baseline, and 18- and 36-month follow-up assessments. Compared with amyloid-negative healthy older adults, amyloid-positive healthy older adults, and amyloid-positive individuals with mild cognitive impairment and Alzheimer's disease showed moderate and equivalent decline in verbal and visual episodic memory over 36 months (d's = 0.47-0.51). Relative to amyloid-negative healthy older adults, amyloid-positive healthy older adults showed no decline in non-memory functions, but amyloid-positive individuals with mild cognitive impairment showed additional moderate decline in language, attention and visuospatial function (d's = 0.47-1.12), and amyloid-positive individuals with Alzheimer's disease showed large decline in all aspects of memory and non-memory function (d's = 0.73-2.28). Amyloid negative individuals with mild cognitive impairment did not show any cognitive decline over 36 months. When non-demented individuals (i.e. healthy older adults and adults with mild cognitive impairment) were further dichotomized, high amyloid-positive non-demented individuals showed a greater rate of decline in episodic memory and language when compared with low amyloid positive non-demented individuals. Memory decline does not plateau with increasing disease severity, and decline in non-memory functions increases in amyloid-positive individuals with mild cognitive impairment and Alzheimer's disease. The combined detection of amyloid positivity and objectively-defined decline in memory are reliable indicators of early Alzheimer's disease, and the detection of decline in non-memory functions in amyloid-positive individuals with mild cognitive impairment may assist in determining the level of disease severity in these individuals. Further, these results suggest that grouping amyloid data into at least two categories of abnormality may be useful in determining the disease risk level in non-demented individuals.


Comparative analysis of loop-mediated isothermal amplification (LAMP)-based assays for rapid detection of SARS-CoV-2 genes.
The COVID-19 pandemic caused by SARS-CoV-2 has infected millions worldwide, therefore there is an urgent need to increase our diagnostic capacity to identify infected cases. Although RT-qPCR remains the gold standard for SARS-CoV-2 detection, this method requires specialised equipment in a diagnostic laboratory and has a long turn-around time to process the samples. To address this, several groups have recently reported the development of loop-mediated isothermal amplification (LAMP) as a simple, low cost and rapid method for SARS-CoV-2 detection. Herein we present a comparative analysis of three LAMP-based assays that target different regions of the SARS-CoV-2: ORF1ab RdRP, ORF1ab nsp3 and Gene N. We perform a detailed assessment of their sensitivity, kinetics and false positive rates for SARS-CoV-2 diagnostics in LAMP or RT-LAMP reactions, using colorimetric or fluorescent detection. Our results independently validate that all three assays can detect SARS-CoV-2 in 30 min, with robust accuracy at detecting as little as 1000 RNA copies and the results can be visualised simply by color changes. Incorporation of RT-LAMP with fluorescent detection further increases the detection sensitivity to as little as 100 RNA copies. We also note the shortcomings of some LAMP-based assays, including variable results with shorter reaction time or lower load of SARS-CoV-2, and false positive results in some experimental conditions and clinical saliva samples. Overall for RT-LAMP detection, the ORF1ab RdRP and ORF1ab nsp3 assays have faster kinetics for detection but varying degrees of false positives detection, whereas the Gene N assay exhibits no false positives in 30 min reaction time, which highlights the importance of optimal primer design to minimise false-positives in RT-LAMP. This study provides validation of the performance of LAMP-based assays as a rapid, highly sensitive detection method for SARS-CoV-2, which have important implications in development of point-of-care diagnostics for SARS-CoV-2.


Blocking adhesion to cell and tissue surfaces by the chemisorption of a poly-L-lysine-graft-(poly(ethylene glycol); phenylboronic acid) copolymer.
A family of graft copolymers that can sterically inhibit interactions between biological surfaces was developed. These copolymers contained phenylboronic acid (PBA) groups as saccharide-binding moieties on a poly-(L-lysine) backbone and poly(ethylene glycol) (PEG) grafted as adhesion-resisting side chains. These copolymers spontaneously chemisorbed to a saccharide-containing resin, and this binding was sterically controlled by the PEG grafting ratio. Copolymers with optimal grafting ratios spontaneously assembled on red blood cell surfaces and sterically prevented their agglutination by lectins and by antibodies to blood groups. The simple conjugation scheme created a PBA moiety with a pKa ca. 6, which can bind cis-diols much more strongly at physiological pH than typical PBA moieties, whose pKas are typically greater than 8. These surfactant copolymers can be employed to PEGylate cell or tissue surfaces by simply incubating the surfaces with an aqueous polymer solution, and have many potential applications such as preventing antibody binding to transplanted cells.


Scrotal construction by expansion of labia majora in biological female transsexuals.
Construction of male genitalia in biological female transsexual patients is problematic. We report 3 transsexuals with scrotal construction by tissue expansion of the labia majora. In each patient, tissue expanders were inserted into the labia majora several months before free flap phalloplasty. In 2 patients, the tissue expanders were removed, and testicular implants were placed into the labial pockets. In the other patient, the tissue expanders were left in place. Because of the close embryological relationship between the labia majora and the scrotum, the neoscrotum appears natural in color match, hair pattern, and skin texture. The anatomical position of the labia majora produces a scrotum properly located in relation to surrounding anatomical structures. Excellent aesthetic results can be achieved with this procedure. Based on our experience, we advocate this procedure for scrotal construction in biological female transsexual patients.


Transducin-like enhancer of split proteins, the human homologs of Drosophila groucho, interact with hepatic nuclear factor 3beta.
Members of the hepatic nuclear factor 3 (HNF3) family, including HNF3alpha, HNF3beta, and HNF3gamma, play important roles in embryonic development, the establishment of tissue-specific gene expression, and the regulation of gene expression in differentiated tissues. We found, using the glutathione S-transferase pull-down method, that the transducin-like Enhancer of split (TLE) proteins, which are the human homologs of Drosophila Groucho, directly associate with HNF3beta. Conserved region II of HNF3beta (amino acids 361-388) is responsible for the interaction with TLE1. A mammalian two-hybrid assay was used to confirm that this interaction occurs in vivo. Overexpression of TLE1 in HepG2 and HeLa cells decreases transactivation mediated through the C-terminal domain of HNF3beta, and Grg5, a naturally occurring dominant negative form of Groucho/TLE, also increases the transcriptional activity of this region of HNF3. These results lead us to suggest that TLE proteins could influence the expression of mammalian genes regulated by HNF3.


Synchronous magnetic control of water droplets in bulk ferrofluid.
We present a microfluidic platform for magnetic manipulation of water droplets immersed in bulk oil-based ferrofluid. Although non-magnetic, the droplets are exclusively controlled by magnetic fields without any pressure-driven flow. The fluids are dispensed in a sub-millimeter Hele-Shaw chamber that includes permalloy tracks on its substrate. An in-plane rotating magnetic field magnetizes the permalloy tracks, producing local magnetic gradients, while an orthogonal magnetic field magnetizes the bulk ferrofluid. To minimize the magnetostatic energy of the system, the water droplets are attracted towards the locations on the tracks where the bulk ferrofluid is repelled. Using this technique, we demonstrate synchronous generation and propagation of water droplets, study the kinematics of propagation, and analyze the flow of the bulk ferrofluid. In addition, we show controlled break-up of droplets and droplet-to-droplet interactions. Finally, we discuss future applications owing to the potential biocompatibility of the droplets.


The MDM2 RING-finger domain is required to promote p53 nuclear export.
MDM2 can bind to p53 and promote its ubiquitination and subsequent degradation by the proteasome. Current models propose that nuclear export of p53 is required for MDM2-mediated degradation, although the function of MDM2 in p53 nuclear export has not been clarified. Here we show that MDM2 can promote the nuclear export of p53 in transiently transfected cells. This activity requires the nuclear-export signal (NES) of p53, but not the NES of MDM2. A mutation within the MDM2 RING-finger domain that inhibits p53 ubiquitination also inhibits the ability of MDM2 to promote p53 nuclear export. Finally, inhibition of nuclear export stabilizes wild-type p53 and leads to accumulation of ubiquitinated p53 in the nucleus. Our results indicate that MDM2-mediated ubiquitination, or other activities associated with the RING-finger domain, can stimulate the export of p53 to the cytoplasm through the activity of the p53 NES.


Clinical significance of histological classification of idiopathic interstitial pneumonia.
Patients with idiopathic interstitial pneumonias (IIPs) can be subdivided into groups based on the histological appearance of lung tissue obtained by surgical biopsy. The quantitative impact of histological diagnosis, baseline factors and response to therapy on survival has not been evaluated. Surgical lung biopsy specimens from 168 patients with suspected IIP were reviewed according to the latest diagnostic criteria. The impact of baseline clinical, physiological, radiographic and histological features on survival was evaluated using Cox regression analysis. The predictive value of honeycombing on high-resolution computed tomography (HRCT) as a surrogate marker for usual interstitial pneumonia (UIP) was examined. The response to therapy and survival of 39 patients treated prospectively with high-dose prednisone was evaluated. The presence of UIP was the most important factor influencing mortality. The risk ratio of mortality when UIP was present was 28.46 (95% confidence interval (CI) 5.5-148.0; p=0.0001) after controlling for patient age, duration of symptoms, radiographic appearance, pulmonary physiology, smoking history and sex. Honeycombing on HRCT indicated the presence of UIP with a sensitivity of 90% and specificity of 86%. Patients with nonspecific interstitial pneumonia were more likely to respond or remain stable (9 of 10) compared to patients with UIP (14 of 29) after treatment with prednisone. Patients remaining stable had the best prognosis. The risk ratio of mortality for stable patients compared to nonresponders was 0.32 (95% CI 0.11-0.93; p=0.04) in all patients and 0.33 (95% CI 0.12-0.96; p=0.04) in patients with UIP. The histological diagnosis of usual interstitial pneumonia is the most important factor determining survival in patients with suspected idiopathic interstitial pneumonia. The presence of honeycombing on high-resolution computed tomography is a good surrogate for usual interstitial pneumonia and could be utilized in patients unable to undergo surgical lung biopsy. Patients with nonspecific interstitial pneumonia are more likely to respond or remain stable following a course of prednisone. Patients remaining stable following prednisone therapy have the best prognosis.


Use of the PARIHS Framework for Retrospective and Prospective Implementation Evaluations.
The Promoting Action on Research Implementation in Health Services (PARIHS) framework has been used by implementation researchers to assess factors impacting implementation and to use that information to identify optimal interventions and implementation strategies. In this paper, two studies are presented demonstrating the utility of PARIHS as a tool for retrospective and prospective evaluation of implementation in the health care setting. Descriptive case study. A qualitative consensus process was used to evaluate provider perceptions of PARIHS constructs of evidence, context, and facilitation and their subelements which were scored on a continuum of low to high. The first example demonstrates retrospective use of PARIHS which provided insight into the factors contributing to variations in implementation across sites in an ongoing program. Evidence was strong (high), whereas context noted some challenges in culture and measurement (mixed), and the presence of dedicated program facilitators was positive but dual roles limited their ability to fully support implementation (mixed). The second example demonstrates prospective use of PARIHS for evaluation which gathered information about intervention sites for the purposes of selecting implementation strategies responsive to site needs. Evidence supporting the intervention was limited (low), context noted that limited awareness of the intervention was a challenge (low), and that a strong internal facilitator supported implementation (high). The descriptive case study presented here underscores the value of a theory-guided approach to implementation and highlights that PARIHS can help implementers understand factors impacting implementation, identify areas for future intervention, and inform selection of strategies to support or enhance implementation efforts.


Morphologic differences, according to etiology, in pigment epithelial detachments by means of en face optical coherence tomography.
To assess morphologic differences in pigment epithelial detachment (PED) with en face optical coherence tomography in central serous chorioretinopathy (CSC) and age-related macular degeneration (AMD). We recruited 30 eyes of 22 patients with PED. Nine eyes had a clinical diagnosis of CSC and 21 had AMD. All patients were assessed with en face optical coherence tomography. Morphologic PED aspects were estimated on C-scans and classified according to shape, inner silhouette, content, wall aspects, wall thickness, and size. Pigment epithelial detachment shape was predominantly circular (88.8%) in CSC and irregular or with multilobular features in AMD (76.2%). The PED inner silhouette had a smooth aspect (88.9%) in CSC and a slightly granular aspect or granular profile in AMD (100%). Clear PED content was the most characteristic feature of CSC (88.9%) but not of AMD. In CSC, PED morphologic wall aspect was uniform or slightly irregular (100%), while in AMD, it was slightly irregular (52.4%) or irregular (47.6%). Pigment epithelial detachment wall thickness and dimensions were larger in AMD than in CSC. Statistically significant differences were observed between CSC and AMD concerning PED inner silhouette, contents, wall aspects, and wall thickness measurements. En face optical coherence tomography scanning is a valuable tool for showing important morphologic differences between CSC and AMD.


Standard and higher dose of olanzapine in patients with schizophrenia or schizoaffective disorder: a randomized, double-blind, fixed-dose study.
The objective of this study was to assess the dose-response relationship of standard and higher doses of olanzapine in a randomized, double-blind, 8-week, fixed-dose study comparing olanzapine 10 (n = 199), 20 (n = 200), and 40 mg/d (n = 200) for patients with schizophrenia or schizoaffective disorder and suboptimal response to current treatment. Patients meeting criteria for antipsychotic treatment resistance were excluded. Dose-response relationship was assessed by linear regression analysis with log-transformed dose (independent variable) and Positive and Negative Syndrome Scale (PANSS) total score (dependent variable). There were no significant dose group differences in patients completing the study (overall, 67.8%). All dose groups showed statistically significant improvement in PANSS total scores from baseline to end point without significant dose-response relationship (P = 0.295). Post hoc analysis of response showed significant interaction between baseline PANSS and dose (P = 0.023), indicating better response at higher doses for patients with higher baseline PANSS. There was a significant dose response for mean change in weight (P = 0.003) with significant difference between the 10- and 40-mg-dose groups (P = 0.002; 1.9 [10 mg/d], 2.3 [20 mg/d], and 3.0 kg [40 mg/d]). There was a significant dose response for change in prolactin (P < 0.001) with a significant difference between each group (-10.5 [10 mg/d], -1.7 [20 mg/d], and 4.9 ng/mL [40 mg/d]; P < or = 0.018). Over 8 weeks, non-treatment-resistant patients with schizophrenia or schizoaffective disorder responded to all 3 doses of olanzapine, without a statistically significant dose-response relationship, suggesting that for many patients with schizophrenia or schizoaffective disorder, particularly those who are mildly or moderately ill, 10 mg/d should be the initial dose of choice.


Inter-Professionalism in Health Care Post-graduate specialization: an innovative Laboratory.
Inter- professional Collaboration (IPC) is an important component of a well-functioning healthcare system. It is linked to improvements in patient safety and case management, optimal use of the skills of each healthcare team member and provision of better health services. Inter- professional Education (IPE), is one key factor in the development of positive behaviors useful for IPC: the basic and post-basic training are key moments to raise awareness, train and help implement the IPC. Aim of this paper is to present and evaluate the use of an innovative laboratory of Consensus Conference implemented in the Nursing Post-graduate specialization at the University of Parma to train students to IPC. An Innovative Laboratory inspired by of the Consensus Conference (CC) methodology on the "Integrated Narrative Nursing Assessment" was designed. Three Post-graduate specialization courses were involved and assigned to different tasks in the CC, according to the characteristics of the specializations. Strengths and weaknesses of the methodology were analyzed. Strengths: students' engagement in their competencies building, and the acquisition inter-professional collaboration skills. Weaknesses: the lack of time to develop the whole process, and the need of a deeper guidance in the scientific production. Although the methodology have to be continuously improved through practice, this experimental Laboratory reached the aim of offering a real experience of IPC to the students. They really collaborated with different professionals to reach a common goal and being already considered an expert.


Direct correlation between overproduction of guanosine 3',5'-bispyrophosphate (ppGpp) and penicillin tolerance in Escherichia coli.
The penicillin tolerance exhibited by amino acid-deprived Escherichia coli has been previously proposed to be a consequence of the stringent response. Evidence indicating that penicillin tolerance is directly attributable to guanosine 3',5'-bispyrophosphate (ppGpp) overproduction and not to some other effect of amino acid deprivation is now presented. Accumulation of ppGpp in the absence of amino acid deprivation was achieved by the controlled overexpression of the cloned relA gene, which encodes ppGpp synthetase I. The overproduction of ppGpp resulted in the inhibition of both peptidoglycan and phospholipid synthesis and in penicillin tolerance. The minimum concentration of ppGpp required to establish these phenomena was determined to be 870 pmol per mg (dry weight) of cells. This represented about 70% of the maximum level of ppGpp accumulated during the stringent response. Penicillin tolerance and the inhibition of peptidoglycan synthesis were both suppressed when ppGpp accumulation was prevented by treatment with chloramphenicol, an inhibitor of ppGpp synthetase I activation. Glycerol-3-phosphate acyltransferase, the product of plsB, was recently identified as the main site of ppGpp inhibition in phospholipid synthesis (R. J. Health, S. Jackowski, and C. O. Rock, J. Biol. Chem. 269:26584-26590, 1994). The overexpression of the cloned plsB gene reversed the penicillin tolerance conferred by ppGpp accumulation. This result supports previous observations indicating that the membrane-associated events in peptidoglycan metabolism were dependent on ongoing phospholipid synthesis. Interestingly, treatment with beta-lactam antibiotics by itself induced ppGpp accumulation, but the maximum levels attained were insufficient to confer penicillin tolerance.


Modulation of prion formation, aggregation, and toxicity by the actin cytoskeleton in yeast.
Self-perpetuating protein aggregates transmit prion diseases in mammals and heritable traits in yeast. De novo prion formation can be induced by transient overproduction of the corresponding prion-forming protein or its prion domain. Here, we demonstrate that the yeast prion protein Sup35 interacts with various proteins of the actin cortical cytoskeleton that are involved in endocytosis. Sup35-derived aggregates, generated in the process of prion induction, are associated with the components of the endocytic/vacuolar pathway. Mutational alterations of the cortical actin cytoskeleton decrease aggregation of overproduced Sup35 and de novo prion induction and increase prion-related toxicity in yeast. Deletion of the gene coding for the actin assembly protein Sla2 is lethal in cells containing the prion isoforms of both Sup35 and Rnq1 proteins simultaneously. Our data are consistent with a model in which cytoskeletal structures provide a scaffold for generation of large aggregates, resembling mammalian aggresomes. These aggregates promote prion formation. Moreover, it appears that the actin cytoskeleton also plays a certain role in counteracting the toxicity of the overproduced potentially aggregating proteins.


Brief mindfulness training reduces salivary IL-6 and TNF-α in young women with depressive symptomatology.
Pro-inflammatory cytokines have been implicated in the pathophysiology and maintenance of depression. This study investigated the effects of a brief mindfulness intervention on salivary pro-inflammatory correlates of depression (IL-6, TNF-α) and self-reported symptoms of depression in college women. Sixty-four females with a cut score of ≥16 on the Center for Epidemiological Studies for Depression Scale (CES-D) were assigned to a 4-week mindfulness-based intervention (MBI; N = 31) or a contact-control group (N = 33). For both groups, salivary cytokines and depressive symptoms were assessed at baseline and posttreatment. For the mindfulness group only, salivary cytokines were also assessed at a 3-month follow-up. Both groups showed similar reductions in depression. However, MBI (vs. control) predicted greater reductions in IL-6 and TNF-α; changes in IL-6 were sustained at 3-month follow-up. Higher baseline depressive symptoms predicted greater reductions in inflammation in the mindfulness group. MBIs may reduce inflammatory immune markers commonly implicated in depression. Individuals with greater depressive symptoms may benefit more from mindfulness training. Although reductions in salivary cytokines in the mindfulness condition were not attributable to changes in depressive symptoms, future work should examine the possibility that such reductions are protective against the development of future depressive episodes. (PsycINFO Database Record


Patient and family perceptions of hospice services: 'I knew they weren't like hospitals'.
The vision for palliative care service provision in New Zealand is for all people who are dying and their families to have timely access to culturally appropriate, quality palliative care services. An Auckland hospice's records show that the ethnically diverse population statistics were not reflected in the referrals for hospice services. The aim of this research was to gain a patient-and-their-family perspective on the hospice, including exploration of components of service care that could be improved for various cultural groups. Patients currently under the care of the hospice and family members were recruited from hospice records. Semi-structured interviews were conducted to explore the emerging issues. The study collected data from a purposive sample of 18 palliative care patients or carer family members, ranging in age from 39 to 81 years, who reflected the ethnic diversity of the population of the region. Interviewing was carried out by an experienced research assistant and continued until data saturation was reached. Four key themes emerged-hospice personnel's approach to patients, quality of service, cultural barriers, and strategies for future improvement. It was determined that the latter two were the most significant to address in this article. The study revealed the need for information-giving and education, including public profiling of the hospice to strengthen community involvement. Strategies to reduce ethnic disparities include strengthening the awareness of, and access to, services by connecting with cultural groups through churches, community and specific cultural media.


Genetic variants associated with warfarin dose in African-American individuals: a genome-wide association study.
VKORC1 and CYP2C9 are important contributors to warfarin dose variability, but explain less variability for individuals of African descent than for those of European or Asian descent. We aimed to identify additional variants contributing to warfarin dose requirements in African Americans. We did a genome-wide association study of discovery and replication cohorts. Samples from African-American adults (aged ≥18 years) who were taking a stable maintenance dose of warfarin were obtained at International Warfarin Pharmacogenetics Consortium (IWPC) sites and the University of Alabama at Birmingham (Birmingham, AL, USA). Patients enrolled at IWPC sites but who were not used for discovery made up the independent replication cohort. All participants were genotyped. We did a stepwise conditional analysis, conditioning first for VKORC1 -1639G→A, followed by the composite genotype of CYP2C9*2 and CYP2C9*3. We prespecified a genome-wide significance threshold of p<5×10(-8) in the discovery cohort and p<0·0038 in the replication cohort. The discovery cohort contained 533 participants and the replication cohort 432 participants. After the prespecified conditioning in the discovery cohort, we identified an association between a novel single nucleotide polymorphism in the CYP2C cluster on chromosome 10 (rs12777823) and warfarin dose requirement that reached genome-wide significance (p=1·51×10(-8)). This association was confirmed in the replication cohort (p=5·04×10(-5)); analysis of the two cohorts together produced a p value of 4·5×10(-12). Individuals heterozygous for the rs12777823 A allele need a dose reduction of 6·92 mg/week and those homozygous 9·34 mg/week. Regression analysis showed that the inclusion of rs12777823 significantly improves warfarin dose variability explained by the IWPC dosing algorithm (21% relative improvement). A novel CYP2C single nucleotide polymorphism exerts a clinically relevant effect on warfarin dose in African Americans, independent of CYP2C9*2 and CYP2C9*3. Incorporation of this variant into pharmacogenetic dosing algorithms could improve warfarin dose prediction in this population. National Institutes of Health, American Heart Association, Howard Hughes Medical Institute, Wisconsin Network for Health Research, and the Wellcome Trust.


Severe Isotype-Matched Immunosuppression (IMI) as a Potential Risk Factor for Progression of MGUS Patients.
Monoclonal gammopathy of undetermined significance (MGUS) precedes multiple myeloma in virtually every case. However, only a small percentage will progress and at very different rates. In addition, recent data have suggested that MGUS is associated with other comorbidities including infections, suggesting impaired immune function in some MGUS patients. Therefore, we aimed at assessing the value of isotype-matched immunosuppression (IMI; e.g., suppression of an IgAκ in an IgAλ patient), a type of immunosuppression more specific than classical immunoparesis (IP; e.g., IgG and/or IgM suppression in an IgA patient), as a prognostic marker for MGUS progression. The Hevylite assay was used to assess IMI and immunoglobulin ratios in 307 serum samples from a cohort of 248 MGUS patients. Follow-up clinical records were available for 154 individuals. A greater incidence of IMI (51%) over classical IP (37%) was observed, although both show a progressive increase with higher risk groups. Survival analysis of 154 patients showed that severe IMI (>50% suppression) differentiates 2 groups with significantly different time to progression (P = 0.024) while severe IP does not (P = 0.48). Also, a combination of severe IMI and involved monoclonal immunoglobulin >1.5g/dL by Hevylite (both variables found to be independent prognostic markers in multivariate analysis) identified a group of patients with a median time to progression 6-fold shorter than the remaining group (P < 0.0001). These findings indicate a possible role for IMI in the malignant transformation of MGUS patients and a potential utility as a new risk factor.


[Pulmonary Embolism in Portugal: Epidemiology and In-Hospital Mortality].
In Portugal, the epidemiology of acute pulmonary embolism is poorly understood. In this study, we sought to characterize the pulmonary embolism from the hospital data and evaluate its in-hospital mortality and respective prognostic factors. The study used diagnostic related groups data from National Health System hospitals from 2003 to 2013 and National Statistics Institute population data to establish the evolution of admissions with the diagnosis of pulmonary embolism, their inhospital mortality rates and the population incidence rates. Diagnosis-related group microdata were used in a logit regression modeling in-hospital mortality as a function of individual characteristics and context variables. Between 2003 and 2013 there were 35,200 episodes of hospitalization in patients with 18 or more years in which one of the diagnoses was pulmonary embolism (primary diagnosis in 67% of cases). The estimated incidence rate in 2013 was 35/100,000 population (≥ 18 years). Between 2003 and 2013, the annual number of episodes kept increasing, but the in-hospital mortality rate decreased (from 31.8% to 17% for all cases and from 25% to 11.2% when pulmonary embolism was the main diagnosis). The probability of death decreases when there is a computerized tomography scan registry or when patients are females and increases with age and the presence of co-morbidities. In the last decade there was an increased incidence of pulmonary embolism likely related to an increased number of dependents and bedridden. However, there was a in-hospital mortality reduction of such size that the actual mortality in the general population was reduced. One possible explanation is that there has been an increase in episodes of pulmonary embolism with incrementally lower levels of severity, due to the greater capacity of diagnosis of less severe cases. Another possible explanation is greater effectiveness of hospital care. According to the logistic regression analysis, improvements in hospital care effectiveness in recent years are primarily responsible for the mortality reduction. About 79% of the reduction of in-hospital mortality of pulmonary embolism between 2003 and 2013 can be attributed to greater effectiveness of hospital care and the rest to the favorable change in patient characteristics associated with risk of death.


Modifiable environmental exposure and risk of rheumatoid arthritis-current evidence from genetic studies.
Rheumatoid arthritis (RA) is a multifactorial chronic autoimmune disease, which involves a complex interplay of environmental triggers and genetic components in its etiology. It has been shown that genetics only explain about half of the liability to develop RA, leaving a large room for non-genetic factors. Indeed, several environmental exposures including smoking, drinking, obesity, and dietary patterns (and more) have been identified to be associated with RA risk, yet the observational nature of conventional epidemiological investigation hampers causal inference, as the validity of results could be plagued by measurement error, confounding, and/or reverse causality. Mendelian randomization (MR) is a novel statistical approach that uses genetic variants as instrumental variables (IV) to make causal inferences from observational data. The current genetic discoveries in the many heritable and modifiable human complex traits have provided an exceptional opportunity to evaluate a putative causal relationship between exposure and outcome in the absence of high-quality experimental or intervention studies, through a MR design. In the current review, we detail the contribution of MR studies hitherto conducted for modifiable environmental exposures with the risk of RA to understand the role of these factors in RA pathogenesis. We start with a brief introduction of each study, follow by a summarization of shortcomings and conclude by highlighting future directions. The application of MR design in the field of rheumatology remains limited. Only a few MR studies have examined the causal roles of vitamin D, cigarette smoking, alcohol consumption, coffee consumption, and levels of education in RA, where, no consistent evidence for a causal relationship has been found. Most studies lacked sensitivity analyses to verify MR model assumptions and to guarantee the validity of results. Almost all studies are likely to bias the strength of association towards a null value, since they used IVs from earlier GWAS(s) of exposures with a small sample size (i.e., few genetic markers). As the magnitudes of GWAS expand rapidly, additional trait-associated loci have been discovered. Incorporating these loci would greatly improve the strength of genetic instruments, as well as both the accuracy and precision of MR estimates. To conclude, there is a need for an update and a huge space for improvement of future MR studies in RA.


Aspects of social support associated with depression at hospitalization and follow-up assessment among cardiac patients.
High levels of depressive symptoms have been shown to affect the morbidity, mortality, and functioning of patients with myocardial infarction (MI). Findings have shown that social support is associated with depression in both patient and community samples. This study examined various aspects of social support as they relate to depressive symptoms in patients with MI, both in the hospital and 2 weeks later. As part of the Enhancing Recovery in Coronary Heart Disease (ENRICHD) pilot study, measures of perceived social support, social networks, social support received, and social conflict were administered to 196 patients with MI. These patients also were administered the Beck Depression Inventory and the Hamilton Rating Scale for Depression. Depression was reassessed 2 weeks later. Relations between social support indicators and the depression measures were examined. The prevalence of depression symptoms was high, especially among poorer and younger patients. There was modest improvement across time. Patients with high social support scores, particularly those reflecting perceived support, had lower scores on depression measures at baseline. High levels of perceived support and low social conflict at baseline were associated with less follow-up depression, as measured by the Beck cognitive scale, but not the Beck somatic scale nor the Hamilton scale. There were few associations with measures of social networks and received support. Social support indicators were differentially related to depression among patients with MI while in the hospital and 2 weeks later. The pattern of associations also depended on the measure of depression. A broad assessment strategy of both social support and depression is needed for a full understanding of their interrelations.


The role of dipeptidylpeptidase-4 inhibitors in management of cardiovascular disease in diabetes; focus on linagliptin.
Multiple population based analyses have demonstrated a high incidence of cardiovascular disease (CVD) and cardiovascular (CV) mortality in subjects with T2DM that reduces life expectancy by as much as 15 years. Importantly, the CV system is particularly sensitive to the metabolic and immune derangements present in obese pre-diabetic and diabetic individuals; consequently, CV dysfunction is often the initial CV derangement to occur and promotes the progression to end organ/tissue damage in T2DM. Specifically, diabetic CVD can manifest as microvascular complications, such as nephropathy, retinopathy, and neuropathy, as well as, macrovascular impairments, including ischemic heart disease, peripheral vascular disease, and cerebrovascular disease. Despite some progress in prevention and treatment of CVD, mainly via blood pressure and dyslipidemia control strategies, the impact of metabolic disease on CV outcomes is still a major challenge and persists in proportion to the epidemics of obesity and diabetes. There is abundant pre-clinical and clinical evidence implicating the DPP-4-incretin axis in CVD. In this regard, linagliptin is a unique DPP-4 inhibitor with both CV and renal safety profiles. Moreover, it exerts beneficial CV effects beyond glycemic control and beyond class effects. Linagliptin is protective for both macrovascular and microvascular complications of diabetes in preclinical models, as well as clinical models. Given the role of endothelial-immune cell interactions as one of the key events in the initiation and progression of CVD, linagliptin modulates these cell-cell interactions by affecting two important pathways involving stimulation of NO signaling and potent inhibition of a key immunoregulatory molecule.


Secretion of fibrinolytic enzymes facilitates human mesenchymal stem cell invasion into fibrin clots.
Adult human mesenchymal stem cells (hMSC) are involved in wound healing and regeneration of mesodermal tissue, but the underlying homing mechanisms are not well understood. Fibrin clot formation is associated with most wound healing processes and potentially guides the recruitment of hMSC. The objective of this study is the investigation of a fibrinolytic capacity, which is required for hMSC to migrate into a wounded tissue and thus to contribute to tissue regeneration. Using RT-PCR, semiquantitative real-time PCR and ELISA, we detected key components of the fibrinolytic cascade, including the urokinase plasminogen activator (uPA) and its receptor (uPAR), the tissue plasminogen activator (tPA) and the plasminogen activator inhibitor (PAI), suggesting a strong fibrinolytic activity of hMSC. To test this activity in a functional assay, we cultured fibrin-embedded hMSC in vitro for 7 days. The cells efficiently dissolved the surrounding fibrin mesh into the fibrin degradation products, the fibrinopeptides. The fibrinolytic activity of hMSC and human dermal fibroblasts, known to be critically involved in skin wound extracellular matrix remodeling, was similar. Our results suggest that a high intrinsic fibrinolytic capacity of hMSC mediates the invasion into a fibrin clot of a wounded tissue.


Possible modes of dissemination of the amphibian chytrid Batrachochytrium dendrobatidis in the environment.
Amphibian chytridiomycosis caused by Batrachochytrium dendrobatidis has spread at an alarming rate over large distances throughout sensitive frog populations in eastern Australia, Central America and New Zealand. Infected amphibians and contaminated water are implicated in translocation, but other vectors are unknown. Through in vitro studies we show that potential means of translocation may be moist soil and bird feathers. B. dendrobatidis survived for up to 3 mo in sterile, moist river sand with no other nutrients added. B. dendrobatidis attached to and grew on sterile feathers and were able to be transported by feathers to establish new cultures in media, surviving between 1 and 3 h of drying between transfers. If these in vitro results are valid in the natural environment, the findings raise the possibilities that B. dendrobatidis may be translocated by movement of moist river sand and that birds may carry the amphibian chytrid between frog habitats. However, further studies using sand and feathers containing normal microflora are essential.


3D fiber-deposited scaffolds for tissue engineering: influence of pores geometry and architecture on dynamic mechanical properties.
One of the main issues in tissue engineering is the fabrication of scaffolds that closely mimic the biomechanical properties of the tissues to be regenerated. Conventional fabrication techniques are not sufficiently suitable to control scaffold structure to modulate mechanical properties. Within novel scaffold fabrication processes 3D fiber deposition (3DF) showed great potential for tissue engineering applications because of the precision in making reproducible 3D scaffolds, characterized by 100% interconnected pores with different shapes and sizes. Evidently, these features also affect mechanical properties. Therefore, in this study we considered the influence of different structures on dynamic mechanical properties of 3DF scaffolds. Pores were varied in size and shape, by changing fibre diameter, spacing and orientation, and layer thickness. With increasing porosity, dynamic mechanical analysis (DMA) revealed a decrease in elastic properties such as dynamic stiffness and equilibrium modulus, and an increase of the viscous parameters like damping factor and creep unrecovered strain. Furthermore, the Poisson's ratio was measured, and the shear modulus computed from it. Scaffolds showed an adaptable degree of compressibility between sponges and incompressible materials. As comparison, bovine cartilage was tested and its properties fell in the fabricated scaffolds range. This investigation showed that viscoelastic properties of 3DF scaffolds could be modulated to accomplish mechanical requirements for tailored tissue engineered applications.


Diet, nutrition and the prevention of hypertension and cardiovascular diseases.
Cardiovascular diseases (CVD) are growing contributors to global disease burdens, with epidemics of CVD advancing across many regions of the world which are experiencing a rapid health transition. Diet and nutrition have been extensively investigated as risk factors for major cardiovascular diseases like coronary heart disease (CHD) and stroke and are also linked to other cardiovascular risk factors like diabetes, high blood pressure and obesity. The interpretation of evidence needs to involve a critical appraisal of methodological issues related to measurement of exposures, nature of outcome variables, types of research design and careful separation of cause, consequence and confounding as the basis for observed associations. Adequate evidence is available, from studies conducted within and across populations, to link several nutrients, minerals, food groups and dietary patterns with an increased or decreased risk of CVD. Dietary fats associated with an increased risk of CHD include trans-fats and saturated fats, while polyunsaturated fats are known to be protective. Dietary sodium is associated with elevation of blood pressure, while dietary potassium lowers the risk of hypertension and stroke. Regular frequent intake of fruits and vegetables is protective against hypertension, CHD and stroke. Composite diets (such as DASH diets, Mediterranean diet, 'prudent' diet) have been demonstrated to reduce the risk of hypertension and CHD. Sufficient knowledge exists to recommend nutritional interventions, at both population and individual levels, to reduce cardiovascular risk. That knowledge should now be translated into policies which promote healthy diets and discourage unhealthy diets. This requires coordinated action at the level of governments, international organizations, civil society and responsible sections of the food industry.


Enriched expression of GluD1 in higher brain regions and its involvement in parallel fiber-interneuron synapse formation in the cerebellum.
Of the two members of the δ subfamily of ionotropic glutamate receptors, GluD2 is exclusively expressed at parallel fiber-Purkinje cell (PF-PC) synapses in the cerebellum and regulates their structural and functional connectivity. However, little is known to date regarding cellular and synaptic expression of GluD1 and its role in synaptic circuit formation. In the present study, we investigated this issue by producing specific and sensitive histochemical probes for GluD1 and analyzing cerebellar synaptic circuits in GluD1-knock-out mice. GluD1 was widely expressed in the adult mouse brain, with high levels in higher brain regions, including the cerebral cortex, striatum, limbic regions (hippocampus, nucleus accumbens, lateral septum, bed nucleus stria terminalis, lateral habenula, and central nucleus of the amygdala), and cerebellar cortex. In the cerebellar cortex, GluD1 mRNA was expressed at the highest level in molecular layer interneurons and its immunoreactivity was concentrated at PF synapses on interneuron somata. In GluD1-knock-out mice, the density of PF synapses on interneuron somata was significantly reduced and the size and number of interneurons were significantly diminished. Therefore, GluD1 is common to GluD2 in expression at PF synapses, but distinct from GluD2 in neuronal expression in the cerebellar cortex; that is, GluD1 in interneurons and GluD2 in PCs. Furthermore, GluD1 regulates the connectivity of PF-interneuron synapses and promotes the differentiation and/or survival of molecular layer interneurons. These results suggest that GluD1 works in concert with GluD2 for the construction of cerebellar synaptic wiring through distinct neuronal and synaptic expressions and also their shared synapse-connecting function.