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The discovery of biomarkers, essential for successful drug development, is often hindered by the limited availability of tissue samples, typically obtained through core needle biopsies. Standard 'omics platforms can consume significant amounts of tissue, forcing scientist to trade off spatial context for high-plex assays, such as genome-wide assays. While bulk gene expression approaches and standard single-cell transcriptomics have been valuable in defining various molecular and cellular mechanisms, they do not retain spatial context. As such, they have limited power in resolving tissue heterogeneity and cell-cell interactions. Current spatial transcriptomics platforms offer limited transcriptome coverage and have low throughput, restricting the number of samples that can be analyzed daily or even weekly. While the Digital Spatial Profiling (DSP) method does not provide single-cell resolution, it presents a significant advancement by enabling scalable whole transcriptome and ultrahigh-plex protein analysis from distinct tissue compartments and structures using a single tissue slide. These capabilities overcome significant constraints in biomarker analysis in solid tissue specimens. These advancements in tissue profiling play a crucial role in deepening our understanding of disease biology and in identifying potential therapeutic targets and biomarkers. To enhance the use of spatial biology tools in drug discovery and development, the DSP Scientific Consortium has created best practices guidelines. These guidelines, built on digital spatial profiling data and expertise, offer a practical framework for designing spatial studies and using current and future spatial biology platforms. The aim is to improve tissue analysis in all research areas supporting drug discovery and development.

The use of resilience in social practice has evolved from a theoretical framework at the intersection between individuals and their social ecology. Critics argue this theory still results in policies and practices that are too individualised, with the potential for negative social consequences. This paper further critiques contemporary understanding of resilience theory and its application. It juxtaposes complex systems theory with a social inequalities oriented resilience practice. This provides a paradoxical approach. It is acknowledged that state and public policy decisions and actions can be anti-resilient, undermining community and social resilience that already exists in the form of social relationships, self-organisation and co-production. Nevertheless, collective social resilience also illustrates the potential of local and service user organisations to contribute to an overall transformational change process., Competing Interests: Declaration of conflicting interestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Automated delivery of therapy in virtual reality (VR) has the potential to be used for smoking cessation. Most obviously, it could be used to practise and establish alternative reactions to smoking cues. The first step in treatment development is to show that VR environments can trigger sufficient cravings in smokers. We evaluated a new VR public house outdoor scenario with 100 individuals who smoked daily. Participants were randomly assigned to the VR scenario with smoking cues or a neutral experience in VR. The VR experiences were presented in a standalone VR headset. Before and after VR, we collected self-reported craving scores for cigarettes and alcohol using the Tobacco Craving Questionnaire (TCQ) and visual analogue scales (VAS). Physiological data were also collected. Compared to the neutral condition, exposure to the smoking cues led to a large increase in craving for a cigarette (TCQ β = 11.44, p < 0.0001, Cohen's d = 1.10) and also a moderate increase in craving for alcohol ( β = 0.7 , p = 0.017 , d = 0.50 ) . There were no significant physiological differences between the two conditions. These results provide good evidence that VR experiences can elicit strong craving for cigarettes. The programming can be part of developing a new VR cognitive therapy to help people reduce smoking., (© 2024. The Author(s).)

Rhythmicity is a cornerstone of behavioral and biological processes, especially metabolism, yet the mechanisms behind metabolite cycling remain elusive. This study uncovers a robust oscillation in key metabolite pathways downstream of glucose in humans. A purpose-built 13 C 6 -glucose isotope tracing platform was used to sample Drosophila every 4h and probe these pathways, revealing a striking peak in biosynthesis shortly after lights-on in wild-type flies. A hyperactive mutant ( fumin ) demonstrates increased Krebs cycle labelling and dawn-specific glycolysis labelling. Surprisingly, neither underlying feeding rhythms nor the presence of food availability explain the rhythmicity of glucose processing across genotypes, suggesting a robust internal mechanism for metabolic control of glucose processing. These results align with clinical data highlighting detrimental effects of mistimed energy intake. Our approach offers a unique insight into the dynamic range of daily metabolic processing and provides a mechanistic foundation for exploring circadian metabolic homeostasis in disease contexts., Competing Interests: Declaration of Interest The authors declare no competing interests.

Background: Low self-confidence in patients with psychosis is common. This can lead to higher symptom severity, withdrawal from activities, and low psychological well-being. There are effective psychological techniques to improve positive self-beliefs but these are seldom provided in psychosis services. With young people with lived experience of psychosis we developed a scalable automated VR therapy to enhance positive-self beliefs., Aims: The aim was to conduct a proof of concept clinical test of whether the new VR self-confidence therapy (Phoenix) may increase positive self-beliefs and psychological well-being., Method: Twelve young patients with non-affective psychosis and with low levels of positive self-beliefs participated. Over 6 weeks, patients were provided with a stand-alone VR headset so that they could use Phoenix at home and were offered weekly psychologist meetings. The outcome measures were the Oxford Positive Self Scale (OxPos), Brief Core Schema Scale, and Warwick-Edinburgh Well-being Scale (WEMWBS). Satisfaction, adverse events and side-effects were assessed., Results: Eleven patients provided outcome data. There were very large end-of-treatment improvements in positive self-beliefs (OxPos mean difference = 32.3; 95% CI: 17.3, 47.3; Cohen's d =3.0) and psychological well-being (WEMWBS mean difference = 11.2; 95% CI: 8.0, 14.3; Cohen's d =1.5). Patients rated the quality of the VR therapy as: excellent ( n =9), good ( n =2), fair ( n =0), poor ( n =0). An average of 5.3 ( SD =1.4) appointments were attended., Conclusions: Uptake of the VR intervention was high, satisfaction was high, and side-effects extremely few. There were promising indications of large improvements in positive self-beliefs and psychological well-being. A randomized controlled clinical evaluation is warranted.

Family prevention programs that enhance mental health, wellness, and resilience-while simultaneously addressing violence and alcohol and other drug (AOD) abuse-among Indigenous families are scarce. This gap in culturally grounded and community-based programs creates a critical need to develop and evaluate the efficacy of such prevention programs. This article fills this gap, with the purpose of describing the structure and content of the Weaving Healthy Families (WHF) program, a culturally grounded and community-based program aimed at preventing violence and AOD use while promoting mental health, resilience, and wellness in Indigenous families. The focus then turns to how to approach this process of developing and implementing the program in a culturally grounded and community-based way., Competing Interests: Declaration of Conflicting Interests The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Steroid hormones are attractive candidates for transmitting long-range signals to affect behavior. These lipid-soluble molecules derived from dietary cholesterol easily penetrate the brain and act through nuclear hormone receptors (NHRs) that function as transcription factors. To determine the extent to which NHRs affect sleep:wake cycles, we knocked down each of the 18 highly conserved NHRs found in Drosophila adults and report that the ecdysone receptor (EcR) and its direct downstream NHR Eip75B (E75) act in glia to regulate the rhythm and amount of sleep. Given that ecdysone synthesis genes have little to no expression in the fly brain, ecdysone appears to act as a long-distance signal and our data suggest that it enters the brain more at night. Anti-EcR staining localizes to the cortex glia in the brain and functional screening of glial subtypes revealed that EcR functions in adult cortex glia to affect sleep. Cortex glia are implicated in lipid metabolism, which appears to be relevant for actions of ecdysone as ecdysone treatment mobilizes lipid droplets (LDs), and knockdown of glial EcR results in more LDs. In addition, sleep-promoting effects of exogenous ecdysone are diminished in lsd-2 mutant flies, which are lean and deficient in lipid accumulation. We propose that ecdysone is a systemic secreted factor that modulates sleep by stimulating lipid metabolism in cortex glia., Competing Interests: YL, PH, SZ, ZY, AS No competing interests declared, (© 2023, Li et al.)

Previous research in England has suggested a medical acceptance of Community Treatment Order (CTO) coercion, but none have explored the importance of social influences on practitioner decision making. A sample of 181 practitioners with Mental Health Act responsibilities completed an online survey. They recorded their perspectives about the influence of medical and social items. Sixteen questionnaire items were subdivided into three operational aspects: discharge, renewal, and recall. Medical item averages scored significantly higher as influences than social items (medical items mean = 4.43; social items mean = 3.58; t = - 19.38, p = 0.001). The influence of separate medical and social factors was evidenced by exploratory factor analysis when related to discharge and renewal, but not for recall, where items divided into factors that resembled 'risk' and 'disengagement'. Participants' scores showed no statistically significant difference for a practitioner's professional allegiance. The findings demonstrate that social influences are a homogeneous influence on managing CTOs, in addition to the higher scoring influence of medical factors., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Local recurrences in early gastric cancers (EGCs) after complete endoscopic submucosal dissection (ESD) remain problematic. Here, we investigated the spatially sequential molecular changes in various cancer-related proteins along the axis of the histologically clear but recurrent resection margins (TRM) to determine the appropriate tumor-free margin distance and potential molecular risk markers related to local recurrence. Five eligible patients with recurrent EGCs after complete ESD were selected from 548 EGC patients. The specimens, including recurrent resection margin axis, were divided into 5 zones. Digital spatial profiling assay was performed to quantify the expression level of 31 cancer-related proteins along each zone. p-Chk1 level was significantly reduced in TRM zone than non-recurrent resection margin. The expression of p44/42 ERK and p-Chk1 were significantly decreased along the lateral axis of the recurrent resection margin, with no significance toward the normal zone, which may suggest that p44/42 ERK and p-Chk1 may be involved in the recurrent side compared to non-recurrent margin. Although we could not evaluate more than 5.5 mm, the significant linear decreases in p44/42 ERK and p-Chk1 were maintained until at least 5.5 mm from the tumor zone in the TRM direction. We estimated the possible margin distance using scatterplots and linear regression analyses, which also showed the estimated distance more than 5.5 mm. In conclusion, the p-Chk1 and p44/42 ERK may be potential candidates of molecular risk markers that may be related to the local recurrence after complete ESD, and a tumor-free distance of 5.5 mm is not enough for safety margin., (Copyright © 2021 The Authors. Published by Elsevier GmbH.. All rights reserved.)

Abstract: Although severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is not a proven foodborne pathogen, the COVID-19 pandemic has put the food system on alert, and food safety has been identified as an important pillar in mitigating the crisis. Therefore, an understanding of how popular media are used as a vital disseminator of food safety and health information for the public is more important than ever. YouTube deserves particular attention as one of the most highly trafficked Web sites on the Internet, especially because YouTube has been blamed during the pandemic for spreading misleading or untrustworthy information that contradicts validated information. This study was conducted to evaluate the food safety information and practices circulating on YouTube during the COVID-19 pandemic and the alignment of these practices with recommendations from government agencies. A search for videos on YouTube was conducted using the key words "food and COVID-19," "food safety and COVID-19," and "groceries and COVID-19." After applying a series of inclusive and exclusive criteria, 85 videos from the United States and Canada were evaluated. More than half (69%) of the videos presented hand washing procedures, 26% showed kitchen disinfection, and most (86%) showed take-out food or grocery store practices. Multiple produce washing procedures were also shown throughout videos. Food was not considered hazardous in 39% of the videos, but 24% mentioned that food packaging is potentially hazardous. Most videos cited government agencies and had a host or guest who was a health care professional, professor, or expert. Three videos were not aligned with a government agency's guideline or information cited; two were presented by a health care professional. These findings reveal the need to develop educational interventions that increase YouTube video host and guest awareness of social media use as a tool for food safety dissemination and the need to provide trustworthy information., (Copyright ©, International Association for Food Protection.)

Objective: Insomnia is a clinically verified nicotine withdrawal symptom. As nicotine is a stimulant, it is plausible that smoking at night could disturb sleep more than smoking at earlier times of the day, but this remains empirically unclear. This study examined smoking status and its associations with insomnia severity and sleep duration while considering the potential role of smoking time., Methods: Data were derived from the Sleep and Healthy Activity Diet Environment and Socialization study, a community-based study of 1007 adults (n nonsmokers  = 818; n smokers  = 189) aged 22-60 from the Philadelphia area. Smoking status and time of smoking were self-reported. Insomnia was assessed with the Insomnia Severity Index and categorized as none, mild, and moderate-to-severe. Sleep duration was assessed with one item from the National Health and Nutrition Examination Survey and categorized as very short, short, normal, and long. Ordinal and multinomial logistic regressions were used to determine the association of smoking status including smoking time with insomnia severity and sleep duration controlling for sociodemographic covariates., Results: Compared to nonsmoking, smoking was associated with experiencing increased insomnia (odds ratio = 2.5, 95% confidence interval [CI] 1.9, 3.4, P < .001) as well as very short (relative risk ratio = 1.9, 95% CI 1.1, 3.3) and short (relative risk ratio = 1.5, 95% CI 1.0, 2.3) sleep (vs normal sleep duration). Night-time smoking was significantly associated with greater insomnia and shorter sleep duration., Conclusions: Findings provide evidence that smoking is associated with increased insomnia severity and shorter sleep duration, particularly nightly smoking. Sleep health should be considered in smoking cessation efforts., (Copyright © 2020 National Sleep Foundation. Published by Elsevier Inc. All rights reserved.)

Background: In melanoma patients, microscopic tumor in the sentinel lymph-node biopsy (SLN) increases the risk of distant metastases, but the transition from tumor in the SLN to metastatic disease remains poorly understood., Methods: Fluorescent staining for CD3, CD20, CD11c, and DNA was performed on SLN tissue and matching primary tumors. Regions of interest (ROI) were then chosen geometrically (e.g., tumor) or by fluorescent cell subset markers (e.g., CD11c). Each ROI was further analyzed using NanoString Digital Spatial Profiling high-resolution multiplex profiling. Digital counts for 59-panel immune-related proteins were collected and normalized to account for system variation and ROI area., Results: Tumor regions of SLNs had variable infiltration of CD3 cells among patients. The patient with overall survival (OS) > 8 years had the most CD11c- and CD3-expressing cells infiltrating the SLN tumor region. All patients had CD11c (dendritic cell, DC) infiltration into the SLN tumor region. Selecting ROI by specific cell subtype, we compared protein expression of CD11c cells between tumor and non-tumor/normal tissue SLN regions. Known markers of DC activation such as CD86, HLA-DR, and OX40L were lowest on CD11c cells within SLN tumor for the patient with OS < 1 year and highest on the patient with OS > 8 years., Conclusion: We demonstrate the feasibility of profiling the protein expression of CD11c cells within the SLN tumor. Identifying early regulators of melanoma control when the disease is microscopically detected in the SLN is beneficial and requires follow-up studies in a larger cohort of patients.

Sleep remains a major mystery of biology, with little understood about its basic function. One of the most commonly proposed functions of sleep is the consolidation of memory 1-3 . However, as conditions such as starvation require the organism to be awake and active 4 , the ability to switch to a memory consolidation mechanism that is not contingent on sleep may confer an evolutionary advantage. Here we identify an adaptive circuit-based mechanism that enables Drosophila to form sleep-dependent and sleep-independent memory. Flies fed after appetitive conditioning needed increased sleep for memory consolidation, but flies starved after training did not require sleep to form memories. Memory in fed flies is mediated by the anterior-posterior α'/β' neurons of the mushroom body, while memory under starvation is mediated by medial α'/β' neurons. Sleep-dependent and sleep-independent memory rely on distinct dopaminergic neurons and corresponding mushroom body output neurons. However, sleep and memory are coupled such that mushroom body neurons required for sleep-dependent memory also promote sleep. Flies lacking Neuropeptide F display sleep-dependent memory even when starved, suggesting that circuit selection is determined by hunger. This plasticity in memory circuits enables flies to retain essential information in changing environments.

Axons in the mature central nervous system (CNS) fail to regenerate after axotomy, partly due to the inhibitory environment constituted by reactive glial cells producing astrocytic scars, chondroitin sulfate proteoglycans, and myelin debris. We investigated this inhibitory milieu, showing that it is reversible and depends on glial metabolic status. We show that glia can be reprogrammed to promote morphological and functional regeneration after CNS injury in Drosophila via increased glycolysis. This enhancement is mediated by the glia derived metabolites: L-lactate and L-2-hydroxyglutarate (L-2HG). Genetically/pharmacologically increasing or reducing their bioactivity promoted or impeded CNS axon regeneration. L-lactate and L-2HG from glia acted on neuronal metabotropic GABA B receptors to boost cAMP signaling. Local application of L-lactate to injured spinal cord promoted corticospinal tract axon regeneration, leading to behavioral recovery in adult mice. Our findings revealed a metabolic switch to circumvent the inhibition of glia while amplifying their beneficial effects for treating CNS injuries., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 Elsevier Inc. All rights reserved.)

N/A., (© 2020 IOP Publishing Ltd.)

Compared to non-smokers, smokers have an increased risk for poor sleep quality, which could undermine confidence to quit. This study examined whether baseline sleep quality was associated with quit-day smoking self-efficacy among smokers enrolled in a smoking cessation trial. Treatment-seeking low-income smokers were randomized to either a low-intensity physical activity integrated with standard smoking cessation intervention or standard care cessation only. Poor sleep quality was significantly associated with lower quit-day (week 4) smoking self-efficacy ( β  = -0.61; standard error = 8.1; p  = .03). Over half the samples (53%) reported poor sleep quality, thus addressing baseline sleep quality is an important consideration in smoking cessation programs.

The Lesser Antilles, in the Eastern Caribbean, were long considered to have only two species in the genus Iguana Laurenti 1768: the Lesser Antillean iguana Iguana delicatissima, which is endemic to parts of the Lesser Antilles, and the Common green iguana Iguana iguana, which also occurs throughout Central and South America. No subspecies are currently recognised. However, herpetologists and reptile collectors have pointed out strong physical differences between some of the island populations of Iguana iguana and those from the continent. Drawing on both morphological and genetic data, this paper describes two subspecies of the Common green iguana Iguana iguana from the southern Lesser Antilles, specifically the countries of Saint Lucia Iguana iguana sanctaluciae and Iguana iguana insularis from St Vincent the Grenadines, and Grenada. The form on the island of Saint Vincent has not been identified. The new subspecies are described based on the following unique combination of characters: Presence of high median and medium to small lateral horns on the snout; Small subtympanic plate not exceeding 20% of the eardrum size; Two or three scales of decreasing size anterior to the subtympanic plate; Fewer than ten small to medium triangular gular spikes; Medium sized dewlap; Low number of small to medium dispersed nuchal tubercles; Dark brown iris, with the white of the eye visible; Oval, prominent nostril; Short and relatively flat head; High dorsal spines; No swelling of the jowls in reproductively active males.                Iguana iguana sanctaluciae has in adults vertical black stripes on body and tail and a black dewlap whereas Iguana iguana insularis is pale grey or creamy white in adults.                Both subspecies are globally threatened by unsustainable hunting (including the pet trade) and by invasive alien species,     including hybridization from invasive iguanas from South and Central America (I. iguana iguana and I. rhinolopha, considered here as full species) that have become established in all three countries. The authors call for stronger measures to conserve the remaining purebred Iguana i. insularis and Iguana i. sanctaluciae ssp. nov. throughout their ranges and for further research to identify other cryptic species and subspecies of Iguana in the Lesser Antilles.

Background: Facioscapulohumeral muscular dystrophy 1 (FSHD1) has an autosomal dominant pattern of inheritance and primarily affects skeletal muscle. The genetic cause of FSHD1 is contraction of the D4Z4 macrosatellite array on chromosome 4 alleles associated with a permissive haplotype causing infrequent sporadic expression of the DUX4 gene. Epigenetically, the contracted D4Z4 array has decreased cytosine methylation and an open chromatin structure. Despite these genetic and epigenetic changes, the majority of FSHD myoblasts are able to repress DUX4 transcription. In this study we hypothesized that histone modifications distinguish DUX4 expressing and non-expressing cells from the same individuals., Results: FSHD myocytes containing the permissive 4qA haplotype with a long terminal D4Z4 unit were sorted into DUX4 expressing and non-expressing groups. We found similar CpG hypomethylation between the groups of FSHD-affected cells suggesting that CpG hypomethylation is not sufficient to trigger DUX4 expression. A survey of histone modifications present at the D4Z4 region during cell lineage commitment revealed that this region is bivalent in FSHD iPS cells with both H3K4me3 activating and H3K27me3 repressive marks present, making D4Z4 poised for DUX4 activation in pluripotent cells. After lineage commitment, the D4Z4 region becomes univalent with H3K27me3 in FSHD and non-FSHD control myoblasts and a concomitant increase in H3K4me3 in a small fraction of cells. Chromatin immunoprecipitation (ChIP) for histone modifications, chromatin modifier proteins and chromatin structural proteins on sorted FSHD myocytes revealed that activating H3K9Ac modifications were ~ fourfold higher in DUX4 expressing FSHD myocytes, while the repressive H3K27me3 modification was ~ fourfold higher at the permissive allele in DUX4 non-expressing FSHD myocytes from the same cultures. Similarly, we identified EZH2, a member of the polycomb repressive complex involved in H3K27 methylation, to be present more frequently on the permissive allele in DUX4 non-expressing FSHD myocytes., Conclusions: These results implicate PRC2 as the complex primarily responsible for DUX4 repression in the setting of FSHD and H3K9 acetylation along with reciprocal loss of H3K27me3 as key epigenetic events that result in DUX4 expression. Future studies focused on events that trigger H3K9Ac or augment PRC2 complex activity in a small fraction of nuclei may expose additional drug targets worthy of study.

Loss of the Neurofibromatosis 1 (Nf1) protein, neurofibromin, in Drosophila disrupts circadian rhythms of locomotor activity without impairing central clock function, suggesting effects downstream of the clock. However, the relevant cellular mechanisms are not known. Leveraging the discovery of output circuits for locomotor rhythms, we dissected cellular actions of neurofibromin in recently identified substrates. Herein, we show that neurofibromin affects the levels and cycling of calcium in multiple circadian peptidergic neurons. A prominent site of action is the pars intercerebralis (PI), the fly equivalent of the hypothalamus, with cell-autonomous effects of Nf1 in PI cells that secrete DH44. Nf1 interacts genetically with peptide signaling to affect circadian behavior. We extended these studies to mammals to demonstrate that mouse astrocytes exhibit a 24-hr rhythm of calcium levels, which is also attenuated by lack of neurofibromin. These findings establish a conserved role for neurofibromin in intracellular signaling rhythms within the nervous system., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

An ab initio study of the effects of implicit and explicit hosts on the excited state properties of pentacene and its nitrogen-based derivatives has been performed using ground state density functional theory (DFT), time-dependent DFT, and ΔSCF. We observe a significant solvatochromic redshift in the excitation energy of the lowest singlet state (S 1 ) of pentacene from inclusion in a p-terphenyl host compared to vacuum; for an explicit host consisting of six nearest neighbour p-terphenyls, we obtain a redshift of 65 meV while a conductor-like polarisable continuum model (CPCM) yields a 78 meV redshift. Comparison is made between the excitonic properties of pentacene and four of its nitrogen-based analogs, 1,8-, 2,9-, 5,12-, and 6,13-diazapentacene with the latter found to be the most distinct due to local distortions in the ground state electronic structure. We observe that a CPCM is insufficient to fully understand the impact of the host due to the presence of a mild charge-transfer (CT) coupling between the chromophore and neighbouring p-terphenyls, a phenomenon which can only be captured using an explicit model. The strength of this CT interaction increases as the nitrogens are brought closer to the central acene ring of pentacene.

Background: Facioscapulohumeral muscular dystrophy (FSHD) is most commonly inherited in an autosomal dominant pattern and caused by the abnormal expression of DUX4 in skeletal muscle. The DUX4 transcription factor has DNA binding domains similar to several paired class homeotic transcription factors, but only myogenic factors PAX3 and PAX7 rescue cell viability when co-expressed with DUX4 in mouse myoblasts. This observation suggests competition for DNA binding sites in satellite cells might limit muscle repair and may be one aspect of DUX4-associated myotoxicity. The competition hypothesis requires that DUX4 and PAX3/7 be expressed in the same cells at some point during development or in adult tissues. We modeled myogenesis using human isogenic iPS and ES cells and examined expression patterns of DUX4, PAX3, and PAX7 to determine if conditions that promote PAX3 and PAX7 expression in cell culture also promote DUX4 expression in the same cells., Methods: Isogenic iPSCs were generated from human fibroblasts of two FSHD-affected individuals with somatic mosaicism. Clones containing the shortened FSHD-causing D4Z4 array or the long non-pathogenic array were isolated from the same individuals. We also examined myogenesis in commercially available hES cell lines derived from FSHD-affected and non-affected embryos. DUX4, PAX3, and PAX7 messenger RNAs (mRNAs) were quantified during a 40-day differentiation protocol, and antibodies were used to identify cell types in different stages of differentiation to determine if DUX4 and PAX3 or PAX7 are present in the same cells., Results: Human iPS and ES cells differentiated into skeletal myocytes as evidenced by Titin positive multinucleated fibers appearing toward the end of a 40-day differentiation protocol. PAX3 and PAX7 were expressed at similar times during differentiation, and DUX4 positive nuclei were seen at terminal stages of differentiation in cells containing the short D4Z4 arrays. Nuclei that expressed both DUX4 and PAX3, or DUX4 and PAX7 were not observed after examining immunostained nuclei at five different time points during myogenic differentiation of pluripotent cells., Conclusions: We conclude that DUX4, PAX3, and PAX7 have distinct expression patterns during myogenic differentiation of stem cells. Our findings are consistent with the hypothesis that muscle damage in FSHD is due to DUX4-mediated toxicity causing destruction of terminally differentiated myofibers. While these studies examine DUX4, PAX3, and PAX7 expression patterns during stem cell myogenesis, they should not be generalized to tissue repair in adult muscle tissue.

The solvatochromic shift, as well as the change in colour of the simple organic dye nile red, is studied in two polar and two non-polar solvents in the context of large-scale time-dependent density-functional theory (TDDFT) calculations treating large parts of the solvent environment from first principles. We show that an explicit solvent representation is vital to resolve absorption peak shifts between nile red in n-hexane and toluene, as well as acetone and ethanol. The origin of the failure of implicit solvent models for these solvents is identified as being due to the strong solute-solvent interactions in form of π-stacking and hydrogen bonding in the case of toluene and ethanol. We furthermore demonstrate that the failures of the computationally inexpensive Perdew-Burke-Ernzerhof (PBE) functional in describing some features of the excited state potential energy surface of the S 1 state of nile red can be corrected for in a straightforward fashion, relying only on a small number of calculations making use of more sophisticated range-separated hybrid functionals. The resulting solvatochromic shifts and predicted colours are in excellent agreement with experiment, showing the computational approach outlined in this work to yield very robust predictions of optical properties of dyes in solution.

In this chapter we describe the workflow we use for labeled quantitative proteomics analysis using tandem mass tags (TMT) starting with the sample preparation and ending with the multivariate analysis of the resulting data. We detail the step-by-step process from sample processing, labeling, fractionation, and data processing using Proteome Discoverer through to data analysis and interpretation in the context of a multi-run experiment. The final analysis and data interpretation rely on an R package we call TMTPrepPro, which are deployed on a local GenePattern server, and used for generating various outputs which are also outlined herein.

Re-circulating underwater breathing apparatus (rebreathers) have become increasingly popular amongst sport divers. In comparison to open-circuit scuba, rebreathers are complex life support equipment that incorporates many inherent failure modes and potential for human error. This individually or in combination can lead to an inappropriate breathing gas. Analysis of rebreather diving incidents suggests that inappropriate breathing gas is the most prevalent disabling agent. This can result in spontaneous loss of consciousness (LoC), water aspiration and drowning. Protecting the airway by maintaining the diver/rebreather oral interface may delay water aspiration following LoC underwater; the possibility of a successful rescue is, thus, increased. One means of protecting the airway following LoC underwater is the use of a full-face mask (FFM). However, such masks are complex and expensive; therefore, they have not been widely adopted by the sport diving community. An alternative to the FFM used extensively throughout the global military diving community is the mouthpiece retaining strap (MRS). A recent study documented 54 LoC events in military rebreather diving with only three consequent drownings; all divers were reported to be using a MRS. Even allowing for the concomitant use of a tethered diving partner system in most cases, the low number of fatalities in this large series is circumstantially supportive of the efficacy of the MRS. Despite drowning featuring as a final common pathway in the vast majority of rebreather fatalities, the MRS has not been widely adopted by the sport rebreather diving community.

Correction for 'The biology of environmental stress: molecular biomarkers in Sydney rock oysters (Saccostrea glomerata)' by D. A. Raftos et al., Environ. Sci.: Processes Impacts, 2016, 18, 1129-1139.

This review describes our recent work on environmental stress in Sydney rock oysters, focusing on the identification of molecular biomarkers for ecotoxicological analysis. We begin by describing the environmental pressures facing coastal estuaries in Australia, with particular reference to Sydney Harbour. After providing that context, we summarise our transcriptional and proteomic analyses of Sydney rock oysters responding to chemical contamination and other forms of environmental stress. This work has shown that the intracellular processes of oysters are highly responsive to environmental threats. Our data agree with the broader literature, which suggests that there is a highly conserved intracellular stress response in oysters involving a limited number of biological processes. We conclude that many effective molecular markers for environmental biomonitoring are likely to lie within these biological pathways.

It is rarely the case that a single electron affects the behaviour of several hundred thousands of atoms. Here we demonstrate a phenomenon where this happens. The key role is played by topological insulators-materials that have surface states protected by time-reversal symmetry. Such states are delocalized over the surface and are immune to its imperfections in contrast to ordinary insulators. For topological insulators, the effects of these surface states will be more strongly pronounced in the case of nanoparticles. Here we show that under the influence of light a single electron in a topologically protected surface state creates a surface charge density similar to a plasmon in a metallic nanoparticle. Such an electron can act as a screening layer, which suppresses absorption inside the particle. In addition, it can couple phonons and light, giving rise to a previously unreported topological particle polariton mode. These effects may be useful in the areas of plasmonics, cavity electrodynamics and quantum information.

Experimental techniques for electron energy loss spectroscopy (EELS) combine high energy resolution with high spatial resolution. They are therefore powerful tools for investigating the local electronic structure of complex systems such as nanostructures, interfaces and even individual defects. Interpretation of experimental electron energy loss spectra is often challenging and can require theoretical modelling of candidate structures, which themselves may be large and complex, beyond the capabilities of traditional cubic-scaling density functional theory. In this work, we present functionality to compute electron energy loss spectra within the onetep linear-scaling density functional theory code. We first demonstrate that simulated spectra agree with those computed using conventional plane wave pseudopotential methods to a high degree of precision. The ability of onetep to tackle large problems is then exploited to investigate convergence of spectra with respect to supercell size. Finally, we apply the novel functionality to a study of the electron energy loss spectra of defects on the (1 0 1) surface of an anatase slab and determine concentrations of defects which might be experimentally detectable.

Temperature is an important factor influencing the distribution of marine picocyanobacteria. However, molecular responses contributing to temperature preferences are poorly understood in these important primary producers. We compared the temperature acclimation of a tropical Synechococcus strain WH8102 with temperate strain BL107 at 18 °C relative to 22 °C and examined their global protein expression, growth patterns, photosynthetic efficiency and lipid composition. Global protein expression profiles demonstrate the partitioning of the proteome into major categories: photosynthesis (>40%), translation (10-15%) and membrane transport (2-8%) with distinct differences between and within strains grown at different temperatures. At low temperature, growth and photosynthesis of strain WH8102 was significantly decreased, while BL107 was largely unaffected. There was an increased abundance of proteins involved in protein biosynthesis at 18 °C for BL107. Each strain showed distinct differences in lipid composition with higher unsaturation in strain BL107. We hypothesize that differences in membrane fluidity, abundance of protein biosynthesis machinery and the maintenance of photosynthesis efficiency contribute to the acclimation of strain BL107 to low temperature. Additional proteins unique to BL107 may also contribute to this strain's improved fitness at low temperature. Such adaptive capacities are likely important factors favoring growth of temperate strains over tropical strains in high latitude niches.

In this work we study the solvatochromic shift of a selected low-energy excited state of alizarin in water by using a linear-scaling implementation of large-scale time-dependent density functional theory (TDDFT). While alizarin, a small organic dye, is chosen as a simple example of solute-solvent interactions, the findings presented here have wider ramifications for the realistic modeling of dyes, paints, and pigment-protein complexes. We find that about 380 molecules of explicit water need to be considered in order to yield an accurate representation of the solute-solvent interaction and a reliable solvatochromic shift. By using a novel method of constraining the TDDFT excitation vector, we confirm that the origin of the slow convergence of the solvatochromic shift with system size is due to two different effects. The first factor is a strong redshift of the excitation due to an explicit delocalization of a small fraction of the electron and the hole from the alizarin onto the water, which is mainly confined to within a distance of 7 Å from the alizarin molecule. The second factor can be identified as long-range electrostatic influences of water molecules beyond the 7 Å region on the ground-state properties of alizarin. We also show that these electrostatic influences are not well reproduced by a QM/MM model, suggesting that full QM studies of relatively large systems may be necessary in order to obtain reliable results.

We report a linear-scaling density functional theory (DFT) study of the structure, wall-polarization absolute band-alignment and optical absorption of several, recently synthesized, open-ended imogolite (Imo) nanotubes (NTs), namely single-walled (SW) aluminosilicate (AlSi), SW aluminogermanate (AlGe), SW methylated aluminosilicate (AlSi-Me), and double-walled (DW) AlGe NTs. Simulations with three different semi-local and dispersion-corrected DFT-functionals reveal that the NT wall-polarization can be increased by nearly a factor of four going from SW-AlSi-Me to DW-AlGe. Absolute vacuum alignment of the NT electronic bands and comparison with those of rutile and anatase TiO2 suggest that the NTs may exhibit marked propensity to both photo-reduction and hole-scavenging. Characterization of the NTs' band-separation and optical properties reveal the occurrence of (near-)UV inside-outside charge-transfer excitations, which may be effective for electron-hole separation and enhanced photocatalytic activity. Finally, the effects of the NTs' wall-polarization on the absolute alignment of electron and hole acceptor states of interacting water (H2O) molecules are quantified and discussed.

This manuscript describes a protocol at the University of Kentucky that allows a translational research team to collect human myocardium that can be used for biological research. We have gained a great deal of practical experience since we started this protocol in 2008, and we hope that other groups might be able to learn from our endeavors. To date, we have procured ~4000 samples from ~230 patients. The tissue that we collect comes from organ donors and from patients who are receiving a heart transplant or a ventricular assist device because they have heart failure. We begin our manuscript by describing the importance of human samples in cardiac research. Subsequently, we describe the process for obtaining consent from patients, the cost of running the protocol, and some of the issues and practical difficulties that we have encountered. We conclude with some suggestions for other researchers who may be considering starting a similar protocol.

We present a solution of the full time-dependent density-functional theory (TDDFT) eigenvalue equation in the linear response formalism exhibiting a linear-scaling computational complexity with system size, without relying on the simplifying Tamm-Dancoff approximation (TDA). The implementation relies on representing the occupied and unoccupied subspaces with two different sets of in situ optimised localised functions, yielding a very compact and efficient representation of the transition density matrix of the excitation with the accuracy associated with a systematic basis set. The TDDFT eigenvalue equation is solved using a preconditioned conjugate gradient algorithm that is very memory-efficient. The algorithm is validated on a small test molecule and a good agreement with results obtained from standard quantum chemistry packages is found, with the preconditioner yielding a significant improvement in convergence rates. The method developed in this work is then used to reproduce experimental results of the absorption spectrum of bacteriochlorophyll in an organic solvent, where it is demonstrated that the TDA fails to reproduce the main features of the low energy spectrum, while the full TDDFT equation yields results in good qualitative agreement with experimental data. Furthermore, the need for explicitly including parts of the solvent into the TDDFT calculations is highlighted, making the treatment of large system sizes necessary that are well within reach of the capabilities of the algorithm introduced here. Finally, the linear-scaling properties of the algorithm are demonstrated by computing the lowest excitation energy of bacteriochlorophyll in solution. The largest systems considered in this work are of the same order of magnitude as a variety of widely studied pigment-protein complexes, opening up the possibility of studying their properties without having to resort to any semiclassical approximations to parts of the protein environment.

The left ventricle (LV) of the heart is composed of a complex organization of cardiac muscle fibers, which contract to generate force and pump blood into the body. It has been shown that both the orientation and contractile strength of these myofibers vary across the ventricular wall. The hypothesis of the current study is that the transmural distributions of myofiber orientation and contractile strength interdependently impact LV pump function. In order to quantify these interactions a finite element (FE) model of the LV was generated, which incorporated transmural variations. The influences of myofiber orientation and contractile strength on the Starling relationship and the end-systolic (ES) apex twist of the LV were assessed. The results suggest that reductions in contractile strength within a specific transmural layer amplified the effects of altered myofiber orientation in the same layer, causing greater changes in stroke volume (SV). Furthermore, when the epicardial myofibers contracted the strongest, the twist of the LV apex was greatest, regardless of myofiber orientation. These results demonstrate the important role of transmural distribution of myocardial contractile strength and its interplay with myofiber orientation. The coupling between these two physiologic parameters could play a critical role in the progression of heart failure.

This article provides an overview of cognitive behavioral therapy (CBT) for insomnia and depression. Included is a discussion of how CBT for insomnia affects depression symptoms and how CBT for depression affects insomnia symptoms. The extant literature is reviewed on ways that depression/insomnia comorbidity moderates CBT response. The article concludes with an introduction to cognitive behavioral social rhythm therapy, a group therapy that integrates tenets of CBT for both disorders., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Social rhythms, also known as daily routines (e.g. exercise, of school or work, recreation, social activities), have been identified as potential time cues to help to regulate the biological clock. Past research has shown links between regularity and healthy sleep. This study examined the regularity and frequency of daytime activities in a clinical insomnia population and a good sleeper comparison group. Participants (N = 69) prospectively monitored their sleep and daily activities for a 2-week period. Although participants with insomnia and good sleepers had similar levels of activity, relative to good sleepers, those with insomnia were less regular in their activities. Findings from this study add to the growing number of studies that highlight the relative importance of the regularity of daytime activities on sleep. Accordingly, future research should test treatment components that focus on regulating daytime activities, which would likely improve treatment outcomes.

Heart failure is associated with pump dysfunction and remodeling but it is not yet known if the condition affects different transmural regions of the heart in the same way. We tested the hypotheses that the left ventricles of non-failing human hearts exhibit transmural heterogeneity of cellular level contractile properties, and that heart failure produces transmural region-specific changes in contractile function. Permeabilized samples were prepared from the sub-epicardial, mid-myocardial, and sub-endocardial regions of the left ventricular free wall of non-failing (n=6) and failing (n=10) human hearts. Power, an in vitro index of systolic function, was higher in non-failing mid-myocardial samples (0.59±0.06μWmg(-1)) than in samples from the sub-epicardium (p=0.021) and the sub-endocardium (p=0.015). Non-failing mid-myocardial samples also produced more isometric force (14.3±1.33kNm(-2)) than samples from the sub-epicardium (p=0.008) and the sub-endocardium (p=0.026). Heart failure reduced power (p=0.009) and force (p=0.042) but affected the mid-myocardium more than the other transmural regions. Fibrosis increased with heart failure (p=0.021) and mid-myocardial tissue from failing hearts contained more collagen than matched sub-epicardial (p<0.001) and sub-endocardial (p=0.043) samples. Power output was correlated with the relative content of actin and troponin I, and was also statistically linked to the relative content and phosphorylation of desmin and myosin light chain-1. Non-failing human hearts exhibit transmural heterogeneity of contractile properties. In failing organs, region-specific fibrosis produces the greatest contractile deficits in the mid-myocardium. Targeting fibrosis and sarcomeric proteins in the mid-myocardium may be particularly effective therapies for heart failure., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Despite voluminous research on the acid oxidation of carbon nanotubes (CNTs), there is a distinct lack of experimental results showing distributions of functional groups at the nanometre length scale. Here, functional peaks have been mapped across individual multi-walled CNTs with low-dose, monochromated electron energy-loss spectroscopy (EELS) in the scanning transmission electron microscope (STEM). Density functional theory simulations show that the EELS features are consistent with oxygenated functional groups, most likely carboxyl moieties.

The timing of spawning and hatching, larval durations and growth exhibited by juvenile brill Scophthalmus rhombus captured along the Irish west coast were estimated using otolith microstructure analysis. Scophthalmus rhombus were estimated to have hatched between February and May, with fish settling onto nursery grounds between March and June. Fish collected later on in the season exhibited higher otolith growth rates in comparison to earlier collected fish. This is the first study to describe the early life history of a commercially valuable but understudied flatfish species., (© 2013 The Fisheries Society of the British Isles.)

The purpose of this study was to identify and explain changes in ventricular and cellular function that contribute to aging-associated cardiovascular disease in aging F344 rats. Three groups of female F344 rats, aged 6, 18, and 22 mo, were studied. Echocardiographic measurements in isoflurane-anesthetized animals showed an increase in peak left ventricular torsion between the 6- and the 18-mo-old groups that was partially reversed in the 22-mo-old animals (P < 0.05). Epicardial, midmyocardial, and endocardial myocytes were subsequently isolated from the left ventricles of each group of rats. Unloaded sarcomere shortening and Ca(2+) transients were then measured in these cells (n = >75 cells for each of the nine age-region groups). The decay time of the Ca(2+) transient and the time required for 50% length relaxation both increased with age but not uniformly across the three regions (P < 0.02). Further analysis revealed a significant shift in the transmural distribution of these properties between 18 and 22 mo of age, with the largest changes occurring in epicardial myocytes. Computational modeling suggested that these changes were due in part to slower Ca(2+) dissociation from troponin in aging epicardial myocytes. Subsequent biochemical assays revealed a >50% reduction in troponin I phosphoprotein content in 22-mo-old epicardium relative to the other regions. These data suggest that between 18 and 22 mo of age (before the onset of heart failure), F344 rats display epicardial-specific myofilament-level modifications that 1) break from the progression observed between 6 and 18 mo and 2) coincide with aberrant patterns of cardiac torsion.

We present an implementation of time-dependent density-functional theory (TDDFT) in the linear response formalism enabling the calculation of low energy optical absorption spectra for large molecules and nanostructures. The method avoids any explicit reference to canonical representations of either occupied or virtual Kohn-Sham states and thus achieves linear-scaling computational effort with system size. In contrast to conventional localised orbital formulations, where a single set of localised functions is used to span the occupied and unoccupied state manifold, we make use of two sets of in situ optimised localised orbitals, one for the occupied and one for the unoccupied space. This double representation approach avoids known problems of spanning the space of unoccupied Kohn-Sham states with a minimal set of localised orbitals optimised for the occupied space, while the in situ optimisation procedure allows for efficient calculations with a minimal number of functions. The method is applied to a number of medium sized organic molecules and a good agreement with traditional TDDFT methods is observed. Furthermore, linear scaling of computational cost with system size is demonstrated on (10,0) carbon nanotubes of different lengths.

The extremely elongated morphology of fungal hyphae is dependent on the cell's ability to assemble and maintain polarized growth machinery over multiple cell cycles. The different morphologies of the fungus Candida albicans make it an excellent model organism in which to study the spatiotemporal requirements for constitutive polarized growth and the generation of different cell shapes. In C. albicans, deletion of the landmark protein Rsr1 causes defects in morphogenesis that are not predicted from study of the orthologous protein in the related yeast Saccharomyces cerevisiae, thus suggesting that Rsr1 has expanded functions during polarized growth in C. albicans. Here, we show that Rsr1 activity localizes to hyphal tips by the differential localization of the Rsr1 GTPase-activating protein (GAP), Bud2, and guanine nucleotide exchange factor (GEF), Bud5. In addition, we find that Rsr1 is needed to maintain the focused localization of hyphal polarity structures and proteins, including Bem1, a marker of the active GTP-bound form of the Rho GTPase, Cdc42. Further, our results indicate that tip-localized Cdc42 clusters are associated with the cell's ability to express a hyphal transcriptional program and that the ability to generate a focused Cdc42 cluster in early hyphae (germ tubes) is needed to maintain hyphal morphogenesis over time. We propose that in C. albicans, Rsr1 "fine-tunes" the distribution of Cdc42 activity and that self-organizing (Rsr1-independent) mechanisms of polarized growth are not sufficient to generate narrow cell shapes or to provide feedback to the transcriptional program during hyphal morphogenesis.

MaineHealth adopts six metrics to assess its impact on population health.

Background: Posterior cruciate ligament injuries can occur as isolated ligament ruptures or in association with the multiligament-injured knee. Delayed reconstruction, at 2-3 weeks post-injury, is predominantly recommended for posterior cruciate ligament tears in the multiligament-injured knee. While acute bone and soft tissue avulsion patterns of injury can be amenable to repair, the described techniques have been associated with some difficulties attaching the avulsed ligament., Description of Technique: As part of a reconstruction/repair of a multiligament-injured knee, we performed arthroscopic primary repair of the posterior cruciate ligament by passing Bunnell-type stitches into the substance of the ligament using a reloadable suture passer. We then passed the sutures through drill holes into the femoral footprint of the ligament and tied them over a bony bridge., Patients and Methods: We retrospectively reviewed three patients with posterior cruciate ligament tears associated with a multiligament-injured knee. All patients had posterior cruciate ligament soft tissue avulsions or "peel off" injuries diagnosed by MRI. The described repair technique was used to repair the posterior cruciate ligament avulsion. Minimum followup was 64 months (mean, 68 months; range, 64-75 months). ROM, stability testing, and functional outcome scores (Lysholm and modified Cincinnati) were recorded., Results: Mean ROM was 0° to 127°. Posterior drawer testing was negative in all three patients. The mean Lysholm score was 92 and the mean modified Cincinnati score was 94. Followup MRI confirmed ligament healing in all patients., Conclusions: We believe arthroscopic posterior cruciate ligament repair for soft tissue peel off injuries is a technique that, when applied to carefully selected patients, may be helpful to the surgeon treating patients with a multiligament-injured knee., Level of Evidence: Level IV, therapeutic study. See Guidelines for Authors for a complete description of levels of evidence.