Elastin is Localised to the Interfascicular Matrix of Energy Storing Tendons and Becomes Increasingly Disorganised With Ageing

Tendon is composed of fascicles bound together by the interfascicular matrix (IFM). Energy storing tendons are more elastic and extensible than positional tendons; behaviour provided by specialisation of the IFM to enable repeated interfascicular sliding and recoil. With ageing, the IFM becomes stiffer and less fatigue resistant, potentially explaining why older tendons become more injury-prone. Recent data indicates enrichment of elastin within the IFM, but this has yet to be quantified. We hypothesised that elastin is more prevalent in energy storing than positional tendons, and is mainly localised to the IFM. Further, we hypothesised that elastin becomes disorganised and fragmented, and decreases in amount with ageing, especially in energy storing tendons. Biochemical analyses and immunohistochemical techniques were used to determine elastin content and organisation, in young and old equine energy storing and positional tendons. Supporting the hypothesis, elastin localises to the IFM of energy storing tendons, reducing in quantity and becoming more disorganised with ageing. These changes may contribute to the increased injury risk in aged energy storing tendons. Full understanding of the processes leading to loss of elastin and its disorganisation with ageing may aid in the development of treatments to prevent age related tendinopathy

Why don't hospital staff activate the rapid response system (RRS)? How frequently is it needed and can the process be improved?

Abstract Background The rapid response system (RRS) is a process of accessing help for health professionals when a patient under their care becomes severely ill. Recent studies and meta-analyses show a reduction in cardiac arrests by a one-third in hospitals that have introduced a rapid response team, although the effect on overall hospital mortality is less clear. It has been suggested that the difficulty in establishing the benefit of the RRS has been due to implementation difficulties and a reluctance of clinical staff to call for additional help. This assertion is supported by the observation that patients continue to have poor outcomes in our institution despite an established RRS being available. In many of these cases, the patient is often unstable for many hours or days without help being sought. These poor outcomes are often discovered in an ad hoc fashion, and the real numbers of patients who may benefit from the RRS is currently unknown. This study has been designed to answer three key questions to improve the RRS: estimate the scope of the problem in terms of numbers of patients requiring activation of the RRS; determine cognitive and socio-cultural barriers to calling the Rapid Response Team; and design and implement solutions to address the effectiveness of the RRS. Methods The extent of the problem will be addressed by establishing the incidence of patients who meet abnormal physiological criteria, as determined from a point prevalence investigation conducted across four hospitals. Follow-up review will determine if these patients subsequently require intensive care unit or critical care intervention. This study will be grounded in both cognitive and socio-cultural theoretical frameworks. The cognitive model of situation awareness will be used to determine psychological barriers to RRS activation, and socio-cultural models of interprofessional practice will be triangulated to inform further investigation. A multi-modal approach will be taken using reviews of clinical notes, structured interviews, and focus groups. Interventions will be designed using a human factors analysis approach. Ongoing surveillance of adverse outcomes and surveys of the safety climate in the clinical areas piloting the interventions will occur before and after implementation

Protocol for a randomised controlled trial investigating self-help email messages for sub-threshold depression: the Mood Memos study

<p>Abstract</p> <p>Background</p> <p>Sub-threshold depression is common, impairs functioning, and increases the risk of developing major depression. Although psychological treatments have been investigated for sub-threshold depression, they are costly. A less costly alternative could be an educational health promotion campaign about effective self-help for depression symptoms. The aim of the study is to test the efficacy of a low-cost email-based mental health promotion campaign in changing self-help behaviour and preventing more severe depression in adults with sub-threshold depression.</p> <p>Methods/Design</p> <p>The project is a randomised controlled trial of an automated preventive email-intervention aimed at people with sub-threshold depression. Adults aged 18+ with sub-threshold depression (as measured with the Patient Health Questionnaire-9), who are not already receiving professional treatment for depression, are eligible for admission to the study. Internet users will sign up via the study website <url>http://www.moodmemos.com</url> and be randomly allocated to receive emails twice weekly for six weeks containing either self-help coping advice or general information about depression as a control. Outcomes will be assessed at the start, midpoint, and end of the intervention, as well as six months later. Outcomes assessed include symptoms, incidence of major depression, psychological distress, social and occupational functioning, coping strategies, and coping self-efficacy. The primary hypothesis is that the Mood Memo emails containing coping strategies will reduce depression symptoms and be better at preventing major depression than the control emails that contain general information about depression.</p> <p>Discussion</p> <p>Promotion of actions an individual can take to prevent physical disease is a technique often used in public health. This study applies this approach to mental health, and explores whether a low-cost, easily disseminated email-based campaign can improve self-help coping behaviour and prevent depression in adults with sub-threshold depression.</p> <p>Trial Registration</p> <p>Australia and New Zealand Clinical Trials Register (ANZCTR): <a href="http://www.anzctr.org.au/ACTRN12609000925246.aspx">ACTRN12609000925246</a></p

Rapid assembly of customized TALENs into multiple

Transcriptional activator-like effector nucleases (TALENs) have become a powerful tool for genome editing. Here we present an efficient TALEN assembly approach in which TALENs are assembled by direct Golden Gate ligation into Gateway® Entry vectors from a repeat variable di-residue (RVD) plasmid array. We constructed TALEN pairs targeted to mouse Ddx3 subfamily genes, and demonstrated that our modified TALEN assembly approach efficiently generates accurate TALEN moieties that effectively introduce mutations into target genes. We generated "user friendly" TALEN Entry vectors containing TALEN expression cassettes with fluorescent reporter genes that can be efficiently transferred via Gateway (LR) recombination into different delivery systems. We demonstrated that the TALEN Entry vectors can be easily transferred to an adenoviral delivery system to expand application to cells that are difficult to transfect. Since TALENs work in pairs, we also generated a TALEN Entry vector set that combines a TALEN pair into one PiggyBac transposon-based destination vector. The approach described here can also be modified for construction of TALE transcriptional activators, repressors or other functional domains. © 2013 Zhang et al

The Effect of Sensory Uncertainty Due to Amblyopia (Lazy Eye) on the Planning and Execution of Visually-Guided 3D Reaching Movements

Background: Impairment of spatiotemporal visual processing in amblyopia has been studied extensively, but its effects on visuomotor tasks have rarely been examined. Here, we investigate how visual deficits in amblyopia affect motor planning and online control of visually-guided, unconstrained reaching movements. Methods: Thirteen patients with mild amblyopia, 13 with severe amblyopia and 13 visually-normal participants were recruited. Participants reached and touched a visual target during binocular and monocular viewing. Motor planning was assessed by examining spatial variability of the trajectory at 50–100 ms after movement onset. Online control was assessed by examining the endpoint variability and by calculating the coefficient of determination (R 2) which correlates the spatial position of the limb during the movement to endpoint position. Results: Patients with amblyopia had reduced precision of the motor plan in all viewing conditions as evidenced by increased variability of the reach early in the trajectory. Endpoint precision was comparable between patients with mild amblyopia and control participants. Patients with severe amblyopia had reduced endpoint precision along azimuth and elevation during amblyopic eye viewing only, and along the depth axis in all viewing conditions. In addition, they had significantly higher R 2 values at 70 % of movement time along the elevation and depth axes during amblyopic eye viewing. Conclusion: Sensory uncertainty due to amblyopia leads to reduced precision of the motor plan. The ability to implemen

N-Acetylglucosamine Induces White to Opaque Switching, a Mating Prerequisite in Candida albicans

To mate, the fungal pathogen Candida albicans must undergo homozygosis at the mating-type locus and then switch from the white to opaque phenotype. Paradoxically, opaque cells were found to be unstable at physiological temperature, suggesting that mating had little chance of occurring in the host, the main niche of C. albicans. Recently, however, it was demonstrated that high levels of CO2, equivalent to those found in the host gastrointestinal tract and select tissues, induced the white to opaque switch at physiological temperature, providing a possible resolution to the paradox. Here, we demonstrate that a second signal, N-acetylglucosamine (GlcNAc), a monosaccharide produced primarily by gastrointestinal tract bacteria, also serves as a potent inducer of white to opaque switching and functions primarily through the Ras1/cAMP pathway and phosphorylated Wor1, the gene product of the master switch locus. Our results therefore suggest that signals produced by bacterial co-members of the gastrointestinal tract microbiota regulate switching and therefore mating of C. albicans

Single-nucleotide polymorphism discovery by high-throughput sequencing in sorghum

<p>Abstract</p> <p>Background</p> <p>Eight diverse sorghum (<it>Sorghum bicolor </it>L. Moench) accessions were subjected to short-read genome sequencing to characterize the distribution of single-nucleotide polymorphisms (SNPs). Two strategies were used for DNA library preparation. Missing SNP genotype data were imputed by local haplotype comparison. The effect of library type and genomic diversity on SNP discovery and imputation are evaluated.</p> <p>Results</p> <p>Alignment of eight genome equivalents (6 Gb) to the public reference genome revealed 283,000 SNPs at ≥82% confirmation probability. Sequencing from libraries constructed to limit sequencing to start at defined restriction sites led to genotyping 10-fold more SNPs in all 8 accessions, and correctly imputing 11% more missing data, than from semirandom libraries. The SNP yield advantage of the reduced-representation method was less than expected, since up to one fifth of reads started at noncanonical restriction sites and up to one third of restriction sites predicted <it>in silico </it>to yield unique alignments were not sampled at near-saturation. For imputation accuracy, the availability of a genomically similar accession in the germplasm panel was more important than panel size or sequencing coverage.</p> <p>Conclusions</p> <p>A sequence quantity of 3 million 50-base reads per accession using a <it>Bsr</it>FI library would conservatively provide satisfactory genotyping of 96,000 sorghum SNPs. For most reliable SNP-genotype imputation in shallowly sequenced genomes, germplasm panels should consist of pairs or groups of genomically similar entries. These results may help in designing strategies for economical genotyping-by-sequencing of large numbers of plant accessions.</p

HPV16 oncogene expression levels during early cervical carcinogenesis are determined by the balance of epigenetic chromatin modifications at the integrated virus genome.

In cervical squamous cell carcinomas, high-risk human papillomavirus (HRHPV) DNA is usually integrated into host chromosomes. Multiple integration events are thought to be present within the cells of a polyclonal premalignant lesion and the features that underpin clonal selection of one particular integrant remain poorly understood. We previously used the W12 model system to generate a panel of cervical keratinocyte clones, derived from cells of a low-grade premalignant lesion naturally infected with the major HRHPV type, HPV16. The cells were isolated regardless of their selective advantage and differed only by the site of HPV16 integration into the host genome. We used this resource to test the hypothesis that levels of HPV16 E6/E7 oncogene expression in premalignant cells are regulated epigenetically. We performed a comprehensive analysis of the epigenetic landscape of the integrated HPV16 DNA in selected clones, in which levels of virus oncogene expression per DNA template varied ~6.6-fold. Across the cells examined, higher levels of virus expression per template were associated with more open chromatin at the HPV16 long control region, together with greater loading of chromatin remodelling enzymes and lower nucleosome occupancy. There were higher levels of histone post-translational modification hallmarks of transcriptionally active chromatin and lower levels of repressive hallmarks. There was greater abundance of the active/elongating form of the RNA polymerase-II enzyme (RNAPII-Ser2P), together with CDK9, the component of positive transcription elongation factor b complex responsible for Ser2 phosphorylation. The changes observed were functionally significant, as cells with higher HPV16 expression per template showed greater sensitivity to depletion and/or inhibition of histone acetyltransferases and CDK9 and less sensitivity to histone deacetylase inhibition. We conclude that virus gene expression per template following HPV16 integration is determined through multiple layers of epigenetic regulation, which are likely to contribute to selection of individual cells during cervical carcinogenesis.This work was supported by Cancer Research UK (Programme Grant A13080); the Medical Research Council; The Pathological Society of Great Britain and Ireland (E.L.A.K.); and the Agency for Science, Technology and Research, Singapore (Q.Y.A).This is the final version of the article. It first appeared from Nature Publishing Group via http://dx.doi.org/10.1038/onc.2016.

The Set3/Hos2 Histone Deacetylase Complex Attenuates cAMP/PKA Signaling to Regulate Morphogenesis and Virulence of Candida albicans

Candida albicans, like other pleiomorphic fungal pathogens, is able to undergo a reversible transition between single yeast-like cells and multicellular filaments. This morphogenetic process has long been considered as a key fungal virulence factor. Here, we identify the evolutionarily conserved Set3/Hos2 histone deacetylase complex (Set3C) as a crucial repressor of the yeast-to-filament transition. Cells lacking core components of the Set3C are able to maintain all developmental phases, but are hypersusceptible to filamentation-inducing signals, because of a hyperactive cAMP/Protein Kinase A signaling pathway. Strikingly, Set3C-mediated control of filamentation is required for virulence in vivo, since set3Δ/Δ cells display strongly attenuated virulence in a mouse model of systemic infection. Importantly, the inhibition of histone deacetylase activity by trichostatin A exclusively phenocopies the absence of a functional Set3C, but not of any other histone deacetylase gene. Hence, our work supports a paradigm for manipulating morphogenesis in C. albicans through alternative antifungal therapeutic strategies