Actin filament organization in the fish keratocyte lamellipodium.
From recent studies of locomoting fish keratocytes it was proposed that the dynamic turnover of actin filaments takes place by a nucleation-release mechanism, which predicts the existence of short (less than 0.5 microns) filaments throughout the lamellipodium (Theriot, J. A., and T. J. Mitchison. 1991. Nature (Lond.). 352:126-131). We have tested this model by investigating the structure of whole mount keratocyte cytoskeletons in the electron microscope and phalloidin-labeled cells, after various fixations, in the light microscope. Micrographs of negatively stained keratocyte cytoskeletons produced by Triton extraction showed that the actin filaments of the lamellipodium are organized to a first approximation in a two-dimensional orthogonal network with the filaments subtending an angle of around 45 degrees to the cell front. Actin filament fringes grown onto the front edge of keratocyte cytoskeletons by the addition of exogenous actin showed a uniform polarity when decorated with myosin subfragment-1, consistent with the fast growing ends of the actin filaments abutting the anterior edge. A steady drop in filament density was observed from the mid-region of the lamellipodium to the perinuclear zone and in images of the more posterior regions of lower filament density many of the actin filaments could be seen to be at least several microns in length. Quantitative analysis of the intensity distribution of fluorescent phalloidin staining across the lamellipodium revealed that the gradient of filament density as well as the absolute content of F-actin was dependent on the fixation method. In cells first fixed and then extracted with Triton, a steep gradient of phalloidin staining was observed from the front to the rear of the lamellipodium. With the protocol required to obtain the electron microscope images, namely Triton extraction followed by fixation, phalloidin staining was, significantly and preferentially reduced in the anterior part of the lamellipodium. This resulted in a lower gradient of filament density, consistent with that seen in the electron microscope, and indicated a loss of around 45% of the filamentous actin during Triton extraction. We conclude, first that the filament organization and length distribution does not support a nucleation release model, but is more consistent with a treadmilling-type mechanism of locomotion featuring actin filaments of graded length. Second, we suggest that two layers of filaments make up the lamellipodium; a lower, stabilized layer associated with the ventral membrane and an upper layer associated with the dorsal membrane that is composed of filaments of a shorter range of lengths than the lower layer and which is mainly lost in Triton.


Metal ion trafficking in earthworms. Identification of a cadmium-specific metallothionein.
Exposure to cadmium poses a considerable risk to human health and environmental safety. Earthworms reside in the most contaminated sites on earth, displaying a phenomenal tolerance to toxic heavy metals. They exhibit a distinct metabolic pathway that allows the bio-accumulation of cadmium to yield body burdens in excess of 1/1000th of total dry body weight, a most impressive figure by any standard. However, the precise molecular mechanism underlying this phenomenon remains to be unraveled. This study meets this challenge by fully characterizing the major metal-binding protein in earthworms, namely the two isoforms of metallothionein. Chemical analysis of recombinant protein showed that although both isoforms bind equimolar amounts of cadmium (6 mol), wMT-2 is more stable during proton competition. Furthermore, isoform-specific transcript analysis demonstrated that only wMT-2 is responsive to cadmium in a dose and temporal manner. The specific sequestration of cadmium to wMT-2 protein was confirmed in situ using polyclonal antisera. The latter also provided the means for mapping the cellular and intracellular distribution of metallothionein, thus yielding a holistic insight into its involvement in cadmium transit during absorption, storage, and excretion. The structure-function relationship of wMT-2 and its role in cadmium detoxification through sequestration and compartmentalization is discussed.


μCB-seq: microfluidic cell barcoding and sequencing for high-resolution imaging and sequencing of single cells.
Single-cell RNA sequencing (scRNA-seq) enables the investigation of complex biological processes in multicellular organisms with high resolution. However, many phenotypic features that are critical to understanding the functional role of cells in a heterogeneous tissue or organ are not directly encoded in the genome and therefore cannot be profiled with scRNA-seq. Quantitative optical microscopy has long been a powerful approach for characterizing diverse cellular phenotypes including cell morphology, protein localization, and chemical composition. Combining scRNA-seq with optical imaging has the potential to provide comprehensive single-cell analysis, allowing for functional integration of gene expression profiling and cell-state characterization. However, it is difficult to track single cells through both measurements; therefore, coupling current scRNA-seq protocols with optical measurements remains a challenge. Here, we report microfluidic cell barcoding and sequencing (μCB-seq), a microfluidic platform that combines high-resolution imaging and sequencing of single cells. μCB-seq is enabled by a novel fabrication method that preloads primers with known barcode sequences inside addressable reaction chambers of a microfluidic device. In addition to enabling multi-modal single-cell analysis, μCB-seq improves gene detection sensitivity, providing a scalable and accurate method for information-rich characterization of single cells.


Depression care in the United States: too little for too few.
To determine the prevalence and adequacy of depression care among different ethnic and racial groups in the United States. Collaborative Psychiatric Epidemiology Surveys (CPES) data were analyzed to calculate nationally representative estimates of depression care. The 48 coterminous United States. Household residents 18 years and older (N = 15 762) participated in the study. Past-year depression pharmacotherapy and psychotherapy using American Psychiatric Association guideline-concordant therapies. Depression severity was assessed with the Quick Inventory of Depressive Symptomatology Self-Report. Primary predictors were major ethnic/racial groups (Mexican American, Puerto Rican, Caribbean black, African American, and non-Latino white) and World Mental Health Composite International Diagnostic Interview criteria for 12-month major depressive episode. Mexican American and African American individuals meeting 12-month major depression criteria consistently and significantly had lower odds for any depression therapy and guideline-concordant therapies despite depression severity ratings not significantly differing between ethnic/racial groups. All groups reported higher use of any past-year psychotherapy and guideline-concordant psychotherapy compared with pharmacotherapy; however, Caribbean black and African American individuals reported the highest proportions of this use. Few Americans with recent major depression have used depression therapies and guideline-concordant therapies; however, the lowest rates of use were found among Mexican American and African American individuals. Ethnic/racial differences were found despite comparable depression care need. More Americans with recent major depression used psychotherapy over pharmacotherapy, and these differences were most pronounced among Mexican American and African American individuals. This report underscores the importance of disaggregating ethnic/racial groups and depression therapies in understanding and directing efforts to improve depression care in the United States.


Isolation, characterization and therapeutic potential assessment of bacteriophages virulent to Staphylococcus aureus associated with goat mastitis.
In the present study, the therapeutic potential of bacteriophages virulent to Staphylococcus aureus associated with goat mastitis were isolated, identified and assessed. Staphylococcus aureus (host or indicator bacterium) was isolated from a goat suffering from clinical mastitis. Based on cultural, morphological, biochemical tests and amplification of S. aureus specific thermonuclease gene in PCR, the identity of the organism was confirmed as S. aureus. Bacteriophages were isolated from soil and faecal samples (n=42) collected from different parts of the Mathura district in Uttar Pradesh (India), and their identity was confirmed by amplification of the bacteriophage-specific endolysin gene fragment in PCR. The thermal tolerance study revealed that all phage isolates were stable at 30 and 40°C with 100% lytic efficacy and their activities reduced to 62-80% at 50°C declining sharply at 60°C with less than 5% efficacy. Likewise, at pH = 6.5 and 7.5, the survivability of all isolates was 100% which reduced to 70-79% and 84-91% at pH = 5.5 and 8.5, respectively. All isolates were stable up to 3 months at 37°C, and for 16 months at 4°C but the stability of their respective endolysins only lasted for 12-23 days at 37°C and 6 months at 4°C. Three of the bacteriophage isolates, S. aureus phage/CIRG/1, S. aureus phage/CIRG/4 and S. aureus phage/CIRG/5 exhibited lytic activity against over 80% of the staphylococcal isolates. The results of the present study provide insight for the use of lytic bacteriophages for therapeutic interventions against multi-drug-resistant S. aureus inducing mastitis in goats.


Infrared microspectroscopy: a multiple-screening platform for investigating single-cell biochemical perturbations upon prion infection.
Prion diseases are a group of fatal neurodegenerative disorders characterized by the accumulation of prions in the central nervous system. The pathogenic prion (PrP(Sc)) possesses the capability to convert the host-encoded cellular isoform of the prion protein, PrP(C), into nascent PrP(Sc). The present work aims at providing novel insight into cellular response upon prion infection evidenced by synchrotron radiation infrared microspectroscopy (SR-IRMS). This non-invasive, label-free analytical technique was employed to investigate the biochemical perturbations undergone by prion infected mouse hypothalamic GT1-1 cells at the cellular and subcellular level. A decrement in total cellular protein content upon prion infection was identified by infrared (IR) whole-cell spectra and validated by bicinchoninic acid assay and single-cell volume analysis by atomic force microscopy (AFM). Hierarchical cluster analysis (HCA) of IR data discriminated between infected and uninfected cells and allowed to deduce an increment of lysosomal bodies within the cytoplasm of infected GT1-1 cells, a hypothesis further confirmed by SR-IRMS at subcellular spatial resolution and fluorescent microscopy. The purpose of this work, therefore, consists of proposing IRMS as a powerful multiscreening platform, drawing on the synergy with conventional biological assays and microscopy techniques in order to increase the accuracy of investigations performed at the single-cell level.


Intersections and Non-Intersections: A Protocol for Identifying Pedestrian Crash Risk Locations in GIS.
Intersection and non-intersection locations are commonly used as spatial units of analysis for modeling pedestrian crashes. While both location types have been previously studied, comparing results is difficult given the different data and methods used to identify crash-risk locations. In this study, a systematic and replicable protocol was developed in GIS (Geographic Information System) to create a consistent spatial unit of analysis for use in pedestrian crash modelling. Four publicly accessible datasets were used to identify unique intersection and non-intersection locations: Roadway intersection points, roadway lanes, legal speed limits, and pedestrian crash records. Two algorithms were developed and tested using five search radii (ranging from 20 to 100 m) to assess the protocol reliability. The algorithms, which were designed to identify crash-risk locations at intersection and non-intersection areas detected 87.2% of the pedestrian crash locations (r: 20 m). Agreement rates between algorithm results and the crash data were 94.1% for intersection and 98.0% for non-intersection locations, respectively. The buffer size of 20 m generally showed the highest performance in the analyses. The present protocol offered an efficient and reliable method to create spatial analysis units for pedestrian crash modeling. It provided researchers a cost-effective method to identify unique intersection and non-intersection locations. Additional search radii should be tested in future studies to refine the capture of crash-risk locations.


Serum miR-22 as potential non-invasive predictor of poor clinical outcome in newly diagnosed, uniformly treated patients with diffuse large B-cell lymphoma: an explorative pilot study.
Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous group of tumors, with aggressive clinical course that renders prognostication and choice of treatment strategy difficult. Chemo-immunotherapy with rituximab, cyclophosphamide, doxorubicin, vincristine, prednisone (R-CHOP) is the current first-line treatment. MicroRNAs (miRNAs) are under investigation as novel diagnostic and prognostic biomarkers in several malignancies, including malignant lymphomas. While tissue miRNAs in DLBCL patients have been extensively studied as biomarkers, only few reports to date have evaluated the role of circulating/serum miRNAs as potential prognostic factors. Here circulating/serum miRNAs, including miR-22, were investigated as potential non-invasive biomarkers, with the aim of a better prognostic stratification of DLBCL patients. MiRNAs were selected by global expression profile of serum miRNAs of DLBCL patients, The Cancer Genome Atlas (TCGA) analysis and literature research. Serum and tissues miRNA expression profile in de novo DLBCL patients, consecutively enrolled for this study, were detected by quantitative real-time polymerase chain reaction. Relative expression was calculated using the comparative Ct method. Statistical significance was determined using the Mann-Whitney rank sum and Fisher's exact test. Survival analysis was conducted through the use of Kaplan-Meier method. Spearman's Rho was applied to study the correlation between miRNA distributions and days to first relapse. Experimentally validated miRNA-target interactions were assessed by miRTarBase database. Negative miRNA-mRNA correlation was evaluated in TCGA DLBCL dataset. Pathway analysis was performed by the functional annotation clustering DAVID tool. We showed a significant modulation of serum miR-22 after R-CHOP treatment compared with basal values but no difference between baseline serum miRNAs values of DLBCL patients and healthy controls. High expression level of serum miR-22 in DLBCL at diagnosis (n = 36) is associated with a worse PFS and is independent of the currently used clinical prognostic index. Integrative and pathways analysis of miR-22 identified target genes involved in different important pathways such as p53 signaling. Our data suggest that miR-22 is of potential interest as non-invasive biomarker to predict clinical outcome in DLBCL patients. Characterization of miR-22 pathways can pave the way to the development of targeted therapy approaches for specific subgroups of DLBCL patients.


Tyrosine phosphorylation of cortactin by the FAK-Src complex at focal adhesions regulates cell motility.
Cell migration plays an important role in many physiological and pathological processes, including immune cell chemotaxis and cancer metastasis. It is a coordinated process that involves dynamic changes in the actin cytoskeleton and its interplay with focal adhesions. At the leading edge of a migrating cell, it is the re-arrangement of actin and its attachment to focal adhesions that generates the driving force necessary for movement. However, the mechanisms involved in the attachment of actin filaments to focal adhesions are still not fully understood. Signaling by the FAK-Src complex plays a crucial role in regulating the formation of protein complexes at focal adhesions to which the actin filaments are attached. Cortactin, an F-actin associated protein and a substrate of Src kinase, was found to interact with FAK through its SH3 domain and the C-terminal proline-rich regions of FAK. We found that the autophosphorylation of Tyr(397) in FAK, which is necessary for FAK activation, was not required for the interaction with cortactin, but was essential for the tyrosine phosphorylation of the associated cortactin. At focal adhesions, cortactin was phosphorylated at tyrosine residues known to be phosphorylated by Src. The tyrosine phosphorylation of cortactin and its ability to associate with the actin cytoskeleton were required in tandem for the regulation of cell motility. Cell motility could be inhibited by truncating the N-terminal F-actin binding domains of cortactin or by blocking tyrosine phosphorylation (Y421/466/475/482F mutation). In addition, the mutant cortactin phosphorylation mimic (Y421/466/475/482E) had a reduced ability to interact with FAK and promoted cell motility. The promotion of cell motility by the cortactin phosphorylation mimic could also be inhibited by truncating its N-terminal F-actin binding domains. Our results suggest that cortactin acts as a bridging molecule between actin filaments and focal adhesions. The cortactin N-terminus associates with F-actin, while its C-terminus interacts with focal adhesions. The tyrosine phosphorylation of cortactin by the FAK-Src complex modulates its interaction with FAK and increases its turnover at focal adhesions to promote cell motility.


Hepatotoxicity of 6-mercaptopurine (6-MP) and Azathioprine (AZA) in adult IBD patients.
6-Mercaptopurine (6-MP) and azathioprine (AZA) are effective in the treatment of IBD; however, drug-induced hepatotoxicity has been reported in 10-15% of pediatric patients and has been associated with the 6-MP metabolite 6-methylmercaptopurine ribonucleotide (6-MMPR) at levels >5,700 pmol/8 x 10(8) RBC. The aim of this study was to assess the prevalence of 6-MP/AZA hepatotoxicity and its correlation with serum 6-MMPR levels in adult IBD patients. Aminotransferases, bilirubin, and 6-MP metabolite levels were measured in 173 adult IBD patients treated with 6-MP or AZA from November 2002 to December 2003. Hepatotoxicity was defined as AST and/or ALT >2x upper limit of normal or cholestasis. Eight patients (4.6%) met criteria for a diagnosis of 6-MP/AZA-induced hepatotoxicity. The mean 6-MMPR level in these 8 patients was 10,537 pmol/8 x 10(8) RBC versus 3,452 pmol/8 x 10(8) RBC in the nonhepatotoxic group (P < 0.001). Risk of hepatotoxicity above the third quartile (6-MMPR > 5,300) was 5 times that below the third quartile (11.4%vs 2.3%, P < 0.05); however, nearly 90% of all patients with 6-MMPR > 5,300 pmol/8 x 10(8) RBC had no hepatotoxicity, while almost 40% of subjects with hepatotoxicity had 6-MMPR levels below this cutoff. 6-MP/AZA-induced hepatotoxicity is uncommon in the adult population. Although hepatotoxicity is associated with higher mean 6-MMPR levels, the sensitivity and specificity of 6-MMPR for drug-induced hepatotoxicity was poor. Monitoring liver tests in patients on 6-MP/AZA is suggested, and dose reduction or cessation of 6-MP/AZA, even with high 6-MMPR levels, should be reserved for patients with elevated aminotransferases.


Hemolysates reduce iron released from transferrin.
Transferrin donates iron to reticulocytes as follows: it binds to a receptor on the reticulocyte surface; the complex is endocytosed; both irons are released and the transferrin is recycled to the cell exterior. It has been proposed that the trigger for iron release after transferrin endocytosis is acidification of the endocytic vesicle. But this could account for removal of only one of transferrin's two irons, since only one of the irons is labile at acid pH. Moreover, iron continues to be removed from transferrin when acidification of the vesicle is blocked by a chloride-transport inhibitor. Thus a detailed explanation of iron removal from transferrin remains elusive. In earlier work we showed that iron can be removed from transferrin by whole hemolysates and also by the combined action of hemoglobin and ATP at pH 7. We now show that the iron released from transferrin by hemolysates, and by hemoglobin and ATP, is in the Fe(II) oxidation state. We also show that ADP and DPG can substitute for ATP and that NADH and NADPH can substitute for the hemoglobin, although with these substitutions Fe(II) is generated less efficiently. The reductive release of iron from transferrin is rapid enough to account for all the iron processed by a young reticulocyte. We speculate that transferrin iron may be reduced to Fe(II) before reaching the mitochondria.


Interactions of emotion and anxiety on visual working memory performance.
It is a widely observed finding that emotion and anxiety interact; highly stressed or anxious individuals show robust attentional biases towards external negative information. More generally, research has suggested that exposure to threatening stimuli, as well as the experience of acute stress, also may impair top-down attentional control and working memory. In the current study, we investigated how the influence of emotion and anxiety may interact to influence working memory performance. Participants were required to encode the orientation of four simple shapes, eight, or four shapes while filtering out four other irrelevant shapes from memory. Before memory displays, an irrelevant neutral or fearful face cue also was presented. Memory performance was found to interact with self-reported state anxiety and cue valence; on neutral cue trials, state anxiety was negatively correlated with performance. This effect was absent following a fear cue. In addition, filtering efficiency was negatively associated with state anxiety solely following a fear cue. Our findings suggest that state anxiety's influence to visual working memory can be strongly modulated by external signals to threat. Most crucially, rather than anxious individuals having greater difficulty rejecting external threatening information, we observed that external threat may in its own right generally impair filtering efficiency in anxious individuals.


Imaging Cellular Proliferation in Prostate Cancer with Positron Emission Tomography.
Prostate cancer remains a major public health problem worldwide. Imaging plays an important role in the assessment of disease at all its clinical phases, including staging, restaging after definitive therapy, evaluation of therapy response, and prognostication. Positron emission tomography with a number of biologically targeted radiotracers has been demonstrated to have potential diagnostic and prognostic utility in the various clinical phases of this prevalent disease. Given the remarkable biological heterogeneity of prostate cancer, one major unmet clinical need that remains is the non-invasive imaging-based characterization of prostate tumors. Accurate tumor characterization allows for image-targeted biopsy and focal therapy as well as facilitates objective assessment of therapy effect. PET in conjunction with radiotracers that track the thymidine salvage pathway of DNA synthesis may be helpful to fulfill this necessity. We review briefly the preclinical and pilot clinical experience with the two major cellular proliferation radiotracers, [(18)F]-3'-deoxy-3'-fluorothymidine and [(18)F]-2'-fluoro-5-methyl-1-beta-D-arabinofuranosyluracil in prostate cancer.


Differential requirements for survival and proliferation of CD8 naïve or memory T cells.
The requisite molecular interactions for CD8 T cell memory were determined by comparison of monoclonal naïve and memory CD8(+) T cells bearing the T cell receptor (TCR) for the HY antigen. Naïve T cells required only the right major histocompatibility complex (MHC) class I-restricting molecule to survive; to expand, they also needed antigen. In contrast, for survival, memory cells did not require the restricting MHC allele, but needed only a nonspecific class I; for expansion the correct class I, but not antigen, was required. Thus, maintenance of CD8 T cell memory still required TCR-MHC class I interactions, but memory T cells may have a lower functional activation threshold that facilitates secondary responses.


Cluster failure: Why fMRI inferences for spatial extent have inflated false-positive rates.
The most widely used task functional magnetic resonance imaging (fMRI) analyses use parametric statistical methods that depend on a variety of assumptions. In this work, we use real resting-state data and a total of 3 million random task group analyses to compute empirical familywise error rates for the fMRI software packages SPM, FSL, and AFNI, as well as a nonparametric permutation method. For a nominal familywise error rate of 5%, the parametric statistical methods are shown to be conservative for voxelwise inference and invalid for clusterwise inference. Our results suggest that the principal cause of the invalid cluster inferences is spatial autocorrelation functions that do not follow the assumed Gaussian shape. By comparison, the nonparametric permutation test is found to produce nominal results for voxelwise as well as clusterwise inference. These findings speak to the need of validating the statistical methods being used in the field of neuroimaging.


Significant changes in systolic blood pressure post vectored upper cervical adjustment vs resting control groups: a possible effect of the cervicosympathetic and/or pressor reflex.
To determine whether a vectored adjustment of the atlas in patients identified as demonstrating signs of upper cervical joint dysfunction would cause lowering of blood pressure in comparison with resting controls. Test 1: controlled clinical trial with a treatment (adjustment) group and a control (resting) group. Test 2: controlled clinical trial with subjects serving as their own controls. Private chiropractic practice. Test 1: Forty established patients demonstrating signs of upper cervical subluxation/joint dysfunction and 40 established patients without such signs. Test 2: Thirty established patients demonstrating signs of upper cervical subluxation/joint dysfunction. Specific, vectored upper cervical (atlas) adjustment or similarly positioned resting. Prerest, postrest, and postadjustment systolic, diastolic, and pulse rates as recorded through use of a digital oscillometric sphygmomanometer. In test 1, subjects receiving adjustment had a significant (P <.001) decrease in systolic blood pressure whereas resting subjects did not. Intergroup comparison of the treatment (adjustment) and control (resting) groups demonstrated a significant difference (P <.001). A greater pre/post drop in systolic pressure was associated with greater age and higher initial systolic pressure. In test 2, the pre/postrest change in systolic blood pressure was not significant. The systolic blood pressure changed significantly (P <.001) from postrest readings to postadjustment readings. The results indicate that palpation and vectored atlas adjustment causes a significant decrease in systolic blood pressure in patients with putative upper cervical subluxation/joint dysfunction in comparison with resting controls. Similar results were also demonstrated when subjects acted as their own controls. The lack of randomization, blinding, and a manipulated control group are factors that weaken these findings. The sudden drop in systolic pressure is proposed to be due to stimulation of the cervicosympathetic reflex or moderation of muscle tone and elimination of the effects of the pressor reflex.


Effect of selenium supplementation on changes in HbA1c: Results from a multiple-dose, randomized controlled trial.
To investigate the effect of selenium supplementation at different dose levels on changes in HbA1c after 6 months and 2 years in a population of low selenium status. The Denmark PRECISE study was a single-centre, randomized, double-blinded, placebo-controlled, multi-arm, parallel clinical trial with four groups. In total, 491 volunteers aged 60 to 74 years were randomly assigned to treatment with 100, 200 or 300 μg selenium/day as selenium-enriched yeast or placebo-yeast. HbA1c measurements were available for 489 participants at baseline, 435 at 6 months, and 369 after 2 years of selenium supplementation. Analyses were performed by intention to treat. The mean (SD) age, plasma-selenium concentration, and blood HbA1c at baseline were 66.1 (4.1) years, 86.5 (16.3) ng/g and 36.6 (7.0) mmol/mol, respectively. During the initial 6-month intervention period, mean HbA1c (95% CI) decreased by 1.5 (-2.8 to -0.2) mmol/mol for 100 μg/d of selenium supplementation and by 0.7 (-2.0 to 0.6) mmol/mol for the 200 and 300 μg/d groups compared with placebo (P = 0.16 for homogeneity of changes across the four groups). After 2 years of selenium supplementation, HbA1c had decreased significantly in all treatment groups, with no difference between active treatment and placebo. Selenium supplementation in an elderly European population of low selenium status did not significantly affect HbA1c levels after 2 years. Our findings corroborate a possible U-shaped response of selenium supplementation on glucose metabolism.


Molecular cloning, characterization, and mutagenesis of a pel gene from Pseudomonas syringae pv. lachyrmans encoding a member of the Erwinia chrysanthemi pelADE family of pectate lyases.
The pelS gene from Pseudomonas syringae pv. lachrymans 859 was cloned by heterologous expression in nonpectolytic P. syringae pv. syringae BUVS1, using genomic DNA libraries constructed with two novel broad-host-range cosmid vectors, pCPP34 and pCPP47. Screening of P. syringae pv. syringae transconjugants for the ability to pit pectate media at pH 6.0 and 8.5 yielded several overlapping clones of the same DNA region. Ultrathin-layer isoelectric focusing gels, activity-stained with diagnostically buffered substrate overlays, revealed that this region encoded a single pectate lyase (PelS) with a pI of 9.4. pelS was subcloned from cosmid pCPP5020 and sequenced, revealing it to encode a member of the Erwinia chrysanthemi PelADE family, with highest similarity to Pseudomonas viridiflava PelV. A pelS probe hybridized at high stringency in DNA gel blots with total DNA from P. syringae pv. lachrymans strains 859 and Pla5, P. syringae pv. tabaci, P. syringae pv. phaseolicola, P. syringae pv. glycinea, P. fluorescens (marginalis), P. viridiflava, and Xanthomonas campestris pv. campestris, but not with P. syringae pv. pisi, P. syringae pv. syringae, P. syringae pv. tomato, P. syringae pv. papulans, E. chrysanthemi, or Ralstonia (Pseudomonas or Burkholderia) solanacearum. The PelS sequence revealed an N-terminal signal peptide, whose processing in Escherichia coli was confirmed by protein sequence analysis. PelS was similar to E. chrysanthemi PelE in its substrate preference and ability to reduce the viscosity of pectate and to macerate potato tuber tissue. A pelS:: omega Kmr mutation was marker-exchanged into P. syringae pv. lachrymans Pla5, pelS was also subcloned into the broad-host-range expression vector pML122 under control of the vector nptII promoter, and then transformed into P. syringae pv. lachrymans Pla5 to produce a strain overproducing PelS. Necrotic lesions developed in cotyledons following inoculation with all of the P. syringae pv. lachrymans Pla5 derivatives, regardless of their Pel phenotype. However, only cotyledons infected with pelS+ strains showed evidence of maceration and yielded Pel activity upon extraction. In contrast, pelS+ P. syringae pv. syringae BUVS1(pCPP5020) produced no symptoms in cucumber cotyledons. Thus, PelS in P. syringae pv. lachrymans appears to alter the final symptoms in infected cucumber cotyledons without contributing to pathogenicity or altering host range.


A randomised controlled trial to assess the clinical effectiveness and cost-effectiveness of alternative treatments to Inhibit VEGF in Age-related choroidal Neovascularisation (IVAN).
Bevacizumab (Avastin®, Roche), which is used in cancer therapy, is the 'parent' molecule from which ranibizumab (Lucentis®, Novartis) was derived for the treatment of neovascular age-related macular degeneration (nAMD). There were reports in the literature on the effectiveness of bevacizumab in treating nAMD, but no trials. The cost per dose of bevacizumab is about 5-10% that of ranibizumab. This trial was a head-to-head comparison of these two drugs. To compare the clinical effectiveness and cost-effectiveness of ranibizumab and bevacizumab, and two treatment regimens, for nAMD. Multicentre, factorial randomised controlled trial with within-trial cost-utility and cost-minimisation analyses from the perspective of the UK NHS. Participants, health professionals and researchers were masked to allocation of drug but not regimen. Computer-generated random allocations to combinations of ranibizumab or bevacizumab, and continuous or discontinuous regimen, were stratified by centre, blocked and concealed. Twenty-three ophthalmology departments in NHS hospitals. Patients ≥ 50 years old with active nAMD in the study eye with best corrected distance visual acuity (BCVA) ≥ 25 letters measured on a Early Treatment of Diabetic Retinopathy Study (ETDRS) chart. Previous treatment for nAMD, long-standing disease, lesion diameter > 6000 µm, thick blood at the fovea and any other confounding ocular disease were exclusion criteria. One eye per participant was studied; the fellow eye was treated according to usual care, if required. Ranibizumab and bevacizumab were procured commercially. Doses were ranibizumab 0.5 mg or bevacizumab 1.25 mg. The repackaged bevacizumab was quality assured. All participants were treated at visits 0, 1 and 2. Participants randomised to the continuous regimen were treated monthly thereafter. Participants randomised to the discontinuous regimen were not retreated after visit 2 unless pre-specified criteria for active disease were met. If retreatment was needed, monthly injections over 3 months were mandated. The primary outcome was BCVA. The non-inferiority margin was 3.5 letters. Secondary outcomes were contrast sensitivity; near visual acuity; reading index; neovascular lesion morphology; generic and disease-specific patient-reported outcomes, including macular disease-specific quality of life; survival free from treatment failure; resource use; quality-adjusted life-years (QALYs); and development of new geographic atrophy (GA) (outcome added during the trial). Results are reported for the study eye, except for patient-reported outcomes. Between 27 March 2008 and 15 October 2010, 610 participants were allocated and treated (314 ranibizumab, 296 bevacizumab; at 3 months, 305 continuous, 300 discontinuous). After 2 years, bevacizumab was neither non-inferior nor inferior to ranibizumab [-1.37 letters, 95% confidence interval (CI) -3.75 to +1.01 letters] and discontinuous treatment was neither non-inferior nor inferior to continuous treatment (-1.63 letters, 95% CI -4.01 to +0.75 letters). Lesion thickness at the fovea was similar by drug [geometric mean ratio (GMR) 0.96, 95% CI 0.90 to 1.03; p = 0.24] but 9% less with continuous treatment (GMR 0.91, 95% CI 0.85 to 0.97; p = 0.004). Odds of developing new GA during the trial were similar by drug [odds ratio (OR) 0.87, 95% CI 0.61 to 1.25; p = 0.46] but significantly higher with continuous treatment (OR 1.47, 95% CI 1.03 to 2.11; p = 0.033). Safety outcomes did not differ by drug but mortality was lower with continuous treatment (OR 0.47, 95% CI 0.22 to 1.03; p = 0.05). Continuous ranibizumab cost £3.5M per QALY compared with continuous bevacizumab; continuous bevacizumab cost £30,220 per QALY compared with discontinuous bevacizumab. These results were robust in sensitivity analyses. Ranibizumab and bevacizumab have similar efficacy. Discontinuing treatment and restarting when required results in slightly worse efficacy. Safety was worse with discontinuous treatment, although new GA developed more often with continuous treatment. Ranibizumab is not cost-effective, although it remains uncertain whether or not continuous bevacizumab is cost-effective compared with discontinuous bevacizumab at £20,000 per QALY threshold. Future studies should focus on the ocular safety of the two drugs, further optimisation of treatment regimens and criteria for stopping treatment. Current Controlled Trials ISRCTN92166560. This project was funded by the NIHR Health Technology Assessment programme and will be published in full in Health Technology Assessment; Vol. 19, No. 78. See the NIHR Journals Library website for further project information.


A phase I and pharmacological study of topotecan infused over 30 minutes for five days in patients with refractory acute leukemia.
The principal objectives of this study were to determine the feasibility of escalating doses of the hydrophilic topoisomerase I (topo I) inhibitor topotecan (TPT) as a 30-min infusion daily for 5 days in adults with refractory or relapsed acute leukemia and to study the pharmacokinetic behavior of high doses of TPT and pharmacodynamic determinants of TPT activity. Fourteen patients received 27 courses of TPT at doses ranging from 3.5 to 5.75 mg/m2/day every 3 weeks. A constellation of unusual adverse effects, consisting of high fever, rigors, precipitous anemia, and hyperbilirubinemia, was the principal dose-limiting toxicity of high doses of TPT on this schedule. These toxicities were consistently intolerable at the 5.75 mg/m2/day dose level; however, they were neither severe nor common at lower doses. Although the precise etiology of these effects is not known, high doses of TPT may induce acute hemolytic reactions in this patient population. Severe, albeit transient, mucositis was experienced by two of eight patients in 2 of 17 courses at the next lower dose level, 4.5 mg/m2/day, which was determined to be the maximum tolerated dose and the dose recommended for further trials. The pharmacokinetic behavior of TPT at high doses was not dose dependent and resembled that at lower doses. In view of preclinical data suggesting that TPT sensitivity might correlate with topo I levels, topo I content in leukemia blasts was assessed by Western blotting. Variations in topo I content were observed. Moreover, strong correlations were evident between topo I content and two markers of proliferation, proliferating cell nuclear antigen and nuclear protein B23, raising the possibility that differences in topo I content observed among various leukemia specimens might reflect differences in the proliferating fractions of cells in various leukemia samples. Although complete clearance of circulating leukemia blasts occurred in most courses, neither sustained responses nor hematopoietic recovery were observed in the heavily pretreated, poor-risk patients enrolled in this study, and it was not possible to correlate these differences in topo I content with clinical response. These results indicate that substantial dose escalation of TPT as a 30-minute infusion for a 5-day schedule above myelosuppressive doses is feasible in adults with refractory or relapsed leukemias; however, further development of alternate high-dose schedules in leukemia may be warranted in view of the nature of the dose-limiting toxicity and the lack of sustained clinical responses in this preliminary investigation.


Establishment of HLA-DR4 transgenic mice for the identification of CD4+ T cell epitopes of tumor-associated antigens.
Reports have shown that activation of tumor-specific CD4(+) helper T (Th) cells is crucial for effective anti-tumor immunity and identification of Th-cell epitopes is critical for peptide vaccine-based cancer immunotherapy. Although computer algorithms are available to predict peptides with high binding affinity to a specific HLA class II molecule, the ability of those peptides to induce Th-cell responses must be evaluated. We have established HLA-DR4 (HLA-DRA*01:01/HLA-DRB1*04:05) transgenic mice (Tgm), since this HLA-DR allele is most frequent (13.6%) in Japanese population, to evaluate HLA-DR4-restricted Th-cell responses to tumor-associated antigen (TAA)-derived peptides predicted to bind to HLA-DR4. To avoid weak binding between mouse CD4 and HLA-DR4, Tgm were designed to express chimeric HLA-DR4/I-E(d), where I-E(d) α1 and β1 domains were replaced with those from HLA-DR4. Th cells isolated from Tgm immunized with adjuvant and HLA-DR4-binding cytomegalovirus-derived peptide proliferated when stimulated with peptide-pulsed HLA-DR4-transduced mouse L cells, indicating chimeric HLA-DR4/I-E(d) has equivalent antigen presenting capacity to HLA-DR4. Immunization with CDCA155-78 peptide, a computer algorithm-predicted HLA-DR4-binding peptide derived from TAA CDCA1, successfully induced Th-cell responses in Tgm, while immunization of HLA-DR4-binding Wilms' tumor 1 antigen-derived peptide with identical amino acid sequence to mouse ortholog failed. This was overcome by using peptide-pulsed syngeneic bone marrow-derived dendritic cells (BM-DC) followed by immunization with peptide/CFA booster. BM-DC-based immunization of KIF20A494-517 peptide from another TAA KIF20A, with an almost identical HLA-binding core amino acid sequence to mouse ortholog, successfully induced Th-cell responses in Tgm. Notably, both CDCA155-78 and KIF20A494-517 peptides induced human Th-cell responses in PBMCs from HLA-DR4-positive donors. Finally, an HLA-DR4 binding DEPDC1191-213 peptide from a new TAA DEPDC1 overexpressed in bladder cancer induced strong Th-cell responses both in Tgm and in PBMCs from an HLA-DR4-positive donor. Thus, the HLA-DR4 Tgm combined with computer algorithm was useful for preliminary screening of candidate peptides for vaccination.


Are Canadian clinicians providing consistent sport-related concussion management advice?
To compare the knowledge and use of recommendations for the management of sport-related concussion (SRC) among sport and exercise medicine physicians (SEMPs) and emergency department physicians (EDPs) to assess the success of SRC knowledge transfer across Canada. A self-administered, multiple-choice survey accessed via e-mail by SEMPs and EDPs. The survey had been assessed for content validity. Canada. The survey was completed between May and July 2012 by SEMPs who had passed the diploma examination of the Canadian Academy of Sport and Exercise Medicine and by EDPs who did not hold this diploma. Knowledge and identification of sources of concussion management information, use of concussion diagnosis strategies, and whether physicians use common and consistent terminology when explaining cognitive rest strategies to patients after an SRC. There was a response rate of 28% (305 of 1085). The SEMP and EDP response rates were 41% (147 of 360) and 22% (158 of 725), respectively. Of the responses, 41% of EDPs and 3% of SEMPs were unaware of any consensus statements on concussion in sport; 74% of SEMPs used the Sport Concussion Assessment Tool, version 2 (SCAT2), "usually or always," whereas 88% of EDPs never used the SCAT2. When queried about how cognitive rest could best be achieved after an SRC, no consistent answer was documented. Differences and a lack of consistency in the implementation of recommendations for SRC patients were identified for SEMPs and EDPs. It appears that the SCAT2 is used more in the SEMP setting than in the emergency context. Further knowledge transfer efforts and research should address the barriers to achieving more consistent advice given by physicians who attend SRC patients.


PTSD symptom severity and sensitivity to blood, injury, and mutilation in U.S. army special operations soldiers.
Sensitivity to blood, injury, and mutilation (SBIM) may increase risk for posttraumatic stress disorder (PTSD), given that traumatic events often involve actual or perceived threat of bodily harm to oneself and/or others, including exposure to blood and other mutilation-related stimuli. A self-report questionnaire was administered to male, active duty, U.S. Army Special Operations Command soldiers who had deployed to Iraq and Afghanistan (n =694 males). We first used exploratory factor analysis to examine whether the 30-item Mutilation Questionnaire (Klorman et al., 1974) comprised a unitary measure of SBIM, finding that 10 of the items form a cohesive SBIM factor. Summed, those 10 SBIM items had a significant bivariate correlation with PTSD symptom severity. In a multiple regression analysis that included demographic characteristics and lifetime trauma exposure, SBIM was positively associated with PTSD symptom severity. Other significant multivariate predictors were high lifetime trauma exposure and junior enlisted rank. When trait neuroticism was added to the model to test the robustness of these findings, the association of SBIM with PTSD symptom severity remained significant. The results suggest that SBIM may be a risk factor for PTSD in male soldiers. Further research is warranted to improve measurement and understanding of SBIM.


Synthesis of 1,3-Dienes via a Sequential Suzuki-Miyaura Coupling/Palladium-Mediated Allene Isomerization Sequence.
We report a facile method for the synthesis of 1,3-dienes by a sequential process consisting of a palladium-catalyzed, base-free, Suzuki-Miyaura coupling/isomerization sequence. This sequence couples boronic acids with propargyl alcohols, generating the requisite allene in situ, followed by conversion of the unactivated allene to its 1,3-diene via a hydro-palladation/dehydro-palladation process. This process is general for a range of boronic acids, including boronic acids with electron-donating and -withdrawing groups, as well as heteroarylboronic acids. Key to this process is the boric acid byproduct of the base-free Suzuki-Miyauru coupling, which generates the required palladium-hydrido complex [H-Pd(II)-OB(OH)2] required for the isomerization.


Thinning pine plantations to reestablish oak openings species in northwestern Ohio.
Globally the area in forest plantations is rising by 2% annually, increasing the importance of plantations for production of human goods and services and for ecological functions such as carbon storage and biodiversity conservation. Specifically in the Great Lakes states and provinces of Midwestern North America, thousands of hectares of pine plantations were established in the early and mid-1900s to revegetate abandoned agricultural fields that had replaced mixed-species forests and oak-prairie ecosystems. Plantation establishment also was intended to bolster the timber base. Management priorities have shifted, with many resource managers currently seeking to manage existing plantations for promoting mixed-species ecosystems. The purpose of this study was to assess plant succession and the reestablishment of oak savanna and prairie species after thinning 14 plantations of Pinus resinosa and strobus in northwestern Ohio, USA. Thinning reduced tree basal area by an average of 75%. Plant communities were sampled on 0.05-ha plots one and 3 years after thinning and compared to 10 unthinned control plantations. By 3 years after thinning, thinned plots contained 2-3 times more species and 14 times more plant cover than control plots. The species composition of colonizing plants was most strongly correlated with residual pine basal area and soil variables related to drainage (e.g., sand concentration, available water capacity). Although plant composition was dominated by widespread colonizers such as Erechtites hieraciifolia, the coefficient of conservatism (indicative of species of more intact, undisturbed communities) significantly increased on thinned plots from year 1 to 3. This finding, coupled with the presence of four rare, state-listed Ohio species whose eight plot occurrences all were on thinned plots, suggests that plant composition is moving towards species typifying more high-quality savanna and prairie habitats.


A Pilot SMART for Developing an Adaptive Treatment Strategy for Adolescent Depression.
This pilot study was conducted to assess the feasibility and acceptability of 4 adaptive treatment strategies (ATSs) for adolescent depression to plan for a subsequent full-scale clinical trial. The ATSs aim to address 2 questions that arise when personalizing treatment: (a) For adolescents treated with Interpersonal Psychotherapy for depressed adolescents (IPT-A; Mufson et al., 2004 ), at what time point should therapists make the determination that the adolescent is not likely to respond if the initial treatment plan is continued (week 4 or week 8)? (b) For adolescents who are judged to need their treatment augmented, should the therapist increase the number of IPT-A sessions or add pharmacotherapy (fluoxetine)? A 16-week pilot sequential multiple assignment randomized trial (SMART) was conducted with 32 adolescents (M age = 14.9) who had a diagnosis of major depressive disorder, dysthymic disorder, or depressive disorder not otherwise specified. Adolescents were primarily female (75%) and Caucasian (84.4%). Data regarding the feasibility and acceptability of the study and treatment procedures and treatment response rates were collected. Week 4 was the more feasible and acceptable decision point for assessing need for a change to treatment. Adolescents, parents, and therapists reported a range of attitudes about medication and more intensive therapy as treatment options. Results from the pilot study have yielded additional research questions for the full-scale SMART and will improve our ability to successfully conduct the trial.


Genomic resolution of an aggressive, widespread, diverse and expanding meningococcal serogroup B, C and W lineage.
Neisseria meningitidis is a leading cause of meningitis and septicaemia. The hyperinvasive ST-11 clonal complex (cc11) caused serogroup C (MenC) outbreaks in the US military in the 1960s and UK universities in the 1990s, a global Hajj-associated serogroup W (MenW) outbreak in 2000-2001, and subsequent MenW epidemics in sub-Saharan Africa. More recently, endemic MenW disease has expanded in South Africa, South America and the UK, and MenC cases have been reported among European and North American men who have sex with men (MSM). Routine typing schemes poorly resolve cc11 so we established the population structure at genomic resolution. Representatives of these episodes and other geo-temporally diverse cc11 meningococci (n = 750) were compared across 1546 core genes and visualised on phylogenetic networks. MenW isolates were confined to a distal portion of one of two main lineages with MenB and MenC isolates interspersed elsewhere. An expanding South American/UK MenW strain was distinct from the 'Hajj outbreak' strain and a closely related endemic South African strain. Recent MenC isolates from MSM in France and the UK were closely related but distinct. High resolution 'genomic' multilocus sequence typing is necessary to resolve and monitor the spread of diverse cc11 lineages globally.


Associations between prenatal sunshine exposure and birth outcomes in China.
This paper is one of the first to examine the associations between prenatal sunshine exposure and birth outcomes, specifically the incidence of low birth weight (LBW) and small for gestational age (SGA), based on a nationally representative birth record dataset in China. During the sample period in the 1990s, migration was limited in rural China, allowing us to address the identification challenges, like residential sorting and avoidance behaviors. We found a nonlinear relationship between the length of sunlight and birth outcomes. In particular, prenatal exposure to increasing sunshine was associated with a reduction in the incidence of LBW and SGA, especially in the second trimester during pregnancy. This finding was consistent with the clinical evidence suggesting positive effects of sunshine on birth outcomes via obtaining vitamin D or relieving maternal stress.


Structural insights into the origins of DNA polymerase fidelity.
DNA polymerases discriminate from a pool of structurally similar molecules to insert the correct nucleotide to preserve Watson-Crick base pairing rules. The ability to choose between "right and wrong" is highly dependent on the identity of the polymerase. Because naturally occurring polymerases with divergent fidelities insert incorrect nucleotides with comparable efficiencies, fidelity is primarily governed by the ability to insert the correct nucleotide. DNA polymerases generally bind the correct nucleotide with similar affinities, but low-fidelity polymerases insert correct nucleotides more slowly than higher fidelity enzymes. A comparison of crystallographic ternary substrate complexes of DNA polymerases from five families exhibiting a range of nucleotide insertion rates reveals possible structural features that lead to rapid, efficient, and faithful DNA synthesis.


Bacteriology of hidradenitis suppurativa/acne inversa: A review.
Given that follicular papules and pustules, as well as nodules and abscesses, are the clinical hallmarks of hidradenitis suppurativa (HS), an infectious, bacterial pathway has been suspected in the pathogenesis of this chronic, inflammatory condition. Elucidating the behavior and role of bacterial species in HS and their interaction with cutaneous innate immunity will provide more insight into the pathophysiology of this condition. This review of prospective investigations suggests a synergistic relationship between impaired innate immunity and microbial factors in the etiology of HS.


Alternative splicing of a multi-drug transporter from Pseudoperonospora cubensis generates an RXLR effector protein that elicits a rapid cell death.
Pseudoperonospora cubensis, an obligate oomycete pathogen, is the causal agent of cucurbit downy mildew, a foliar disease of global economic importance. Similar to other oomycete plant pathogens, Ps. cubensis has a suite of RXLR and RXLR-like effector proteins, which likely function as virulence or avirulence determinants during the course of host infection. Using in silico analyses, we identified 271 candidate effector proteins within the Ps. cubensis genome with variable RXLR motifs. In extending this analysis, we present the functional characterization of one Ps. cubensis effector protein, RXLR protein 1 (PscRXLR1), and its closest Phytophthora infestans ortholog, PITG_17484, a member of the Drug/Metabolite Transporter (DMT) superfamily. To assess if such effector-non-effector pairs are common among oomycete plant pathogens, we examined the relationship(s) among putative ortholog pairs in Ps. cubensis and P. infestans. Of 271 predicted Ps. cubensis effector proteins, only 109 (41%) had a putative ortholog in P. infestans and evolutionary rate analysis of these orthologs shows that they are evolving significantly faster than most other genes. We found that PscRXLR1 was up-regulated during the early stages of infection of plants, and, moreover, that heterologous expression of PscRXLR1 in Nicotiana benthamiana elicits a rapid necrosis. More interestingly, we also demonstrate that PscRXLR1 arises as a product of alternative splicing, making this the first example of an alternative splicing event in plant pathogenic oomycetes transforming a non-effector gene to a functional effector protein. Taken together, these data suggest a role for PscRXLR1 in pathogenicity, and, in total, our data provide a basis for comparative analysis of candidate effector proteins and their non-effector orthologs as a means of understanding function and evolutionary history of pathogen effectors.


Continuously Tunable 250 GHz Gyrotron with a Double Disk Window for DNP-NMR Spectroscopy.
In this paper, we describe the design and experimental results from the rebuild of a 250 GHz gyrotron used for Dynamic Nuclear Polarization enhanced Nuclear Magnetic Resonance spectroscopy on a 380 MHz spectrometer. Tuning bandwidth of approximately 2 GHz is easily achieved at a fixed magnetic field of 9.24 T and a beam current of 95 mA producing an average output power of >10 W over the entire tuning band. This tube incorporates a double disk output sapphire window in order to maximize the transmission at 250.58 GHz. DNP Signal enhancement of >125 is achieved on a 13C-Urea sample using this gyrotron.


AKT1 polymorphisms are associated with risk for metabolic syndrome.
Converging lines of evidence suggest that AKT1 is a major mediator of the responses to insulin,insulin-like growth factor 1 (IGF1), and glucose. AKT1 also plays a key role in the regulation of both muscle cell hypertrophy and atrophy. We hypothesized that AKT1 variants may play a role in the endophenotypes that makeup metabolic syndrome. We studied a 12-kb region including the first exon of the AKT1 gene for association with metabolic syndrome-related phenotypes in four study populations [FAMUSS cohort (n = 574; age 23.7 ± 5.7 years), Strong Heart Study (SHS) (n = 2,134; age 55.5 ± 7.9 years), Dynamics of Health, Aging and Body Composition (Health ABC) (n = 3,075; age 73.6 ± 2.9 years), and Studies of a Targeted Risk Reduction Intervention through Defined Exercise (STRRIDE)(n = 175; age 40–65 years)]. We identified a three SNP haplotype that we call H1, which represents the ancestral alleles eles at the three loci and H2, which represents the derived alleles at the three loci. In young adult European Americans (FAMUSS), H1 was associated with higher fasting glucose levels in females. In middle age Native Americans (SHS), H1 carriers showed higher fasting insulin and HOMA in males, and higher BMI in females. Inolder African-American and European American subjects(Health ABC) H1 carriers showed a higher incidence of metabolic syndrome. Homozygotes for the H1 haplotype showed about twice the risk of metabolic syndrome in both males and females (p < 0.001). In middle-aged European Americans with insulin resistance (STRRIDE) studied by intravenous glucose tolerance test (IVGTT), H1 carriers showed increased insulin resistance due to the Sg component (p = 0.021). The 12-kb haplotype is a risk factor for metabolic syndrome and insulin resistance that needs to be explored in further populations.


Is vision function related to physical functional ability in older adults?
To assess the relationship between a broad range of vision functions and measures of physical performance in older adults. Cross-sectional study. Population-based cohort of community-dwelling older adults, subset of an on-going longitudinal study. Seven hundred eighty-two adults aged 55 and older (65% of living eligible subjects) had subjective health measures and objective physical performance evaluated in 1989/91 and again in 1993/95 and a battery of vision functions tested in 1993/95. Comprehensive battery of vision tests (visual acuity, contrast sensitivity, effects of illumination level, contrast and glare on acuity, visual fields with and without attentional load, color vision, temporal sensitivity, and the impact of dimming light on walking ability) and physical function measures (self-reported mobility limitations and observed measures of walking, rising from a chair and tandem balance). The failure rate for all vision functions and physical performance measures increased exponentially with age. Standard high-contrast visual acuity and standard visual fields showed the lowest failure rates. Nonstandard vision tests showed much higher failure rates. Poor performance on many individual vision functions was significantly associated with particular individual measures of physical performance. Using constructed combination vision variables, significant associations were found between spatial vision, field integrity, binocularity and/or adaptation, and each of the functional outcomes. Vision functions other than standard visual acuity may affect day-to-day functioning of older adults. Additional studies of these other aspects of vision and how they can be treated or rehabilitated are needed to determine whether these aspects play a role in strategies for reducing disability in older adults.


Interactions between plasma concentrations of folate and markers of vitamin B(12) status with cognitive performance in elderly people not exposed to folic acid fortification: the Hordaland Health Study.
A combination of high folate with low vitamin B12 plasma status has been associated with cognitive impairment in a population exposed to mandatory folic acid fortification. The objective of the present study was to examine the interactions between plasma concentrations of folate and vitamin B12 markers in relation to cognitive performance in Norwegian elderly who were unexposed to mandatory or voluntary folic acid fortification. Cognitive performance was assessed by six cognitive tests in 2203 individuals aged 72-74 years. A combined score was calculated using principal component analysis. The associations of folate concentrations, vitamin B12 markers (total vitamin B12, holotranscobalamin (holoTC) and methylmalonic acid (MMA)) and their interactions in relation to cognitive performance were evaluated by quantile regression and least-squares regression, adjusted for sex, education, apo-ɛ4 genotype, history of CVD/hypertension and creatinine. Cross-sectional analyses revealed an interaction (P= 0·009) between plasma concentrations of folate and vitamin B12 in relation to cognitive performance. Plasma vitamin B12 concentrations in the lowest quartile ( < 274 pmol/l) combined with plasma folate concentrations in the highest quartile (>18·5 nmol/l) were associated with a reduced risk of cognitive impairment compared with plasma concentrations in the middle quartiles of both vitamins (OR 0·22, 95 % CI 0·05, 0·92). The interaction between folate and holoTC or MMA in relation to cognitive performance was not significant. In conclusion, this large study population unexposed to mandatory folic acid fortification showed that plasma folate, but not plasma vitamin B12, was associated with cognitive performance. Among the elderly participants with vitamin B12 concentrations in the lower range, the association between plasma folate and cognitive performance was strongest.


LYSOZYME IN EPIPHYSEAL CARTILAGE : IV. Embryonic Chick Cartilage Lysozyme-Its Localization and Partial Characterization.
THE LOCALIZATION OF CHICK EMBRYONIC LYSOZYME WAS DETERMINED BY TWO TECHNIQUES: by studying the rate of release from the tissue during sequential enzymatic digestion and by immunocytochemistry. Both techniques indicate that, in this tissue, lysozyme is primarily extra-cellular. Cartilage lysozyme was isolated and partially characterized and found to be identical with egg white lysozyme in its immunologic and enzymatic behavior. In addition, a method for the isolation of large numbers of viable chondrocytes is described.


The microbiology of chronic osteomyelitis: prevalence of resistance to common empirical anti-microbial regimens.
This study describes the microbiological spectrum of chronic osteomyelitis and so guides the choice of empirical antibiotics for this condition. We performed a prospective review of a 166 prospective patient series of chronic osteomyelitis from Oxford, UK in which a standardised surgical sampling protocol was used. Staphylococcus aureus was most commonly isolated (32%) amongst a wide range of organisms including gram negative bacilli, anaerobes and coagulase negative staphylococci. Low grade pathogens were not confined to patients with a history of metalwork, a high proportion of cases were polymicrobial (29%) and culture negative cases were common (28%). No clear predictors of causative organism could be established. Many isolates were found to be resistant to commonly used empirical anti-microbial regimens. The wide range of causative organisms and degree of resistance to commonly used anti-microbials supports the importance of extensive intra-operative sampling and provides important information to guide clinicians' choice of empirical antibiotics.


Telavancin for hospital-acquired pneumonia: clinical response and 28-day survival.
U.S. Food and Drug Administration draft guidance for future antibiotic clinical trials of bacterial nosocomial pneumonia recommends the use of diagnostic criteria according to American Thoracic Society/Infectious Diseases Society of America (ATS/IDSA) guidelines and the use of a primary endpoint of 28-day all-cause mortality. The effect of applying these guidelines on outcomes of phase III nosocomial pneumonia studies of telavancin was evaluated in a post hoc analysis. ATS/IDSA criteria were applied in a blind fashion to the original all-treated (AT) group. Clinical cure rates at final follow-up were determined in the refined AT and clinically evaluable (CE) groups (ATS/IDSA-AT and ATS/IDSA-CE, respectively). The exploratory endpoint of 28-day survival was evaluated for the ATS/IDSA-AT group. Noninferiority of telavancin versus vancomycin was demonstrated, with similar cure rates in the ATS/IDSA-AT (59% versus 59%) and ATS/IDSA-CE (83% versus 80%) groups. Cure rates favored telavancin in ATS/IDSA-CE patients where Staphylococcus aureus was the sole pathogen (86% versus 75%). Overall, 28-day survival rates were similar in the telavancin (76%) and vancomycin (77%) groups but lower in telavancin-treated patients with preexisting moderate-to-severe renal impairment (creatinine clearance [CLCR] of <50 ml/min). Telavancin should be administered to patients with moderate-to-severe renal impairment only if treatment benefit outweighs the risk or if no suitable alternatives are available.


Blood compatibility of enveloped viruses.
A significant limitation to the use of viruses as systemic vectors is the susceptibility of the vector to inactivation and clearance by various blood components. Despite much focus on antibodies as the primary neutralizing molecules in blood, other mechanisms inactivate and clear virus particles from the bloodstream in both naïve and pre-immune hosts. This review provides an overview of the major blood components that interact with enveloped viruses. The mechanisms of action of these blood components by which virus particles are inactivated are also discussed. In addition, important blood components that act as barriers to the systemic delivery of therapeutic viruses are identified, and recent advances in overcoming these barriers are highlighted. Particular attention is given to the field of oncolytic virotherapy in which adequate intravenous virus delivery is critical for therapeutic success.


Determinants of compliance to antenatal micronutrient supplementation and women's perceptions of supplement use in rural Nepal.
We examined factors affecting compliance to antenatal micronutrient supplementation and women's perceptions of supplement use. Randomized controlled supplementation trial of four alternative combinations of micronutrients given during pregnancy through to 3 months postpartum. Women were visited twice weekly to monitor compliance and to replenish tablets by female study workers. At 6 weeks postpartum women with live births (n 4096) were interviewed regarding their perceptions of the supplement. Median compliance calculated as percentage of total eligible doses received by women was high (84 %). Rural southern Nepal. Pregnant women. Women with high compliance (above the median of 84 %) were likely to be older, less educated, poorer, undernourished, belong to lower caste and of Pahadi (hill) ethnicity compared with women with low compliance (at or below the median of 84 %). Smoking and drinking alcohol in the past week during pregnancy were strongly associated with low compliance. The major reason for irregular intake was forgetting to take supplements. A higher proportion of the high compliers liked taking the supplements but only half of them were willing to purchase them in the future. A large proportion of women (91 %) perceived a benefit from taking the supplement such as improved strength and health, whereas only about 10 % perceived any side-effects which were not a major barrier to compliance. The present analysis highlights that poor, undernourished, uneducated women can have high compliance to antenatal supplementation if they are supplied with the tablets and reminded to take them regularly, and counselled about side-effects.


Identification of small RNAs in Francisella tularensis.
Regulation of bacterial gene expression by small RNAs (sRNAs) have proved to be important for many biological processes. Francisella tularensis is a highly pathogenic Gram-negative bacterium that causes the disease tularaemia in humans and animals. Relatively little is known about the regulatory networks existing in this organism that allows it to survive in a wide array of environments and no sRNA regulators have been identified so far. We have used a combination of experimental assays and in silico prediction to identify sRNAs in F. tularensis strain LVS. Using a cDNA cloning and sequencing approach we have shown that F. tularensis expresses homologues of several sRNAs that are well-conserved among diverse bacteria. We have also discovered two abundant putative sRNAs that share no sequence similarity or conserved genomic context with any previously annotated regulatory transcripts. Deletion of either of these two loci led to significant changes in the expression of several mRNAs that likely include the cognate target(s) of these sRNAs. Deletion of these sRNAs did not, however, significantly alter F. tularensis growth under various stress conditions in vitro, its replication in murine cells, or its ability to induce disease in a mouse model of F. tularensis infection. We also conducted a genome-wide in silico search for intergenic loci that suggests F. tularensis encodes several other sRNAs in addition to the sRNAs found in our experimental screen. Our findings suggest that F. tularensis encodes a significant number of non-coding regulatory RNAs, including members of well conserved families of structural and housekeeping RNAs and other poorly conserved transcripts that may have evolved more recently to help F. tularensis deal with the unique and diverse set of environments with which it must contend.


Neural correlates of dyadic interaction during infancy.
This study examines the electrophysiological correlates of dyadic interaction in 14-month-old infants. Infants were presented with three conditions of live stimuli. A baseline condition involved the observation of moving geometric shapes. In a second condition the infant observed an adult performing movements that were not in reference to the infant and were not within the infant's proficient motor repertoire, such as dancing or hopping. A third condition involved face to face dyadic interactions in the context of an imitative game, where the infant and adult engaged in copying each other's hand and facial actions. Motor activity by the infant was controlled between conditions by synchronizing EEG with video frames of action. Infant EEG data was then edited offline to match in motor intensity across conditions. We expected mu rhythm (6-9 Hz) suppression during dyadic interactions relative to the amount of mu present in the baseline condition. This prediction was confirmed. We also observed suppressed mu in the interaction condition relative to the observation condition. The mu rhythm results suggest that infants at 14 months may well utilize a functioning mirroring system during dyadic interactions.


Economic trust in young children.
Mutually beneficial interactions often require trust that others will reciprocate. Such interpersonal trust is foundational to evolutionarily unique aspects of human social behaviour, such as economic exchange. In adults, interpersonal trust is often assessed using the 'trust game', in which a lender invests resources in a trustee who may or may not repay the loan. This game captures two crucial elements of economic exchange: the potential for greater mutual benefits by trusting in others, and the moral hazard that others may betray that trust. While adults across cultures can trust others, little is known about the developmental origins of this crucial cooperative ability. We developed the first version of the trust game for use with young children that addresses these two components of trust. Across three experiments, we demonstrate that 4- and 6-year-olds recognize opportunities to invest in others, sharing more when reciprocation is possible than in a context measuring pure generosity. Yet, children become better with age at investing in trustworthy over untrustworthy partners, indicating that this cooperative skill emerges later in ontogeny. Together, our results indicate that young children can engage in complex economic exchanges involving judgements about interpersonal trust and show increasing sensitivity to appropriate partners over development.


Molecular monitoring of the Leu-164 mutation of dihydrofolate reductase in a highly sulfadoxine/pyrimethamine-resistant area in Africa.
The selection of point mutation at codon 164 (from isoleucine to leucine) of the dihydrofolate reductase (DHFR) enzyme in Plasmodium falciparum is associated with high sulfadoxine /pyrimethamine (SP) resistance. Using the yeast expression system that allows the detection of dhfr allele present at low level, the presence of this mutation had previously been reported between 1998-1999 in Muheza, Tanzania, an area of high SP resistance. Eighty five P. falciparum isolates, obtained from the same area between 2002 and 2003, were analysed for the presence of Leu-164 mutation, using standard protocol based on PCR-RFLP. None of the isolates had the Leu-164 mutation.


Assessment of T cell response to novel Mycobacterium tuberculosis synthetic overlapping peptides mixtures (Rv2659 and Rv2660) and ESAT-6 in Egyptian patients.
Tuberculosis (TB) remains one of the major challenges to the global public health. The most powerful tools in any TB control program are prompt diagnosis and successful treatment of patients with active contagious disease. For almost 100 years the tuberculin skin test (TST) has been used to support the diagnosis of active and latent TB infection. The TST has a number of limitations, most notable low specificity in BCG vaccinated individuals due to cross-reactive components in PPD and the Mycobacterium bovis BCG vaccine strain and an intensive search for new and more specific diagnostic antigens has therefore been ongoing. The current diagnostic techniques utilize production of Interferon-gamma (IFN-γ) in response to novel M. tuberculosis (MTB) synthetic overlapping peptides mixtures to detect MTB infection. The aim of this study was to evaluate human immune responses to two novel Mycobacterium tuberculosis latency associated antigens Rv2659 Pepmix and Rv2660 Pepmix in comparison with ESAT-6 Pepmix. We compared the production of IFN-γ by ELISA following overnight stimulation with the antigens among the different groups of our study, TST negative healthy subjects (n = 16), TST positive healthy subjects (n = 16) and active pulmonary TB patients (n = 30). Our results showed that in TB patients, a positive IFN-γ response was observed to ESAT-6 by 73% of the donors, 47% responded to Rv2659 and 57% responded to Rv2660 when compared to TST negative controls. In conclusion, the ESAT-6 pepmix is recognized in a greater proportion of TB patients compared to Rv2659 and Rv2660, and levels of IFN-γ in response to ESAT-6 are higher than the levels observed in response to Rv2659 and Rv2660.


Fibrisoma limi gen. nov., sp. nov., a filamentous bacterium isolated from tidal flats.
An orange-pigmented, Gram-staining-negative, non-motile, filament-forming, rod-shaped bacterium (BUZ 3(T)) was isolated from a coastal mud sample from the North Sea (Fedderwardersiel, Germany) and characterized taxonomically using a polyphasic approach. According to 16S rRNA gene sequence data, it belonged to the family Cytophagaceae, exhibiting low 16S rRNA gene sequence similarity (<90 %) with members of the genera Spirosoma, Rudanella and Fibrella. The DNA G+C content was 52.0 mol%. The major fatty acids were summed feature 3 (comprising C(16 : 1)ω7c and/or iso-C(15 : 0) 2-OH), C(16 : 1)ω5c and iso-C(17 : 0) 3-OH. The major polar lipids consisted of phosphatidylethanolamine and several aminolipids. On the basis of phenotypic, chemotaxonomic and phylogenetic data, it is proposed that strain BUZ 3(T) represents a novel genus and species, for which the name Fibrisoma limi gen. nov., sp. nov. is proposed. The type strain is BUZ 3(T) ( = DSM 22564(T) = CCUG 58137(T)).


Morphological effects of mesenchymal stem cells and pulsed ultrasound on condylar growth in rats: a pilot study.
The aim of this study was to assess and describe the morphological effects of an intra-articular iniection of Mesenchymal Stem Cells (MSCs) and/or Low Intensity Pulsed Ultrasound (LIPUS) stimulation on the mandibular condyles of growing rats, using cone beam computed tomography (CBCT) and histology. Twenty-six young (23-day-old) rats were divided into 5 groups identified as LIPUS-stimulated (20 minutes daily using 50 mW/cm2, 1MHz, 0.2 millisecond pulses), MSCs injected (1 x 10(5) cells/kg), LIPUS + MSCs, medium inlected, and untreated controls. All treatments were performed in the left temporomandibular joint of each rat (TMJs). At day 21, CBCTs were obtained for cephalometric analysis and 3D reconstructions. After animal sacrifice, left and right TMJ sections were histologically prepared and examined. The Wilcoxon sign rank test and the Kruskal-Wallis 2 test were applied for statistical comparison. Imaging results showed that left condyles were wider in all LIPUS-treated groups (p < 0.05), while the LIPUS-only group had a greater left sagittal condylar length. LIPUS-treated groups displayed a lower midline shift to the right (p < 0.02). No significant differences were observed in the MSC group. Bone marrow morphology and vascularity differed between the groups as LIPUS-treated groups exhibited increased vascularity in the erosive cartilage zone. It was established that LIPUS and MSC application to the TMJ region of growing rats favoured transverse condylar growth, while LIPUS application alone may enhance sagittal condylar development.The MSC injection model had little effect on sagittal condylar growth.


Whole-tree nonstructural carbohydrate storage and seasonal dynamics in five temperate species.
Despite the importance of nonstructural carbohydrates (NSC) for growth and survival in woody plants, we know little about whole-tree NSC storage. The conventional theory suggests that NSC reserves will increase over the growing season and decrease over the dormant season. Here, we compare storage in five temperate tree species to determine the size and seasonal fluctuation of whole-tree total NSC pools as well as the contribution of individual organs. NSC concentrations in the branches, stemwood, and roots of 24 trees were measured across 12 months. We then scaled up concentrations to the whole-tree and ecosystem levels using allometric equations and forest stand inventory data. While whole-tree total NSC pools followed the conventional theory, sugar pools peaked in the dormant season and starch pools in the growing season. Seasonal depletion of total NSCs was minimal at the whole-tree level, but substantial at the organ level, particularly in branches. Surprisingly, roots were not the major storage organ as branches stored comparable amounts of starch throughout the year, and root reserves were not used to support springtime growth. Scaling up NSC concentrations to the ecosystem level, we find that commonly used, process-based ecosystem and land surface models all overpredict NSC storage.


The 2000 Centers for Disease Control and Prevention growth charts: several insights after 8 years.
This paper explores three issues related to the 2000 Centers for Disease Control and Prevention growth charts. First, it clarifies the methods that were used to create the charts as it has become apparent that the smoothing techniques have been somewhat misunderstood. The techniques included smoothing-selected percentiles between and including the 3rd and 97th percentiles and then approximating these smoothed curves using a procedure to provide the transformation parameters, lambda, mu, and sigma. Only the selected percentiles were used in this process due to small sample sizes beyond these percentiles. Second, given the concern that the infant charts were created with relatively few data points in the first few months of life, it compares the original observed percentiles with percentiles that include newly available US national data for the first few months of life. Third, it discusses the issues that arise if a 99th percentile is extrapolated based on the lambda, mu, and sigma parameters. The 99th percentile of the body mass index-for-age chart has been recommended to identify extremely obese children, yet the 97th percentile is the highest available percentile on the Centers for Disease Control and Prevention growth charts.


Mental disorders in elderly suicide.
The purpose of this study was to investigate the prevalence and comorbidity of current mental disorders defined by DSM-III-R among elderly suicide victims and to compare them with the mental disorders among younger victims. Using a psychological autopsy method, we collected comprehensive data on all suicides in Finland during 1 year. Retrospective Axis I-III consensus diagnoses were assigned to a random sample consisting of 43 victims aged 60 years or over and 186 victims aged under 60 from the nationwide suicide population. At least one Axis I diagnosis was made for 91% of the elderly victims. Major depression as the principal diagnosis was more common among the elderly victims. Almost all elderly female victims were major depressives. Psychiatric comorbidity was more common among elderly male than among elderly female victims. More of the elderly victims (88%) than the younger (36%) received Axis III diagnoses. Suicide among the elderly without a diagnosable mental disorder and somatopsychiatric comorbidity seems to be rare.


Inactivation of bacteria from contaminated streams in Limpopo, South Africa by silver- or copper-nanoparticle paper filters.
There is an urgent need for inexpensive point-of-use methods to purify drinking water in developing countries to reduce the incidence of illnesses caused by waterborne pathogens. Previously, our work showed the deactivation of laboratory-cultured bacteria by percolation through a thick paper sheet containing either silver (Ag) or copper (Cu) nanoparticles (NP). In this study, these paper filters containing AgNPs or CuNPs have been tested with water sourced from contaminated streams in Limpopo, South Africa. Following the percolation of the contaminated stream water through the metal nanoparticle (MNP) papers, the water quality of the filtered effluent was evaluated with respect to the colony counts of total coliform and E. coli bacteria, turbidity, and either silver or copper ions. Influent total coliform bacteria concentrations from the stream water in Limpopo ranged from 250 CFU/100 mL to 1,750,000 CFU/100 mL. With the less contaminated stream water (250 - 15,000 CFU/100 mL), both AgNP and CuNP papers showed complete inactivation of the coliform bacteria. With the surface water with higher coliform bacteria levels (500,000 - 1,000,000 CFU/100 mL), both the AgNP and CuNP papers showed similar results with a slightly higher bacteria reduction of log10 5.1 for the AgNP papers than the log10 4.8 reduction for the CuNP papers. E. coli results followed similar trends. For most water purification experiments, the metal release from the sheets was minimal, with values under 0.1 ppm for Ag and 1.0 ppm for Cu (the current US EPA and WHO drinking water limits for Ag and Cu, respectively). These results show good potential for the use of paper embedded with silver and/or copper nanoparticles as effective point-of-use water purifiers.


Resveratrol supplementation confers neuroprotection in cortical brain tissue of nonhuman primates fed a high-fat/sucrose diet.
Previous studies have shown positive effects of long-term resveratrol (RSV) supplementation in preventing pancreatic beta cell dysfunction, arterial stiffening and metabolic decline induced by high-fat/high-sugar (HFS) diet in nonhuman primates. Here, the analysis was extended to examine whether RSV may reduce dietary stress toxicity in the cerebral cortex of the same cohort of treated animals. Middle-aged male rhesus monkeys were fed for 2 years with HFS alone or combined with RSV, after which whole-genome microarray analysis of cerebral cortex tissue was carried out along with ELISA, immunofluorescence, and biochemical analyses to examine markers of vascular health and inflammation in the cerebral cortices. A number of genes and pathways that were differentially modulated in these dietary interventions indicated an exacerbation of neuroinflammation (e.g., oxidative stress markers, apoptosis, NF-κB activation) in HFS-fed animals and protection by RSV treatment. The decreased expression of mitochondrial aldehyde dehydrogenase 2, dysregulation in endothelial nitric oxide synthase, and reduced capillary density induced by HFS stress were rescued by RSV supplementation. Our results suggest that long-term RSV treatment confers neuroprotection against cerebral vascular dysfunction during nutrient stress.


Homeopathic medicines for adverse effects of cancer treatments.
Homeopathic medicines are used by patients with cancer, often alongside conventional treatment. Cancer treatments can cause considerable morbidity and one of the reasons patients use homeopathic medicines is to help with adverse effects. Evaluate effectiveness and safety of homeopathic medicines used to prevent or treat adverse effects of cancer treatments. The following were searched up to November 2008: Cochrane PaPaS Trials Register; Cochrane Central Register of Controlled Trials (CENTRAL); MEDLINE; EMBASE; CINAHL; BNI; CancerLIT; AMED; CISCOM; Hom-Inform; SIGLE; National Research Register; Zetoc; www.controlled-trials.com; http://clinicaltrials.gov; Liga Medicorum Homeopathica Internationalis (LMHI, Liga) conference proceedings; reference lists of relevant studies were checked; and homeopathic manufacturers, leading researchers and practitioners were contacted. Randomised controlled trials (RCTs) of homeopathic medicines in participants with a clinical or histological diagnosis of cancer where the intervention was aimed at preventing or treating symptoms associated with cancer treatments. All age groups, and all stages of disease were included. Two review authors independently assessed studies for inclusion and two review authors extracted data. Three review authors independently assessed trial quality using the Delphi List and the Cochrane Collaboration's tool for assessing risk of bias. Disagreements were resolved by consensus. Where available, data were extracted for analysis. Eight controlled trials (seven placebo controlled and one trial against an active treatment) with a total of 664 participants met the inclusion criteria. Three studied adverse effects of radiotherapy, three studied adverse effects of chemotherapy and two studied menopausal symptoms associated with breast cancer treatment.Two studies with low risk of bias demonstrated benefit: one with 254 participants demonstrated superiority of topical calendula over trolamine (a topical agent not containing corticosteroids) for prevention of radiotherapy-induced dermatitis, and another with 32 participants demonstrated superiority of Traumeel S (a proprietary complex homeopathic medicine) over placebo as a mouthwash for chemotherapy-induced stomatitis. Two other studies reported positive results, although the risk of bias was unclear, and four further studies reported negative results.No serious adverse effects or interactions were reported attributable to the homeopathic medicines used. This review found preliminary data in support of the efficacy of topical calendula for prophylaxis of acute dermatitis during radiotherapy and Traumeel S mouthwash in the treatment of chemotherapy-induced stomatitis. These trials need replicating. There is no convincing evidence for the efficacy of homeopathic medicines for other adverse effects of cancer treatments. Further research is required.


Metal-based imaging agents: progress towards interrogating neurodegenerative disease.
Central nervous system (CNS) neurodegeneration is defined by a complex series of pathological processes that ultimately lead to death. The precise etiology of these disorders remains unknown. Recent efforts show that a mechanistic understanding of the malfunctions underpinning disease progression will prove requisite in developing new treatments and cures. Transition metals and lanthanide ions display unique characteristics (i.e., magnetism, radioactivity, and luminescence), often with biological relevance, allowing for direct application in CNS focused imaging modalities. These techniques include positron emission tomography (PET), single-photon emission computed tomography (SPECT), magnetic resonance imaging (MRI), and luminescent-based imaging (LumI). In this Tutorial Review, we have aimed to highlight the various metal-based imaging techniques developed in the effort to understand the pathophysiological processes associated with neurodegeneration. Each section has been divided so as to include an introduction to the particular imaging technique in question. This is then followed by a summary of key demonstrations that have enabled visualization of a specific neuropathological biomarker. These strategies have either exploited the high binding affinity of a receptor for its corresponding biomarker or a specific molecular transformation caused by a target species, all of which produce a concomitant change in diagnostic signal. Advantages and disadvantages of each method with perspectives on the utility of molecular imaging agents for understanding the complexities of neurodegenerative disease are discussed.


Depressed Colorectal Cancer: A New Paradigm in Early Colorectal Cancer.
In contrast to most colorectal carcinomas arising from pedunculated or sessile protruded adenomas, submucosal-invasive (pT1) colorectal carcinoma exhibiting a depressed surface (hereinafter, "depressed colorectal carcinoma," identified by means of high-definition endoscopy) is considered to be derived from depressed precursors. We hypothesized that depressed colorectal neoplasms have unique clinicopathological features different that are different from those of protruded and flat colorectal neoplasms. We classified 27,129 colorectal neoplasms (909 pT1 carcinomas and 26,220 adenomas) resected between 2001 and 2017 into depressed (211 carcinomas and 109 adenomas), flat (304 carcinomas and 11,246 adenomas), and protruded subtypes (394 carcinomas and 14,865 adenomas) and compared their clinicopathological features. As exploratory analyses of pT1 carcinomas, we conducted whole-exome sequencing for 19 depressed and 8 protruded subtypes and RNA sequencing for 8 depressed and 8 protruded subtypes. pT1 carcinomas were more common in depressed lesions (66%) than in protruded (2.6%) and flat lesions (2.6%) (P < 0.001). Compared with nondepressed pT1 carcinomas, depressed pT1 carcinomas were positively correlated with lymphovascular invasion, tumor budding, and massive submucosal invasion and inversely correlated with the presence of an adenoma component (all P < 0.001). Depressed adenomas were more likely to contain high-grade dysplasia than nondepressed adenomas (49% vs 11%, P < 0.001). A KRAS mutation was observed only in one of the 19 depressed pT1 carcinomas. Relative to protruded carcinomas, depressed carcinomas generally exhibited higher expression of genes related to angiogenesis and epithelial-mesenchymal transition. Depressed colorectal neoplasms may harbor a unique combination of malignant histopathological phenotypes and molecular features.


Cohort studies around the world: methodologies, research questions and integration to address the emerging global epidemic of chronic diseases.
With ageing populations, increased economic prosperity and the ensuing lifestyle changes, there has been a dramatic increase in the burden of chronic non-communicable diseases in countries of the developing world. The distribution of risk factors for chronic diseases among populations in developing countries has traditionally been very different from that in their Western counterparts, thus resulting in considerable variation in disease distribution in these settings. However, with the increase in globalization along with rapid advancements in technology, many developing countries are now faced with the challenge of a dual disease burden, battling existing communicable infectious diseases as well as the emerging epidemic of non-communicable chronic diseases. This paper highlights the need for multiple cohort studies on chronic diseases around the world, and explores some of the challenges in establishing and maintaining these studies in resource-constrained settings.


Rheumatoid arthritis and tabetic arthropathy: physiopathological link or simple association?
To the best of our knowledge, the association of rheumatoid arthritis and tabetic arthropathy has never been described before in the literature. We report here a first observation. We report the case of a 50-year-old man, treated for syphilitic arthritis evolving for 4 years, who presented with a table of rheumatoid arthritis. The diagnosis of rheumatoid arthritis was established according to the 2010 American College of Rheumatology/European League Against Rheumatism criteria (ACR/EULAR). The treatment was based on weekly injection of methotrexate and a symptomatic treatment by corticosteroid. The association of rheumatoid arthritis and tabetic arthropathy is rare, to our knowledge this is the first case reported. This case reminds us that a neuropathic arthropathy as tabetic arthropathy, although it is rare, can be associated in a sporadic or exceptional way with other rheumatic disease like rheumatoid arthritis. A physiopathological link between the both diseases remains to be proved.


Ethical dilemmas concerning autonomy when persons with dementia wish to live at home: a qualitative, hermeneutic study.
Caring for people with dementia living in their own homes is a challenging care issue that raises ethical dilemmas of how to balance autonomy with their safety and well-being. The theoretical framework for this study consisted of the concepts of autonomy, beneficence, non-maleficence, paternalism and from the ethics of care. The aim of this study was to explore ethical dilemmas concerning autonomy that were identified when persons with dementia wished to live at home. This Norwegian study had a qualitative, hermeneutic design and was based on nine cases. Each case consisted of of a triad: the person with dementia, the family carer and the professional caregiver. Inclusion criteria for the persons with dementia were: (1) 67 years or older (2) diagnosed with dementia (3) Clinical Dementia Rating score 2 i.e. dementia of moderate degree (4) able to communicate verbally and (5) expressed a wish to live at home. The family carers and professional caregivers registered in the patients' records were included in the study. An interview guide was used in interviews with family carers and professional caregivers. Field notes were written after participant observation of interactions between persons with dementia and professional caregivers during morning care or activities at a day care centre. By means of deductive analysis, autonomy-related ethical dilemmas were identified. The final interpretation was based on perspectives from the theoretical framework. The analysis revealed three main ethical dilemmas: When the autonomy of the person with dementia conflicted with (1) the family carer's and professional caregiver's need to prevent harm (non-maleficence) (2) the beneficence of family carers and professional caregivers (3) the autonomy of the family carer. In order to remain living in their own homes, people with dementia accepted their dependence on others in order to uphold their actual autonomy and live in accordance with their identified values. Paternalism could be justified in light of beneficence and non-maleficence and within an ethics of care.


A four-helix bundle DNA nanostructure with binding pockets for pyrimidine nucleotides.
Designed DNA nanostructures of impressive size have been described, but designed structures of the size of protein enzymes that bind organic ligands with high specificity are rare. Here we report a four-helix motif consisting of three synthetic strands with 65 base pairs and 165 nucleotides in total that folds well. Furthermore, we show that in the interior of this small folded DNA nanostructure, cavities can be set up that bind pyrimidine nucleotides with micromolar affinity. Base-specific binding for both thymidine and cytidine derivatives is demonstrated. The binding affinity depends on the position in the structure, as expected for recognition beyond simple base pairing. The folding motif reported here can help to expand DNA nanotechnology into the realm of selective molecular recognition that is currently dominated by protein-based enzymes and receptors.