A synthetic glycodendropeptide induces methylation changes on regulatory T cells linked to tolerant responses in anaphylactic-mice.

Introduction: Lipid transfer proteins (LTPs) are allergens found in a wide range of plant-foods. Specifically, Pru p 3, the major allergen of peach, is commonly responsible for severe allergic reactions. The need for new alternatives to conventional food allergy treatments, like restrictive diets, suggests allergen immunotherapy as a promising option. It has been demonstrated that sublingual immunotherapy (SLIT) with synthetic glycodendropeptides, such as D1ManPrup3, containing mannose and Pru p 3 peptides induced tolerance in mice and that the persistence of this effect depends on treatment dose (2nM or 5nM). Moreover, it produces changes associated with differential gene expression and methylation profile of dendritic cells, as well as phenotypical changes in regulatory T cells (Treg). However, there are no works addressing the study of epigenetic changes in terms of methylation in the cell subsets that sustain tolerant responses, Treg. Therefore, in this work, DNA methylation changes in splenic-Treg from Pru p 3 anaphylactic mice were evaluated. Methods: It was performed by whole genome bisulphite sequencing comparing SLIT-D1ManPrup3 treated mice: tolerant (2nM D1ManPrup3), desensitized (5nM D1ManPrup3), and sensitized but not treated (antigen-only), with anaphylactic mice. Results: Most of the methylation changes were found in the gene promoters from both SLIT-treated groups, desensitized (1,580) and tolerant (1,576), followed by the antigen-only (1,151) group. Although tolerant and desensitized mice showed a similar number of methylation changes, only 445 genes were shared in both. Remarkably, interesting methylation changes were observed on the promoter regions of critical transcription factors for Treg function like Stat4, Stat5a, Stat5b, Foxp3, and Gata3. In fact, Foxp3 was observed exclusively as hypomethylated in tolerant group, whereas Gata3 was only hypomethylated in the desensitized mice. Discussion: In conclusion, diverse D1ManPrup3 doses induce different responses (tolerance or desensitization) in mice, which are reflected by differential methylation changes in Tregs.

Corrigendum: MicroRNAs as potential regulators of immune response networks in asthma and chronic obstructive pulmonary disease.

[This corrects the article DOI: 10.3389/fimmu.2020.608666.].

Basophil Activation Test Utility as a Diagnostic Tool in LTP Allergy.

Plant-food allergy is an increasing problem, with nonspecific lipid transfer proteins (nsLTPs) triggering mild/severe reactions. Pru p 3 is the major sensitizer in LTP food allergy (FA). However, in vivo and in vitro diagnosis is hampered by the need for differentiating between asymptomatic sensitization and allergy with clinical relevance. The basophil activation test (BAT) is an ex vivo method able to identify specific IgE related to the allergic response. Thus, we aimed to establish the value of BAT in a precise diagnosis of LTP-allergic patients. Ninety-two individuals with peach allergy sensitized to LTP, Pru p 3, were finally included, and 40.2% of them had symptoms to peanut (n = 37). In addition, 16 healthy subjects were recruited. BAT was performed with Pru p 3 and Ara h 9 (peanut LTP) at seven ten-fold concentrations, and was evaluated by flow cytometry, measuring the percentage of CD63 (%CD63+) and CD203c (%CD203chigh) cells, basophil allergen threshold sensitivity (CD-Sens), and area under the dose−response curve (AUC). Significant changes in BAT parameters (%CD63+ and %CD203chigh) were found between the controls and patients. However, comparisons for %CD63+, %CD203chigh, AUC, and CD-Sens showed similar levels among patients with different symptoms. An optimal cut-off was established from ROC curves, showing a significant positive percentage of BAT in patients compared to controls and great values of sensitivity (>87.5%) and specificity (>85%). In addition, BAT showed differences in LTP-allergic patients tolerant to peanut using its corresponding LTP, Ara h 9. BAT can be used as a potential diagnostic tool for identifying LTP allergy and for differentiating peanut tolerance, although neither reactivity nor sensitivity can distinguish the severity of the clinical symptoms.

Role of miR-185-5p as modulator of periostin synthesis and smooth muscle contraction in asthma.

Asthma is a chronic respiratory disease produced by an aberrant immune response that originates with breathing difficulties and cough, through airway remodeling. The above pathophysiological events of asthma emerge the regulators of effectors, like epigenetics, which include microRNAs (miRNAs) who perform post-transcriptional regulation, controlling diverse pathways in respiratory diseases. The objective of the study was to determine how miR-185-5p regulates the secretion of periostin by airway structural cells, and smooth muscle cells contraction, both related to airway remodeling in asthma. We used miR-185-5p mimic and inhibitors in bronchial smooth muscle cells (BSMCs) and small airway epithelial cells (SAECs) from healthy subjects. Gene expression and protein levels of periostin (POSTN), CDC42, and RHOA were analyzed by RT-PCR and ELISA/Western blot, respectively. BSMC contractility was analyzed using cell-embedded collagen gels and measurement of intracellular calcium was performed using Fura-2. Additionally, miR-185-5p and periostin expression were evaluated in sputum from healthy and asthmatics. From these experiments, we observed that miR-185-5p modulation regulates periostin mRNA and protein in BSMCs and SAECs. A tendency for diminished miR-185-5p expression and higher periostin levels was seen in sputum cells from asthmatics compared to healthy, with an inverse correlation observed between POSTN and miR-185-5p. Inhibition of miR-185-5p produced higher BSMCs contraction induced by histamine. Calcium mobilization was not modified by miR-185-5p, showing that miR-185-5p role in BSMC contractility is performed by regulating CDC42 and RhoA pro-contractile factors instead. In conclusion, miR-185-5p is a modulator of periostin secretion by airway structural cells and of smooth muscle contraction, which can be related to asthma pathophysiology, and thus, might be a promising therapeutic target.

Epigenetics in Food Allergy and Immunomodulation.

Food allergy (FA) is an increasing problem worldwide and, over recent years, its prevalence is rising in developed countries. Nowadays, the immunological and cellular processes that occur in the allergic reactions are not fully understood, which hampers the development of in vitro diagnostic tools and further treatment options. Moreover, allergic diseases could be reinforced by environmental exposure and genetic modifications. Gene expression can be controlled by different epigenetic mechanisms like DNA methylation, histone modifications, and microRNAs. In addition, several environmental factors such as dietary components (vitamin D, butyrate, folic acid) are able to regulate this epigenetic mechanism. All these factors produce modifications in immune genes that could alter the development and function of immune cells, and therefore the etiology of the disease. Furthermore, these epigenetic mechanisms have also an influence on immunomodulation, which could explain sustained responsiveness or unresponsiveness during immunotherapy due to epigenetic modifications in key genes that induce tolerance in several FA. Thus, in this review we focus on the different epigenetic mechanisms that occur in FA and on the influence of several dietary components in these gene modifications.

Eosinophils and Chronic Respiratory Diseases in Hospitalized COVID-19 Patients.

BACKGROUND: Studies on the role of eosinophils in coronavirus disease 2019 (COVID-19) are scarce, though available findings suggest a possible association with disease severity. Our study analyzes the relationship between eosinophils and COVID-19, with a focus on disease severity and patients with underlying chronic respiratory diseases. METHODS: We performed a retrospective analysis of 3018 subjects attended at two public hospitals in Madrid (Spain) with PCR-confirmed SARS-CoV-2 infection from January 31 to April 17, 2020. Patients with eosinophil counts less than 0.02×109/L were considered to have eosinopenia. Individuals with chronic respiratory diseases (n=384) were classified according to their particular underlying condition, i.e., asthma, chronic pulmonary obstructive disease, or obstructive sleep apnea. RESULTS: Of the 3018 patients enrolled, 479 were excluded because of lack of information at the time of admission. Of 2539 subjects assessed, 1396 patients presented an eosinophil count performed on admission, revealing eosinopenia in 376 cases (26.93%). Eosinopenia on admission was associated with a higher risk of intensive care unit (ICU) or respiratory intensive care unit (RICU) admission (OR:2.21; 95%CI:1.42-3.45; p<0.001) but no increased risk of mortality (p>0.05). CONCLUSIONS: Eosinopenia on admission conferred a higher risk of severe disease (requiring ICU/RICU care), but was not associated with increased mortality. In patients with chronic respiratory diseases who develop COVID-19, age seems to be the main risk factor for progression to severe disease or death.

New Insights in Therapy for Food Allergy.

Food allergy is an increasing problem worldwide, with strict avoidance being classically the only available reliable treatment. The main objective of this review is to cover the latest information about the tools available for the diagnosis and treatment of food allergies. In recent years, many efforts have been made to better understand the humoral and cellular mechanisms involved in food allergy and to improve the strategies for diagnosis and treatment. This review illustrates IgE-mediated food hypersensitivity and provides a current description of the diagnostic strategies and advances in different treatments. Specific immunotherapy, including different routes of administration and new therapeutic approaches, such as hypoallergens and nanoparticles, are discussed in detail. Other treatments, such as biologics and microbiota, are also described. Therefore, we conclude that although important efforts have been made in improving therapies for food allergies, including innovative approaches mainly focusing on efficacy and safety, there is an urgent need to develop a set of basic and clinical results to help in the diagnosis and treatment of food allergies.

Clinical and inflammatory characteristics of patients with asthma in the Spanish MEGA project cohort.

INTRODUCTION: The MEGA (MEchanism underlying the Genesis and evolution of Asthma) project is a multicenter cohort study carried out in eight Spanish hospitals, gathering clinical, physiological, and molecular data from patients with asthma and multimorbidities in order to gain insight into the different physiopathological mechanisms involved in this disorder. MATERIAL AND METHODS: We report the baseline clinical and physiological characteristics and biomarker measures of adult participants in the project with the aim of better understanding the natural history and underlying mechanisms of asthma as well as the associated multimorbidities across different levels of severity. We carried out a detailed clinical examination, pulmonary function testing, measurement of fractional exhaled nitric oxide (FeNO), blood counts, induced sputum, skin prick tests, chest computed tomography scan, asthma questionnaires, and multimorbidity assessment in 512 asthmatic patients. RESULTS: When compared to patients with milder disease, severe asthmatic patients showed greater presence of symptoms, more exacerbations, lower asthma control, increased airflow obstruction, and higher frequency of chronic rhinosinusitis with nasal polyps, severe rhinitis, anxiety and depression, gastroesophageal reflux, and bronchiectasis. CONCLUSION: The MEGA project succeeded in recruiting a high number of asthma patients, especially those with severe disease, who showed lower control and higher frequency of multimorbidities.

Changes in Serum MicroRNAs after Anti-IL-5 Biological Treatment of Severe Asthma.

There is currently enough evidence to think that miRNAs play a role in several key points in asthma, including diagnosis, severity of the disease, and response to treatment. Cells release different types of lipid double-membrane vesicles into the extracellular microenvironment, including exosomes, which function as very important elements in intercellular communication. They are capable of distributing genetic material, mRNA, mitochondrial DNA, and microRNAs (miRNAs). Serum miRNA screening was performed in order to analyze possible changes in serum miRNAs in 10 patients treated with reslizumab and 6 patients with mepolizumab after 8 weeks of treatment. The expression of miR-338-3p was altered after treatment (p < 0.05), although no significant differences between reslizumab and mepolizumab were found. Bioinformatic analysis showed that miR-338-3p regulates important pathways in asthma, such as the MAPK and TGF-ß signaling pathways and the biosynthesis/degradation of glucans (p < 0.05). However, it did not correlate with an improvement in lung function. MiRNA-338-3p could be used as a biomarker of early response to reslizumab and mepolizumab in severe eosinophilic asthmatic patients. In fact, this miRNA could be involved in airway remodeling, targeting genes related to MAPK and TGF-ß signaling pathways.

Serum microRNAs as Tool to Predict Early Response to Benralizumab in Severe Eosinophilic Asthma.

Severe eosinophilic asthma poses a serious health and economic problem, so new therapy approaches have been developed to control it, including biological drugs such as benralizumab, which is a monoclonal antibody that binds to IL-5 receptor alpha subunit and depletes peripheral blood eosinophils rapidly. Biomarkers that predict the response to this drug are needed so that microRNAs (miRNAs) can be useful tools. This study was performed with fifteen severe eosinophilic asthmatic patients treated with benralizumab, and serum miRNAs were evaluated before and after treatment by semi-quantitative PCR (qPCR). Patients showed a clinical improvement after benralizumab administration. Additionally, deregulation of miR-1246, miR-5100 and miR-338-3p was observed in severe asthmatic patients after eight weeks of therapy, and a correlation was found between miR-1246 and eosinophil counts, including a number of exacerbations per year in these severe asthmatics. In silico pathway analysis revealed that these three miRNAs are regulators of the MAPK signaling pathway, regulating target genes implicated in asthma such as NFKB2, NFATC3, DUSP1, DUSP2, DUSP5 and DUSP16. In this study, we observed an altered expression of miR-1246, miR-5100 and miR-338-3p after eight weeks of benralizumab administration, which could be used as early response markers.

Serum carotenoids and Pediatric Metabolic Index predict insulin sensitivity in Mexican American children.

High concentrations of carotenoids are protective against cardiometabolic risk traits (CMTs) in adults and children. We recently showed in non-diabetic Mexican American (MA) children that serum α-carotene and ß-carotene are inversely correlated with obesity measures and triglycerides and positively with HDL cholesterol and that they were under strong genetic influences. Additionally, we previously described a Pediatric Metabolic Index (PMI) that helps in the identification of children who are at risk for cardiometabolic diseases. Here, we quantified serum lycopene and ß-cryptoxanthin concentrations in approximately 580 children from MA families using an ultraperformance liquid chromatography-photodiode array and determined their heritabilities and correlations with CMTs. Using response surface methodology (RSM), we determined two-way interactions of carotenoids and PMI on Matsuda insulin sensitivity index (ISI). The concentrations of lycopene and ß-cryptoxanthin were highly heritable [h2 = 0.98, P = 7 × 10-18 and h2 = 0.58, P = 1 × 10-7]. We found significant (P ≤ 0.05) negative phenotypic correlations between ß-cryptoxanthin and five CMTs: body mass index (- 0.22), waist circumference (- 0.25), triglycerides (- 0.18), fat mass (- 0.23), fasting glucose (- 0.09), and positive correlations with HDL cholesterol (0.29). In contrast, lycopene only showed a significant negative correlation with fasting glucose (- 0.08) and a positive correlation with HDL cholesterol (0.18). Importantly, we found that common genetic influences significantly contributed to the observed phenotypic correlations. RSM showed that increased serum concentrations of α- and ß-carotenoids rather than that of ß-cryptoxanthin or lycopene had maximal effects on ISI. In summary, our findings suggest that the serum carotenoids are under strong additive genetic influences and may have differential effects on susceptibility to CMTs in children.

Exosomes: A Key Piece in Asthmatic Inflammation.

Asthma is a chronic disease of the airways that has an important inflammatory component. Multiple cells are implicated in asthma pathogenesis (lymphocytes, eosinophils, mast cells, basophils, neutrophils), releasing a wide variety of cytokines. These cells can exert their inflammatory functions throughout extracellular vesicles (EVs), which are small vesicles released by donor cells into the extracellular microenvironment that can be taken up by recipient cells. Depending on their size, EVs can be classified as microvesicles, exosomes, or apoptotic bodies. EVs are heterogeneous spherical structures secreted by almost all cell types. One of their main functions is to act as transporters of a wide range of molecules, such as proteins, lipids, and microRNAs (miRNAs), which are single-stranded RNAs of approximately 22 nucleotides in length. Therefore, exosomes could influence several physiological and pathological processes, including those involved in asthma. They can be detected in multiple cell types and biofluids, providing a wealth of information about the processes that take account in a pathological scenario. This review thus summarizes the most recent insights concerning the role of exosomes from different sources (several cell populations and biofluids) in one of the most prevalent respiratory diseases, asthma.

Association of Antioxidant Vitamins A, C, E and Carotenoids with Cognitive Performance over Time: A Cohort Study of Middle-Aged Adults.

Carotenoids may strengthen the association of antioxidant vitamins A, C, and E with favorable cognitive outcomes over time, though a few prospective studies have examined this hypothesis. We evaluated the longitudinal data from 1251 participants in the Healthy Aging in Neighborhoods of Diversity across the Life Span (HANDLS) study (Age at visit 1 in 2004-2009 (v1): 30-65 years). Vitamins A, C, and E dietary intakes and total and individual dietary carotenoids were computed using two 24-h recalls at v1. Cognitive tests, covering global mental status and domains of memory/learning, attention, psychomotor speed, visuo-spatial, language/verbal, and executive function were conducted at v1 and/or v2 (2009-2013); mean ± SD follow-up: 4.66 ± 0.93 years. Mixed-effects linear regression models detected an interaction between vitamin E and total (and individual) carotenoids for three of 11 cognitive tests at v1, with only one meeting the statistical significance upon multiple testing correction whereby vitamin E was linked with greater verbal memory performance in the uppermost total carotenoid tertile (γ0a = +0.26 ± 0.08, p = 0.002), a synergism largely driven by carotenoid lycopene. Vitamins A and C showed no consistent interactions with carotenoids. In conclusion, we provide partial evidence for synergism between vitamin E and carotenoids in relation to better baseline cognitive performance, pending further studies with time-dependent exposures and randomized trials directly examining this synergism.

The Genomics and Metagenomics of Asthma Severity (GEMAS) Study: Rationale and Design.

Asthma exacerbations are a major contributor to the global disease burden, but no significant predictive biomarkers are known. The Genomics and Metagenomics of Asthma Severity (GEMAS) study aims to assess the role of genomics and the microbiome in severe asthma exacerbations. Here, we present the design of GEMAS and the characteristics of patients recruited from March 2018 to March 2020. Different biological samples and demographic and clinical variables were collected from asthma patients recruited by allergy and pulmonary medicine units in several hospitals from Spain. Cases and controls were defined by the presence/absence of severe asthma exacerbations in the past year (oral corticosteroid use, emergency room visits, and/or asthma-related hospitalizations). A total of 137 cases and 120 controls were recruited. After stratifying by recruitment location (i.e., Canary Islands and Basque Country), cases and controls did not differ for most demographic and clinical variables (p > 0.05). However, cases showed a higher proportion of characteristics inherent to asthma exacerbations (impaired lung function, severe disease, uncontrolled asthma, gastroesophageal reflux, and use of asthma medications) compared to controls (p < 0.05). Similar results were found after stratification by recruitment unit. Thereby, asthma patients enrolled in GEMAS are balanced for potential confounders and have clinical characteristics that support the phenotype definition. GEMAS will improve the knowledge of potential biomarkers of asthma exacerbations.

Biochemical and Hematological Correlates of Elevated Homocysteine in National Surveys and a Longitudinal Study of Urban Adults.

Elevated blood homocysteine (Hcy) among middle-aged adults can increase age-related disease risk, possibly through other biochemical and hematological markers. We selected markers for hyperhomocysteinemia among middle-aged adults, studied time-dependent Hcy-marker associations and computed highly predictive indices of hyperhomocysteinemia, with cross-sectional and longitudinal validations. We used data from the National Health and Nutrition Examination Survey (NHANES III, phase 2, nmax = 4000), the NHANES 1999-2006 (nmax = 10,151) and pooled NHANES (cross-sectional validation). Longitudinal validation consisted of mixed-effects linear regression models (Hcy predicting markers' annual rates of change), applied to the Healthy Aging in Neighborhoods of Diversity Across the Life Span (HANDLS, n = 227-244 participants, k = 2.4 repeats/participant, Agebase: 30-65 years) data. Machine learning detected nine independent markers for Hcy > 14 µmol/L (NHANES III, phase 2): older age; lower folate and B-12 status; higher serum levels of creatinine, uric acid, alkaline phosphatase, and cotinine; mean cell hemoglobin and red cell distribution widths (RDW); results replicated in the 1999-2006 NHANES [AUC = 0.60-0.80]. Indices combining binary markers increased elevated Hcy odds by 6.9-7.5-fold. In HANDLS, first-visit Hcy predicted annual increase in creatinine, RDW and alkaline phosphatase, with third-visit index (2013-2018) directly predicting Hcy (2004-2009). We provide evidence of the internal and external validity of indices composed of several biomarkers that are strongly associated with elevated Hcy.

MicroRNAs as Potential Regulators of Immune Response Networks in Asthma and Chronic Obstructive Pulmonary Disease.

Chronic respiratory diseases (CRDs) are an important factor of morbidity and mortality, accounting for approximately 6% of total deaths worldwide. The main CRDs are asthma and chronic obstructive pulmonary disease (COPD). These complex diseases have different triggers including allergens, pollutants, tobacco smoke, and other risk factors. It is important to highlight that although CRDs are incurable, various forms of treatment improve shortness of breath and quality of life. The search for tools that can ensure accurate diagnosis and treatment is crucial. MicroRNAs (miRNAs) are small non-coding RNAs and have been described as promising diagnostic and therapeutic biomarkers for CRDs. They are implicated in multiple processes of asthma and COPD, regulating pathways associated with inflammation, thereby showing that miRNAs are critical regulators of the immune response. Indeed, miRNAs have been found to be deregulated in several biofluids (sputum, bronchoalveolar lavage, and serum) and in both structural lung and immune cells of patients in comparison to healthy subjects, showing their potential role as biomarkers. Also, miRNAs play a part in the development or termination of histopathological changes and comorbidities, revealing the complexity of miRNA regulation and opening up new treatment possibilities. Finally, miRNAs have been proposed as prognostic tools in response to both conventional and biologic treatments for asthma or COPD, and miRNA-based treatment has emerged as a potential approach for clinical intervention in these respiratory diseases; however, this field is still in development. The present review applies a systems biology approach to the understanding of miRNA regulatory networks in asthma and COPD, summarizing their roles in pathophysiology, diagnosis, and treatment.

Systemic inflammation is associated with depressive symptoms differentially by sex and race: a longitudinal study of urban adults.

Systemic inflammation may influence trajectories of depressive symptoms over time, perhaps differentially by sex and race. Inflammatory markers and the Center for Epidemiologic Studies-Depression scale [total score: CES-Dtotal and four distinctive domains: somatic complaints, depressed affect, positive affect and interpersonal problems] were examined among African-American (AA) and White urban adults participating in the Healthy Aging in Neighborhoods of Diversity across the Life Span (HANDLS) study [2004-2013, Agebase:30-64 y, mean ± SD follow-up time: 4.64 ± 0.93 y, N = 150 (with cytokine data) to N = 1,767 (with other inflammatory markers)]. Findings suggest that serum concentrations of high-sensitivity C-reactive protein (hsCRP), z-inflammation composite score [ICS, combining elevated hsCRP and ESR with low serum albumin and iron], and serum interleukin (IL) 1ß were positively associated with ΔCES-Dtotal (Δ: annual rate of increase) among Whites only. IL-12 was directly related to ΔCES-Dtotal among men and AA. The race-specific associations of hsCRP, ICS, IL-1ß and the sex-specific association of IL-12 with ΔCES-Dtotal were replicated for the "depressed affect" domain. Similarly, among men, lower serum albumin and higher ICS were linked with higher baseline "somatic complaints". IL-10 among AA and IL-12 among men were inversely related to Δ"positive affect", while "interpersonal problems" were cross-sectionally associated with IL-6 among AA and IL-10 among Whites. Finally, baseline ICS was positively associated with incident "elevated depressive symptoms" (EDS: CES-Dtotal ≥ 16) among AA (HR = 1.28, 95% CI: 1.04-1.56, P = 0.017). Overall, systemic inflammation was directly linked to increased depressive symptoms over time and at baseline, differentially across sex and race groups. More longitudinal research is needed to replicate our findings.