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The effects of patient habitus (e.g., breast attenuation in women and diaphragmatic attenuation in men) have long been recognized as factors that reduce the accuracy of myocardial perfusion imaging. Although it has long been assumed that patient obesity effects accuracy, this has never been formally evaluated. We studied the effects of patient obesity, defined as a body mass index (BMI) > or = 30, on 607 patients who underwent exercise thallium-201 single-photon emission computed tomography (SPECT). Because the effects of obesity are most likely mediated through increased photon attenuation and scatter, we also evaluated the effects of other markers of patient size: body surface area (BSA) and patient weight. Accuracy was determined by performing quantitative analysis and measuring the area under the receiver operating characteristic curve (AUC). Obesity was associated with significantly lower accuracy (AUC 0.86 +/- 0.03 vs 0.92 +/- 0.02, p <0.05) despite similar estimates of maximal coronary blood flow (as estimated by heart rate and rate-pressure product at peak exercise) and severity of coronary disease. There were no significant differences attributable to either patient weight or BSA. Weight and BSA correlated significantly with left ventricular chamber size whereas BMI did not. We conclude that the accuracy of quantitative SPECT thallium-201 is significantly reduced by patient obesity and that although BSA and weight are also associated with increased attenuation, they have no effect on accuracy, which is most likely due to the compensating effects of increased chamber size. [ABSTRACT FROM AUTHOR]

Background: The COVID-19 pandemic has catalyzed a move from face-to-face to digital delivery of services by hospitals and primary care. However, little is known about the impact of digital transformation on organizations supporting unpaid caregivers. Since the start of the COVID-19 pandemic, the value of care provided by such informal caregivers is estimated to be £111 billion (US$ 152.7 billion) in England., Objective: This study aims to analyze service uptake patterns (including digital service options) over the pandemic period in an English caregivers' support organization covering a population of 0.98 million; measure changes in organizational performance, service efficiency, and quality; and identify the views of caregivers on service provision and future digital delivery., Methods: This was a retrospective analysis of the use of digital versus nondigital support services (January 2019 to June 2021) by caregivers in city and rural geographic areas. We compared organizational performance and service quality indicators for 2 financial years (2019-2020 and 2020-2021). A survey was conducted to identify barriers and facilitators to digital service uptake, the computer proficiency of caregivers (the Computer Proficiency Questionnaire, 12-item version), and preferences for future digital service provision. Quantitative data were analyzed using Stata 13 (StataCorp LLC). Thematic analysis was used for open-text survey responses., Results: The number of caregivers registered with the organization rose from 14,817 in 2019 to 20,237 in 2021. Monthly contacts rose from 1929 to 6741, with remote contacts increasing from 48.89% (943/1929) to 86.68% (5843/6741); distinctive patterns were observed for city versus rural caregivers. There was an increase in one-to-one contacts (88.8%) and caregiver assessments (20.9%), with no expansion in staffing. Service quality indicators showed an improvement in 5 of 8 variables (all P<.05). The 152 carers completing the survey had similar demographics to all registered caregivers. The Computer Proficiency Questionnaire, 12-item version, mean score of 25.61 (SD 4.40) indicated relatively high computer proficiency. The analysis of open-text responses identified a preference for the organization to continue to offer face-to-face services as well as web-based options. The digital services that were the most highly rated were carers' well-being assessments, support needs checks, and peer support groups., Conclusions: Our findings show that staff in the caregiver support organization were agile in adapting their services to digital delivery while dealing with increased numbers of registered clients and higher monthly contacts, all without obvious detriment to service quality. Caregivers indicated a preference for blended services, even while recording high computer proficiency. Considering the economic importance of unpaid caregivers, more attention should be given to organizations funded to provide support for them and to the potential for technology to enhance caregivers' access to, and engagement with, such services., (©Ala Szczepura, Amir Jahan Khan, Deidre Wild, Sara Nelson, Sonja Woodhouse, Mark Collinson. Originally published in JMIR Aging (https://aging.jmir.org), 13.05.2024.)

The major cause for skin cancer is the excessive and unprotected exposure to ultraviolet radiation (UVR), which can be prevented by engaging in sun protection behaviours. As longitudinal studies on both adolescents' and adults' performances of sun protection behaviours are limited, the current study aimed to investigate changes in sun protection in these population segments in Western Australia, a region with high annual UVR. During six summer seasons (2015/16 to 2020/21), cross-sectional surveys were conducted among 1806 adolescents (14 to 17 years old) and 1808 adults (18-45 years old), investigating the frequency of five sun protection behaviours (wearing clothing, applying sunscreen, wearing a hat, wearing sunglasses, and seeking shade) and sun avoidance (staying indoors). Over the six-year period, staying indoors increased in both groups. Among adolescents, a decrease in wearing clothing and sunglasses and an increase in seeking shade was demonstrated, and hat and sunscreen use remained relatively stable. Among adults, an increase in sunscreen use was shown, whereas all other sun-related behaviours remained consistent over the six-year period. The results from this study can provide directions for health communications focusing on improving sun protection behaviours among both adolescent and adult populations.

The thalamus is the principal information hub of the vertebrate brain, with essential roles in sensory and motor information processing, attention, and memory. The complex array of thalamic nuclei develops from a restricted pool of neural progenitors. We apply longitudinal single-cell RNA sequencing and regional abrogation of Sonic hedgehog (Shh) to map the developmental trajectories of thalamic progenitors, intermediate progenitors, and post-mitotic neurons as they coalesce into distinct thalamic nuclei. These data reveal that the complex architecture of the thalamus is established early during embryonic brain development through the coordinated action of four cell differentiation lineages derived from Shh-dependent and -independent progenitors. We systematically characterize the gene expression programs that define these thalamic lineages across time and demonstrate how their disruption upon Shh depletion causes pronounced locomotor impairment resembling infantile Parkinson's disease. These results reveal key principles of thalamic development and provide mechanistic insights into neurodevelopmental disorders resulting from thalamic dysfunction., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Conditions have been identified in which phenolic aldoximes and ketoximes of the types used in commercial solvent extraction processes can be doubly deprotonated and generate polynuclear Cu complexes with lower extractant:Cu molar ratios than those found in commercial operations. Electrospray mass spectrometry has provided an insight into the solution speciation in extraction experiments and has identified conditions to allow isolation and characterization of polynuclear Cu-complexes. Elevation of pH is effective in enhancing the formation of trinuclear complexes containing planar {Cu 3 -μ 3 -O} 4+ or {Cu 3 -μ 3 -OH} 5+ units. DFT calculations suggest that such trinuclear complexes are more stable than other polynuclear species. Solid structures of complexes formed by a salicylaldoxime with a piperidino substituent ortho to the phenolic OH group ( L 9 H 2 ) contain two trinuclear units in a supramolecular assembly, {[Cu 3 OH( L 9 H) 3 (ClO 4 )](ClO 4 )} 2 , formed by H-bonding between the central {Cu 3 -μ 3 -OH} 5+ units and oxygen atoms in the ligands of an adjacent complex. Whilst the lower ligand:Cu molar ratios provide more efficient Cu-loading in solvent extraction processes, the requirement to raise the pH of the aqueous phase to achieve this will make it impractical in most commercial operations because extraction will be accompanied by the precipitation (as oxyhydroxides) of Fe(III) which is present in significant quantities in feed solutions generated by acid leaching of most Cu ores.

A notable number of acute lymphoblastic leukemia (ALL) patients develop CD19-positive relapse within 1 year after receiving chimeric antigen receptor (CAR) T cell therapy. It remains unclear if the long-term response is associated with the characteristics of CAR T cells in infusion products, hindering the identification of biomarkers to predict therapeutic outcomes. Here, we present 101,326 single-cell transcriptomes and surface protein landscape from the infusion products of 12 ALL patients. We observed substantial heterogeneity in the antigen-specific activation states, among which a deficiency of T helper 2 function was associated with CD19-positive relapse compared with durable responders (remission, >54 months). Proteomic data revealed that the frequency of early memory T cells, rather than activation or coinhibitory signatures, could distinguish the relapse. These findings were corroborated by independent functional profiling of 49 patients, and an integrative model was developed to predict the response. Our data unveil the molecular mechanisms that may inform strategies to boost specific T cell function to maintain long-term remission.

Health diagnostics of wildlife have historically relied on the evaluation of select serum biomarkers and the identification of a contaminant or pathogen burden within specific tissues as an indicator of a level of insult. However, these approaches fail to measure the physiological reaction of the individual to stressors, thus limiting the scope of interpretation. Gene-based health diagnostics provide an opportunity for an alternate, whole-system, or holistic assessment of health, not only in individuals or populations but potentially in ecosystems. Seabirds are among the most threatened marine taxonomic groups in the world, with ~25% of this species currently listed as threatened or considered of special concern; among seabirds, the penguins (Family Spheniscidae) are the most threatened seabird Family. We used gene expression to develop baseline physiological indices for wild penguins in the Falkland-Malvinas Islands, and captive zoo penguins. We identified the almost complete statistical separation of penguin groups (gentoo Detroit Zoo, gentoo Falkland-Malvinas Islands, rockhopper Detroit Zoo, and rockhopper Falkland-Malvinas Islands) based on gene expression profiles. Implementation of long-term longitudinal studies would allow for the assessment of temporal increases or decreases of select transcripts and would facilitate interpretation of the drivers of change.

Background: Associative transcriptomics has been used extensively in Brassica napus to enable the rapid identification of markers correlated with traits of interest. However, within the important vegetable crop species, Brassica oleracea, the use of associative transcriptomics has been limited due to a lack of fixed genetic resources and the difficulties in generating material due to self-incompatibility. Within Brassica vegetables, the harvestable product can be vegetative or floral tissues and therefore synchronisation of the floral transition is an important goal for growers and breeders. Vernalisation is known to be a key determinant of the floral transition, yet how different vernalisation treatments influence flowering in B. oleracea is not well understood., Results: Here, we present results from phenotyping a diverse set of 69 B. oleracea accessions for heading and flowering traits under different environmental conditions. We developed a new associative transcriptomics pipeline, and inferred and validated a population structure, for the phenotyped accessions. A genome-wide association study identified miR172D as a candidate for the vernalisation response. Gene expression marker association identified variation in expression of BoFLC.C2 as a further candidate for vernalisation response., Conclusions: This study describes a new pipeline for performing associative transcriptomics studies in B. oleracea. Using flowering time as an example trait, it provides insights into the genetic basis of vernalisation response in B. oleracea through associative transcriptomics and confirms its characterisation as a complex G x E trait. Candidate leads were identified in miR172D and BoFLC.C2. These results could facilitate marker-based breeding efforts to produce B. oleracea lines with more synchronous heading dates, potentially leading to improved yields., (© 2021. The Author(s).)

The transcription factor Rora has been shown to be important for the development of ILC2 and the regulation of ILC3, macrophages and Treg cells. Here we investigate the role of Rora across CD4+ T cells in general, but with an emphasis on Th2 cells, both in vitro as well as in the context of several in vivo type 2 infection models. We dissect the function of Rora using overexpression and a CD4-conditional Rora-knockout mouse, as well as a RORA-reporter mouse. We establish the importance of Rora in CD4+ T cells for controlling lung inflammation induced by Nippostrongylus brasiliensis infection, and have measured the effect on downstream genes using RNA-seq. Using a systematic stimulation screen of CD4+ T cells, coupled with RNA-seq, we identify upstream regulators of Rora, most importantly IL-33 and CCL7. Our data suggest that Rora is a negative regulator of the immune system, possibly through several downstream pathways, and is under control of the local microenvironment., Competing Interests: The authors have read the journal’s policy and have the following competing interests: JF was an employee of Microsoft Research Cambridge at the time the study was conducted, but is no longer an employee of the company. EN is an employee of Aleph Labs, but was not employed by the company at the time the study was conducted. GK is an employee of AstraZeneca, but was not employed by the company at the time the study was conducted. This does not alter our adherence to PLOS ONE policies on sharing data and materials. There are no patents, products in development or marketed products associated with this research to declare.

Background: Autologous T cells engineered to express a chimeric antigen receptor (CAR) specific for CD19 molecule have transformed the therapeutic landscape in patients with highly refractory leukemia and lymphoma, and the use of donor-generated allogeneic CAR T is paving the way for further breakthroughs in the treatment of cancer. However, it remains unknown how the intrinsic heterogeneities of these engineered cells mediate therapeutic efficacy and whether allogeneic products match the effectiveness of autologous therapies., Methods: Using single-cell mRNA sequencing in conjunction with CITE-seq, we performed multiomics characterization of CAR T cells generated from healthy donor and patients with acute lymphoblastic leukemia. CAR T cells used in this study were manufactured at the University of Pennsylvania through lentiviral transduction with a CD19-4-1BB-CD3ζ construct. Besides the baseline condition, we engineered NIH-3T3 cells with human CD19 or mesothelin expression to conduct ex vivo antigen-specific or non-antigen stimulation of CAR T cells through 6-hour coculture at a 1:1 ratio., Results: We delineated the global cellular and molecular CAR T landscape and identified that transcriptional CAR tonic signaling was regulated by a mixture of early activation, exhaustion signatures, and cytotoxic activities. On CD19 stimulation, we illuminated the disparities of CAR T cells derived from different origins and found that donor CAR T had more pronounced activation level in correlation with the upregulation of major histocompatibility complex class II genes compared with patient CAR T cells. This finding was independently validated in additional datasets from literature. Furthermore, GM-CSF( CSF2 ) expression was found to be associated with functional gene productions, but it induced little impact on the CAR T activation., Conclusions: Through integrated multiomics profiling and unbiased canonical pathway analyses, our results unveil heterogeneities in the transcriptional, phenotypic, functional, and metabolic profiles of donor and patient CAR T cells, providing mechanistic basis for ameliorating clinical outcomes and developing next-generation 'off- the-shelf' allogeneic products., Competing Interests: Competing interests: RF is a cofounder of IsoPlexis, Singleron Biotechnologies and AtlasXomics and a member of their scientific advisory boards with financial interests, which could affect or have the perception of affecting the author’s objectivity. The interests of RF were reviewed and managed by Yale University Provost’s Office in accordance with the University’s conflict of interest policies., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Comparative transcriptomics can be used to translate an understanding of gene regulatory networks from model systems to less studied species. Here, we use RNA-Seq to determine and compare gene expression dynamics through the floral transition in the model species Arabidopsis thaliana and the closely related crop Brassica rapa . We find that different curve registration functions are required for different genes, indicating that there is no single common 'developmental time' between Arabidopsis and B. rapa . A detailed comparison between Arabidopsis and B. rapa and between two B. rapa accessions reveals different modes of regulation of the key floral integrator SOC1 , and that the floral transition in the B. rapa accessions is triggered by different pathways. Our study adds to the mechanistic understanding of the regulatory network of flowering time in rapid cycling B. rapa and highlights the importance of registration methods for the comparison of developmental gene expression data., Competing Interests: The authors declare no significant competing financial, professional or personal interests which might influence the performance or presentation of this study., (© The Author(s), 2021. Published by Cambridge University Press in association with The John Innes Centre 2021.)

Highly multiplexed immunohistochemistry (mIHC) enables the staining and quantification of dozens of antigens in a tissue section with single-cell resolution. However, annotating cell populations that differ little in the profiled antigens or for which the antibody panel does not include specific markers is challenging. To overcome this obstacle, we have developed an approach for enriching mIHC images with single-cell RNA sequencing data, building upon recent experimental procedures for augmenting single-cell transcriptomes with concurrent antigen measurements. Spatially-resolved Transcriptomics via Epitope Anchoring (STvEA) performs transcriptome-guided annotation of highly multiplexed cytometry datasets. It increases the level of detail in histological analyses by enabling the systematic annotation of nuanced cell populations, spatial patterns of transcription, and interactions between cell types. We demonstrate the utility of STvEA by uncovering the architecture of poorly characterized cell types in the murine spleen using published cytometry and mIHC data of this organ., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).)

Flowering time is a key adaptive and agronomic trait. In Arabidopsis, natural variation in expression levels of the floral repressor FLOWERING LOCUS C (FLC) leads to differences in vernalization. In Brassica napus there are nine copies of FLC. Here, we study how these multiple FLC paralogues determine vernalization requirement as a system. We collected transcriptome time series for Brassica napus spring, winter, semi-winter, and Siberian kale crop types. Modelling was used to link FLC expression dynamics to floral response following vernalization. We show that relaxed selection pressure has allowed expression of FLC paralogues to diverge, resulting in variation of FLC expression during cold treatment between paralogues and accessions. We find that total FLC expression dynamics best explains differences in cold requirement between cultivars, rather than expression of specific FLC paralogues. The combination of multiple FLC paralogues with different expression dynamics leads to rich behaviour in response to cold and a wide range of vernalization requirements in B. napus. We find evidence for different strategies to determine the response to cold in existing winter rapeseed accessions., (© 2020 The Authors New Phytologist © 2020 New Phytologist Foundation.)

Purpose: Liquid biopsy specimen genomic profiling is integrated in non-small-cell lung cancer (NSCLC) guidelines; however, data on the clinical relevance for ALK /ROS1 alterations are scarce. We evaluated the clinical utility of a targeted amplicon-based assay in a large prospective cohort of patients with ALK/ROS1 -positive NSCLC and its impact on outcomes., Patients and Methods: Patients with advanced ALK/ROS1 -positive NSCLC were prospectively enrolled in the study by researchers at eight French institutions. Plasma samples were analyzed using InVisionFirst-Lung and correlated with clinical outcomes., Results: Of the 128 patients included in the study, 101 were positive for ALK and 27 for ROS1 alterations. Blood samples (N = 405) were collected from 29 patients naïve for treatment with tyrosine kinase inhibitors (TKI) or from 375 patients under treatment, including 105 samples collected at disease progression (PD). Sensitivity was 67% (n = 18 of 27) for ALK/ROS1 fusion detection. Higher detection was observed for ALK fusions at TKI failure (n = 33 of 74; 46%) versus in patients with therapeutic response (n = 12 of 109; 11%). ALK -resistance mutations were detected in 22% patients (n = 16 of 74) overall; 43% of the total ALK -resistance mutations identified occurred after next-generation TKI therapy. ALK G1202R was the most common mutation detected (n = 7 of 16). Heterogeneity of resistance was observed. ROS1 G2032R resistance was detected in 30% (n = 3 of 10). The absence of circulating tumor DNA mutations at TKI failure was associated with prolonged median overall survival (105.7 months). Complex ALK -resistance mutations correlated with poor overall survival (median, 26.9 months v NR for single mutation; P = .003) and progression-free survival to subsequent therapy (median 1.7 v 6.3 months; P = .003)., Conclusion: Next-generation, targeted, amplicon-based sequencing for liquid biopsy specimen profiling provides clinically relevant detection of ALK/ROS1 fusions in TKI-naïve patients and allows for the identification of resistance mutations in patients treated with TKIs. Liquid biopsy specimens from patients treated with TKIs may affect clinical outcomes and capture heterogeneity of TKI resistance, supporting their role in selecting sequential therapy., Competing Interests: The following represents disclosure information provided by authors of this manuscript. All relationships are considered compensated unless otherwise noted. Relationships are self-held unless noted. I = Immediate Family Member, Inst = My Institution. Relationships may not relate to the subject matter of this manuscript. For more information about ASCO’s conflict of interest policy, please refer to www.asco.org/rwc or ascopubs.org/po/author-center. Open Payments is a public database containing information reported by companies about payments made to US-licensed physicians (Open Payments). Laura MezquitaConsulting or Advisory Role: Roche, Roche Diagnostics, Takeda Speakers’ Bureau: Roche, Bristol-Myers Squibb, Tecnofarma Travel, Accommodations, Expenses: Roche, Bristol-Myers SquibbAurélie SwalduzHonoraria: Roche, Bristol-Myers Squibb, Takeda, AZD Consulting or Advisory Role: Eli Lilly, Pfizer, Bristol-Myers Squibb Travel, Accommodations, Expenses: Takeda, Pfizer, Boehringer Ingelheim, RocheKaren HowarthEmployment: Inivata Stock and Other Ownership Interests: Inivata Research Funding: Inivata Patents, Royalties, Other Intellectual Property: Patent pendingDavid PlanchardHonoraria: Prime Oncology, Peer CME Consulting or Advisory Role: AstraZeneca, Boehringer Ingelheim, Bristol-Myers Squibb, Novartis, Roche, Pfizer, MSD Oncology, Celgene, MedImmune, BeiGene Research Funding: AstraZeneca/MedImmune (Inst),, Bristol-Myers Squibb (Inst),, Boehringer Ingelheim (Inst), Eli Lilly (Inst), Merck (Inst), Novartis (Inst), Pfizer (Inst), Roche (Inst), Sanofi/Aventis (Inst), Taiho Pharmaceutical (Inst), Daiichi Sankyo (Inst), AbbVie (Inst)Gonzalo RecondoConsulting or Advisory Role: Roche, Amgen, Pfizer Travel, Accommodations, Expenses: AstraZeneca, PfizerFrank De KievitEmployment: Inivata Travel, Accommodations, Expenses: InivataVincent PlagnolEmployment: Inivata, Genomics Stock and Other Ownership Interests: Inivata, Genomics Patents, Royalties, Other Intellectual Property: Inivata patentsEtienne RouleauHonoraria: AstraZeneca (Inst), Roche (Inst), Bristol-Myers Squibb (Inst), AstraZeneca Research Funding: AstraZeneca (Inst) Travel, Accommodations, Expenses: AstraZeneca, Bristol-Myers SquibbPierre FournelConsulting or Advisory Role: Bristol-Myers Squibb, Amgen, MSD Oncology, Eli Lilly, AstraZeneca, Pfizer, AbbVie, Takeda, Roche, AstraZeneca, Amgen, Eli Lilly, Pfizer, Bristol-Myers-Squibb, MSD Oncology Research Funding: Bristol-Myers Squibb (Inst), AstraZeneca (Inst), Amgen (Inst), Pfizer (Inst), Pfizer/EMD Serono (Inst), Ipsen (Inst) Travel, Accommodations, Expenses: Novartis, Roche, Eli Lilly, Pfizer, Boehringer Ingelheim, AstraZeneca, Amgen, Bristol-Myers Squibb, MSD OncologyCaroline CaramellaHonoraria: Bristol-Myers Squibb Honoraria: MSD Oncology, PfizerClaire TissotConsulting or Advisory Role: Roche, Bristol-Myers Squibb, AstraZeneca, MSD OncologyJulien AdamConsulting or Advisory Role: Roche, Bristol-Myers Squibb, AstraZeneca, Merck Sharp & Dohme Research Funding: Pierre Fabre (Inst), Merck Sharp & Dohme (Inst)Samuel WoodhouseEmployment: Inivata Patents, Royalties, Other Intellectual Property: Patents related to fusion technology used by Inivata and in this articleEdouard AuclinHonoraria: Sanofi Genzyme, MundipharmaJordi RemonConsulting or Advisory Role: Pfizer, Bristol-Myers Squibb, Boehringer Ingelheim, MSD Oncology, AstraZeneca, Roche, Inivata, OSE ImmunotherapeuticsClive MorrisEmployment: Inivata Leadership: Inivata Stock and Other Ownership Interests: InivataEmma GreenEmployment: Inivata Stock and Other Ownership Interests: InivataChristophe MassardConsulting or Advisory Role: Amgen, Astellas Pharma, AstraZeneca, Bayer, BeiGene, Bristol-Myers Squibb, Celgene, Debiopharm Group, Roche, Ipsen, Janssen, Eli Lilly, MSD Oncology, Novartis, Pfizer, Sanofi, Orion, Tahio, Blueprint Medicines, Innate Pharma, PharmaMarMaurice PérolConsulting or Advisory Role: Eli Lilly, Roche, Pfizer, AstraZeneca, Boehringer Ingelheim, Merck Sharp & Dohme, Bristol-Myers Squibb, Novartis, Amgen, Takeda, Chugai Pharma Research Funding: AstraZeneca (Inst), Roche (Inst), Takeda (Inst) Travel, Accommodations, Expenses: AstraZeneca, Roche, Bristol-Myers Squibb, Merck Sharp & Dohme, PfizerBenjamin BesseResearch Funding: AstraZeneca (Inst), Pfizer (Inst), Eli Lilly (Inst), Onxeo (Inst), Bristol-Myers Squibb (Inst), Inivata (Inst), AbbVie (Inst), Amgen (Inst), Biogen (Inst), Blueprint Medicines (Inst), Celgene (Inst), GlaxoSmithKline (Inst), Ignyta (Inst), Ipsen (Inst), Merck (Inst), MSD Oncology (Inst), Nektar (Inst), PharmaMar (Inst), Sanofi (Inst), Spectrum Pharmaceuticals (Inst), Takeda (Inst), Tiziana Therapeutics (Inst)Pierre SaintignyHonoraria: HTG Molecular Diagnostics, Inivata, ArcherDx, Bristol-Myers Squibb, Roche Molecular Diagnostics, Roche, AstraZeneca, Novartis, Bristol-Myers Squibb Foundation, Illumina No other potential conflicts of interest were reported., (© 2020 by American Society of Clinical Oncology.)

Claudins regulate paracellular permeability in different tissues. The claudin-binding domain of Clostridium perfringens enterotoxin (cCPE) is a known modulator of a claudin subset. However, it does not efficiently bind to claudin-1 (Cldn1). Cldn1 is a pharmacological target since it is (i) an essential co-receptor for hepatitis C virus (HCV) infections and (ii) a key element of the epidermal barrier limiting drug delivery. In this study, we investigated the potential of a Cldn1-binding cCPE mutant (i) to inhibit HCV entry into hepatocytes and (ii) to open the epidermal barrier. Inhibition of HCV infection by blocking of Cldn1 with cCPE variants was analyzed in the Huh7.5 hepatoma cell line. A model of reconstructed human epidermis was used to investigate modulation of the epidermal barrier by cCPE variants. In contrast to cCPEwt, the Cldn1-binding cCPE-S305P/S307R/S313H inhibited infection of Huh7.5 cells with HCV in a dose-dependent manner. In addition, TJ modulation by cCPE variant-mediated targeting of Cldn1 and Cldn4 opened the epidermal barrier in reconstructed human epidermis. cCPE variants are potent claudin modulators. They can be applied for mechanistic in vitro studies and might also be used as biologics for therapeutic claudin targeting including HCV treatment (host-targeting antivirals) and improvement of drug delivery.

Background: Members of the ZFP36 family of RNA-binding proteins regulate gene expression post-transcriptionally by binding to AU-rich elements in the 3'UTR of mRNA and stimulating mRNA degradation. The proteins within this family target different transcripts in different tissues. In particular, ZFP36 targets myogenic transcripts and may have a role in adult muscle stem cell quiescence. Our study examined the requirement of ZFP36L1 and ZFP36L2 in adult muscle cell fate regulation., Methods: We generated single and double conditional knockout mice in which Zfp36l1 and/or Zfp36l2 were deleted in Pax7-expressing cells. Immunostained muscle sections were used to analyse resting skeletal muscle, and a cardiotoxin-induced injury model was used to determine the regenerative capacity of muscle., Results: We show that ZFP36L1 and ZFP36L2 proteins are expressed in satellite cells. Mice lacking the two proteins in Pax7-expressing cells have reduced body weight and have reduced skeletal muscle mass. Furthermore, the number of satellite cells is reduced in adult skeletal muscle and the capacity of this muscle to regenerate following muscle injury is diminished., Conclusion: ZFP36L1 and ZFP36L2 act redundantly in myogenesis. These findings add further intricacy to the regulation of the cell fate of Pax7-expressing cells in skeletal muscle by RNA-binding proteins.

Background: Reconstruction of executable mechanistic models from single-cell gene expression data represents a powerful approach to understanding developmental and disease processes. New ambitious efforts like the Human Cell Atlas will soon lead to an explosion of data with potential for uncovering and understanding the regulatory networks which underlie the behaviour of all human cells. In order to take advantage of this data, however, there is a need for general-purpose, user-friendly and efficient computational tools that can be readily used by biologists who do not have specialist computer science knowledge., Results: The Single Cell Network Synthesis toolkit (SCNS) is a general-purpose computational tool for the reconstruction and analysis of executable models from single-cell gene expression data. Through a graphical user interface, SCNS takes single-cell qPCR or RNA-sequencing data taken across a time course, and searches for logical rules that drive transitions from early cell states towards late cell states. Because the resulting reconstructed models are executable, they can be used to make predictions about the effect of specific gene perturbations on the generation of specific lineages., Conclusions: SCNS should be of broad interest to the growing number of researchers working in single-cell genomics and will help further facilitate the generation of valuable mechanistic insights into developmental, homeostatic and disease processes.

Aims: This study aimed to examine the relationships between gastroparesis symptom severity, illness perceptions, coping styles, quality of life (QoL), and psychological distress in patients with gastroparesis, guided by the common sense model., Methods: One hundred and seventy-nine adults with gastroparesis (165 females, 14 males; mean age 41.82 years) completed an online questionnaire. The Gastroparesis Cardinal Symptom Index was used to measure gastroparesis symptom severity, QoL was explored using the PAGI-QOL, illness perceptions were measured using the Brief Illness Perception Questionnaire, the Carver Brief COPE scale assessed coping styles, and psychological distress was investigated using the DASS21., Results: Structural equation modeling resulted in a final model with excellent fit. Gastroparesis symptom severity directly influenced illness perceptions (β = .52, p < .001) and QoL (β = .30, p < .001). Illness perceptions directly influenced maladaptive coping (β = - .64, p < .001), psychological distress (β = - .32, p < .001), and QoL (β = .30, p = .01). Maladaptive coping directly influenced psychological distress (β = .62, p < .001), which in turn had a direct influence on QoL (β = - .38, p < .001)., Conclusions: The final model showed that the influence of gastroparesis symptom severity on psychological distress was fully mediated by illness perceptions, while the influence on QoL was partially mediated by illness perceptions. The study provides guidance for the development of psychological interventions targeted toward improving mediating psychological factors.

Introduction: Detection and monitoring of circulating tumor DNA (ctDNA) is rapidly becoming a diagnostic, prognostic and predictive tool in cancer patient care. A growing number of gene targets have been identified as diagnostic or actionable, requiring the development of reliable technology that provides analysis of multiple genes in parallel. We have developed the InVision™ liquid biopsy platform which utilizes enhanced TAm-Seq™ (eTAm-Seq™) technology, an amplicon-based next generation sequencing method for the identification of clinically-relevant somatic alterations at low frequency in ctDNA across a panel of 35 cancer-related genes., Materials and Methods: We present analytical validation of the eTAm-Seq technology across two laboratories to determine the reproducibility of mutation identification. We assess the quantitative performance of eTAm-Seq technology for analysis of single nucleotide variants in clinically-relevant genes as compared to digital PCR (dPCR), using both established DNA standards and novel full-process control material., Results: The assay detected mutant alleles down to 0.02% AF, with high per-base specificity of 99.9997%. Across two laboratories, analysis of samples with optimal amount of DNA detected 94% mutations at 0.25%-0.33% allele fraction (AF), with 90% of mutations detected for samples with lower amounts of input DNA., Conclusions: These studies demonstrate that eTAm-Seq technology is a robust and reproducible technology for the identification and quantification of somatic mutations in circulating tumor DNA, and support its use in clinical applications for precision medicine.

Circulating tumor DNA (ctDNA) analysis is being incorporated into cancer care; notably in profiling patients to guide treatment decisions. Responses to targeted therapies have been observed in patients with actionable mutations detected in plasma DNA at variant allele fractions (VAFs) below 0.5%. Highly sensitive methods are therefore required for optimal clinical use. To enable objective assessment of assay performance, detailed analytical validation is required. We developed the InVisionFirst™ assay, an assay based on enhanced tagged amplicon sequencing (eTAm-Seq™) technology to profile 36 genes commonly mutated in non-small cell lung cancer (NSCLC) and other cancer types for actionable genomic alterations in cell-free DNA. The assay has been developed to detect point mutations, indels, amplifications and gene fusions that commonly occur in NSCLC. For analytical validation, two 10mL blood tubes were collected from NSCLC patients and healthy volunteer donors. In addition, contrived samples were used to represent a wide spectrum of genetic aberrations and VAFs. Samples were analyzed by multiple operators, at different times and using different reagent Lots. Results were compared with digital PCR (dPCR). The InVisionFirst assay demonstrated an excellent limit of detection, with 99.48% sensitivity for SNVs present at VAF range 0.25%-0.33%, 92.46% sensitivity for indels at 0.25% VAF and a high rate of detection at lower frequencies while retaining high specificity (99.9997% per base). The assay also detected ALK and ROS1 gene fusions, and DNA amplifications in ERBB2, FGFR1, MET and EGFR with high sensitivity and specificity. Comparison between the InVisionFirst assay and dPCR in a series of cancer patients showed high concordance. This analytical validation demonstrated that the InVisionFirst assay is highly sensitive, specific and robust, and meets analytical requirements for clinical applications.

Aims and Objectives: To build on the understanding of how individuals experience gastroparesis, how gastroparesis impacts on their lives and how they adapt to living with gastroparesis., Background: Gastroparesis is a neurogastroenterological disorder associated with increased psychological distress and reduced quality of life. Research shows that gastroparesis poses a significant burden across many facets of life; however, less is known about how individuals cope and adapt to living with the condition., Design: The study employed an interpretive phenomenological approach with semistructured interviews and thematic analysis., Methods: Ten gastroparesis patients were interviewed over the telephone (n = 8), Skype (n = 1) or face-to-face (n = 1). All interviews were audio-recorded and transcribed., Results: Key themes identified: (1) frustration, (2) identity and (3) coping and adaptation. Gastroparesis patients experience significant frustration around their diagnostic journey, being misunderstood and the burden of living with the illness. Patients differed in how they identified with the illness, and this appeared to be associated with adaptation and whether they remained socially engaged., Conclusions: Gastroparesis is associated with significant frustration and burden; however, some patients adapt to living with the condition more effectively than others. Identity appears to play an important role in this relationship. Support aimed at fostering a health-focused and resilient identity may assist gastroparesis patients in adaptation., Relevance to Clinical Practice: The findings of this study can help nurses and other healthcare professionals better understand the experience of living with gastroparesis and the factors that help patients best adapt to living with the condition. Nurses can help promote resilience in patients by discussing the importance of being health-focused rather than illness-focused. Nurses can also support patients by helping them problem-solve issues that may arise around social eating and remaining socially engaged., (© 2017 John Wiley & Sons Ltd.)

Individual human epidermal cells differ in their self-renewal ability. To uncover the molecular basis for this heterogeneity, we performed genome-wide pooled RNA interference screens and identified genes conferring a clonal growth advantage on normal and neoplastic (cutaneous squamous cell carcinoma, cSCC) human epidermal cells. The Hippo effector YAP was amongst the top positive growth regulators in both screens. By integrating the Hippo network interactome with our data sets, we identify WW-binding protein 2 (WBP2) as an important co-factor of YAP that enhances YAP/TEAD-mediated gene transcription. YAP and WPB2 are upregulated in actively proliferating cells of mouse and human epidermis and cSCC, and downregulated during terminal differentiation. WBP2 deletion in mouse skin results in reduced proliferation in neonatal and wounded adult epidermis. In reconstituted epidermis YAP/WBP2 activity is controlled by intercellular adhesion rather than canonical Hippo signalling. We propose that defective intercellular adhesion contributes to uncontrolled cSCC growth by preventing inhibition of YAP/WBP2.

Aim: To systematically review literature addressing three key psychologically-oriented controversies associated with gastroparesis., Methods: A comprehensive search of PubMed, CINAHL, and PsycINFO databases was performed to identify literature addressing the relationship between gastroparesis and psychological factors. Two researchers independently screened all references. Inclusion criteria were: an adult sample of gastroparesis patients, a quantitative methodology, and at least one of the following: (1) evaluation of the prevalence of psychopathology; (2) an outcome measure of anxiety, depression, or quality of life; and (3) evidence of a psychological intervention. Case studies, review articles, and publications in languages other than English were excluded from the current review., Results: Prevalence of psychopathology was evaluated by three studies ( n = 378), which found that combined anxiety/depression was present in 24% of the gastroparesis cohort, severe anxiety in 12.4%, depression in 21.8%-23%, and somatization in 50%. Level of anxiety and depression was included as an outcome measure in six studies ( n = 1408), and while limited research made it difficult to determine the level of anxiety and depression in the cohort, a clear positive relationship with gastroparesis symptom severity was evident. Quality of life was included as an outcome measure in 11 studies ( n = 2076), with gastroparesis patients reporting lower quality of life than population norms, and a negative relationship between quality of life and symptom severity. One study assessed the use of a psychological intervention for gastroparesis patients ( n = 120) and found that depression and gastric function were improved in patients who received psychological intervention, however the study had considerable methodological limitations., Conclusion: Gastroparesis is associated with significant psychological distress and poor quality of life. Recommendations for future studies and the development of psychological interventions are provided., Competing Interests: Conflict-of-interest statement: The authors have no conflicting interests to declare.

Background: Distress is regarded as the sixth vital sign in cancer care, but few studies describe distress in patients undergoing radiation therapy., Objectives: The purpose of this study was to assess distress levels among patients with breast or prostate cancer undergoing radiation therapy and investigate which problems contribute to patients' distress levels., Methods: A retrospective medical record review was conducted for 217 patients with breast or prostate cancer at a midwestern community cancer center. Demographic data, distress scores, and problems or concerns from the patient-completed Distress Thermometer and associated Problem List were collected. Descriptive and bivariate statistics were calculated., Findings: The average distress of patients with breast cancer was significantly higher than that of patients with prostate cancer, and patients with breast cancer reported more problems than those with prostate cancer.

Competing Interests: Competing interests: None declared.

Background: Rapid technological innovation for the generation of single-cell genomics data presents new challenges and opportunities for bioinformatics analysis. One such area lies in the development of new ways to train gene regulatory networks. The use of single-cell expression profiling technique allows the profiling of the expression states of hundreds of cells, but these expression states are typically noisier due to the presence of technical artefacts such as drop-outs. While many algorithms exist to infer a gene regulatory network, very few of them are able to harness the extra expression states present in single-cell expression data without getting adversely affected by the substantial technical noise present., Results: Here we introduce BTR, an algorithm for training asynchronous Boolean models with single-cell expression data using a novel Boolean state space scoring function. BTR is capable of refining existing Boolean models and reconstructing new Boolean models by improving the match between model prediction and expression data. We demonstrate that the Boolean scoring function performed favourably against the BIC scoring function for Bayesian networks. In addition, we show that BTR outperforms many other network inference algorithms in both bulk and single-cell synthetic expression data. Lastly, we introduce two case studies, in which we use BTR to improve published Boolean models in order to generate potentially new biological insights., Conclusions: BTR provides a novel way to refine or reconstruct Boolean models using single-cell expression data. Boolean model is particularly useful for network reconstruction using single-cell data because it is more robust to the effect of drop-outs. In addition, BTR does not assume any relationship in the expression states among cells, it is useful for reconstructing a gene regulatory network with as few assumptions as possible. Given the simplicity of Boolean models and the rapid adoption of single-cell genomics by biologists, BTR has the potential to make an impact across many fields of biomedical research.

Advances in synthetic genomics are now well underway in yeasts due to the low cost of synthetic DNA. These new capabilities also bring greater need for quantitating the presence, loss and rearrangement of loci within synthetic yeast genomes. Methods for achieving this will ideally; i) be robust to industrial settings, ii) adhere to a global standard and iii) be sufficiently rapid to enable at-line monitoring during cell growth. The methylotrophic yeast Pichia pastoris (P. pastoris) is increasingly used for industrial production of biotherapeutic proteins so we sought to answer the following questions for this particular yeast species. Is time-consuming DNA purification necessary to obtain accurate end-point polymerase chain reaction (e-pPCR) and quantitative PCR (qPCR) data? Can the novel linear regression of efficiency qPCR method (LRE qPCR), which has properties desirable in a synthetic biology standard, match the accuracy of conventional qPCR? Does cell cultivation scale influence PCR performance? To answer these questions we performed e-pPCR and qPCR in the presence and absence of cellular material disrupted by a mild 30s sonication procedure. The e-pPCR limit of detection (LOD) for a genomic target locus was 50pg (4.91×10(3) copies) of purified genomic DNA (gDNA) but the presence of cellular material reduced this sensitivity sixfold to 300pg gDNA (2.95×10(4) copies). LRE qPCR matched the accuracy of a conventional standard curve qPCR method. The presence of material from bioreactor cultivation of up to OD600=80 did not significantly compromise the accuracy of LRE qPCR. We conclude that a simple and rapid cell disruption step is sufficient to render P. pastoris samples of up to OD600=80 amenable to analysis using LRE qPCR which we propose as a synthetic biology standard., (Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Differentiation of lymphocytes is frequently accompanied by cell cycle changes, interplay that is of central importance for immunity but is still incompletely understood. Here, we interrogate and quantitatively model how proliferation is linked to differentiation in CD4+ T cells., Results: We perform ex vivo single-cell RNA-sequencing of CD4+ T cells during a mouse model of infection that elicits a type 2 immune response and infer that the differentiated, cytokine-producing cells cycle faster than early activated precursor cells. To dissect this phenomenon quantitatively, we determine expression profiles across consecutive generations of differentiated and undifferentiated cells during Th2 polarization in vitro. We predict three discrete cell states, which we verify by single-cell quantitative PCR. Based on these three states, we extract rates of death, division and differentiation with a branching state Markov model to describe the cell population dynamics. From this multi-scale modelling, we infer a significant acceleration in proliferation from the intermediate activated cell state to the mature cytokine-secreting effector state. We confirm this acceleration both by live imaging of single Th2 cells and in an ex vivo Th1 malaria model by single-cell RNA-sequencing., Conclusion: The link between cytokine secretion and proliferation rate holds both in Th1 and Th2 cells in vivo and in vitro, indicating that this is likely a general phenomenon in adaptive immunity.

New single-cell technologies readily permit gene expression profiling of thousands of cells at single-cell resolution. In this review, we will discuss methods for visualisation and interpretation of single-cell gene expression data, and the computational analysis needed to go from raw data to predictive executable models of gene regulatory network function. We will focus primarily on single-cell real-time quantitative PCR and RNA-sequencing data, but much of what we cover will also be relevant to other platforms, such as the mass cytometry technology for high-dimensional single-cell proteomics.

There is increasing evidence that adult attachment plays a role in the development and perseverance of symptoms of posttraumatic stress disorder (PTSD). This meta-analysis aims to synthesise this evidence and investigate the relationship between adult attachment styles and PTSD symptoms. A random-effects model was used to analyse 46 studies (N=9268) across a wide range of traumas. Results revealed a medium association between secure attachment and lower PTSD symptoms (ρˆ=-.27), and a medium association, in the opposite direction, between insecure attachment and higher PTSD symptoms (ρˆ=.26). Attachment categories comprised of high levels of anxiety most strongly related to PTSD symptoms, with fearful attachment displaying the largest association (ρˆ=.44). Dismissing attachment was not significantly associated with PTSD symptoms. The relationship between insecure attachment and PTSD was moderated by type of PTSD measure (interview or questionnaire) and specific attachment category (e.g. secure, fearful). Results have theoretical and clinical significance., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Reconstruction of the molecular pathways controlling organ development has been hampered by a lack of methods to resolve embryonic progenitor cells. Here we describe a strategy to address this problem that combines gene expression profiling of large numbers of single cells with data analysis based on diffusion maps for dimensionality reduction and network synthesis from state transition graphs. Applying the approach to hematopoietic development in the mouse embryo, we map the progression of mesoderm toward blood using single-cell gene expression analysis of 3,934 cells with blood-forming potential captured at four time points between E7.0 and E8.5. Transitions between individual cellular states are then used as input to develop a single-cell network synthesis toolkit to generate a computationally executable transcriptional regulatory network model of blood development. Several model predictions concerning the roles of Sox and Hox factors are validated experimentally. Our results demonstrate that single-cell analysis of a developing organ coupled with computational approaches can reveal the transcriptional programs that underpin organogenesis.

Objective: To describe ophthalmic examination findings and standard diagnostic test results in 2 penguin species., Animals Studied: Macaroni & Southern Rockhopper Penguins., Procedure: Complete ophthalmic examinations including Schirmer tear test (STT), modified phenol red thread test (PTT), tonometry, and echobiometry were performed on penguins housed at the Detroit Zoo. Mean and standard deviation of ophthalmic tests are reported and compared for significance using two sample t-tests with significance set at P < 0.05. Correlations between variables were assessed using Pearson's correlation coefficient., Results: Cataracts were the most common finding, present in 64% of Macaroni Penguins, and 68% of Rockhopper Penguins. There were anterior segment anomalies in all eyes with cataracts consistent with lens-induced uveitis. The mean modified PTT for the Macaronis was 24.7 ± 6.37 mm/15 s and 25.1 ± 7.07 mm/15 s in the Rockhoppers. The mean STT value for the Macaronis was 12.1 ± 5.43 mm/min and 11.0 ± 3.96 mm/min in the Rockhoppers. Mean intraocular pressure (IOP) for the Macaronis was 21.9 ± 7.05 mmHg measured by applanation tonometry and 29.1 ± 7.16 mmHg using rebound tonometry. The Rockhoppers had a mean IOP of 20.0 ± 5.77 mmHg and 24.1 ± 5.09 mmHg for applanation and rebound tonometry, respectively. In both populations, there was a significant difference in IOP measurement between the two instruments. In the Macaroni penguins, the presence of cataracts correlated significantly with increased age and lower IOP readings. Anterior chamber distance and axial globe length were significantly greater in males than in females in both penguin species., (© 2013 American College of Veterinary Ophthalmologists.)

Metastatic colonization of distant organs underpins the majority of human-cancer-related deaths, including deaths from head and neck squamous cell carcinoma (HNSCC). We report that miR-203, a miRNA that triggers differentiation in multilayered epithelia, inhibits multiple postextravasation events during HNSCC lung metastasis. Inducible reactivation of miR-203 in already established lung metastases reduces the overall metastatic burden. Using an integrated approach, we reveal that miR-203 inhibits metastasis independently of its effects on differentiation. In vivo genetic reconstitution experiments show that miR-203 inhibits lung metastasis by suppressing the prometastatic activities of three factors involved in cytoskeletal dynamics (LASP1), extracellular matrix remodeling (SPARC), and cell metabolism (NUAK1). Expression of miR-203 and its downstream effectors correlates with HNSCC overall survival outcomes, indicating the therapeutic potential of targeting this signaling axis., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

Transcription factors (TFs) act within wider regulatory networks to control cell identity and fate. Numerous TFs, including Scl (Tal1) and PU.1 (Spi1), are known regulators of developmental and adult haematopoiesis, but how they act within wider TF networks is still poorly understood. Transcription activator-like effectors (TALEs) are a novel class of genetic tool based on the modular DNA-binding domains of Xanthomonas TAL proteins, which enable DNA sequence-specific targeting and the manipulation of endogenous gene expression. Here, we report TALEs engineered to target the PU.1-14kb and Scl+40kb transcriptional enhancers as efficient new tools to perturb the expression of these key haematopoietic TFs. We confirmed the efficiency of these TALEs at the single-cell level using high-throughput RT-qPCR, which also allowed us to assess the consequences of both PU.1 activation and repression on wider TF networks during developmental haematopoiesis. Combined with comprehensive cellular assays, these experiments uncovered novel roles for PU.1 during early haematopoietic specification. Finally, transgenic mouse studies confirmed that the PU.1-14kb element is active at sites of definitive haematopoiesis in vivo and PU.1 is detectable in haemogenic endothelium and early committing blood cells. We therefore establish TALEs as powerful new tools to study the functionality of transcriptional networks that control developmental processes such as early haematopoiesis., (© 2014. Published by The Company of Biologists Ltd.)

Hematopoiesis represents one of the paradigmatic systems for studying stem cell biology, but our understanding of how the hematopoietic system develops during embryogenesis is still incomplete. While many lessons have been learned from studying the mouse embryo, embryonic stem cells have come to the fore as an alternative and more tractable model to recapitulate hematopoietic development. Here we review what is known about the embryonic origin of blood from these complementary systems and how transcription factor networks regulate the emergence of hematopoietic tissue from the mesoderm. Furthermore, we have performed an integrated analysis of genome-wide microarray and ChIP-seq data sets from mouse embryos and embryonic stem (ES) cell lines deficient in key regulators and demonstrate how this type of analysis can be used to reconstruct regulatory hierarchies that both confirm existing regulatory linkages and suggest additional interactions., (© 2013.)

Stem cell function is essential for the maintenance of adult tissue homeostasis. Controlling the balance between self-renewal and differentiation is crucial to maintain a receptive satellite cell pool capable of responding to growth and regeneration cues. The mitogen-activated protein kinase p38α has been implicated in the regulation of these processes but its influence in adult muscle remains unknown. Using conditional satellite cell p38α knockout mice we have demonstrated that p38α restricts excess proliferation in the postnatal growth phase while promoting timely myoblast differentiation. Differentiation was still able to occur in the p38α-null satellite cells, however, but was delayed. An absence of p38α resulted in a postnatal growth defect along with the persistence of an increased reservoir of satellite cells into adulthood. This population was still capable of responding to cardiotoxin-induced injury, resulting in complete, albeit delayed, regeneration, with further enhancement of the satellite cell population. Increased p38γ phosphorylation accompanied the absence of p38α, and inhibition of p38γ ex vivo substantially decreased the myogenic defect. We have used genome-wide transcriptome analysis to characterize the changes in expression that occur between resting and regenerating muscle, and the influence p38α has on these expression profiles. This study provides novel evidence for the fundamental role of p38α in adult muscle homeostasis in vivo., (Copyright © 2013 AlphaMed Press.)

Tissue generation and repair requires a stepwise process of cell fate restriction to ensure that adult stem cells differentiate in a timely and appropriate manner. A crucial role has been implicated for Polycomb-group (PcG) proteins and the H3K27me3 repressive histone mark in coordinating the transcriptional programmes necessary for this process, but the targets and developmental timing for this repression remain unclear. To address these questions, we generated novel genome-wide maps of H3K27me3 and H3K4me3 in freshly isolated muscle stem cells. These data, together with the analysis of two conditional Ezh2-null mouse strains, identified a critical proliferation phase in which Ezh2 activity is essential. Mice lacking Ezh2 in satellite cells exhibited decreased muscle growth, severely impaired regeneration and reduced stem cell number, due to a profound failure of the proliferative progenitor population to expand. Surprisingly, deletion of Ezh2 after the onset of terminal differentiation did not impede muscle repair or homeostasis. Using these knockout models and the RNA-Seq and ChIP-Seq datasets, we show that Ezh2 does not regulate the muscle differentiation process in vivo. These results emphasise the lineage and cell-type-specific functions of Ezh2 and Polycomb repressive complex 2.

Background: Fibrates have been prescribed for decades as 'broad-spectrum' lipid modifying agents that can improve plasma levels of triglycerides, high-density lipoprotein cholesterol, and triglyceride-rich lipoproteins, including very low- and intermediate-density lipoproteins. Fibrates are variably effective in lowering low-density cholesterol levels. Available fibrates include gemfibrozil, fenofibrate, bezafibrate, etiofibrate and ciprofobrate; only fenofibrate and gemfibrozil are available in Australia., Methods: Members of the Queensland Lipid Group provided consensus grades of recommendations for the clinical use of fibrates based on PubMed searches, product information, and personal clinical experience., Results: Fibrates are well tolerated, and the combination of fenofibrate with statins appears to be safer than gemfibrozil, particularly with regard to adverse effects on muscle. Evidence has been provided recently for the efficacy of fenofibrate in reducing microvascular complications in diabetic patients, including progression of retinopathy, progression of microalbuminuria and nephropathy, development of sensory neuropathy, and leg amputation. Macrovascular benefits appear to be confined to those with reduced high-density lipoprotein cholesterol and/or increased triglyceride levels, and the relationship of microvascular benefits of fenofibrate to baseline lipid levels is variable and requires further assessment., Conclusions:   Indications for fibrate therapy may be extended in the future to include protection from both macro- and micro-vascular disease, particularly in diabetic patients and patients with residual dyslipidaemia in spite of statin therapy. We provide recommendations on the use of fibrates in clinical practice to highlight these potential indications., (© 2012 The Authors. International Journal of Evidence-Based Healthcare © 2012 The Joanna Briggs Institute.)

This article examines how the nature of competition between brands in a therapeutic category changes after generic entry and provide a framework for analyzing the effect of generic entry on consumer welfare that takes into account the generic free riding problem. It demonstrates that changes in competition along dimensions other than retail price--such as competition in research and development efforts and in promotional activities--may, in certain situations, result in generic entry having an overall negative impact on consumer welfare.