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Junji Lin,1 Yunfeng Li,2 Haijun Tian,2 Michael J Goodman,1 Susan Gabriel,2 Tara Nazareth,2 Stuart J Turner,2,3 Stephen Arcona,2 Kristijan H Kahler21Department of Pharmacotherapy, University of Utah, Salt Lake City, UT, USA; 2Health Economics and Outcomes Research, Novartis Pharmaceuticals Corporation, East Hanover, NJ, USA; 3Ernest Mario School of Pharmacy, Rutgers University, New Brunswick, NJ, USA Background: Patients with chronic obstructive pulmonary disease (COPD) are at increased risk for lung infections and other pathologies (eg, pneumonia); however, few studies have evaluated the impact of pneumonia on health care resource utilization and costs in this population. The purpose of this study was to estimate health care resource utilization and costs among COPD patients with newly acquired pneumonia compared to those without pneumonia. Methods: A retrospective claims analysis using Truven MarketScan® Commercial and Medicare databases was conducted. COPD patients with and without newly acquired pneumonia diagnosed between January 1, 2004 and September 30, 2011 were identified. Propensity score matching was used to create a 1:1 matched cohort. Patient demographics, comorbidities (measured by Charlson Comorbidity Index), and medication use were evaluated before and after matching. Health care resource utilization (ie, hospitalizations, emergency room [ER] and outpatient visits), and associated health care costs were assessed during the 12-month follow-up. Logistic regression was conducted to evaluate the risk of hospitalization and ER visits, and gamma regression models and two-part models compared health care costs between groups after matching. Results: In the baseline cohort (N=467,578), patients with newly acquired pneumonia were older (mean age: 70 versus [vs] 63 years) and had higher Charlson Comorbidity Index scores (3.3 vs 2.6) than patients without pneumonia. After propensity score matching, the pneumonia cohort was nine times more likely to have a hospitalization (odds ratio; 95% confidence intervals [CI] =9.2; 8.9, 9.4) and four times more likely to have an ER visit (odds ratio; 95% CI =4.4; 4.3, 4.5) over the 12-month follow-up period compared to the control cohort. The estimated 12-month mean hospitalization costs ($14,353 [95% CI: $14,037–$14,690]), outpatient costs ($6,891 [95% CI: $6,706–$7,070]), and prescription drug costs ($1,104 [95% CI: $1,054–$1,142]) were higher in the pneumonia cohort than in the control cohort. Conclusion: This study demonstrated elevated health care resource use and costs in patients with COPD after acquiring pneumonia compared to those without pneumonia. Keywords: COPD, health care costs, cost effectiveness/economics, outcomes research

© 2018 Author(s). Diamond formation in polystyrene (C8H8)n, which is laser-compressed and heated to conditions around 150 GPa and 5000 K, has recently been demonstrated in the laboratory [Kraus et al., Nat. Astron. 1, 606-611 (2017)]. Here, we show an extended analysis and comparison to first-principles simulations of the acquired data and their implications for planetary physics and inertial confinement fusion. Moreover, we discuss the advanced diagnostic capabilities of adding high-quality small angle X-ray scattering and spectrally resolved X-ray scattering to the platform, which shows great prospects of precisely studying the kinetics of chemical reactions in dense plasma environments at pressures exceeding 100 GPa.

Background: Despite the substantial burden of asthma-related emergency department (ED) visits, there have been no recent multicenter efforts to characterize this high-risk population. Objective: We aimed to characterize patients with asthma according to their frequency of ED visits and to identify factors associated with frequent ED visits. Methods: A multicenter chart review study of 48 EDs across 23 US states. We identified ED patients ages 18 to 54 years with acute asthma during 2011 and 2012. Primary outcome was frequency of ED visits for acute asthma in the past year, excluding the index ED visit. Results: Of the 1890 enrolled patients, 863 patients (46%) had 1 or more (frequent) ED visits in the past year. Specifically, 28% had 1 to 2 visits, 11% had 3 to 5 visits, and 7% had 6 or more visits. Among frequent ED users, guideline-recommended management was suboptimal. For example, of patients with 6 or more ED visits, 85% lacked evidence of prior evaluation by an asthma specialist, and 43% were not treated with inhaled corticosteroids. In a multivariable model, significant predictors of frequent ED visits were public insurance, no insurance, and markers for chronic asthma severity (all P < .05). Stronger associations were found among those with a higher frequency of asthma-related ED visits (eg, 6 or more ED visits). Conclusion: This multicenter study of US adults with acute asthma demonstrated many frequent ED users and suboptimal preventive management in this high-risk population. Future reductions in asthma morbidity and associated health care utilization will require continued efforts to bridge these majorgaps in asthma care.

The paper describes the settings for an official dialogue between religious communities, including the Catholic Church, and the institutions of the European Union. It goes on to describe some important and necessary contributions that a Christian vision can make to the European project. Finally it considers how far certain styles (rather than content) of Catholic discourse can impede the reception of this vision, and so render effective dialogue more difficult. [ABSTRACT FROM AUTHOR]

BACKGROUND: Tumor necrosis factor alpha (TNF-alpha) mediates inflammation. High TNF-alpha production has adverse effects during disease. Polymorphisms in the TNF-alpha and lymphotoxin alpha genes influence TNF-alpha production. Fish oil suppresses TNF-alpha production and has variable antiinflammatory effects on disease. OBJECTIVE: We examined the relation between TNF-alpha and lymphotoxin alpha genotypes and the ability of dietary fish oil to suppress TNF-alpha production by peripheral blood mononuclear cells (PBMCs) in healthy men. DESIGN: Polymorphisms in the TNF-alpha (TNF*1 and TNF*2) and lymphotoxin alpha (TNFB*1 and TNFB*2) genes were determined in 111 healthy young men. TNF-alpha production by endotoxin-stimulated PBMCs was measured before and 12 wk after dietary supplementation with fish oil (6 g/d). RESULTS: Homozygosity for TNFB*2 was 2.5 times more frequent in the highest than in the lowest tertile of inherent TNF-alpha production. The percentage of subjects in whom fish oil suppressed TNF-alpha production was lowest (22%) in the lowest tertile and doubled with each ascending tertile. In the highest and lowest tertiles, mean TNF-alpha production decreased by 43% (P < 0.05) and increased by 160% (P < 0.05), respectively. In the lowest tertile of TNF-alpha production, only TNFB*1/TNFB*2 heterozygous subjects were responsive to the suppressive effect of fish oil. In the middle tertile, this genotype was 6 times more frequent than the other lymphotoxin alpha genotypes among responsive individuals. In the highest tertile, responsiveness to fish oil appeared unrelated to lymphotoxin alpha genotype. CONCLUSION: The ability of fish oil to decrease TNF-alpha production is influenced by inherent TNF-alpha production and by polymorphisms in the TNF-alpha and lymphotoxin alpha genes. Copyright © 2002 American Society for Clinical Nutrition [ABSTRACT FROM AUTHOR]

AIDS helplines provide an important public health service in disseminating information on HIV/AIDS, sexual risk behaviors, testing information, and situations that do not confer the risk of HIV acquisition. The Alaskan AIDS Assistance Association (AAAA) maintained a telephone helpline for the state of Alaska. This paper evaluated data collection efforts at this community-based organization. Data concerning the demographics of callers and topics and patterns of helpline utilization for the years 1991,1992,1995, and 1996 were investigated. Differences were found in topics of inquiry with respect to caller gender and time of day in which the call was logged, using correspondence analysis. Men were more likely to request information on HIV testing and safe-sex practices. Women were more likely to request information on other topics such as household transmission of HIV and information on transmission of HIV to children in settings such as childcare centers and schools. Knowledge of caller demographics and concerns has been used by AAAA to improve helpline services and volunteer training. Revelations concerning missing data have resulted in changes to data collection procedures. [ABSTRACT FROM AUTHOR]

Background: Cigarette smoke has both pro-inflammatory and immunosuppressive effects. Both active and passive cigarette smoke exposure are linked to an increased incidence and severity of respiratory virus infections, but underlying mechanisms are not well defined. We hypothesized, based on prior gene expression profiling studies, that upregulation of pro-inflammatory mediators by short term smoke exposure would be protective against a subsequent influenza infection.Methods: BALB/c mice were subjected to whole body smoke exposure with 9 cigarettes/day for 4 days. Mice were then infected with influenza A (H3N1, Mem71 strain), and analyzed 3 and 10 days later (d3, d10). These time points are the peak and resolution (respectively) of influenza infection.Results: Inflammatory cell influx into the bronchoalveolar lavage (BALF), inflammatory mediators, proteases, histopathology, viral titres and T lymphocyte profiles were analyzed. Compared to smoke or influenza alone, mice exposed to smoke and then influenza had more macrophages, neutrophils and total lymphocytes in BALF at d3, more macrophages in BALF at d10, lower net gelatinase activity and increased activity of tissue inhibitor of metalloprotease-1 in BALF at d3, altered profiles of key cytokines and CD4+ and CD8+ T lymphocytes, worse lung pathology and more virus-specific, activated CD8+ T lymphocytes in BALF. Mice smoke exposed before influenza infection had close to 10-fold higher lung virus titres at d3 than influenza alone mice, although all mice had cleared virus by d10, regardless of smoke exposure. Smoke exposure caused temporary weight loss and when smoking ceased after viral infection, smoke and influenza mice regained significantly less weight than smoke alone mice.Conclusion: Smoke induced inflammation does not protect against influenza infection.In most respects, smoke exposure worsened the host response to influenza. This animal model may be useful in studying how smoke worsens respiratory viral infections. [ABSTRACT FROM AUTHOR]

Activation of CD8 + T cells enable them to control virus infections and tumors. This process involves the differentiation of naïve CD8 + T cells into effector and memory states, driven by specific transcription factors (TFs). Previously, we have shown that Granzyme A (Gzma) induction in activated CD8 + T cells depends on Gata3 and the establishment of a permissive chromatin landscape at the Gzma locus. Interestingly, Gzma expression is independent of IL-4 signaling, which typically upregulates Gata3 in CD4 + T cells, suggesting an alternative pathway for Gata3 induction. Here we demonstrate that Notch signals during CD8 + T cell activation promote Gzma expression. Inhibition of Notch signaling or loss of the Notch transactivator Rbp-j leads to reduced Gzma expression, with transcriptionally repressive chromatin at the Gzma locus. The genome targets of Gata3 differ in effector CD8 + T cells activated with IL-4 compared with those activated with Notch signals or isolated after IAV infection. This indicates that the signals received during CD8 + T cell activation can alter the chromatin landscape, affecting Gata3 function. Furthermore, Gata3 deficiency results in reduced IAV-specific CD8 + T cell responses and decreased Gzma expression, although the Gzma locus maintains a permissive chromatin landscape. These findings suggest that Notch signals received by virus-specific CD8 + T cells prepare the chromatin landscape for Gata3 binding to CD8 + lineage-specific gene loci, promoting effective CD8 + T cell immunity., (© 2025 The Author(s). Immunology & Cell Biology published by John Wiley & Sons Australia, Ltd on behalf of the Australian and New Zealand Society for Immunology, Inc.)

Core-shell nanoparticles can exhibit strongly enhanced performances in electro-, photo- and thermal catalysis. Lattice strain plays a key role in this and is induced by the mismatch between the crystal structure of the core and the shell metal. However, investigating the impact of lattice strain has been challenging due to the lack of a material system in which lattice strain can be controlled systematically, hampering further progress in the field of core-shell catalysis. In this work, we achieve such a core-shell nanoparticle system through the colloidal synthesis of trimetallic Pt-shell Au 1- x Cu x -core nanoparticles. Our seed-mediated growth methodology yields well-defined Au 1- x Cu x -cores, tunable in composition from 0 at% Cu to 77 at% Cu, and monodisperse in size. Subsequent overgrowth results in uniform, epitaxially grown Pt-shells with a controlled thickness of ∼3 atomic layers. By employing a multi-technique characterization strategy combining X-ray diffraction, electron diffraction and aberration corrected electron microscopy, we unravel the atomic structure of the trimetallic system on a single nanoparticle-, ensemble- and bulk scale level, and we unambiguously demonstrate the controlled variation of strain in the Pt-shell from -3.62% compressive-, to +3.79% tensile strain, while retaining full control over all other structural characteristics of the system.

Unavoidable water formation during the reduction of solid catalyst precursors has long been known to influence the nanoparticle size and dispersion in the active catalyst. This in situ transmission electron microscopy study provides insight into the influence of water vapor at the nanoscale on the nucleation and growth of the nanoparticles (2-16 nm) during the reduction of a nickel phyllosilicate catalyst precursor under H 2 /Ar gas at 700 °C. Water suppresses and delays nucleation, but counterintuitively increases the rate of particle growth. After full reduction is achieved, water vapor significantly enhances Ostwald ripening which in turn increases the likelihood of particle coalescence. This study proposes that water leads to formation of mobile nickel hydroxide species, leading to faster rates of particle growth during and after reduction., (© 2024 The Authors. Small published by Wiley‐VCH GmbH.)

Objectives: Despite vaccination strategies, people with chronic kidney disease, particularly kidney transplant recipients (KTRs), remained at high risk of poor COVID-19 outcomes. We assessed serological responses to the three-dose COVID-19 vaccine schedule in KTRs and people on dialysis, as well as seroresponse predictors and the relationship between responses and breakthrough infection., Methods: Plasma from 30 KTRs and 17 people receiving dialysis was tested for anti-Spike receptor binding domain (RBD) IgG and neutralising antibodies (NAb) to the ancestral and Omicron BA.2 variant after Doses 2 and 3 of vaccination., Results: After three doses, KTRs achieved lower anti-Spike RBD IgG levels ( P  < 0.001) and NAb titres than people receiving dialysis ( P  = 0.002). Seropositive cross-reactive Omicron neutralisation levels were achieved in 11/27 (40.7%) KTRs and 11/14 (78.6%) dialysis recipients. ChAdOx1/viral-vector vaccine type, higher mycophenolate dose (> 1 g per day) and lower absolute B-cell counts predicted poor serological responses in KTRs. ChAdOx-1 vaccine type and higher monocyte counts were negative predictors in dialysis recipients. Among ancestral NAb seroresponders, higher NAb levels positively correlated with higher Omicron neutralisation ( R  = 0.9, P  < 0.001). More KTRs contracted SARS-CoV-2 infection (14/30; 47%) than dialysis recipients (5/17; 29%) and had more severe disease. Those with breakthrough infections had significantly lower median interdose incremental change in anti-Spike RBD IgG and ancestral NAb titres., Conclusion: Serological responses to COVID-19 vaccines in KTRs lag behind their dialysis counterparts. KTRs remained at high risk of breakthrough infection after their primary vaccination schedule underlining their need for booster doses, strict infection prevention measures and close surveillance., Competing Interests: MCvZ is an inventor on a patent related to this work. The other authors have no conflicts of interest to declare., (© 2024 The Author(s). Clinical & Translational Immunology published by John Wiley & Sons Australia, Ltd on behalf of Australian and New Zealand Society for Immunology, Inc.)

The close bond that can exist between humans and their dogs is an important aspect of the evolutionary, economic, and social connections between the two species. There is a need for a better understanding of the place of the dog within the human-dog bond and on ways the human-dog bond affects dog welfare. We conducted a scoping review to investigate to what extent and in what ways dog welfare is addressed in the research literature on the human-dog bond. We identified 706 publications on the human-dog bond from across the globe that were published from 2012 to 2023. We found that 246 of these 706 publications had a focus on dog welfare. Our review showed that the interplay of characteristics and backgrounds of owners/handlers and their dogs was linked to dog welfare in multiple, both positive and negative, ways. Our review is limited by the fact that most of the research that we reviewed involved pet dogs and in majority came from Western, Educated, Industrialized, Rich, Democratic (WEIRD) societies. There is a need for a better understanding of how the human-dog bond affects the welfare of working, assistance, and service dogs.

A long-standing paradigm of arc geochemistry is that the trace element compositions of arc lavas arise from two compositionally distinct slab components: an aqueous dehydration fluid from the subducting igneous ocean crust that transports "fluid-mobile" elements, such as barium (Ba), and a sediment melt that supplies thorium (Th) and the light rare earth elements. This two-component framework has been widely called upon to explain global geochemical trends as well as geochemical variations within individual arcs, such as the Marianas. Here, we show that this paradigm is inconsistent with mass balance, due to the low Ba contents of igneous ocean crust, and with experimental data, which show that aqueous fluids from the igneous oceanic crust would be too dilute to substantially affect arc compositions. Observations previously attributed to the sediment melt/igneous-crust-fluid hypothesis are better explained by diverse subducting sediment compositions coupled with ambient mantle wedge heterogeneity, both globally and for the Marianas.

Speciation is often driven by selective processes like those associated with viability, mate choice, or local adaptation, and "speciation genes" have been identified in many eukaryotic lineages. In contrast, neutral processes are rarely considered as the primary drivers of speciation, especially over short evolutionary timeframes. Here, we describe a rapid vertebrate speciation event driven primarily by genetic drift. The White Sands pupfish ( Cyprinodon tularosa ) is endemic to New Mexico's Tularosa Basin where the species is currently managed as two Evolutionarily significant units (ESUs) and is of international conservation concern (Endangered). Whole-genome resequencing data from each ESU showed remarkably high and uniform levels of differentiation across the entire genome (global F ST ≈ 0.40). Despite inhabiting ecologically dissimilar springs and streams, our whole-genome analysis revealed no discrete islands of divergence indicative of strong selection, even when we focused on an array of candidate genes. Demographic modeling of the joint allele frequency spectrum indicates the two ESUs split only ~4 to 5 kya and that both ESUs have undergone major bottlenecks within the last 2.5 millennia. Our results indicate the genome-wide disparities between the two ESUs are not driven by divergent selection but by neutral drift due to small population sizes, geographic isolation, and repeated bottlenecks. While rapid speciation is often driven by natural or sexual selection, here we show that isolation and drift have led to speciation within a few thousand generations. We discuss these evolutionary insights in light of the conservation management challenges they pose., Competing Interests: Competing interests statement:The authors declare no competing interest.

Background and Objectives: Pathogenic variants in GRIN2A are associated with a spectrum of epilepsy-aphasia syndromes (EASs). Seizures as well as speech and language disorders occur frequently but vary widely in severity, both between individuals and across the life span. The link between this phenotypic spectrum and brain characteristics is unknown. Specifically, altered brain networks at the root of speech and language deficits remain to be identified. Patients with pathogenic variants in GRIN2A offer an opportunity to interrogate the impact of glutamate receptor dysfunction on brain development., Methods: We characterized brain anomalies in individuals with pathogenic GRIN2A variants and EASs, hypothesizing alterations in perisylvian speech-language regions and the striatum. We compared structural MRI data from 10 individuals (3 children and 7 adults, 3 female) with pathogenic GRIN2A variants with data from age-matched controls (N = 51 and N = 203 in a secondary analysis). We examined cortical thickness and volume in 4 a priori hypothesized speech and language regions (inferior frontal, precentral, supramarginal, and superior temporal) and across the whole brain. Subcortical structures (hippocampus, basal ganglia, thalamus) and the corpus callosum were also compared., Results: Individuals with pathogenic GRIN2A variants showed increased thickness and volume in the posterior part of Broca's area (inferior frontal gyrus, pars opercularis). For thickness, the effects were bilateral but more pronounced in the left (large effect size, η 2 = 0.37) than the right (η 2 = 0.12) hemisphere. Both volume and thickness were also higher in the bilateral superior temporal region while the supramarginal region showed increased thickness only. Whole-brain analyses confirmed left-sided thickness increases in Broca's area, with additional increases in the occipital and superior frontal cortices bilaterally. Hippocampal volume was reduced in the left hemisphere. There were no age-dependent effects or corpus callosum group differences., Discussion: Anomalies in perisylvian regions, with largest differences in Broca's area, suggest an altered development of classical speech-language networks in GRIN2A -related EAS. Left hippocampal reduction suggests a role for this structure in early speech and language development and is consistent with GRIN2A gene expression in that region. Overall, elucidating the neural correlates of EAS provides insights into the impact of GRIN2A dysfunction, opening avenues for targeted intervention in developmental syndromes with compromised speech-language development., Competing Interests: The authors report no relevant disclosures. Go to Neurology.org/NG for full disclosures., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.)

Background: Point mutations in histone variant H3.3 (H3.3K27M, H3.3G34R) and the H3.3-specific ATRX/DAXX chaperone complex are frequent events in pediatric gliomas. These H3.3 point mutations affect many chromatin modifications but the exact oncogenic mechanisms are currently unclear. Histone H3.3 is known to localize to nuclear compartments known as promyelocytic leukemia (PML) nuclear bodies, which are frequently mutated and confirmed as oncogenic drivers in acute promyelocytic leukemia., Results: We find that the pediatric glioma-associated H3.3 point mutations disrupt the formation of PML nuclear bodies and this prevents differentiation down glial lineages. Similar to leukemias driven by PML mutations, H3.3-mutated glioma cells are sensitive to drugs that target PML bodies. We also find that point mutations in IDH1/2-which are common events in adult gliomas and myeloid leukemias-also disrupt the formation of PML bodies., Conclusions: We identify PML as a contributor to oncogenesis in a subset of gliomas and show that targeting PML bodies is effective in treating these H3.3-mutated pediatric gliomas., (© 2023. The Author(s).)

Stuttering is a common speech disorder that interrupts speech fluency and tends to cluster in families. Typically, stuttering is characterized by speech sounds, words or syllables which may be repeated or prolonged and speech that may be further interrupted by hesitations or 'blocks'. Rare variants in a small number of genes encoding lysosomal pathway proteins have been linked to stuttering. We studied a large four-generation family in which persistent stuttering was inherited in an autosomal dominant manner with disruption of the cortico-basal-ganglia-thalamo-cortical network found on imaging. Exome sequencing of three affected family members revealed the PPID c.808C>T (p.Pro270Ser) variant that segregated with stuttering in the family. We generated a Ppid p.Pro270Ser knock-in mouse model and performed ex vivo imaging to assess for brain changes. Diffusion-weighted MRI in the mouse revealed significant microstructural changes in the left corticospinal tract, as previously implicated in stuttering. Quantitative susceptibility mapping also detected changes in cortico-striatal-thalamo-cortical loop tissue composition, consistent with findings in affected family members. This is the first report to implicate a chaperone protein in the pathogenesis of stuttering. The humanized Ppid murine model recapitulates network findings observed in affected family members., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The differentiation of naive CD8 + T lymphocytes into cytotoxic effector and memory CTL results in large-scale changes in transcriptional and phenotypic profiles. Little is known about how large-scale changes in genome organization underpin these transcriptional programs. We use Hi-C to map changes in the spatial organization of long-range genome contacts within naive, effector, and memory virus-specific CD8 + T cells. We observe that the architecture of the naive CD8 + T cell genome is distinct from effector and memory genome configurations, with extensive changes within discrete functional chromatin domains associated with effector/memory differentiation. Deletion of BACH2, or to a lesser extent, reducing SATB1 DNA binding, within naive CD8 + T cells results in a chromatin architecture more reminiscent of effector/memory states. This suggests that key transcription factors within naive CD8 + T cells act to restrain T cell differentiation by actively enforcing a unique naive chromatin state., Competing Interests: Declaration of interests A.W.G. is a member of the scientific advisory board of ArsenalBio. No funding from ArsenalBio was provided for this work., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Glucocorticoids (GCs) are potent anti-inflammatory agents and are broadly used in treating rheumatoid arthritis (RA) patients, albeit with adverse side effects associated with long-term usage. The negative consequences of GC therapy provide an impetus for research into gaining insights into the molecular mechanisms of GC action. We have previously reported that granulocyte-macrophage colony-stimulating factor (GM-CSF)-induced CCL17 has a non-redundant role in inflammatory arthritis. Here, we provide molecular evidence that GCs can suppress GM-CSF-mediated upregulation of IRF4 and CCL17 expression via downregulating JMJD3 expression and activity. In mouse models of inflammatory arthritis, GC treatment inhibited CCL17 expression and ameliorated arthritic pain-like behavior and disease. Significantly, GC treatment of RA patient peripheral blood mononuclear cells ex vivo resulted in decreased CCL17 production. This delineated pathway potentially provides new therapeutic options for the treatment of many inflammatory conditions, where GCs are used as an anti-inflammatory drug but without the associated adverse side effects., Competing Interests: JAH had a consulting role with GlaxoSmithKline., (© 2023 The Author(s).)

Understanding nanoparticle growth is crucial to increase the lifetime of supported metal catalysts. In this study, we employ in situ gas-phase transmission electron microscopy to visualize the movement and growth of ensembles of tens of nickel nanoparticles supported on carbon for CO 2 hydrogenation at atmospheric pressure (H 2 :CO 2 = 4:1) and relevant temperature (450 °C) in real time. We observe two modes of particle movement with an order of magnitude difference in velocity: fast, intermittent movement ( v max = 0.7 nm s -1 ) and slow, gradual movement ( v average = 0.05 nm s -1 ). We visualize the two distinct particle growth mechanisms: diffusion and coalescence, and Ostwald ripening. The diffusion and coalescence mechanism dominates at small interparticle distances, whereas Ostwald ripening is driven by differences in particle size. Strikingly, we demonstrate an interplay between the two mechanisms, where first coalescence takes place, followed by fast Ostwald ripening due to the increased difference in particle size. Our direct visualization of the complex nanoparticle growth mechanisms highlights the relevance of studying nanoparticle growth in supported nanoparticle ensembles under reaction conditions and contributes to the fundamental understanding of the stability in supported metal catalysts.

Cytokines that signal via STAT1 and STAT3 transcription factors instruct decisions affecting tissue homeostasis, antimicrobial host defense, and inflammation-induced tissue injury. To understand the coordination of these activities, we applied RNA sequencing, chromatin immunoprecipitation sequencing, and assay for transposase-accessible chromatin with high-throughput sequencing to identify the transcriptional output of STAT1 and STAT3 in peritoneal tissues from mice during acute resolving inflammation and inflammation primed to drive fibrosis. Bioinformatics focused on the transcriptional signature of the immunomodulatory cytokine IL-6 in both settings and examined how profibrotic IFN-γ-secreting CD4+ T cells altered the interpretation of STAT1 and STAT3 cytokine cues. In resolving inflammation, STAT1 and STAT3 cooperated to drive stromal gene expression affecting antimicrobial immunity and tissue homeostasis. The introduction of IFN-γ-secreting CD4+ T cells altered this transcriptional program and channeled STAT1 and STAT3 to a previously latent IFN-γ activation site motif in Alu-like elements. STAT1 and STAT3 binding to this conserved sequence revealed evidence of reciprocal cross-regulation and gene signatures relevant to pathophysiology. Thus, we propose that effector T cells retune the transcriptional output of IL-6 by shaping a regulatory interplay between STAT1 and STAT3 in inflammation., (Copyright © 2023 The Authors.)

Objectives: Influenza causes significant morbidity and mortality, especially in high-risk populations. Although current vaccination regimens are the best method to combat annual influenza disease, vaccine efficacy can be low in high-risk groups, such as haematopoietic stem cell transplant (HSCT) recipients., Methods: We comprehensively assessed humoral immunity, antibody landscapes, systems serology and influenza-specific B-cell responses, together with their phenotypes and isotypes, to the inactivated influenza vaccine (IIV) in HSCT recipients in comparison to healthy controls., Results: Inactivated influenza vaccine significantly increased haemagglutination inhibition (HAI) titres in HSCT recipients, similar to healthy controls. Systems serology revealed increased IgG1 and IgG3 antibody levels towards the haemagglutinin (HA) head, but not to neuraminidase, nucleoprotein or HA stem. IIV also increased frequencies of total, IgG class-switched and CD21 lo CD27 + influenza-specific B cells, determined by HA probes and flow cytometry. Strikingly, 40% of HSCT recipients had markedly higher antibody responses towards A/H3N2 vaccine strain than healthy controls and showed cross-reactivity to antigenically drifted A/H3N2 strains by antibody landscape analysis. These superior humoral responses were associated with a greater time interval after HSCT, while multivariant analyses revealed the importance of pre-existing immune memory. Conversely, in HSCT recipients who did not respond to the first dose, the second IIV dose did not greatly improve their humoral response, although 50% of second-dose patients reached a seroprotective HAI titre for at least one of vaccine strains., Conclusions: Our study demonstrates efficient, although time-dependent, immune responses to IIV in HSCT recipients, and provides insights into influenza vaccination strategies targeted to immunocompromised high-risk groups., Competing Interests: MK has acted as a consultant for Sanofi group of companies but not at the time the research was conducted. All other authors have declared that no conflict of interest exists., (© 2023 The Authors. Clinical & Translational Immunology published by John Wiley & Sons Australia, Ltd on behalf of Australian and New Zealand Society for Immunology, Inc.)

Although the importance of genome organization for transcriptional regulation of cell-fate decisions and function is clear, the changes in chromatin architecture and how these impact effector and memory CD8 + T cell differentiation remain unknown. Using Hi-C, we studied how genome configuration is integrated with CD8 + T cell differentiation during infection and investigated the role of CTCF, a key chromatin remodeler, in modulating CD8 + T cell fates through CTCF knockdown approaches and perturbation of specific CTCF-binding sites. We observed subset-specific changes in chromatin organization and CTCF binding and revealed that weak-affinity CTCF binding promotes terminal differentiation of CD8 + T cells through the regulation of transcriptional programs. Further, patients with de novo CTCF mutations had reduced expression of the terminal-effector genes in peripheral blood lymphocytes. Therefore, in addition to establishing genome architecture, CTCF regulates effector CD8 + T cell heterogeneity through altering interactions that regulate the transcription factor landscape and transcriptome., Competing Interests: Declaration of interests A.W.G. is a member of the scientific advisory board of ArsenalBio. S.J.T. is a member of the scientific advisory board for Medicago, Inc., QC, Canada. No funding from Medicago or ArsenalBio was provided for this work., (Copyright © 2023 Elsevier Inc. All rights reserved.)

More than half the world's population is infected with human cytomegalovirus (HCMV), causing congenital birth defects and impacting the immuno-compromised. Many of the >170 HCMV genes remain uncharacterized, and this gap in knowledge limits the development of novel antivirals. In this study, we investigated the essential viral protein UL49 and found it displayed leaky late expression kinetics, and localized to nuclear replication compartments. Cells infected with mutant UL49 virus were unable to produce infectious virions and phenocopied other beta-gamma viral pre-initiation complex (vPIC) subunit (UL79, UL87, UL91, UL92, and UL95) mutant infections. RNA-seq analysis of vPIC mutant infections revealed a consistent diminution of genes encoding capsid subunits, including TRX2/UL85 and MCP/UL86, envelope glycoproteins gM, gL and gO, and egress-associated tegument proteins UL99 and UL103. Therefore, as a member of the vPIC, UL49 serves as a fundamental HCMV effector that governs viral gene transcription required to complete the replication cycle., Competing Interests: The authors have declared that no competing interests exist., (© 2022 The Authors.)

Special AT-binding protein 1 (SATB1) is a chromatin-binding protein that has been shown to be a key regulator of T-cell development and CD4 + T-cell fate decisions and function. The underlying function for SATB1 in peripheral CD8 + T-cell differentiation processes is largely unknown. To address this, we examined SATB1-binding patterns in naïve and effector CD8 + T cells demonstrating that SATB1 binds to noncoding regulatory elements linked to T-cell lineage-specific gene programs, particularly in naïve CD8 + T cells. We then assessed SATB1 function using N-ethyl-N-nitrosourea-mutant mice that exhibit a point mutation in the SATB1 DNA-binding domain (termed Satb1 m1Anu/m1Anu ). Satb1 m1Anu/m1Anu mice exhibit diminished SATB1-binding, naïve, Satb1 m1Anu/m1Anu CD8 + T cells exhibiting transcriptional and phenotypic characteristics reminiscent of effector T cells. Upon activation, the transcriptional signatures of Satb1 m1Anu/m1Anu and wild-type effector CD8 + T cells converged. While there were no overt differences, primary respiratory infection of Satb1 m1Anu/m1Anu mice with influenza A virus (IAV) resulted in a decreased proportion and number of IAV-specific CD8 + effector T cells recruited to the infected lung when compared with wild-type mice. Together, these data suggest that SATB1 has a major role in an appropriate transcriptional state within naïve CD8 + T cells and ensures appropriate CD8 + T-cell effector gene expression upon activation., (© 2022 The Authors. Immunology & Cell Biology published by John Wiley & Sons Australia, Ltd on behalf of Australian and New Zealand Society for Immunology, Inc.)

Despite the great commercial relevance of zinc-promoted copper catalysts for methanol synthesis, the nature of the Cu-ZnO x synergy and the nature of the active Zn-based promoter species under industrially relevant conditions are still a topic of vivid debate. Detailed characterization of the chemical speciation of any promoter under high-pressure working conditions is challenging but specifically hampered by the large fraction of Zn spectator species bound to the oxidic catalyst support. We present the use of weakly interacting graphitic carbon supports as a tool to study the active speciation of the Zn promoter phase that is in close contact with the Cu nanoparticles using time-resolved X-ray absorption spectroscopy under working conditions. Without an oxidic support, much fewer Zn species need to be added for maximum catalyst activity. A 5-15 min exposure to 1 bar H 2 at 543 K only slightly reduces the Zn(II), but exposure for several hours to 20 bar H 2 /CO and/or H 2 /CO/CO 2 leads to an average Zn oxidation number of +(0.5-0.6), only slightly increasing to +0.8 in a 20 bar H 2 /CO 2 feed. This means that most of the added Zn is in a zerovalent oxidation state during methanol synthesis conditions. The Zn average coordination number is 8, showing that this phase is not at the surface but surrounded by other metal atoms (whether Zn or Cu), and indicating that the Zn diffuses into the Cu nanoparticles under reaction conditions. The time scale of this process corresponds to that of the generally observed activation period for these catalysts. These results reveal the speciation of the relevant Zn promoter species under methanol synthesis conditions and, more generally, present the use of weakly interacting graphitic supports as an important strategy to avoid excessive spectator species, thereby allowing us to study the nature of relevant promoter species., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

Over 50% of the world’s population is infected with Human Cytomegalovirus (HCMV). HCMV is responsible for serious complications in the immuno-compromised and is a leading cause of congenital birth defects. The molecular function of many HCMV proteins remains unknown, and a deeper understanding of the viral effectors that modulate virion maturation is required. In this study, we observed that UL34 is a viral protein expressed with leaky late kinetics that localises to the nucleus during infection. Deletion of UL34 from the HCMV genome (ΔUL34) did not abolish the spread of HCMV. Instead, over >100-fold fewer infectious virions were produced, so we report that UL34 is an augmenting gene. We found that ΔUL34 is dispensable for viral DNA replication, and its absence did not alter the expression of IE1, MCP, gB, UL26, UL83, or UL99 proteins. In addition, ΔUL34 infections were able to progress through the replication cycle to form a viral assembly compartment; however, virion maturation in the cytoplasm was abrogated. Further examination of the nucleus in ΔUL34 infections revealed replication compartments with aberrant morphology, containing significantly less assembled capsids, with almost none undergoing subsequent maturation. Therefore, this work lays the foundation for UL34 to be further investigated in the context of nuclear organization and capsid maturation during HCMV infection.

Developmental stuttering is a condition of speech dysfluency, characterized by pauses, blocks, prolongations and sound or syllable repetitions. It affects around 1% of the population, with potential detrimental effects on mental health and long-term employment. Accumulating evidence points to a genetic aetiology, yet gene-brain associations remain poorly understood due to a lack of MRI studies in affected families. Here we report the first neuroimaging study of developmental stuttering in a family with autosomal dominant inheritance of persistent stuttering. We studied a four-generation family, 16 family members were included in genotyping analysis. T1-weighted and diffusion-weighted MRI scans were conducted on seven family members (six male; aged 9-63 years) with two age and sex matched controls without stuttering (n = 14). Using Freesurfer, we analysed cortical morphology (cortical thickness, surface area and local gyrification index) and basal ganglia volumes. White matter integrity in key speech and language tracts (i.e. frontal aslant tract and arcuate fasciculus) was also analysed using MRtrix and probabilistic tractography. We identified a significant age by group interaction effect for cortical thickness in the left hemisphere pars opercularis (Broca's area). In affected family members this region failed to follow the typical trajectory of age-related thinning observed in controls. Surface area analysis revealed the middle frontal gyrus region was reduced bilaterally in the family (all cortical morphometry significance levels set at a vertex-wise threshold of P < 0.01, corrected for multiple comparisons). Both the left and right globus pallidus were larger in the family than in the control group (left P = 0.017; right P = 0.037), and a larger right globus pallidus was associated with more severe stuttering (rho = 0.86, P = 0.01). No white matter differences were identified. Genotyping identified novel loci on chromosomes 1 and 4 that map with the stuttering phenotype. Our findings denote disruption within the cortico-basal ganglia-thalamo-cortical network. The lack of typical development of these structures reflects the anatomical basis of the abnormal inhibitory control network between Broca's area and the striatum underpinning stuttering in these individuals. This is the first evidence of a neural phenotype in a family with an autosomal dominantly inherited stuttering., (© The Author(s) (2021). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Latent HIV-1 provirus in infected individuals on suppressive therapy does not always remain transcriptionally silent. Both HIV-1 LTR and human gene promoter derived transcriptional events can contribute HIV-1 sequences to the mRNA produced in the cell. In addition, chimeric cellular:HIV mRNA can arise through readthrough transcription and aberrant splicing. Using target enrichment coupled to the Illumina Mi-Seq and PacBio RS II platforms, we show that 3' LTR activation is frequent in latently infected cells from both the CCL19-induced primary cell model of HIV-1 latency as well as ex vivo samples. In both systems of latent HIV-1 infection, we detected several chimeric species that were generated via activation of a cryptic splice donor site in the 5' LTR of HIV-1. Aberrant splicing involving the major HIV-1 splice donor sites, SD1 and SD4 disrupts post-transcriptional processing of the gene in which HIV-1 is integrated. In the primary cell model of HIV-1 latency, Tat-encoding sequences are incorporated into the chimeric mRNA transcripts through the use of SD4. Our study unravels clues to the characteristics of HIV-1 integrants that promote formation of chimeric cellular:HIV mRNA and improves the understanding of the HIV-1 RNA footprint in latently infected cells., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Lee, Khoury, Olshansky, Sonza, Carter, McMahon, Stinear, Turner, Lewin and Purcell.)

Single molecule (SM) super-resolution microscopies bypass the diffraction limit of conventional optical techniques and provide excellent spatial resolutions in the tens of nanometers without overly complex microscope hardware. SM imaging using optical astigmatism is an efficient strategy for visualizing subcellular features in 3D with a z -range of up to ∼1 µm per acquisition. This approach however, places high demands on fluorophore brightness and photoswitching resilience meaning that imaging entire cell volumes in 3D using SM super-resolution remains challenging. Here we employ SM astigmatism together with multiplane acquisition to visualize the whole nuclear lamina of COS-7 and T cells in 3D. Nuclear lamina provides structural support to the nuclear envelope and participates in vital nuclear functions including internuclear transport, chromatin organization and gene regulation. Its position at the periphery of the nucleus provides a visible reference of the nuclear boundary and can be used to quantify the spatial distribution of intranuclear components such as histone modifications and transcription factors. We found Alexa Fluor 647, a popular photoswitchable fluorophore, remained viable for over an hour of continuous high laser power exposure, and provided sufficient brightness detectable up to 8 µm deep into a cell, allowing us to capture the entire nuclear lamina in 3D. Our approach provides sufficient super-resolution detail of nuclear lamina morphology to enable quantification of overall nuclear dimensions and local membrane features., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Rozario, Morey, Elliott, Russ, Whelan, Turner and Bell.)

The COVID-19 pandemic has highlighted the need and benefits for all communities to be permitted timely access to on-demand screening for infectious respiratory diseases. This can be achieved with simplified testing approaches and affordable access to core resources. While RT-qPCR-based tests remain the gold standard for SARS-CoV-2 detection due to their high sensitivity, implementation of testing requires high upfront costs to obtain the necessary instrumentation. This is particularly restrictive in low-resource settings. The Ubiquitome Liberty16 system was developed as an inexpensive, portable, battery-operated single-channel RT-qPCR device with an associated iPhone app to simplify assay set-up and data reporting. When coupled with the SalivaDirect protocol for testing saliva samples for SARS-CoV-2, the Liberty16 device yielded a limit of detection (LOD) of 12 SARS-CoV-2 RNA copies/µL, comparable to the upper end of the LOD range for the standard SalivaDirect protocol when performed on larger RT-qPCR instruments. While further optimization may deliver even greater sensitivity and assay speed, findings from this study indicate that small portable devices such as the Liberty16 can deliver reliable results and provide the opportunity to further increase access to gold standard SARS-CoV-2 testing., Competing Interests: ST was an employee of Ubiquitome Limited at the time of this study and PP is the CEO of Ubiquitome Limited. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Yolda-Carr, Thammavongsa, Vega, Turner, Pickering and Wyllie.)

Ineffective antibody-mediated responses are a key characteristic of chronic viral infection. However, our understanding of the intrinsic mechanisms that drive this dysregulation are unclear. Here, we identify that targeting the epigenetic modifier BMI-1 in mice improves humoral responses to chronic lymphocytic choriomeningitis virus. BMI-1 was upregulated by germinal center B cells in chronic viral infection, correlating with changes to the accessible chromatin landscape, compared to acute infection. B cell-intrinsic deletion of Bmi1 accelerated viral clearance, reduced splenomegaly and restored splenic architecture. Deletion of Bmi1 restored c-Myc expression in B cells, concomitant with improved quality of antibody and coupled with reduced antibody-secreting cell numbers. Specifically, BMI-1-deficiency induced antibody with increased neutralizing capacity and enhanced antibody-dependent effector function. Using a small molecule inhibitor to murine BMI-1, we could deplete antibody-secreting cells and prohibit detrimental immune complex formation in vivo. This study defines BMI-1 as a crucial immune modifier that controls antibody-mediated responses in chronic infection., (© 2021. The Author(s), under exclusive licence to Springer Nature America, Inc.)

The NF-κB transcription factor c-Rel is a critical regulator of Treg ontogeny, controlling multiple points of the stepwise developmental pathway. Here, we found that the thymic Treg defect in c-Rel-deficient (cRel -/- ) mice is quantitative, not qualitative, based on analyses of TCR repertoire and TCR signaling strength. However, these parameters are altered in the thymic Treg-precursor population, which is also markedly diminished in cRel -/- mice. Moreover, c-Rel governs the transcriptional programme of both thymic and peripheral Tregs, controlling a core of genes involved with immune signaling, and separately in the periphery, cell cycle progression. Last, the immune suppressive function of peripheral cRel -/- tTregs is diminished in a lymphopenic model of T cell proliferation and is associated with decreased stability of Foxp3 expression. Collectively, we show that c-Rel is a transcriptional regulator that controls multiple aspects of Treg development, differentiation, and function via distinct mechanisms., (© 2021 Wiley-VCH GmbH.)

Chinese hamster ovary (CHO) cells are the leading mammalian cell host employed to produce complex secreted recombinant biotherapeutics such as monoclonal antibodies (mAbs). Metabolic selection marker technologies (e.g. glutamine synthetase (GS) or dihydrofolate reductase (DHFR)) are routinely employed to generate such recombinant mammalian cell lines. Here we describe the development of a selection marker system based on the metabolic requirement of CHO cells to produce proline, and that uses pyrroline-5-carboxylase synthetase (P5CS) to complement this auxotrophy. Firstly, we showed the system can be used to generate cells that have growth kinetics in proline-free medium similar to those of the parent CHO cell line, CHOK1SV GS-KO™ grown in proline-containing medium. As we have previously described how engineering lipid metabolism can be harnessed to enhance recombinant protein productivity in CHO cells, we then used the P5CS selection system to re-engineer lipid metabolism by over-expression of either sterol regulatory element binding protein 1 (SREBF1) or stearoyl CoA desaturase 1 (SCD1). The cells with re-engineered proline and lipid metabolism showed consistent growth and P5CS, SCD1 and SREBF1 expression across 100 cell generations. Finally, we show that the P5CS and GS selection systems can be used together. A GS vector containing the light and heavy chains for a mAb was super-transfected into a CHOK1SV GS-KO™ host over-expressing SCD1 from a P5CS vector. The resulting stable transfectant pools achieved a higher concentration at harvest for a model difficult to express mAb than the CHOK1SV GS-KO™ host. This demonstrates that the P5CS and GS selection systems can be used concomitantly to enable CHO cell line genetic engineering and recombinant protein expression., Competing Interests: AD, SJT, CMJ, GS, and AJR are employed by Lonza Biologics, who developed and license the GS Gene Expression System®. Lonza is the assigned owner of, and CMS, JDB, TJK, and RJY are named inventors on, the filed patent ‘Modulation of lipid metabolism for protein production’, patent number WO2017191165A1. Lonza is assigned owner of, and JDB, CMJ, AJR, JR, GS, CMS and RJY named inventors on, the filed patent ‘Methods of cell selection and modifying cell metabolism’ patent number WO2019152876AA2. The authors declare no other financial or commercial conflict of interest., (© 2021 The Authors.)

How innate and adaptive immune responses work in concert to resolve influenza disease is yet to be fully investigated in one single study. Here, we utilize longitudinal samples from patients hospitalized with acute influenza to understand these immune responses. We report the dynamics of 18 important immune parameters, related to clinical, genetic and virological factors, in influenza patients across different severity levels. Influenza disease correlates with increases in IL-6/IL-8/MIP-1α/β cytokines and lower antibody responses. Robust activation of circulating T follicular helper cells correlates with peak antibody-secreting cells and influenza heamaglutinin-specific memory B-cell numbers, which phenotypically differs from vaccination-induced B-cell responses. Numbers of influenza-specific CD8 + or CD4 + T cells increase early in disease and retain an activated phenotype during patient recovery. We report the characterisation of immune cellular networks underlying recovery from influenza infection which are highly relevant to other infectious diseases.

The generation of effective adaptive T-cell memory is a cardinal feature of the adaptive immune system. The establishment of protective T-cell immunity requires the differentiation of CD8 + T cells from a naive state to one where pathogen-specific memory CD8 + T cells are capable of responding to a secondary infection more rapidly and robustly without the need for further differentiation. The study of factors that determine the fate of activated CD8 + T cells into either effector or memory subsets has a long history. The advent of new technologies is now providing new insights into how epigenetic regulation not only impacts acquisition and maintenance of effector function, but also the maintenance of the quiescent yet primed memory state. There is growing appreciation that rather than distinct subsets, memory T-cell populations may reflect different points on a spectrum between the starting naive T-cell population and a terminally differentiated effector CD8 + T-cell population. Interestingly, there is growing evidence that the molecular mechanisms that underpin the rapid effector function of memory T cells are also observed in innate immune cells such as macrophages and natural killer (NK) cells. This raises an interesting hypothesis that the memory/effector T-cell state represents a default innate-like response to antigen recognition, and that it is the naive state that is the defining feature of adaptive immunity. These issues are discussed., (Copyright © 2021 Cold Spring Harbor Laboratory Press; all rights reserved.)

Mucosal-associated invariant T (MAIT) cells are an innate-like population of unconventional T cells that respond rapidly to microbial metabolite Ags or cytokine stimulation. Because of this reactivity and surface expression of CD45RO + , CD45RA - , and CD127 + , they are described as effector memory cells. Yet, there is heterogeneity in MAIT cell effector response. It is unclear what factors control MAIT cell effector capacity, whether it is fixed or can be modified and if this differs based on whether activation is TCR dependent or independent. To address this, we have taken a systematic approach to examine human MAIT cell effector capacity across healthy individuals in response to ligand and cytokine stimulation. We demonstrate the heterogenous nature of MAIT cell effector capacity and that the ability to produce an effector response is not directly attributable to TCR clonotype or coreceptor expression. Global gene transcription analysis revealed that the MAIT cell effector capacity produced in response to TCR stimulation is associated with increased expression of the epigenetic regulator lysine demethylase 6B (KDM6B). Addition of a KDM6B inhibitor did not alter MAIT cell effector function to Ag or cytokine stimulation. However, addition of the KDM6B cofactor α-ketoglutarate greatly enhanced MAIT cell effector capacity to TCR-dependent stimulation in a partially KDM6B-dependent manner. These results demonstrate that the TCR-dependent effector response of MAIT cells is epigenetically regulated and dependent on the availability of metabolic cofactors., (Copyright © 2021 by The American Association of Immunologists, Inc.)

Naive CD8 + T cell activation results in an autonomous program of cellular proliferation and differentiation. However, the mechanisms that underpin this process are unclear. Here, we profile genome-wide changes in chromatin accessibility, gene transcription, and the deposition of a key chromatin modification (H3K27me3) early after naive CD8 + T cell activation. Rapid upregulation of the histone demethylase KDM6B prior to the first cell division is required for initiating H3K27me3 removal at genes essential for subsequent T cell differentiation and proliferation. Inhibition of KDM6B-dependent H3K27me3 demethylation limits the magnitude of an effective primary virus-specific CD8 + T cell response and the formation of memory CD8 + T cell populations. Accordingly, we define the early spatiotemporal events underpinning early lineage-specific chromatin reprogramming that are necessary for autonomous CD8 + T cell proliferation and differentiation., Competing Interests: Declaration of interests S.R. is currently the founder/chief scientific officer of EpiAxis Therapeutics. This manuscript received no funding from EpiAxis for this work. S.J.T. is a member of the scientific advisory board for Medicago, Inc., QC, Canada. This manuscript received no funding from Medicago for this work., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

CD8 + T cells play a pivotal role in clearing intracellular pathogens and combatting tumours. Upon infection, naïve CD8 + T cells differentiate into effector and memory cells, and this program is underscored by large-scale and coordinated changes in the chromatin architecture and gene expression. Importantly, recent evidence demonstrates that the epigenetic mechanisms that regulate the capacity for rapid effector function of memory T cells are shared by innate immune cells such as natural killer (NK) cells. Thus, it appears that the crucial difference between innate and adaptive immunity is the presence of the naïve state. This important distinction raises an intriguing new hypothesis, that the naïve state was evolutionary installed to restrain a default program of effector and memory differentiation in response to antigen recognition. We argue that the hallmark of adaptive T immunity is therefore the naïve program, which actively maintains CD8 + T cell quiescence until receipt of appropriate activation signals. In this review, we examine the mechanistic control of naïve CD8 + T cell quiescence and summarise the multiple levels of restraint imposed in naïve cells in to limit spontaneous and inappropriate activation. This includes epigenetic mechanisms and transcription factor (TF) regulation of gene expression, in addition to novel inhibitory receptors, abundance of RNA, and protein degradation.

Regulatory elements (REs) consist of enhancers and promoters that occupy a significant portion of the noncoding genome and control gene expression programs either in cis or in trans Putative REs have been identified largely based on their regulatory features (co-occupancy of ESC-specific transcription factors, enhancer histone marks, and DNase hypersensitivity) in mouse embryonic stem cells (mESCs). However, less has been established regarding their regulatory functions in their native context. We deployed cis- and trans- regulatory elements scanning through saturating mutagenesis and sequencing (ctSCAN-SMS) to target elements within the ∼12-kb cis -region ( cis- REs; CREs) of the Oct4 gene locus, as well as genome-wide 2,613 high-confidence trans- REs (TREs), in mESCs. ctSCAN-SMS identified 10 CREs and 12 TREs as novel candidate REs of the Oct4 gene in mESCs. Furthermore, deletions of these candidate REs confirmed that the majority of the REs are functionally active, and CREs are more active than TREs in controlling Oct4 gene expression. A subset of active CREs and TREs physically interact with the Oct4 promoter to varying degrees; specifically, a greater number of active CREs, compared with active TREs, physically interact with the Oct4 promoter. Moreover, comparative genomics analysis reveals that a greater number of active CREs than active TREs are evolutionarily conserved between mice and primates, including humans. Taken together, our study demonstrates the reliability and robustness of ctSCAN-SMS screening to identify critical REs and investigate their roles in the regulation of transcriptional output of a target gene (in this case Oct4 ) in their native context., Competing Interests: Conflict of interest—The authors declare that they have no conflicts of interest with the contents of this article., (© 2020 Canver et al.)

The role unconventional T cells play in protective immunity in humans is unclear. Mucosal-associated invariant T (MAIT) cells are an unconventional T cell subset restricted to the antigen-presenting molecule MR1. Here, we report the discovery of a patient homozygous for a rare Arg31His (R9H in the mature protein) mutation in MR1 who has a history of difficult-to-treat viral and bacterial infections. MR1 R9H was unable to present the potent microbially derived MAIT cell stimulatory ligand. The MR1 R9H crystal structure revealed that the stimulatory ligand cannot bind due to the mutation lying within, and causing structural perturbation to, the ligand-binding domain of MR1. While MR1 R9H could bind and be up-regulated by a MAIT cell inhibitory ligand, the patient lacked circulating MAIT cells. This shows the importance of the stimulatory ligand for MAIT cell selection in humans. The patient had an expanded γδ T cell population, indicating a compensatory interplay between these unconventional T cell subsets., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Liver resident-memory CD8 + T cells (T RM cells) can kill liver-stage Plasmodium -infected cells and prevent malaria, but simple vaccines for generating this important immune population are lacking. Here, we report the development of a fully synthetic self-adjuvanting glycolipid-peptide conjugate vaccine designed to efficiently induce liver T RM cells. Upon cleavage in vivo, the glycolipid-peptide conjugate vaccine releases an MHC I-restricted peptide epitope (to stimulate Plasmodium -specific CD8 + T cells) and an adjuvant component, the NKT cell agonist α-galactosylceramide (α-GalCer). A single dose of this vaccine in mice induced substantial numbers of intrahepatic malaria-specific CD8 + T cells expressing canonical markers of liver T RM cells (CD69, CXCR6, and CD101), and these cells could be further increased in number upon vaccine boosting. We show that modifications to the peptide, such as addition of proteasomal-cleavage sequences or epitope-flanking sequences, or the use of alternative conjugation methods to link the peptide to the glycolipid improved liver T RM cell generation and led to the development of a vaccine able to induce sterile protection in C57BL/6 mice against Plasmodium berghei sporozoite challenge after a single dose. Furthermore, this vaccine induced endogenous liver T RM cells that were long-lived (half-life of ~425 days) and were able to maintain >90% sterile protection to day 200. Our findings describe an ideal synthetic vaccine platform for generating large numbers of liver T RM cells for effective control of liver-stage malaria and, potentially, a variety of other hepatotropic infections., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Naive CD8 + T cell survival in the periphery is critically dependent on tonic TCR signaling through peptide + MHC class I (MHCI) recognition; however, little is known about how natural variation in MHCI levels impacts the naive CD8 + T cell repertoire. Using mice that are hemizygous or homozygous for a single MHCI allele, we showed that despite a reduction in peripheral CD8 + T cell numbers of ∼50% in MHCI hemizygous mice, MHCI levels had no notable impact on the rate of thymic generation or emigration of CD8 single-positive T cells. Moreover, the peripheral T cell repertoire in hemizygous mice showed selective retention of T cell clonotypes with a greater competitive advantage as evidenced by increased expression of CD5 and IL-7Rα. The qualitative superiority of CD8 + T cells retained in hemizygous mice was also seen during influenza A virus infection, in which epitope-specific CD8 + T cells from hemizygous mice had a higher avidity for pMHCI and increased cytokine polyfunctionality, despite a reduced response magnitude. Collectively, this study suggests that natural variation in MHCI expression levels has a notable and biologically relevant impact on the maintenance, but not generation, of the naive CD8 + T cell repertoire., (Copyright © 2020 by The American Association of Immunologists, Inc.)