The Role of Hand1 in the Development of the Lateral Plate Mesoderm in Xenopus

The transcription factor hand1 is expressed in the heart, lateral plate mesoderm (LPM) and neural crest cells during development. As Hand1-null mice die early in embryogenesis, identification of the precise role of Hand1 in development is difficult. In Xenopus, we observed that hand1 expression patterns correlates very closely with development of LPM derivatives, leading us to hypothesize that hand1 is required for normal LPM development. Using hand1 knockdown and overexpression models in Xenopus, development of LPM derivatives were assessed by whole mount in situ hybridization. I found that hand1 is required for proper heart morphogenesis. Furthermore, hand1 is required for the formation of a complex vasculature plexus within embryos, by maintaining early endothelial cell populations however Hand1 is not sufficient to induce endothelial cell differentiation. These findings confirm a conserved role of Hand1 in heart morphogenesis and suggest a new role for Hand1 in development of the vasculature plexus

An experimental model for calcium carbonate urolithiasis in goats

This article is also available at the journal website: https://onlinelibrary.wiley.com/doi/full/10.1111/jvim.15061Background: Calcium carbonate is a common urolith type in small ruminants with no high-yield experimental model to evaluate animal susceptibility or preventative measure response. Hypothesis: That novel plastic winged implants would allow accumulation and quantification of calcium carbonate calculus formation in goats on a high-calcium diet and identify individual variation between goats in the mass of calculi produced. Animals: Eight nonpregnant 3- and 4-year-old Boer-cross does, weighing 22.3–39.5 kg, determined to be healthy based on physical examination, were used in these experiments. Methods: Prospective cohort study for in vivo experimental model development. Implants were placed into the urinary bladder lumen in 8 goats over 2 evaluation periods. The alfalfa-based ration had a total ration Ca : P of 3.29 and 3.84 : 1, respectively. Urine was collected at 0, 28, 56, and 84 days in the 1st experiment; blood and urine at those timepoints in the 2nd experiment. For each evaluation period, the implants were removed 84 days after implantation and weighed. Accumulated calculi mass was calculated and compared between goats and was analyzed for composition. Results: Implant retention was 100% and 86% in the 2 studies. All goats with retained implants accumulated calcium carbonate at a mean implant gain per day across studies ranging from 0.44 to 57.45 mg. Two goats accumulated (0.44–7.65 mg/day and 33.64 & 57.45 mg/day) significantly more urolith material than the cohort across both studies (P5.047). No routine analytes on blood or urine were found to be explanatory for the difference observed. Conclusions and Clinical Importance: These findings form a basis for implant and diet selection for use in future studies of urolithiasis development and for studies regarding individual susceptibility to urolithiasis. KEYWORDS 3D printing, calculogenesis, urinary calculi, urolithFunding information: Department of Large Animal Clinical Sciences, Texas A&M University College of Veterinary Medicin

Farnesol Induces Hydrogen Peroxide Resistance in Candida albicans Yeast by Inhibiting the Ras-Cyclic AMP Signaling Pathway

Farnesol, a Candida albicans cell-cell signaling molecule that participates in the control of morphology, has an additional role in protection of the fungus against oxidative stress. In this report, we show that although farnesol induces the accumulation of intracellular reactive oxygen species (ROS), ROS generation is not necessary for the induction of catalase (Cat1)-mediated oxidative-stress resistance. Two antioxidants, α-tocopherol and, to a lesser extent, ascorbic acid effectively reduced intracellular ROS generation by farnesol but did not alter farnesol-induced oxidative-stress resistance. Farnesol inhibits the Ras1-adenylate cyclase (Cyr1) signaling pathway to achieve its effects on morphology under hypha-inducing conditions, and we demonstrate that farnesol induces oxidative-stress resistance by a similar mechanism. Strains lacking either Ras1 or Cyr1 no longer exhibited increased protection against hydrogen peroxide upon preincubation with farnesol. While we also observed the previously reported increase in the phosphorylation level of Hog1, a known regulator of oxidative-stress resistance, in the presence of farnesol, the hog1/hog1 mutant did not differ from wild-type strains in terms of farnesol-induced oxidative-stress resistance. Analysis of Hog1 levels and its phosphorylation states in different mutant backgrounds indicated that mutation of the components of the Ras1-adenylate cyclase pathway was sufficient to cause an increase of Hog1 phosphorylation even in the absence of farnesol or other exogenous sources of oxidative stress. This finding indicates the presence of unknown links between these signaling pathways. Our results suggest that farnesol effects on the Ras-adenylate cyclase cascade are responsible for many of the observed activities of this fungal signaling molecule

Farnesol and Cyclic AMP Signaling Effects on the Hypha-to-Yeast Transition in Candida Albicans

Candida albicans, a fungal pathogen of humans, regulates its morphology in response to many environmental cues and this morphological plasticity contributes to virulence. Farnesol, an autoregulatory molecule produced by C. albicans, inhibits the induction of hyphal growth by inhibiting adenylate cyclase (Cyr1). The role of farnesol and Cyr1 in controlling the maintenance of hyphal growth has been less clear. Here, we demonstrate that preformed hyphae transition to growth as yeast in response to farnesol and that strains with increased cyclic AMP (cAMP) signaling exhibit more resistance to farnesol. Exogenous farnesol did not induce the hypha-to-yeast transition in mutants lacking the Tup1 or Nrg1 transcriptional repressors in embedded conditions. Although body temperature is not required for embedded hyphal growth, we found that the effect of farnesol on the hypha-to-yeast transition varies inversely with temperature. Our model of Cyr1 activity being required for filamentation is also supported by our liquid assay data, which show increased yeast formation when preformed filaments are treated with farnesol. Together, these data suggest that farnesol can modulate morphology in preformed hyphal cells and that the repression of hyphal growth maintenance likely occurs through the inhibition of cAMP signaling

CRISPR provides acquired resistance against viruses in prokaryotes

Clustered regularly interspaced short palindromic repeats (CRISPR) are a distinctive feature of the genomes of most Bacteria and Archaea and are thought to be involved in resistance to bacteriophage. We found that following viral challenge, bacteria integrated new spacers derived from phage genomic sequences. Removal or addition of particular spacers modified the phage-resistance phenotype of the cell. Thus, CRISPR, together with associated cas genes, provided resistance against phages, whereby specificity is determined by spacer/phage sequence similarity

Allosteric inhibition of a stem cell RNA-binding protein by an intermediary metabolite

Gene expression and metabolism are coupled at numerous levels. Cells must sense and respond to nutrients in their environment, and specialized cells must synthesize metabolic products required for their function. Pluripotent stem cells have the ability to differentiate into a wide variety of specialized cells. How metabolic state contributes to stem cell differentiation is not understood. In this study, we show that RNA-binding by the stem cell translation regulator Musashi-1 (MSI1) is allosterically inhibited by 18-22 carbon omega-9 monounsaturated fatty acids. The fatty acid binds to the N-terminal RNA Recognition Motif (RRM) and induces a conformational change that prevents RNA association. Musashi proteins are critical for development of the brain, blood, and epithelium. We identify stearoyl-CoA desaturase-1 as a MSI1 target, revealing a feedback loop between omega-9 fatty acid biosynthesis and MSI1 activity. We propose that other RRM proteins could act as metabolite sensors to couple gene expression changes to physiological state

The Molecular Basis of Drug Resistance against Hepatitis C Virus NS3/4A Protease Inhibitors

Hepatitis C virus (HCV) infects over 170 million people worldwide and is the leading cause of chronic liver diseases, including cirrhosis, liver failure, and liver cancer. Available antiviral therapies cause severe side effects and are effective only for a subset of patients, though treatment outcomes have recently been improved by the combination therapy now including boceprevir and telaprevir, which inhibit the viral NS3/4A protease. Despite extensive efforts to develop more potent next-generation protease inhibitors, however, the long-term efficacy of this drug class is challenged by the rapid emergence of resistance. Single-site mutations at protease residues R155, A156 and D168 confer resistance to nearly all inhibitors in clinical development. Thus, developing the next-generation of drugs that retain activity against a broader spectrum of resistant viral variants requires a comprehensive understanding of the molecular basis of drug resistance. In this study, 16 high-resolution crystal structures of four representative protease inhibitors - telaprevir, danoprevir, vaniprevir and MK-5172 - in complex with the wild-type protease and three major drug-resistant variants R155K, A156T and D168A, reveal unique molecular underpinnings of resistance to each drug. The drugs exhibit differential susceptibilities to these protease variants in both enzymatic and antiviral assays. Telaprevir, danoprevir and vaniprevir interact directly with sites that confer resistance upon mutation, while MK-5172 interacts in a unique conformation with the catalytic triad. This novel mode of MK-5172 binding explains its retained potency against two multi-drug-resistant variants, R155K and D168A. These findings define the molecular basis of HCV N3/4A protease inhibitor resistance and provide potential strategies for designing robust therapies against this rapidly evolving virus

Staff training in physical interventions: a literature review

BackgroundRestrictive practices are used frequently by frontline staff in a variety of care contexts, including psychiatric hospitals, children’s services, and support services for older adults and individuals with intellectual and developmental disabilities. Physical restraint has been associated with emotional harm, physical injury to staff and consumers, and has even resulted in death of individuals in care environments. Various interventions have been implemented within care settings with the intention of reducing instances of restraint. One of the most common interventions is staff training that includes some physical intervention skills to support staff to manage crisis situations. Despite physical intervention training being used widely in care services, there is little evidence to support the effectiveness and application of physical interventions. This review will examine the literature regarding outcomes of staff training in physical interventions across care sectors.MethodA systematic search was conducted following PRISMA guidelines using Cochrane Database, Medline EBSCO, Medline OVID, PsychINFO, and the Web of Science. Main search keywords were staff training, physical intervention, physical restraint. The MMAT was utilised to provide an analytical framework for the included studies.Results and discussionSeventeen articles have been included in this literature review. The included studies take place in a range of care settings and comprise a wide range of outcomes and designs. The training programmes examined vary widely in their duration, course content, teaching methods, and extent to which physical skills are taught. Studies were of relatively poor quality. Many descriptions of training programmes did not clearly operationalise the knowledge and skills taught to staff. As such, it is difficult to compare course content across the studies. Few papers described physical interventions in sufficient detail. This review demonstrates that, although staff training is a ‘first response’ to managing health and safety in care settings, there is very little evidence to suggest that staff training in physical intervention skills leads to meaningful outcomes

Structural basis for CRISPR RNA-guided DNA recognition by Cascade

The CRISPR (clustered regularly interspaced short palindromic repeats) immune system in prokaryotes uses small guide RNAs to neutralize invading viruses and plasmids. In Escherichia coli, immunity depends on a ribonucleoprotein complex called Cascade. Here we present the composition and low-resolution structure of Cascade and show how it recognizes double-stranded DNA (dsDNA) targets in a sequence-specific manner. Cascade is a 405-kDa complex comprising five functionally essential CRISPR-associated (Cas) proteins (CasA1B2C6D1E1) and a 61-nucleotide CRISPR RNA (crRNA) with 5′-hydroxyl and 2′,3′-cyclic phosphate termini. The crRNA guides Cascade to dsDNA target sequences by forming base pairs with the complementary DNA strand while displacing the noncomplementary strand to form an R-loop. Cascade recognizes target DNA without consuming ATP, which suggests that continuous invader DNA surveillance takes place without energy investment. The structure of Cascade shows an unusual seahorse shape that undergoes conformational changes when it binds target DNA.

Games for health for children - current status and needed research

Videogames for health (G4H) offer exciting, innovative, potentially highly effective methods for increasing knowledge, delivering persuasive messages, changing behaviors, and influencing health outcomes. Although early outcome results are promising, additional research is needed to determine the game design and behavior change procedures that best promote G4H effectiveness and to identify and minimize possible adverse effects. Guidelines for ideal use of different types of G4H by children and adolescents should be elucidated to enhance effectiveness and minimize adverse effects. G4H stakeholders include organizational implementers, policy makers, players and their families, researchers, designers, retailers, and publishers. All stakeholders should be involved in G4H development and have a voice in setting goals to capitalize on their insights to enhance effectiveness and use of the game. In the future, multiple targeted G4H should be available to meet a population's diverse health needs in developmentally appropriate ways. Substantial, consistent, and sophisticated research with appropriate levels of funding is needed to realize the benefits of G4H