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The miR-15/16 family is a highly expressed group of tumor suppressor miRNAs that target a large network of genes in T cells to restrict their cell cycle, memory formation and survival. Upon T cell activation, miR-15/16 are downregulated, allowing rapid expansion of differentiated effector T cells to mediate a sustained immune response. Here, using conditional deletion of miR-15/16 in immunosuppressive regulatory T cells (Tregs) that express FOXP3, we identify new functions of the miR-15/16 family in T cell immunity. miR-15/16 are indispensable to maintain peripheral tolerance by securing efficient suppression by a limited number of Tregs. miR-15/16-deficiency alters Treg expression of critical functional proteins including FOXP3, IL2Rα/CD25, CTLA4, PD-1 and IL7Rα/CD127, and results in accumulation of functionally impaired FOXP3loCD25loCD127hiTregs. Excessive proliferation in the absence of miR-15/16 inhibition of cell cycle programs shifts Treg diversity and produces an effector Treg phenotype characterized by low expression of TCF1, CD25 and CD62L, and high expression of CD44. These Tregs fail to control immune activation of CD4+effector T cells, leading to spontaneous multi-organ inflammation and increased allergic airway inflammation in a mouse model of asthma. Together, our results demonstrate that miR-15/16 expression in Tregs is essential to maintain immune tolerance.HighlightsTreg-specific miR-15/16 expression is essential to prevent systemic tissue inflammationmiR-15/16 restrict Treg proliferation and regulate expression of the key functional Treg molecules FOXP3, IL2Rα, CTLA4, PD-1 and IL7RαmiR-15/16 limit formation of effector Tregs and is necessary for high suppressive capacity

Heart rate variability (HRV) is a psychophysiological variable that is often used in applied analysis techniques to indicate health status because it provides a window into the intrinsic regulation of the autonomic nervous system. However, HRV data analysis methods are varied and complex, which has led to different approaches to data collection, analysis, and interpretation of results. Our scoping review aimed to explore the diverse use of HRV methods in studies designed to assess health outcomes in outdoor free-living contexts. Four database indexes were searched, which resulted in the identification of 17,505 candidate studies. There were 34 studies and eight systematic reviews that met the inclusion criteria. Just over half of the papers referenced the 1996 task force paper that outlined the standards of measurement and physiological interpretation of HRV data, with even fewer adhering to recommended HRV recording and analysis procedures. Most authors reported an increase in parasympathetic (n = 23) and a decrease in systematic nervous system activity (n = 20). Few studies mentioned methods-related limitations and challenges, despite a wide diversity of recording devices and analysis software used. We conclude our review with five recommendations for future research using HRV methods in outdoor and health-related contexts.

Funder: Horizon 2020 Framework Programme, Funder: National Institutes of Health, CD28 provides the prototypical costimulatory signal required for productive T-cell activation. Known molecular consequences of CD28 costimulation are mostly based on studies of protein signaling molecules. The microRNA cluster miR-17∼92 is induced by T cell receptor stimulation and further enhanced by combined CD28 costimulation. We demonstrate that transgenic miR-17∼92 cell-intrinsically largely overcomes defects caused by CD28 deficiency. Combining genetics, transcriptomics, bioinformatics, and biochemical miRNA:mRNA interaction maps we empirically validate miR-17∼92 target genes that include several negative regulators of T cell activation. CD28-deficient T cells exhibit derepressed miR-17∼92 target genes during activation. CRISPR/Cas9-mediated ablation of the miR-17∼92 targets Pten and Nrbp1 in naive CD28-/- CD4+ T cells differentially increases proliferation and expression of the activation markers CD25 and CD44, respectively. Thus, we propose that miR-17∼92 constitutes a central mediator for T cell activation, integrating signals by the TCR and CD28 costimulation by dampening multiple brakes that prevent T cell activation.

The aging brain exhibits a decline in the regenerative populations of neural stem cells (NSCs), which may underlie age-associated defects in sensory and cognitive functions1–6. While mechanisms that restore old NSC function have started to be identified7–23, the role of lipids – especially complex lipids – in NSC aging remains largely unclear. Using lipidomic profiling by mass spectrometry, we identify age-related lipidomic signatures in young and old quiescent NSCs in vitro and in vivo. These analyses reveal drastic changes in several complex membrane lipid classes, including phospholipids and sphingolipids in old NSCs. Moreover, poly-unsaturated fatty acids (PUFAs) strikingly increase across complex lipid classes in quiescent NSCs during aging. Age-related changes in complex lipid levels and side chain composition are largely occurring in plasma membrane lipids, as revealed by lipidomic profiling of isolated plasma membrane vesicles. Experimentally, we find that aging is accompanied by modifications in plasma membrane biophysical properties, with a decrease in plasma membrane order in old quiescent NSCs in vitro and in vivo. To determine the functional role of plasma membrane lipids in aging NSCs, we performed genetic and supplementation studies. Knockout of Mboat2, which encodes a phospholipid acyltransferase, exacerbates age-related lipidomic changes in old quiescent NSCs and impedes their ability to activate. As Mboat2 expression declines with age, Mboat2 deficiency may drive NSC decline during aging. Interestingly, supplementation of plasma membrane lipids derived from young NSCs boosts the ability of old quiescent NSCs to activate. Our work could lead to lipid-based strategies for restoring the regenerative potential of NSCs in old individuals, which has important implications for countering brain decline during aging.

Les policiers exercent un métier où ils sont confrontés à des situations graves et complexes. Plus encore, ils jouent un rôle central et d’une grande importance dans la société; ils assurent la sécurité publique et agissent à titre d’intervenants de première ligne (Leclercq, 2008; Shane, 2010). La presse négative combinée à l’arrivée des nouvelles technologies et leur grande accessibilité représente un enjeu supplémentaire qui peut affecter la santé psychologique des policiers. Depuis quelques années, les policiers font régulièrement la manchette, souvent de manière négative. La médiatisation de ces événements a-t-elle exercé un impact négatif sur la santé psychologique au travail des policiers concernés ? L’étude de Chermak et al. (2006) soutient que les médias représentent souvent la source centrale de perception de la légitimité policière pour les citoyens. La présente étude vise à explorer la relation entre les communications médiatiques et la santé psychologique des policiers. Elle entend utiliser le modèle théorique de Gilbert, Dagenais-Desmarais et Savoie (2011) pour expliquer la santé psychologique au travail. Les communications médiatiques seront discutées selon le modèle général des effets réciproques de la couverture médiatique de Kepplinger (2007). En considérant ce qui précède et puisqu’il s’agit d’une étude qualitative, un échantillonnage par réseau a été réalisé pour recruter les participants. Au total, 12 policiers ont accepté de participer, sur une base volontaire, à une entrevue semi-dirigée. Cette recherche a permis de reconnaître l’influence des communications médiatiques tant sur la santé psychologique des policiers au travail (désengagement, remise en question, consommation excessive de psychotropes, méfiance, état dépressif et perte d’intégrité pour n’en nommer que quelques-uns) que sur leur réseau social de même que sur l’opinion des citoyens envers eux., Law enforcement is a profession where individuals are faced with serious and complex situations. More importantly, they play a pivotal role in our modern society; they provide public safety and act as frontline during crisis (Leclercq, 2008; Shane, 2010). The arrival of new technologies, negative press brought forward through a near infinite number of media outlet and their wide accessibility causes issues that can easily affect the psychological health of a police officer. In recent years, law enforcement has been regularly breaking headlines, often in a negative way. Ergo, does the coverage of these events affect the psychological health at work of the police officers concerned? The study by Chermak and al. (2006) upholds that the media are often the central source of citizens' perception of police legitimacy. In light of these findings, the present study is intended to understand the relationship between media communications of police events and their psychological health. This study intends to use the theoretical model of Gilbert, Dagenais-Desmarais and Savoie (2011) to explain psychological health at work. Media communications will be discussed according to the general model of the reciprocal effects of media coverage by Kepplinger (2007). Considering the above, and since this is a qualitative study, network sampling was used to select the participants; twelve (12) police officers have accepted to participate in this study on a voluntary basis via a semi-structured interview. This res earch allowed to recognize the influence of media communications on the psychological health and well-being of law enforcement officers at work (such as loss of commitment in the workplace and at home, questioning one’s abilities, excessive use of psychoactive drugs, mistrust, depressive state and loss of integrity) as well as on social media and in citizen opinion about them in general.

Objectives Describe Ontario's school-based human papillomavirus (HPV) vaccination program from the perspective of local public health units (PHUs). Methods In 2018, Vaccine Preventable Diseases (VPD) managers at each of Ontario's 35 PHUs were invited to participate in an online survey regarding the organization and delivery of their HPV vaccination program. Questions were asked on the school-based program, training and support of vaccine providers, communication and promotion, assessing coverage rates and perceptions of the program's strengths and challenges. Descriptive statistics were generated for close-ended items. A thematic content analysis was performed for open-ended items. Results Eighteen PHUs (54%, n = 19/35) responded. All responding PHUs provided the HPV vaccine in publicly funded schools but only 6 reported being permitted to provide HPV vaccine in private schools. Fact sheets, Q&As or other written information locally developed by the PHUs were the main tools used to communicate with parents (n = 17), students (n = 13), school personnel (n = 13) and school board officials (n = 9). The most frequently reported barriers were: limited program resources, negative perceptions held by parents and/or school staff regarding the HPV vaccine, logistical issues (e.g., getting the consents forms returned, collaboration with schools for vaccine delivery) and the fact that HPV vaccination is not mandatory under Ontario legislation. Conclusion Local public health units that implement HPV vaccine programs in schools identified logistical barriers, public perceptions about the HPV vaccine and the voluntary nature of the program as the main barriers.

This study investigated whether muscle cooling and its associated effects on skeletal muscle oxidative responses, blood gases, and hormonal concentrations influenced energy metabolism during cycling. Twelve healthy participants (Males: seven; Females: five) performed two steady-state exercise sessions at 70% of ventilatory threshold on a cycle ergometer. Participants completed one session with pre-exercise leg cooling until muscle temperature (Tm) decreased by 6 °C (LCO), and a separate session without cooling (CON). They exercised until Tm returned to baseline and for an additional 30 min. Cardiovascular, respiratory, metabolic, hemodynamic variables, and skeletal muscle tissue oxidative responses were assessed continuously. Venous blood samples were collected to assess blood gases, and hormones. Heart rate, stroke volume, and cardiac output all increased across time but were not different between conditions. VO2 was greater in LCO when muscle temperature was restored until the end of exercise (p

A number of countries have implemented vaccination in pregnancy as a strategy to reduce the burden of influenza and pertussis. The aim of this study was to assess the involvement of Canadian maternity care providers in administration of vaccines to their pregnant patients. A cross-sectional web-based survey was sent to family physicians, obstetricians-gynecologists, midwives, pharmacists, and nurses. A multivariable logistic regression model was used to determine variables independently associated with offering vaccination services in pregnancy in providers’ practice. A total of 1,135 participants participated. Overall, 64% (n = 724) of the participants reported offering vaccines in their practice and 56% (n = 632) reported offering vaccines to pregnant patients. The main reasons reported for not offering vaccination services in pregnancy were the belief that vaccination was outside of the scope of practice; logistical issues around access to vaccines; or lack of staff to administer vaccines. In multivariable analysis, the main factors associated with vaccination of pregnant patients in practices where vaccination services were offered were: providers’ confidence in counseling pregnant patients about vaccines, seeing fewer than 11 pregnant patients on average each week, and being a nurse or a family physician. Although the majority of participants expressed strong support for vaccination during pregnancy, half were not offering vaccination services in their practice. Many were not equipped to offer vaccines in their practice or felt that it was not their role to do so. To enhance vaccine acceptance and uptake in pregnancy, it will be important to address the logistical barriers identified in this study.

RESUME: Selon la litterature, le travail du policier est une profession a risque etant donne son contexte d'intervention souvent urgent et imprevisible (De Soir, Daubechies et Van den Steene, 2012?; cites par Desjardins, 2017). Le metier de policier comporte egalement un double mandat, soit celui de proteger la population tout en faisant regner l'ordre (Oligny, 1991). Ces particularites rendent le travail de policiers plus a risque que d'autres professions sur le plan de la sante mentale (Marchand, 2007). Malgre l'importance accordee a la sante au travail et les risques associes au metier de policier, peu d'etudes empiriques sont repertoriees a ce sujet au sein de cette population. Silveri (2017) a identifie dans son article le soutien social par les collegues et par les superieurs comme faisant partie des ressources les plus importantes pour attenuer les «?effets des exigences de travail sur le stress?». La presente recherche s'interesse plus specifiquement a l'effet des relations interpersonnelles (variable independante) et du soutien social (variable independante) sur la sante psychologique des policiers (variable dependante). Le modele theorique de Gilbert, Dagenais-Desmarais et Savoie (2011) a ete retenu pour definir la variable dependante composee du bien-etre psychologique (BEPT?; serenite, engagement et harmonie sociale) et de la detresse psychologique (DET?; anxiete, desengagement et irritabilite). L'etude suppose que la qualite percue des relations interpersonnelles avec les collegues et le superieur, ainsi que le soutien social, sont relies positivement au bien-etre psychologique au travail. Deuxiemement, l'etude suppose egalement que la qualite des relations et le soutien social sont relies negativement a la detresse psychologique au travail. Un devis correlationnel est utilise pour repondre a la question de recherche. Neuf cent quatre-vingt-dix (990) participants issus du milieu policier quebecois provenant de la province de Quebec composent l'echantillon. Les resultats des analyses de regression, qui abondent dans le meme sens que les hypotheses de recherche, ont demontre un lien significatif entre la qualite des relations percue avec les collegues, le soutien de type conseil et le bien-etre psychologique au travail. Les resultats soutiennent egalement l'importance du developpement du lien de confiance entre le superieur et le policier pour contrer la detresse psychologique au travail. -- Mot(s) cle(s) en francais : Policier, sante psychologique au travail, relations interpersonnelles, soutien social, soutien emotionnel, soutien conseil, bien-etre au travail, detresse au travail. -- ABSTRACT: According to the literature, police work is a profession at risk given its context of urgent and unpredictable intervention (De Soir, Daubechies and Van den Steene, 2012; cited by Desjardins, 2017). The policing profession also has a dual mandate: to protect the public while maintaining order (Oligny, 1991). These features make police work even more risky than other professions in terms of mental health (Marchand, 2007). Despite the importance given to occupational health and the risks associated with being a police officer, there are few empirical studies on this topic about this population. Silveri (2017) has identified, in his article, social support by colleagues and superiors as one of the most important resources to mitigate the "effects of work demands on stress". This research focuses more specifically on the effect of interpersonal relationships (independent variable) and social support (independent variable) on the psychological health of police officers (dependent variable). The theoretical model of Gilbert, Dagenais-Desmarais and Savoie (2011) was used to define the dependent variable composed of psychological well-being (BEPT, serenity, commitment and social harmony) and psychological distress (DET, anxiety, disengagement and irritability). The present study assumes that the perceived quality of interpersonal relationships with colleagues and the superior, as well as social support, are positively related to psychological well-being at work. Second, the study also assumes that the quality of relationships and social support are negatively related to psychological distress at work. A correlational quote is used to answer the search question. Nine hundred and ninety (990) participants from the Quebec policing community from the province of Quebec make up the study's sample. The results of regression analyzes, which abound in the same direction as the research hypotheses, demonstrated a significant link between the perceived quality of relationships with colleagues, counseling-type support and psychological well-being at work. The results also highlight the importance of developing the bond of trust between the superior and the police officer to counter psychological distress at work. -- Mot(s) cle(s) en anglais : Police officer, psychological health at work, interpersonal relationships, social support, emotional support, counseling support, well-being at work, distress at work.

Physical activity plays an important role in maintaining mental and physical health. This study assessed the effect of physical activity monitoring awareness on the physical activity level and subjective self-assessment of physical activity in middle-aged subjects with normal cognitive function (NCF) and mild cognitive impairment (MCI).Thirty-five subjects aged 50-65 years with NCF and MCI were randomised into two experimental groups, each taking part in two one-week intervention periods. Subjects in group A were not aware that their physical activity was monitored in the first week (phase I) and were aware of the monitoring in the second week (phase II), whereas it was the opposite order for group B. Physical activity was assessed using the ActiGraph GT9X accelerometer and International Physical Activity Questionnaire (IPAQ).A total of 32 subjects (MCI: n = 12, NCF: n = 20) completed both intervention periods, with MCI subjects having significantly lower objectively assessed physical activity than NCF participants. Moreover, subjectively assessed physical activity in the MCI group was significantly higher when the participants were unaware of physical activity monitoring. A significant phase-group interaction was found in total (MET-min/d: p = 0.0072; min/d: p = 0.0194) and moderate (MET-min/d: p = 0.0015; min/d: p = 0.0020) physical activity as well as energy expenditure (p = 0.0366) assessed by the IPAQ and in the percentage of sedentary behaviour (p = 0.0330) and the average number of steps (p = 0.0342) assessed by ActiGraph.The awareness of physical activity assessment might decrease the ability to subjectively assess physical activity in subjects with MCI.

BackgroundIn solid tumors, CAR T cell efficacy is limited by off-tumor toxicity and suppression by the tumor microenvironment (TME). AB-X is an integrated circuit T cell (ICT cell) intended for the treatment of ovarian cancer. AB-X includes a transgene cassette with two functional modules: 1) an ”AND” logic gate designed to limit off-tumor toxicity through dual tumor antigen recognition; 2) a dual shRNA-miR to resist TME suppression and improve ICT cell function. The AB-X logic gate consists of a priming receptor that induces expression of an anti-mesothelin (MSLN) CAR upon engagement of a ALPG/P (alkaline phosphatase germ-line/placental). The dual shRNA-miR mediates downregulation of FAS and PTPN2. The AB-X DNA cassette is inserted into the T cell genome at a defined novel genomic site via CRISPR-based gene editing.MethodsDual-antigen specificity of the logic gate was assessed in mice harboring MSLN+ and ALPG/P+MSLN+ K562 tumors established on contralateral flanks. Potency was measured in a subcutaneous MSTO xenograft model. Logic-gated ICT cells were compared with MSLN CAR T cells in both models. In vitro, expansion of ICT cells with the FAS/PTPN2 shRNA-miR was evaluated in a 14 day repetitive stimulation assay (RSA). In vivo, expansion and potency were measured in the MSTO xenograft model. An in vitro FAS cross-linking assay was conducted to assess the impact of FAS knockdown on FAS-mediated apoptosis.ResultsLogic-gated ICT cells demonstrated specific activity against ALPG/P+MSLN+ tumors, but had no effect against MSLN+ tumors in the K562 in vivo specificity model. In addition, logic-gated ICT cells demonstrated greater in vivo potency than MSLN CAR T cells in the MSTO xenograft model. In our RSA, ICT cells containing the FAS/PTPN2 shRNA-miR had 8-fold greater expansion than the MSLN CAR T cells. Enhanced expansion was confirmed in vivo with ICT cells demonstrating >10-fold expansion in tumors and peripheral blood, enabling comparable growth inhibition in MSTO xenografts at less than one quarter the dose of the MSLN CAR T cells. Importantly, PTPN2 knockdown resulted in balanced expansion of all T cell subsets, including CD45RA+, CCR7+ memory cells. Lastly, ICT cells containing the FAS/PTPN2 shRNA-miR were resistant to FAS-mediated apoptosis.ConclusionsAB-X ICT cells specifically recognize ALPG/P+MSLN+ tumors, demonstrate superior potency, expansion, and persistence compared with MSLN CAR T cells, and are resistant to ovarian TME suppression. AB-X will be evaluated in clinical trials for treatment of platinum resistant/refractory ovarian cancer.AcknowledgementsWe would like to acknowledge all of our colleagues at Arsenal Biosciences, without whom this work would not have been possible.

SUMMARY Coordinate control of T cell proliferation, survival, and differentiation are essential for host protection from pathogens and cancer. Long-lived memory cells, whose precursors are formed during the initial immunological insult, provide protection from future encounters, and their generation is the goal of many vaccination strategies. microRNAs (miRNAs) are key nodes in regulatory networks that shape effective T cell responses through the fine-tuning of thousands of genes. Here, using compound conditional mutant mice to eliminate miR-15/16 family miRNAs in T cells, we show that miR-15/16 restrict T cell cycle, survival, and memory T cell differentiation. High throughput sequencing of RNA isolated by cross-linking immuno-precipitation of AGO2 combined with gene expression analysis in miR-15/16-deficient T cells indicates that these effects are mediated through the direct inhibition of an extensive network of target genes within pathways critical to cell cycle, survival, and memory., In Brief Coordinate control of T cell proliferation, survival, and differentiation are essential for effective cell-mediated adaptive immunity. Gagnon et al. define roles for the miR-15/16 family of microRNAs in restricting T cell cycle and long-lived memory T cell accumulation through the direct inhibition of a very large network of target mRNAs., Graphical Abstract

Compared to coding sequences, untranslated regions of the transcriptome are not well conserved, and functional annotation of these sequences is challenging. Global relationships between nucleotide composition of 3′ UTR sequences and their sequence conservation have been appreciated since mammalian genomes were first sequenced, but the functional relevance of these patterns remain unknown. We systematically measured the effect on gene expression of the sequences of more than 25,000 RNA-binding protein (RBP) binding sites in primary mouse T cells using a massively parallel reporter assay. GC-rich sequences were destabilizing of reporter mRNAs and come from more rapidly evolving regions of the genome. These sequences were more likely to be folded in vivo and contain a number of structural motifs that reduced accumulation of a heterologous reporter protein. Comparison of full-length 3′ UTR sequences across vertebrate phylogeny revealed that strictly conserved 3′ UTRs were GC-poor and enriched in genes associated with organismal development. In contrast, rapidly evolving 3′ UTRs tended to be GC-rich and derived from genes involved in metabolism and immune responses. Cell-essential genes had lower GC content in their 3′ UTRs, suggesting a connection between unstructured mRNA noncoding sequences and optimal protein production. By reducing gene expression, GC-rich RBP-occupied sequences act as a rapidly evolving substrate for gene regulatory interactions.

Supplemental Digital Content is available in the text, Objective: To describe physiological responses of mine rescuers during a simulated mine emergency. Methods: Body-worn monitors (n = 74) and core temperature (Tc) capsules (n = 54) assessed heart rate (HR), respiration rate (RR), energy expenditure (EE), oxygen consumption (), Tc and skin temperature (Tskin), by team position and task. A multivariate analysis was performed with team positions, tasks, and measures as factors. Results: HRmean and HRpeak were 78.6% and 94.5%, respectively, of predicted maximum heart rate. Arduous labor tasks elicited higher HR, RR, and than casualty care. Captains exhibited lower HRmean, HRpeak, RR, RRpeak, , Tc, and Tskin compared with other positions. Tc mean exceeded 38.6 °C (n = 14 recorded Tc >39 °C). Conclusions: Captains’ physical loading and heat stress were lowest. Nonetheless, all tasks and positions induced high physical load and heat strain.

Exogenous ketone supplementation and whole-body cooling (WBC) have shown to independently influence exercise metabolism. Whether readily available ketone salts, with and without WBC, would provide similar metabolic benefits during steady-state aerobic and time-trial performances was investigated. Nine active males (VO2peak: 56.3 ± 2.2 mL·kg−1·min−1) completed three single-blind exercise sessions preceded by: (1) ingestion of placebo (CON), (2) ketone supplementation (0.3 g·kg−1 β-OHB) (KET), and (3) ketone supplementation with WBC (KETCO). Participants cycled in steady-state (SS, 60% Wmax) condition for 30-min, immediately followed by a 15-min time trial (TT). Skin and core temperature, cardio-metabolic, and respiratory measures were collected continuously, whereas venous blood samples were collected before and after supplementation, after SS and TT. Venous β-OHB was elevated, while blood glucose was lower, with supplementation vs. CON (p < 0.05). TT power output was not different between conditions (p = 0.112, CON: 190 ± 43.5 W, KET: 185 ± 40.4 W, KETCO: 211 ± 50.7 W). RER was higher during KETCO (0.97 ± 0.09) compared to both CON (0.88 ± 0.04, p = 0.012) and KET (0.88 ± 0.05, p = 0.014). Ketone salt supplementation and WBC prior to short-term exercise sufficiently increase blood β-OHB concentrations, but do not benefit metabolic shifts in fuel utilization or improve time trial performance.

Background : Determinants of COVID-19 vaccine acceptance among healthcare workers (HCW) remains poorly understood. We assessed HCWs’ willingness to be vaccinated and reasons underlying hesitancy. Methods : Cross-sectional survey across 17 healthcare institutions. HCWs eligible for vaccination (Pfizer-BioNTech mRNA) in December 2020 were invited to receive immunization. Multivariate logistic regression was performed to identify predictors of acceptance. Reasons for refusal among those who never intended to be vaccinated (i.e. firm refusers) and those who preferred delaying vaccination (i.e. vaccine hesitants) were assessed. Results : Among 2761 respondents (72% female, average age, 44), 2233 (80.9%) accepted the vaccine. Physicians, environmental services workers and healthcare managers were more likely to accept vaccination compared to nurses. Male sex, age over 50, rehabilitation center workers, and occupational COVID-19 exposure were independently associated with vaccine acceptance by multivariate analysis. Factors for refusal included vaccine novelty, wanting others to receive it first, and insufficient time for decision-making. Among those who declined, 74% reported they may accept future vaccination. Vaccine firm refusers were more likely than vaccine hesitants to distrust pharmaceutical companies and to prefer developing a natural immunity by getting COVID-19. Conclusions : Vaccine hesitancy exists among HCWs. Our findings provide useful information to plan future interventions and improve acceptance.

High-intensity interval exercise (HIIE) has been shown to be more effective than moderate-intensity exercise for increasing acute lipid oxidation and lowering blood lipids during exercise and postprandially. Exercise in cold environments is also known to enhance lipid oxidation; however, the immediate and long-term effects of HIIE exercise in cold are unknown. The purpose of this study was to examine the effects cold stress during HIIE on acute exercise metabolism and postprandial metabolism. Eleven recreationally active individuals (age: 23 ± 3 yr, weight: 80 ± 9.7 kg, V̇O

Metabolic flexibility is compromised in individuals suffering from metabolic diseases, lipo- and glucotoxicity, and mitochondrial dysfunctions. Exercise studies performed in cold environments have demonstrated an increase in lipid utilization, which could lead to a compromised substrate competition, glycotoxic-lipotoxic state, or metabolic inflexibility. Whether metabolic flexibility is altered during incremental maximal exercise to volitional fatigue in a cold environment remains unclear.Ten young healthy participants performed four maximal incremental treadmill tests to volitional fatigue, in a fasted state, in a cold (0 °C) or a thermoneutral (22.0 °C) environment, with and without a pre-exercise ingestion of a 75-g glucose solution. Metabolic flexibility was assessed via indirect calorimetry using the change in respiratory exchange ratio (ΔRER), maximal fat oxidation (ΔMFO), and where MFO occurred along the exercise intensity spectrum (ΔFatMultiple linear mixed-effects regressions revealed an increase in glucose oxidation from glucose ingestion and an increase in lipid oxidation from the cold during exercise (p 0.001). No differences were observed in metabolic flexibility as assessed via ΔRER (0.05 ± 0.03 vs. 0.05 ± 0.03; p = 0.734), ΔMFO (0.21 ± 0.18 vs. 0.16 ± 0.13 g minFollowing glucose loading, metabolic flexibility was unaffected during exercise to volitional fatigue in a cold environment, inducing an increase in lipid oxidation. These results suggest that competing pathways responsible for the regulation of fuel selection during exercise and cold exposure may potentially be mechanistically independent. Whether long-term metabolic influences of high-fat diets and acute lipid overload in cold and warm environments would impact metabolic flexibility remain unclear.

SummaryT cell activation is paramount for productive adaptive immune responses. CD28 is a key clinically targeted immunoregulatory receptor because it provides the prototypical costimulatory signal required for T cell activation. Therefore, a precise understanding of the molecular consequences of CD28 costimulation has direct therapeutic relevance. Here, we uncover that the microRNA cluster miR-17~92 is part of the molecular program triggered by CD28 costimulation and hence T cell activation. Combining genetics, transcriptomics, bioinformatics and biochemical miRNA:mRNA interaction maps we demonstrate that transgenic miR-17~92 can cell-intrinsically largely overcome defects caused by CD28-deficiency. miR-17~92 promotes transcription of a proinflammatory gene signature by enhancing the calcineurin/NFAT pathway. miR-17~92 binds to and represses a network of genes including several negative regulators of T cell activation. Finally, CD28-deficient T cells exhibit derepressed miR-17~92 target genes during activation, demonstrating that this non-coding RNA is required to shape the transcriptome. Thus, we propose that miR-17~92 constitutes a central mediator for T cell activation, integrating signals by the TCR and CD28 costimulation. In this model miR-17~92 facilitates T cell activation by dampening the breaks that prevent T cell activation.

Fat oxidation during exercise is associated with cardio-metabolic benefits, but the extent of which whole-body exercise modality elicits the greatest fat oxidation remains unclear. We investigated the effects of treadmill, elliptical and rowing exercise on fat oxidation in healthy individuals. Nine healthy males participated in three, peak oxygen consumption tests, on a treadmill, elliptical and rowing ergometer. Indirect calorimetry was used to assess maximal oxygen consumption (V̇O(2peak)), maximal fat oxidation (MFO) rates, and the exercise intensity MFO occurred (Fat(max)). Mixed venous blood was collected to assess lactate and blood gases concentrations. While V̇O(2peak) was similar between exercise modalities, MFO rates were higher on the treadmill (mean ± SD; 0.61 ± 0.06 g·min(-1)) compared to both the elliptical (0.41 ± 0.08 g·min(-1), p = 0.022) and the rower (0.40 ± 0.08 g·min(-1), p = 0.017). Fat(max) values were also significantly higher on the treadmill (56.0 ± 6.2 %V̇O(2peak)) compared to both the elliptical (36.8 ± 5.4 %V̇O(2peak), p = 0.049) and rower (31.6 ± 5.0 %V̇O(2peak), p = 0.021). Post-exercise blood lactate concentrations were also significantly lower following treadmill exercise (p = 0.021). Exercising on a treadmill maximizes fat oxidation to a greater extent than elliptical and rowing exercises, and remains an important exercise modality to improve fat oxidation, and consequently, cardio-metabolic health.

Regular physical activity (PA) improves cognitive functions, prevents brain atrophy, and delays the onset of cognitive decline, dementia, and Alzheimer’s disease. Presently, there are no specific recommendations for PA producing positive effects on brain health and little is known on its mediators. PA affects production and release of several peptides secreted from peripheral and central tissues, targeting receptors located in the central nervous system (CNS). This review will provide a summary of the current knowledge on the association between PA and cognition with a focus on the role of (neuro)peptides. For the review we define peptides as molecules with less than 100 amino acids and exclude myokines. Tachykinins, somatostatin, and opioid peptides were excluded from this review since they were not affected by PA. There is evidence suggesting that PA increases peripheral insulin growth factor 1 (IGF-1) levels and elevated serum IGF-1 levels are associated with improved cognitive performance. It is therefore likely that IGF-1 plays a role in PA induced improvement of cognition. Other neuropeptides such as neuropeptide Y (NPY), ghrelin, galanin, and vasoactive intestinal peptide (VIP) could mediate the beneficial effects of PA on cognition, but the current literature regarding these (neuro)peptides is limited.

Vaccination has a huge public health impact. Maintaining vaccine coverage is key to avoid the devastating consequences of resurgence. In the Province of Quebec, vaccine coverage in young children are sub-optimal, mostly due to ambivalence toward vaccine safety and efficacy. We previously conducted a regional study in the Quebec’s Eastern Townships region, the PromoVac Study, to test a new educational intervention, based on motivational interviewing techniques, aimed at promoting infant vaccination. This first study evidenced that the intervention led to a marked increase in mothers’ intention to vaccinate, and vaccine coverage in their infants. The current study protocol aims at scaling up these results at a provincial level using a randomized controlled trial design. This pragmatic, randomized, controlled, parallel-group clinical trial will compare the effectiveness of the motivational interviewing to an educational intervention, including the distribution of an information flyer as standard of care on vaccination coverage in four maternity wards across the Province of Quebec (PromovaQ). Adult mothers of children born in participating maternity wards were recruited between March 2014 and February 2015. Vaccination coverage will be assessed at 3-years of age, thus the trial is expected to be completed in March 2019. Statistical analyses will be conducted under the intention-to-treat principle. Vaccine coverage will be analyzed using Chi-squared distribution testing and logistic regression to identify determinant factors. Secondary outcomes will include vaccine hesitation and intention scores, mother’s knowledge, attitudes and beliefs about immunization, and psychosocial determinants of intention to vaccinate. In the case results of this Provincial RCT be confirmed, serious consideration should then be given by Ministry of Health authorities to the possible implementation of MI-based strategies across provincial maternity wards. To ensure adequate input and secure implementation, study design and results will be reviewed with relevant stakeholders, including the children’s families, and provincial and regional decision-makers. Results will be adapted and shared with all stakeholders. ClinicalTrials.gov NCT02666872 (Retrospectively registered as January 28, 2016).

IF=0.2; International audience; Aircraft seat accessibility for wheelchair users is very complicated. The distance between two seats is very small which creates accommodation deficiencies. Improving air travel experience, and more specifically egress and ingress of aircraft seat could be associated with development of some adaptations. An experimental study was made to propose three removal adaptations on aircraft seat and to collect perceptions of wheelchair users. According to the results the adaptation the most appreciate was when surfaces of the boarding chair and of the aircraft seat were fixed at the same level. The results of this study give perspectives for further researches on the impact of some adaptations which could be provided on aircraft seat. Results might also be used to design products and service to improve sitting transfer.

Objectives To understand the reasons why women present to the Emergency Room (ER) for Early Pregnancy Loss (EPL)-related care, how they perceive care and counseling there, and their overall experience during and after their visit. Study design This qualitative study utilized semi-structured telephone interviews. Participants were recruited in a large urban ER; women who experienced EPL were interviewed by telephone about their experiences 1–3 weeks after their visit. Audio recordings were transcribed and coded by two independent coders. Main outcome measures This qualitative study utilized semi-structured interviews without the use of formal outcome measurement tools. Results Of the sixty-seven women recruited, ten completed the full telephone interview. Interview participants’ responses were grouped into four categories: Feelings about EPL, reasons for going to the ER, experience in the ER, and experience after leaving the ER. Women had mixed feelings about their ER experiences; many reported chaos, lack of information or lack of emotional support, while a few felt informed and supported. Many did not know much about EPL before their experience. Conclusions ER care for women experiencing suspected or confirmed EPL may not be addressing the emotional needs and knowledge gaps of women. Patient education, emotional support, and clear plans for outpatient follow up are critical. Further research is needed to guide interventions to improve care.

Parental decision making about childhood vaccinations is complex and multidimensional. There is a perception that the number of parents having concerns regarding childhood vaccinations has been increasing in Canada. The aim of this study was to explore vaccine hesitancy among Canadian parents and to examine factors associated with a parent’s intention to vaccinate his/her child. Informed by the Theory of Planned Behaviour (TPB) this study assesses potential associations between parents’ knowledge, attitudes and beliefs toward vaccination and their intention to vaccinate their child in the future. A national sample of Canadian parents of children aged 24–59 months (N = 2013) was surveyed using an online survey methodology. Half of the surveyed parents strongly intended to have their child vaccinated in the future. Parents’ information needs and searches as well as parents’ trust in different institutions were associated with intention to vaccinate. Parents who reported having frequently looked for vaccine information, who considered that it was their role as parents to question vaccines, or who had previously experienced difficulty accessing vaccination services were less likely to strongly intend to vaccinate their child in the future. Parents who had a high level of trust in doctors and public health were most likely to strongly intend to vaccinate their child. Results of the multivariate analysis showed that positive attitudes (aOR = 8.0; 95% CI: 6.0, 10.4), higher perceived social support (aOR = 3.0; 95% CI: 2.3, 3.93), and higher perceived behavioural control (aOR = 1.8; 95% CI: 1.4, 2.43) were associated with parents’ intention to vaccinate their child. Findings of this study suggest that trust-building interventions that promote pro-vaccine social norms and that address negative attitudes toward vaccination could enhance vaccine acceptance among Canadian parents.

Purpose Nonadherence to medicines contributes to poor health outcomes, especially for patients with complicated medicine regimens. We examined adherence among patients at a family health center and the impact that barriers to getting medicines and negative beliefs about medicines have on adherence. Methods A survey was administered incorporating the 8-item Morisky Medication Adherence Scale, questions from the Beliefs about Medicine Questionnaire, and questions about patients' external barriers to getting medicines. Low adherence was examined by any external barrier and by higher negative beliefs, adjusting for patient characteristics. Results The convenience sample of 343 participants is demographically representative of the larger population. Among these patients, 54% report low adherence, 51% have at least 1 barrier to adherence, and 52% report more negative than positive beliefs about medicines. When beliefs and barriers are examined together, patients with negative beliefs are 49% less likely to adhere than those with more positive beliefs, whereas barriers show no significant impact on adherence. Conclusions Negative beliefs about medicines are as prevalent in this population as external barriers to accessing medicines, but negative beliefs were more significantly associated with adherence than external barriers. Physicians should identify and address patients' negative beliefs about medicines to improve adherence rates.

MicroRNAs (miRNAs) are a class of short noncoding RNAs that play critical roles in the regulation of a broad range of biological processes. Like transcription factors, miRNAs exert their effects by modulating the expression of networks of genes that operate in common or convergent pathways. CD8(+) T cells are critical agents of the adaptive immune system that provide protection from infection and cancer. Here, we review the important roles of miRNAs in the regulation of CD8(+) T cell biology and provide perspectives on the broader emerging principles of miRNA function.

BackgroundOxidative stress results in lipid, protein, and DNA oxidation, resulting in telomere erosion, chromosomal damage, and accelerated cellular aging. Training promotes healthy metabolic and oxidative profiles whereas the effects of multi-day, prolonged, and continuous exercise are unknown. This study investigated the effects of multi-day prolonged exercise on metabolic and oxidative stress as well as telomere integrity in healthy adults.MethodsFifteen participants performed a 14-day, 260-km, wilderness canoeing expedition (12 males) (EXP) (24 ± 7 years, 72 ± 6 kg, 178 ± 8.0 cm, 18.4 ± 8.4% BF, 47.5 ± 9.3 mlO2 kg–1 min–1), requiring 6–9 h of low- to moderate-intensity exercise daily. Ten controls participated locally (seven males) (CON) (31 ± 11 years, 72 ± 15 kg, 174 ± 10 cm, 22.8 ± 10.0% BF, 47.1 ± 9.0 mlO2 kg–1 min–1). Blood plasma, serum, and mononuclear cells were sampled before and after the expedition to assess hormonal, metabolic, and oxidative changes.ResultsSerum cholesterol, high- and low-density lipoprotein, testosterone, insulin, sodium, potassium, urea, and chloride concentrations were not different between groups, whereas triglycerides, glucose, and creatinine levels were lower following the expedition (p < 0.001). Malondialdehyde and relative telomere length (TL) were unaffected (EXP: 4.2 ± 1.3 vs. CON: 4.1 ± 0.7 μM; p > 0.05; EXP: 1.00 ± 0.48 vs. CON: 0.89 ± 0.28 TS ratio; p = 0.77, respectively); however, superoxidase dismutase activity was greater in the expedition group (3.1 ± 0.4 vs. 0.8 ± 0.5 U ml–1; p < 0.001).ConclusionThese results indicate a modest improvement in metabolic and oxidative profiles with increased superoxidase dismutase levels, suggesting an antioxidative response to counteract the exercise-associated production of free radicals and reactive oxygen species during prolonged exercise, mimicking the effects from long-term training. Although improved antioxidant activity may lead to increased TL, the present exercise stimulus was insufficient to promote a positive cellular aging profile with concordant chromosomal changes in our healthy and young participants.

Fat oxidation in cold environments and carbohydrate (CHO) use in hot environments are increased during exercise at steady-state submaximal workloads. However, the influence of cold and heat on fat and CHO oxidation curves remain unknown. We therefore examined the influence of a cold and warm ambient temperature on fat and CHO oxidation across a wide range of exercise intensities during treadmill and cycle ergometer exercise. Nine, young, healthy, male subjects completed four trials, during which they performed an incremental peak oxygen consumption (⩒O

A mass vaccination campaign with the 4CMenB vaccine (Bexsero®) was launched in a serogroup B endemic area in Quebec. A study was conducted to assess parents’ and adolescents’ opinions about the acceptability of the vaccine before and after the campaign (two telephone surveys). This paper reports the results of the second survey and describes the factors associated with complete and incomplete 4CMenB vaccine status. Overall, 82.5% of children and 58.7% of adolescents were completely vaccinated. A positive association between intention reported prior to the campaign and vaccine receipt reported after the campaign was observed for both children and adolescents. Protection against meningococcal diseases was the main reason reported for those who completed the 4CMenB dose series, while lack of time, interest or information remained one of the main reasons for having refused the vaccine or not having completed the series. About half of the vaccinees reported an adverse event after having received a dose of 4CMenB and pain at the injection site and fever were the most often cited. Neither negative perceptions regarding the safety of the vaccine nor report of adverse events after having received a dose of the vaccine were associated with the vaccine refusal.

Skin and core tissue cooling modulates skeletal muscle oxygenation at rest. Whether tissue cooling also influences the skeletal muscle deoxygenation response during exercise is unclear. We evaluated the effects of skin and core tissue cooling on skeletal muscle blood volume and deoxygenation during sustained walking and running. Eleven male participants walked or ran six times on a treadmill for 60 min in ambient temperatures of 22°C (Neutral), 0°C for skin cooling (Cold 1), and at 0°C following a core and skin cooling protocol (Cold 2). Difference between oxy/deoxygenated haemoglobin ([diffHb]: deoxygenation index) and total haemoglobin content ([tHb]: total blood volume) in the vastus lateralis (VL) muscle was measured continuously. During walking, lower [tHb] was observed at 1 min in Cold 1 and Cold 2 vs. Neutral (P˂0.05). Lower [diffHb] was seen at 1 and 10 min in Cold 2 vs. Neutral by 13.5 ± 1.2 µM and 15.3 ± 1.4 µM and Cold 1 by 10.4 ± 3.1 µM and 11.1 ± 4.1 µM, respectively (P˂0.05). During running, [tHb] was lower in Cold 2 vs. Neutral at 10 min only (P = 0.004). [diffHb] was lower at 1 min in Cold 2 by 11.3 ± 3.1 µM compared to Neutral and by 13.5 ± 2.8 µM compared to Cold 1 (P˂0.001). Core tissue cooling, prior to exercise, induced greater deoxygenation of the VL muscle during the early stages of exercise, irrespective of changes in blood volume. Skin cooling alone, however, did not influence deoxygenation of the VL during exercise.

We sought feedback from elderly patients living with asthma to understand their experience with assuming self-management roles for their asthma in order to inform the design and implementation of a primary care-based strategy that could best support their asthma control.We held six focus groups with a total of 31 English- and Spanish-speaking older adults with a current diagnosis of asthma. Focus groups addressed the effect of asthma on patients' lives and self-management strategies. Transcripts were analyzed using constant comparative techniques.Asthma exerted a consistent effect on patients' physical and psychological well-being. Common barriers to self-care included misuse of controller medications and uncertainty whether shortness of breath, fatigue, and cough were due to their asthma or some other chronic illness. Patients developed coping strategies to continue with daily activities even when experiencing symptoms, but did not recognize attainable asthma quality of life.Asthma had a distinct impact on elderly adults' quality of life; due to their longstanding history with this condition, many patients had accepted these symptoms as a "new normal." Developing strategies to reorient patients' perceptions of the possibilities for managing their illness will be critical to the success of asthma self-management support programs specific to older adults.

Primary prevention of human papillomavirus (HPV) through vaccination is a high priority in Canada's cancer prevention efforts. All Canadian provinces and territories have introduced publicly funded, school-based vaccination programs against HPV, but vaccine uptake remains suboptimal in some jurisdictions. We conducted a descriptive qualitative study to better understand the determinants of low HPV vaccine uptake and identify strategies to enhance vaccine acceptance using the socio-ecological model. In Quebec, interviews and focus groups were held in 2015-2016 with 70 key informants including immunization managers, school nurses, school principals, teachers and parents of Grade 4 students (9 years of age). Our findings showed that HPV vaccine uptake was dependent on many interrelated factors at the individual and interpersonal level (e.g. knowledge and attitudes of the different players involved in the vaccination system), at the community level (e.g. social group values and norms, media coverage around the HPV vaccine), at the organizational level (e.g. allocated resources, information provision, consent process, immunization setting and environment) and at the policy level (e.g. changes in provincial HPV vaccine program). We are using the data collection and interpretation tools and approaches developed by our team and used in Quebec to expand our study to four other provinces (British Columbia, Alberta, Ontario and Nova Scotia). We are conducting environmental scans, semi-structured interviews and a survey to better understand the determinants of low HPV vaccine uptake and identify strategies to enhance vaccine acceptance. Having an in-depth understanding of the determinants of HPV vaccination in school settings is critical in order to identify root causes of the suboptimal vaccine uptake and to develop tailored interventions to address these on both supply- and demand-side issues.

Introduction: Physical activity (PA) has been associated with telomere shortening. The association of PA intensity or volume with telomere length (TL) is nonetheless unclear. The aim of our study was to investigate the associations of exercise intensity and volume with TL in elderly adults from Northern Finland (65° latitude North). Methods: Seven hundred elderly subjects born in 1945 in the Oulu region were investigated. PA was measured during a 2-week period with a wrist-worn accelerometer. In addition, a questionnaire was used to assess sedentary time and to achieve a longitudinal PA history and intensity. Relative telomere lengths (RTL) were determined from frozen whole blood samples using a qPCR-based method. Results: Relative telomere lengths were significantly longer in women than men and negatively correlated with age in both genders (men r = -0.210, p = 0.000, women r = -0.174, and p = 0.000). During the 2-week study period, women took more steps than men (p = 0.001), but the association between steps and RTL was only seen in men (p = 0.05). Total steps taken (r = 0.202 and p = 0.04) and sedentary time (r = -0.247 and p = 0.007) significantly correlated with RTLs in 70-year old subjects. Moderate PA was associated with RTL in subjects with the highest quartile of moderate PA compared to the three lower quartiles (p-values: 0.023 between 4th and 1st, 0.04 between 4th and 2nd, and 0.027 between 4th and 3rd) in the 70-year old subjects. Conclusion: Women had longer RTL and a higher step count compared to men. However, exercise volume and RTL correlated positively only in men. Surprisingly, age correlated negatively with RTL already within an age difference of 2 years. This suggests that telomere attrition rate may accelerate in older age. Moderate physical activity at the time of study was associated with RTL.

International audience; Introduction. While the scientific consensus on the benefits of vaccination is unambiguous, there is a growing proportion of the population that is skeptical about vaccination. The idea that vaccination programs are losing their momentum concerns public health agencies throughout the world. Many studies assessing determinants of vaccine acceptance have been published in the last decade. Areas covered. In this article, we review the existing qualitative literature on parents' attitudes toward childhood vaccination. Studies were included if they: (1) focused on the views, decision-making, or experiences of caregivers (hereafter, referred to as `parents') regarding vaccinations for young children; (2) used qualitative methods for both data and data analysis; (3) were conducted in countries that ranked `very high' on the 2016 United Nations Human Development Index; and (4) had been peer-reviewed. Twenty-two (22) studies met our inclusion criteria and were reviewed, using the socio-ecological model as a conceptual framework. Expert commentary. Parental vaccination decisions are complex and multi-dimensional. Experiences, emotions, routine ways of thinking, information sources, peers/family, risk perceptions, and trust, among other factors, inform parents' attitudes and decision-making processes. Further research is needed in order to design evidence-informed responses to vaccine hesitancy appropriate to the setting, context, and hesitant subgroups.

Objective Influenza vaccine uptake among Canadian pregnant individuals is suboptimal. Failure to incorporate vaccination into routine prenatal care and a lack of recommendations from healthcare providers are recognized as barriers to vaccination. The aim of this study was to assess Canadian maternity care providers’ knowledge, attitudes, and practices regarding influenza vaccination in pregnancy. Methods A cross-sectional Web-based questionnaire was sent during July and August 2017 to family physicians, obstetricians-gynaecologists, midwives, pharmacists, and nurses who care for pregnant individuals. A multivariable logistic regression model was used to determine variables independently associated with providers’ recommendation of the influenza vaccine in pregnancy. Results The analysis included 1061 providers. Most participants (85%) reported being vaccinated against influenza themselves, and 72% reported recommending the influenza vaccine to all of their pregnant patients during the previous influenza season. Participants’ attitudes regarding influenza vaccination during pregnancy were generally positive: 64% strongly agreed that pregnant individuals are at an increased risk of complications from influenza, and 69% strongly agreed that it is safe to vaccinate pregnant individuals against influenza. The main determinants of participants’ recommendations for influenza vaccination to all pregnant patients were following official recommendations on influenza vaccination, discussing vaccines with most or all pregnant individuals seen in their practice, and being vaccinated themselves during the previous influenza season. Conclusion Enhancing influenza vaccine uptake in pregnancy is largely dependent on maternity care providers’ recommendations. This study provides valuable insight on providers’ knowledge, attitudes, and practices.

The majority of genetic variants associated with common human diseases map to enhancers, non-coding elements that shape cell-type-specific transcriptional programs and responses to extracellular cues1–3. Systematic mapping of functional enhancers and their biological contexts is required to understand the mechanisms by which variation in non-coding genetic sequences contributes to disease. Functional enhancers can be mapped by genomic sequence disruption4–6, but this approach is limited to the subset of enhancers that are necessary in the particular cellular context being studied. We hypothesized that recruitment of a strong transcriptional activator to an enhancer would be sufficient to drive target gene expression, even if that enhancer was not currently active in the assayed cells. Here we describe a discovery platform that can identify stimulus-responsive enhancers for a target gene independent of stimulus exposure. We used tiled CRISPR activation (CRISPRa)7 to synthetically recruit a transcriptional activator to sites across large genomic regions (more than 100 kilobases) surrounding two key autoimmunity risk loci, CD69 and IL2RA. We identified several CRISPRa-responsive elements with chromatin features of stimulus-responsive enhancers, including an IL2RA enhancer that harbours an autoimmunity risk variant. Using engineered mouse models, we found that sequence perturbation of the disease-associated Il2ra enhancer did not entirely block Il2ra expression, but rather delayed the timing of gene activation in response to specific extracellular signals. Enhancer deletion skewed polarization of naive T cells towards a pro-inflammatory T helper (TH17) cell state and away from a regulatory T cell state. This integrated approach identifies functional enhancers and reveals how non-coding variation associated with human immune dysfunction alters context-specific gene programs.

During tumorigenesis, a heterotypic interface exists between cancer and stromal cells that can both support and repress tumor growth. In the breast, studies have demonstrated a pro-tumorigenic role for adipocytes. However, the molecular mechanisms by which breast cancer cells coopt adipocytes remain elusive. Studying breast tumors and normal adjacent tissue (NAT) from several patient cohorts, patient-derived xenografts and mouse models, we show that lipolysis and lipolytic signaling are activated in NAT. We investigated the tumor-adipocyte interface and find that functional gap junctions form between breast cancer cells and adipocytes. As a result, cAMP, a critical lipolysis-inducing signaling molecule, is transferred from breast cancer cells to adipocytes and activates lipolysis in a gap junction-dependent manner. We found that gap junction formation depends upon connexin 31 (Cx31), and that Cx31 is essential for breast tumor growth and activation of lipolysisin vivo. Thus, direct tumor cell-adipocyte interaction is critical for tumorigenesis, and may serve as a new therapeutic target in breast cancer.One sentence summaryGap junctions between breast cancer cells and adipocytes transfer cAMP and activate lipolysis in the breast tumor microenvironment.

The clear role of autophagy in human inflammatory diseases such as Crohn disease was first identified by genome-wide association studies and subsequently dissected in multiple mechanistic studies. ATG16L1 has been particularly well studied in knockout and hypomorph settings as well as models recapitulating the Crohn disease–associated T300A polymorphism. Interestingly, ATG16L1 has a single homolog, ATG16L2, which is independently implicated in diseases, including Crohn disease and systemic lupus erythematosus. However, the contribution of ATG16L2 to canonical autophagy pathways and other cellular functions is poorly understood. To better understand its role, we generated and analyzed the first, to our knowledge, ATG16L2 knockout mouse. Our results show that ATG16L1 and ATG16L2 contribute very distinctly to autophagy and cellular ontogeny in myeloid, lymphoid, and epithelial lineages. Dysregulation of any of these lineages could contribute to complex diseases like Crohn disease and systemic lupus erythematosus, highlighting the value of examining cell-specific effects. We also identify a novel genetic interaction between ATG16L2 and epithelial ATG16L1. These findings are discussed in the context of how these genes may contribute distinctly to human disease.

Coordinate control of T cell proliferation, survival, and differentiation are essential for host protection from pathogens and cancer. Long-lived memory cells, whose precursors are formed during the initial immunological insult, provide protection from future encounters and their generation is the goal of many vaccination strategies. microRNAs are key nodes in regulatory networks that shape effective T cell responses through the fine-tuning of thousands of genes. Here, using new compound conditional mutant mice to eliminate miR-15/16 family miRNAs in T cells, we show that miR-15/16 restrict T cell cycle, survival, and memory T cell differentiation. High throughput sequencing of RNA isolated by cross-linking immunoprecipitation of AGO2 combined with gene expression analysis in miR-15/16 deficient T cells indicate that these effects are mediated through the direct inhibition of an extensive network of known and novel target genes within pathways critical to cell cycle, survival, and memory. This study highlights the important role that miRNAs play in shaping the global gene expression program that is required for T cell cycle, survival, and the proper formation of memory cells.