In Situ Characterization of Human Lymphoid Tissue Immune Cells by Multispectral Confocal Imaging and Quantitative Image Analysis; Implications for HIV Reservoir Characterization.

CD4 T cells are key mediators of adaptive immune responses during infection and vaccination. Within secondary lymphoid organs, helper CD4 T cells, particularly those residing in germinal centers known as follicular helper T cells (Tfh), provide critical help to B-cells to promote their survival, isotype switching and selection of high affinity memory B-cells. On the other hand, the important role of Tfh cells for the maintenance of HIV reservoir is well documented. Thus, interrogating and better understanding the tissue specific micro-environment and immune subsets that contribute to optimal Tfh cell differentiation and function is important for designing successful prevention and cure strategies. Here, we describe the development and optimization of eight multispectral confocal microscopy immunofluorescence panels designed for in depth characterization and immune-profiling of relevant immune cells in formalin-fixed paraffin-embedded human lymphoid tissue samples. We provide a comprehensive library of antibodies to use for the characterization of CD4+ T-cells -including Tfh and regulatory T-cells- as well as CD8 T-cells, B-cells, macrophages and dendritic cells and discuss how the resulting multispectral confocal datasets can be quantitatively dissected using the HistoCytometry pipeline to collect information about relative frequencies and immune cell spatial distributions. Cells harboring actively transcribed virus are analyzed using an in-situ hybridization assay for the characterization of HIV mRNA positive cells in combination with additional protein markers (multispectral RNAscope). The application of this methodology to lymphoid tissues offers a means to interrogate multiple relevant immune cell targets simultaneously at increased resolution in a reproducible manner to guide CD4 T-cell studies in infection and vaccination

Large Asymmetric Hypertrophy of Rectus Abdominis Muscle in Professional Tennis Players

Purpose: To determine the volume and degree of asymmetry of the musculus rectus abdominis (RA) in professional tennis players. Methods: The volume of the RA was determined using magnetic resonance imaging (MRI) in 8 professional male tennis players and 6 non-active male control subjects. Results: Tennis players had 58 % greater RA volume than controls (P = 0.01), due to hypertrophy of both the dominant (34% greater volume, P = 0.02) and non-dominant (82 % greater volume, P = 0.01) sides, after accounting for age, the length of the RA muscle and body mass index (BMI) as covariates. In tennis players, there was a marked asymmetry in the development of the RA, which volume was 35 % greater in the non-dominant compared to the dominant side (P,0.001). In contrast, no sideto-side difference in RA volume was observed in the controls (P = 0.75). The degree of side-to-side asymmetry increased linearly from the first lumbar disc to the pubic symphysis (r = 0.97, P,0.001). Conclusions: Professional tennis is associated with marked hypertrophy of the musculus rectus abdominis, which achieves a volume that is 58 % greater than in non-active controls. Rectus abdominis hypertrophy is more marked in the non-dominant than in the dominant side, particularly in the more distal regions. Our study supports the concept that humans can differentially recruit both rectus abdominis but also the upper and lower regions of each muscle. It remains to b

Multilevel human secondary lymphoid immune system compartmentalization revealed by complementary imaging approaches.

Secondary human lymphoid tissue immune reactions take place in a highly coordinated environment with compartmentalization representing a fundamental feature of this organization. In situ profiling methodologies are indispensable for the understanding of this compartmentalization. Here, we propose a complementary experimental approach aiming to reveal different aspects of this process. The analysis of human tonsils, using a combination of single cell phenotypic analysis based on flow cytometry and multiplex imaging and mass spectrometry-based methodologies, revealed a compartmentalized organization at the cellular and molecular levels. More specifically, the skewed distribution of highly specialized immune cell subsets and relevant soluble mediators was accompanied by a compartmentalized localization of several lipids across different anatomical areas of the tonsillar tissue. The performance of such combinatorial experimental approaches could lead to the identification of novel in situ interactions and molecular targets for the in vivo manipulation of lymphoid organ, particularly the germinal center, immune reactions

Validity and reliability of the international fItness scale (IFIS) in preschool children

Objectives Examine the validity and reliability of parent-reported International FItness Scale (IFIS) in preschoolers. Method A cross-sectional study of 3051 Spanish preschoolers (3-5 years). Fitness was measured by PREFIT battery and reported by parents using an adapted version of the IFIS. Waist circumference was evaluated, and the waist-to-height ratio (WHtR) was calculated. Seventy-six parents of randomly selected schoolchildren completed the IFIS twice for a reliability assessment. Results ANCOVA, adjusted for sex, age and WHtR, showed that preschoolers who were scored by their parents as having average-to-very good fitness had better levels of measured physical fitness than those preschoolers who were classified as having very poor/poor fitness levels (18.1laps to 22.1laps vs 15.6laps for cardiorespiratory fitness; 6.6 kg to 7.5 kg vs 5.3 kg for muscular fitness-handgrip-; 71.7 cm to 76.4 cm vs 62.0 cm for muscular fitness-standing long jump-; 17.2s to 16.2s vs 18.2s for speed/agility; and 11.2s to 15.6s vs 8.7s for balance; p < 0.001). The weighted kappa for concordance between parent-reported fitness levels and objective assessment was poor (kappa <= 0.18 for all fitness measures). Overall, the mean values of the abdominal adiposity indicators were significantly lower in high-level fitness categories reported by parents than in low-level fitness categories (p < 0.05). The test-retest reliability ranged from 0.46 to 0.62. Conclusions The reliability of the parent-reported IFIS are acceptable, but the concordance between parents reported and objectively measures fitness levels is poor, suggesting that parents' responses may not be able to correctly classify preschoolers according to their fitness level.The PREFIT project takes place thanks to the funding linked to the Ramon y Cajal grant held by Ortega FB (RYC-2011-09011)

Contribution of Individual and Environmental Factors to Physical Activity Level among Spanish Adults

BACKGROUND: Lack of physical activity (PA) is a major risk for chronic disease and obesity. The main aims of the present study were to identify individual and environmental factors independently associated with PA and examine the relative contribution of these factors to PA level in Spanish adults. METHODOLOGY/PRINCIPAL FINDINGS: A population-based cross-sectional sample of 3,000 adults (18-75 years old) from Gran Canaria (Spain) was selected using a multistage stratified random sampling method. The participants were interviewed at home using a validated questionnaire to assess PA as well as individual and environmental factors. The data were analyzed using bivariate and multivariate logistic regression. One demographic variable (education), two cognitive (self-efficacy and perceived barriers), and one social environmental (organized format) were independently associated with PA in both genders. Odds ratios ranged between 1.76-2.07 in men and 1.35-2.50 in women (both p<0.05). Individual and environmental factors explained about one-third of the variance in PA level. CONCLUSIONS/SIGNIFICANCE: Self-efficacy and perceived barriers were the most significant factors to meet an adequate level of PA. The risk of insufficient PA was twofold greater in men with primary or lesser studies and who are employed. In women, living in rural environments increased the risk of insufficient PA. The promotion of organized PA may be an efficient way to increase the level of PA in the general population. Improvement in the access to sport facilities and places for PA is a prerequisite that may be insufficient and should be combined with strategies to improve self-efficacy and overcome perceived barriers in adulthood

Associations between Screen Time and Physical Activity among Spanish Adolescents

Excessive time in front of a single or several screens could explain a displacement of physical activity. The present study aimed at determining whether screen-time is associated with a reduced level of moderate to vigorous physical activity (MVPA) in Spanish adolescents living in favorable environmental conditions. or more to total screen-time showed a 64% (OR = 0.61, 95% CI, 0.44–0.86) increased risk of failing to achieve the recommended adolescent MVPA level. Participation in organized physical activities and sports competitions were more strongly associated with MVPA than screen-related behaviors.No single screen-related behavior explained the reduction of MVPA in adolescents. However, the total time accumulated through several screen-related behaviors was negatively associated with MVPA level in boys. This association could be due to lower availability of time for exercise as the time devoted to sedentary screen-time activities increases. Participation in organized physical activities seems to counteract the negative impact of excessive time in front of screens on physical activity

The effects of repeated-sprint training on field-based fitness measures: a meta-analysis of controlled and non-controlled trials

Background: Repeated-sprint training appears to be an efficient and practical means for the simultaneous development of different components of fitness relevant to team sports. Objective: Our objective was to systematically review the literature and meta-analyse the effect of repeated-sprint training on a selection of field-based measures of athletic performance, i.e. counter-movement jump, 10 m sprint, 20 m sprint, 30 m sprint, repeated-sprint ability and high-intensity intermittent running performance. Data Sources: The SPORTDiscus, PubMed, MEDLINE and Web of Science databases were searched for original research articles. Search terms included 'repeated-sprint training', 'sprint training', 'aerobic endurance', 'repeated-sprint ability', 'counter-movement jump' and 'sprint performance'. Study Selection: Inclusion criteria included intervention consisting of a series of ≤10 s sprints with ≤60 s recovery; trained participants; intervention duration of 2–12 weeks; field-based fitness measures; running- or cycling-based intervention; published up to, and including, February 2014. Data Extraction: Our final dataset included six trials for counter-movement jump (two controlled trials), eight trials for 10 m sprint, four trials for 20 m sprint (three controlled trials), two trials for 30 m sprint, eight trials for repeated-sprint ability and three trials for high-intensity intermittent running performance. Analyses were conducted using comprehensive meta-analysis software. Uncertainty in the meta-analysed effect of repeated-sprint training was expressed as 95 % confidence limits (CL), along with the probability that the true value of the effect was trivial, beneficial or harmful. Magnitude-based inferences were based on standardised thresholds for small, moderate and large changes of 0.2, 0.6 and 1.2 standard deviations, respectively. Results: Repeated-sprint training had a likely small beneficial effect in non-controlled counter-movement jump trials (effect size 0.33; 95 % CL ±0.30), with a possibly moderate beneficial effect in controlled trials (0.63; 95 % CL ±0.44). There was a very likely small beneficial effect on 10 m sprint time in non-controlled trials (−0.42; 95 % CL ±0.24), with a possibly moderate beneficial effect on 20 m sprint time in non-controlled (−0.49; 95 % CL ±0.46) and controlled (−0.65; 95 % CL ±0.61) trials. Repeated-sprint training had a possibly large beneficial effect on 30 m sprint performance in non-controlled trials (−1.01; 95 % CL ±0.93), with possibly moderate beneficial effects on repeated-sprint ability (−0.62; 95 % CL ±0.25) and high-intensity intermittent running performance (−0.61; 95 % CL ±0.54). Conclusions: Repeated-sprint training can induce small to large improvements in power, speed, repeated-sprint ability and endurance, and may have relevance for training in team sports

Immune Mechanisms of Viral Control in HIV-2 Infection

Human immunodeficiency virus 2 (HIV-2) infection is a zoonosis in which simian immunodeficiency virus from a West African monkey species; the sooty mangabey is thought to have entered the human population on at least eight separate occasions. This has given rise to eight distinct HIV-2 groups, of which only groups A and B have continued to spread among humans; the other clades appear only to have led to single-person infections. Viral control in HIV-2 infection is associated with several distinct features-a high-magnitude cellular immune response directed toward conserved Gag epitopes, an earlier-differentiated CD8 + T cell phenotype with increased polyfunctionality and exceptionally high functional avidity, supported by polyfunctional virus-specific CD4 + T cells, against a background of substantially less extensive immune activation than is seen in human immunodeficiency virus 1 (HIV-1) infection. Emerging as one of the most striking differences from HIV-1 infection is the slower evolution and a possible lower frequency of adaptive immune escape in asymptomatic HIV-2-infected individuals. © 2012 Elsevier Inc. All rights reserved

Visualization of HIV-1 reservoir: an imaging perspective.

The persistence of HIV-1-infected cells, despite the introduction of the combinatorial antiretroviral therapy, is a major obstacle to HIV-1 eradication. Understanding the nature of HIV reservoir will lead to novel therapeutic approaches for the functional cure or eradication of the virus. In this review, we will update the recent development in imaging applications toward HIV-1/simian immunodeficiency virus (SIV) viral reservoirs research and highlight some of their limitations. CD4 T cells are the primary target of HIV-1/SIV and the predominant site for productive and latent reservoirs. This viral reservoir preferentially resides in lymphoid compartments that are difficult to access, which renders sampling and measurements problematical and a hurdle for understanding HIV-1 pathogenicity. Novel noninvasive technologies are needed to circumvent this and urgently help to find a cure for HIV-1. Recent technological advancements have had a significant impact on the development of imaging methodologies allowing the visualization of relevant biomarkers with high resolution and analytical capacity. Such methodologies have provided insights into our understanding of cellular and molecular interactions in health and disease. Imaging of the HIV-1 reservoir can provide significant insights for the nature (cell types), spatial distribution, and the role of the tissue microenvironment for its in vivo dynamics and potentially lead to novel targets for the virus elimination

Human lymph node immune dynamics as driver of vaccine efficacy: an understudied aspect of immune responses.

During the last century, changes in hygiene, sanitation, and the advent of childhood vaccination have resulted in profound reductions in mortality from infectious diseases. Despite this success, infectious diseases remain an enigmatic public health threat, where effective vaccines for influenza, human immunodeficiency virus (HIV), tuberculosis, and malaria, among others remain elusive. In addition to the immune evasion tactics employed by complex pathogens, our understanding of immunopathogenesis and the development of effective vaccines is also complexified by the inherent variability of human immune responses. Lymph nodes (LNs) are the anatomical sites where B cell responses develop. An important, but understudied component of immune response complexity is variation in LN immune dynamics and in particular variation in germinal center follicular helper T cells (Tfh) and B cells which can be impacted by genetic variation, aging, the microbiome and chronic infection. We describe the contribution of genetic variation, aging, microbiome and chronic infection on LN immune dynamics and associated Tfh responses and offers perspective on how inclusion of LN immune subset and cytoarchitecture analyses, along with peripheral blood biomarkers can supplement systems vaccinology or immunology approaches for the development of vaccines or other interventions to prevent infectious diseases