RESUMEN
BACKGROUND: In the USA, genetically admixed populations have the highest asthma prevalence and severe asthma exacerbations rates. This could be explained not only by environmental factors but also by genetic variants that exert ethnic-specific effects. However, no admixture mapping has been performed for severe asthma exacerbations. OBJECTIVE: We sought to identify genetic variants associated with severe asthma exacerbations in Hispanic/Latino subgroups by means of admixture mapping analyses and fine mapping, and to assess their transferability to other populations and potential functional roles. METHODS: We performed an admixture mapping in 1124 Puerto Rican and 625 Mexican American children with asthma. Fine-mapping of the significant peaks was performed via allelic testing of common and rare variants. We performed replication across Hispanic/Latino subgroups, and the transferability to non-Hispanic/Latino populations was assessed in 1001 African Americans, 1250 Singaporeans and 941 Europeans with asthma. The effects of the variants on gene expression and DNA methylation from whole blood were also evaluated in participants with asthma and in silico with data obtained through public databases. RESULTS: Genomewide significant associations of Indigenous American ancestry with severe asthma exacerbations were found at 5q32 in Mexican Americans as well as at 13q13-q13.2 and 3p13 in Puerto Ricans. The single nucleotide polymorphism (SNP) rs1144986 (C5orf46) showed consistent effects for severe asthma exacerbations across Hispanic/Latino subgroups, but it was not validated in non-Hispanics/Latinos. This SNP was associated with DPYSL3 DNA methylation and SCGB3A2 gene expression levels. CONCLUSIONS: Admixture mapping study of asthma exacerbations revealed a novel locus that exhibited Hispanic/Latino-specific effects and regulated DPYSL3 and SCGB3A2.
Asunto(s)
Asma , Hispánicos o Latinos , Adolescente , Humanos , Asma/genética , Estudio de Asociación del Genoma Completo , Hispánicos o Latinos/genética , Polimorfismo de Nucleótido Simple , Estados Unidos/epidemiología , Niño , Americanos MexicanosRESUMEN
BACKGROUND: Severe bronchiolitis is often associated with subsequent respiratory morbidity, mainly recurrent wheezing and asthma. However, the underlying immune mechanisms remain unclear. The main goal of this study was to investigate the association of nasal detection of periostin and thymic stromal lymphopoietin (TSLP) during severe bronchiolitis with the development of asthma at 4 years of age. METHODS: Observational, longitudinal, post-bronchiolitis, hospital-based, follow-up study. Children hospitalized for bronchiolitis between October/2013 and July/2017, currently aged 4 years, included in a previous study to investigate the nasal airway secretion of TSLP and periostin during bronchiolitis, were included. Parents were contacted by telephone, and were invited to a clinical interview based on a structured questionnaire to obtain information on the respiratory evolution. The ISAAC questionnaire for asthma symptoms for 6-7-year-old children, was also employed. RESULTS: A total of 248 children were included (median age 4.4 years). The mean age at admission for bronchiolitis was 3.1 (IQR: 1.5-6.5) months. Overall, 21% had ever been diagnosed with asthma and 37% had wheezed in the last 12 months. Measurable nasal TSLP was detected at admission in 27(11%) cases and periostin in 157(63%). The detection of nasal TSLP was associated with the subsequent prescription of maintenance asthma treatment (p = 0.04), montelukast (p = 0.01), and the combination montelukast/inhaled glucocorticosteroids (p = 0.03). Admissions for asthma tended to be more frequent in children with TSLP detection (p = 0.07). In the multivariate analysis, adjusting for potential confounders, the detection of TSLP remained independently associated with chronic asthma treatment prescription (aOR:2.724; CI 1.051-7.063, p:0.04) and with current asthma (aOR:3.41; CI 1.20-9.66, p:0.02). Nasal detection of periostin was associated with lower frequency of ever use of short-acting beta2-agonists (SABA) (p = 0.04), lower prevalence of current asthma (p = 0.02), less prescription of maintenance asthma treatment in the past 12 months (p = 0.02, respectively). In the multivariate analysis, periostin was associated with lower risk of asthma at 4 years, independently of the atopic status (aOR:0.511 CI 95% 0.284-0.918, p:0.025). CONCLUSIONS: Our results show a positive correlation between nasal TSLP detection in severe bronchiolitis and the presence of current asthma, prescription of asthma maintenance treatment and respiratory admissions up to the age of 4 years. By contrast, we found a protective association between nasal periostin detection and current asthma at 4 years, ever diagnosis of asthma, maintenance asthma treatment prescription, and respiratory admissions.
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Asma , Bronquiolitis , Infecciones por Virus Sincitial Respiratorio , Niño , Preescolar , Humanos , Lactante , Asma/diagnóstico , Asma/tratamiento farmacológico , Asma/epidemiología , Asma/inmunología , Bronquiolitis/complicaciones , Bronquiolitis/diagnóstico , Bronquiolitis/epidemiología , Bronquiolitis/inmunología , Citocinas , Estudios de Seguimiento , Infecciones por Virus Sincitial Respiratorio/epidemiología , Linfopoyetina del Estroma TímicoRESUMEN
Asthma is a heterogeneous inflammatory disease of the airways that causes breathing difficulties, episodes of cough and wheezing, and in more severe cases can greatly diminish quality of life. Epigenetic regulation, including post-transcriptional mediation of microRNAs (miRNAs), is one of the mechanisms behind the development of the range of asthma phenotypes and endotypes. As in every other immune-mediated disease, miRNAs regulate the behavior of cells that shape the airway structure as well as those in charge of the defense mechanisms in the bronchi and lungs, controlling cell survival, growth, proliferation, and the ability of cells to synthesize and secrete chemokines and immune mediators. More importantly, miRNAs are molecules with chemical and biological properties that make them appropriate biomarkers for disease, enabling stratification of patients for optimal drug selection and thereby simplifying clinical management and reducing both the economic burden and need for critical care associated with the disease. In this review, we summarize the roles of miRNAs in asthma and describe how they regulate the mechanisms of the disease. We further describe the current state of miRNAs as biomarkers for asthma phenotyping, endotyping, and treatment selection.
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Asma , MicroARNs , Humanos , MicroARNs/genética , Epigénesis Genética , Calidad de Vida , Asma/diagnóstico , Asma/tratamiento farmacológico , Asma/genética , BiomarcadoresRESUMEN
Clarifying inflammatory processes and categorising asthma into phenotypes and endotypes improves asthma management. Obesity worsens severe asthma and reduces quality of life, although its specific molecular impact remains unclear. We previously demonstrated that hsa-miR-26a-1-3p and hsa-miR-376a-3p, biomarkers related to an inflammatory profile, discriminate eosinophilic from non-eosinophilic asthmatics. We aimed to study hsa-miR-26a-1-3p, hsa-miR-376a-3p, and their target genes in asthmatic subjects with or without obesity to find biomarkers and comprehend obese asthma mechanisms. Lung tissue samples were obtained from asthmatic patients (n = 16) and healthy subjects (n = 20). We measured miRNA expression using RT-qPCR and protein levels (IGF axis) by ELISA in confirmation samples from eosinophilic (n = 38) and non-eosinophilic (n = 39) obese (n = 26) and non-obese (n = 51) asthma patients. Asthmatic lungs showed higher hsa-miR-26a-1-3p and hsa-miR-376a-3p expression than healthy lungs. A study of seven genes regulated by these miRNAs revealed differential expression of IGFBP3 between asthma patients and healthy individuals. In obese asthma patients, we found higher hsa-miR-26a-1-3p and IGF-1R values and lower values for hsa-miR-376a-3p and IGFBP-3. Hsa-miR-26a-1-3p and IGFBP-3 were directly and inversely correlated with body mass index, respectively. Hsa-miR-26a-1-3p and hsa-miR-376a-3p could be used as biomarkers to phenotype patients with eosinophilic and non-eosinophilic asthma in relation to comorbid obesity.
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Asma , MicroARNs , Obesidad , Humanos , Asma/complicaciones , Asma/genética , Biomarcadores , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/genética , MicroARNs/genética , MicroARNs/metabolismo , Obesidad/complicaciones , Obesidad/genética , Fenotipo , Calidad de VidaRESUMEN
Nowadays, microRNAs (miRNAs) are increasingly used as biomarkers due to their potential contribution to the diagnosis and targeted treatment of a range of diseases. The aim of the study was to analyze the miRNA expression profiles in serum and lung tissue from patients with severe asthma treated with oral corticosteroids (OCS) and those without OCS treatment. For this purpose, serum and lung tissue miRNAs of OCS and non-OCS asthmatic individuals were evaluated by miRNAs-Seq, and subsequently miRNA validation was performed using RT-qPCR. Additionally, pathway enrichment analysis of deregulated miRNAs was conducted. We observed altered expression by the next-generation sequencing (NGS) of 11 miRNAs in serum, of which five (hsa-miR-148b-3p, hsa-miR-221-5p, hsa-miR-618, hsa-miR-941, and hsa-miR-769-5p) were validated by RT-qPCR, and three miRNAs in lung tissue (hsa-miR-144-3p, hsa-miR-144-5p, and hsa-miR-451a). The best multivariate logistic regression model to differentiate individuals with severe asthma, treated and untreated with OCS, was to combine the serum miRNAs hsa-miR-221-5p and hsa-miR-769-5p. Expression of hsa-miR-148b-3p and hsa-miR-221-5p correlated with FEV1/FVC (%) and these altered miRNAs act in key signaling pathways for asthma disease and the regulated expression of some genes (FOXO3, PTEN, and MAPK3) involved in these pathways. In conclusion, there are miRNA profiles differentially expressed in OCS-treated individuals with asthma and could be used as biomarkers of OCS treatment.
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Asma , MicroARNs , Humanos , Glucocorticoides/uso terapéutico , MicroARNs/metabolismo , Pulmón/metabolismo , Biomarcadores , Asma/tratamiento farmacológico , Asma/genéticaRESUMEN
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.
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Asma , MicroARNs , Remodelación de las Vías Aéreas (Respiratorias)/genética , Asma/metabolismo , Calcio/metabolismo , Proliferación Celular/genética , Humanos , MicroARNs/genética , MicroARNs/metabolismo , Contracción Muscular/genética , Miocitos del Músculo Liso/metabolismoRESUMEN
BACKGROUND: Asthma exacerbations are a serious public health concern due to high healthcare resource utilization, work/school productivity loss, impact on quality of life, and risk of mortality. The genetic basis of asthma exacerbations has been studied in several populations, but no prior study has performed a multi-ancestry meta-analysis of genome-wide association studies (meta-GWAS) for this trait. We aimed to identify common genetic loci associated with asthma exacerbations across diverse populations and to assess their functional role in regulating DNA methylation and gene expression. METHODS: A meta-GWAS of asthma exacerbations in 4989 Europeans, 2181 Hispanics/Latinos, 1250 Singaporean Chinese, and 972 African Americans analyzed 9.6 million genetic variants. Suggestively associated variants (p ≤ 5 × 10-5 ) were assessed for replication in 36,477 European and 1078 non-European asthma patients. Functional effects on DNA methylation were assessed in 595 Hispanic/Latino and African American asthma patients and in publicly available databases. The effect on gene expression was evaluated in silico. RESULTS: One hundred and twenty-six independent variants were suggestively associated with asthma exacerbations in the discovery phase. Two variants independently replicated: rs12091010 located at vascular cell adhesion molecule-1/exostosin like glycosyltransferase-2 (VCAM1/EXTL2) (discovery: odds ratio (ORT allele ) = 0.82, p = 9.05 × 10-6 and replication: ORT allele = 0.89, p = 5.35 × 10-3 ) and rs943126 from pantothenate kinase 1 (PANK1) (discovery: ORC allele = 0.85, p = 3.10 × 10-5 and replication: ORC allele = 0.89, p = 1.30 × 10-2 ). Both variants regulate gene expression of genes where they locate and DNA methylation levels of nearby genes in whole blood. CONCLUSIONS: This multi-ancestry study revealed novel suggestive regulatory loci for asthma exacerbations located in genomic regions participating in inflammation and host defense.
Asunto(s)
Asma , Estudio de Asociación del Genoma Completo , Asma/genética , Predisposición Genética a la Enfermedad , Hispánicos o Latinos/genética , Humanos , Polimorfismo de Nucleótido Simple , Calidad de VidaRESUMEN
BACKGROUND: Recurrent wheezing (RW) is frequently developed in infants that have suffered bronchiolitis (BCH) during first months of life, but the immune mechanism underlying is not clear. The goal was to analyze the innate immune response that characterizes BCH and RW. METHODS: Ninety-eight and seventy hospitalized infants with BCH or RW diagnosis, respectively, were included. Nasopharyngeal aspirate (NPA) was processed. Cellular pellet was employed to evaluate type 2 innate lymphoid cells (ILC2) by flow cytometry and mRNA expression assays by semi-quantitative real-time PCR (qRT-PCR). In supernatant, twenty-seven pro-inflammatory and immunomodulatory factors, as well as lipid mediators and nitrites, were evaluated by ELISA and Luminex. RESULTS: Bronchiolitis patients showed higher ILC2 percentage compared with RW (P < .05). Also, ST2+ /ILC2 percentage was higher in the BCH group than in the RW group (P < .01). TLR3, IL33, IFNG, IL10, and FLG mRNA levels were significantly increased in BCH vs RW (P < .05). In supernatant, no significant differences were reached, observing similar levels of parameters linked to vascular damage, monocyte activation, and fibroblast growth. Prostaglandin E2 and cysteinyl leukotrienes C4 were evaluated; a significant difference was only found in their ratio. CONCLUSION: Bronchiolitis is associated with elevated nasal percentage of ILC2. This cellular population could be the key element in the differential immune response between BCH and RW which share some mechanisms such us monocyte activation, vascular damage, and fibroblast repair. Lipid mediators could play a role in the evolution of the disease later in life through innate lymphoid cells.
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Bronquiolitis , Inmunidad Innata , Proteínas Filagrina , Humanos , Linfocitos , Ruidos RespiratoriosRESUMEN
Eosinophils are complex granulocytes with the capacity to react upon diverse stimuli due to their numerous and variable surface receptors, which allows them to respond in very different manners. Traditionally believed to be only part of parasitic and allergic/asthmatic immune responses, as scientific studies arise, the paradigm about these cells is continuously changing, adding layers of complexity to their roles in homeostasis and disease. Developing principally in the bone marrow by the action of IL-5 and granulocyte macrophage colony-stimulating factor GM-CSF, eosinophils migrate from the blood to very different organs, performing multiple functions in tissue homeostasis as in the gastrointestinal tract, thymus, uterus, mammary glands, liver, and skeletal muscle. In organs such as the lungs and gastrointestinal tract, eosinophils are able to act as immune regulatory cells and also to perform direct actions against parasites, and bacteria, where novel mechanisms of immune defense as extracellular DNA traps are key factors. Besides, eosinophils, are of importance in an effective response against viral pathogens by their nuclease enzymatic activity and have been lately described as involved in severe acute respiratory syndrome coronavirus SARS-CoV-2 immunity. The pleiotropic role of eosinophils is sustained because eosinophils can be also detrimental to human physiology, for example, in diseases like allergies, asthma, and eosinophilic esophagitis, where exosomes can be significant pathophysiologic units. These eosinophilic pathologies, require specific treatments by eosinophils control, such as new monoclonal antibodies like mepolizumab, reslizumab, and benralizumab. In this review, we describe the roles of eosinophils as effectors and regulatory cells and their involvement in pathological disorders and treatment.
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Eosinófilos/fisiología , Anticuerpos Monoclonales/uso terapéutico , Asma/tratamiento farmacológico , Asma/inmunología , Asma/patología , COVID-19/inmunología , COVID-19/patología , COVID-19/virología , Esofagitis Eosinofílica/tratamiento farmacológico , Esofagitis Eosinofílica/inmunología , Esofagitis Eosinofílica/patología , Eosinófilos/citología , Eosinófilos/inmunología , Exosomas/metabolismo , Trampas Extracelulares/metabolismo , Humanos , Células Plasmáticas/citología , Células Plasmáticas/metabolismo , SARS-CoV-2/aislamiento & purificaciónRESUMEN
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.
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Asma/etiología , Exosomas/fisiología , Inflamación/etiología , Animales , Asma/patología , Comunicación Celular/fisiología , Micropartículas Derivadas de Células/metabolismo , Exosomas/patología , Humanos , Inflamación/patología , Vesículas Secretoras/patología , Vesículas Secretoras/fisiologíaRESUMEN
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.
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Antiasmáticos/uso terapéutico , Asma/tratamiento farmacológico , Interleucina-5/antagonistas & inhibidores , MicroARNs/sangre , Adulto , Antiasmáticos/farmacología , Anticuerpos Monoclonales Humanizados/farmacología , Anticuerpos Monoclonales Humanizados/uso terapéutico , Asma/genética , Biomarcadores/sangre , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Masculino , Persona de Mediana Edad , Resultado del TratamientoRESUMEN
BACKGROUND: Bronchiolitis is the main cause of hospitalization of children younger than 1 year; however, the immune mechanism of bronchiolitis is not completely understood. The aim of this study was to analyze the recovery of immune response after a bronchiolitis episode. METHODS: Forty-nine infants hospitalized with bronchiolitis diagnosis were enrolled. Nasopharyngeal aspirates (NPAs) were processed. Twenty-seven pro-inflammatory biomarkers linked to innate immunity, inflammation, and epithelial damage, as well as nitrites and lipid mediators, were evaluated in the NPA supernatant by ELISA (enzyme-linked immunosorbent assay) and Luminex. Also, 11 genes were analyzed in NPA cells by quantitative PCR. RESULTS: A widespread statistically significant decline of multiple pro-inflammatory parameters and cytokines were detected in the recovery period after respiratory infection: interferon-α2 (IFNα2), IFNγ, interleukin-10 (IL-10), IL-1ß, IL-8, IFN-γ-inducible protein-10, vascular endothelial growth factor, monocyte chemoattractant protein-1, macrophage inflammatory protein-1α (MIP-1α), and MIP-1ß. Supporting these results, a decreased nuclear factor-κB gene expression was observed (P = 0.0116). A significant diminution of cysteinyl leukotriene C4 (LTC4) soluble levels (P = 0.0319) and cyclooxygenase-2 (COX-2) gene expression were observed in the recovery sample. In children classified by post-bronchiolitis wheezing, LTC4 remains elevated in the NPA supernatant. CONCLUSIONS: After bronchiolitis, cytokines and biomarkers linked to innate immune response in NPA decrease significantly in the recovery period accompanied by a drop in LTC4 levels; however, this reduction was lower in infants with post-bronchiolitis wheezing.
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Inmunidad Adaptativa , Bronquiolitis/inmunología , Citocinas/metabolismo , Inmunidad Innata , Leucotrieno C4/metabolismo , Nasofaringe/inmunología , Biomarcadores/metabolismo , Bronquiolitis/diagnóstico , Bronquiolitis/metabolismo , Bronquiolitis/terapia , Citocinas/genética , Regulación hacia Abajo , Femenino , Humanos , Lactante , Masculino , Estudios Prospectivos , Factores de TiempoRESUMEN
OBJECTIVES: Eosinophilic esophagitis (EoE) is a chronic esophageal disease characterized by eosinophilic inflammation. Proton-pump inhibitors (PPI) induce disease remission but no predictive factors of PPI-responsiveness have been identified yet. So, a biomarker must be found to differentiate between responders (PPI-R) and nonresponder patients (PPI-NR) to PPI. Aims were to identify any molecular biomarker that could predict PPI responsiveness and to study molecular remission after PPI therapy. METHODS: This prospective study enrolled 39 controls and 43 pediatric children with EoE from 2 hospitals, and they were treated with esomeprazole for 8 to 12 weeks. After therapy, patients were classified as either PPI-R or PPI-NR. Biopsies were collected and RNA, microRNAs, and proteins were isolated from them, measuring levels by qPCR and Western blot (WB). Also, miRNAs were evaluated in serum. RESULTS: We found several esophageal miRNAs with different expression values between PPI-R and PPI-NR children, which can be used to discriminate them (area under curveâ=â0.90). No useful serum miRNAs were, however, identified. Also, these miRNAs were dysregulated in responder patients before and after PPI therapy. Moreover, we corroborated in this child population, that PPI-R displayed a significant decrease in eotaxin-3, IL-5, IL-13, periostin, and major basic protein (Pâ<â0.05) and a significant increase in filaggrin levels after PPI treatment (Pâ<â0.01). CONCLUSIONS: Esophageal miRNA levels found are able to discriminate between both PPI-R and PPI-NR at baseline, and before and after treatment in PPI-R, so they could be used as biomarkers. Furthermore, we observed clinical and esophageal molecular restoration in PPI-R patients after PPI therapy.
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Esofagitis Eosinofílica , MicroARNs , Inhibidores de la Bomba de Protones , Biomarcadores/análisis , Niño , Esofagitis Eosinofílica/tratamiento farmacológico , Esofagitis Eosinofílica/genética , Proteínas Filagrina , Humanos , Masculino , MicroARNs/metabolismo , Estudios Prospectivos , Inhibidores de la Bomba de Protones/uso terapéuticoRESUMEN
BACKGROUND: Asthma is a syndrome characterized by airway inflammation and obstruction. Due to its heterogeneity, the difficulties in asthma diagnosis and treatment make the discovery of new biomarkers a focus of research. So, we determined the differential miRNA expression of eosinophils between healthy and asthmatic patients and to establish a differentially expressed miRNA profile detectable in sera for use as biomarker. METHODS: MicroRNAs from peripheral eosinophils from healthy and asthmatic subjects were isolated and analyzed by next-generation sequencing and confirmed by quantitative PCR in 29 asthmatics and 10 healthy individuals. The levels of serum miRNAs were performed by quantitative PCR in 138 asthmatics and 39 healthy subjects. Regression analysis and Random Forest models were performed. RESULTS: We found a set of miRNAs whose expression differs between eosinophils from asthmatics and healthy subjects. These miRNAs can classify asthmatics into two clusters that differed in the number of eosinophils and periostin concentration in serum. Some of these miRNAs were also confirmed in sera, as miR-185-5p which discriminates asthmatics from healthy subjects. Together with other two miRNAs, miR-185-5p allowed us to create a logistic regression model to discriminate better both conditions and a Random Forest model that can even sort the asthmatics into intermittent, mild persistent, moderate persistent, and severe persistent asthma. CONCLUSION: Our data show that miRNAs profile in eosinophils can be used as asthma diagnosis biomarker in serum and that this profile is able to rank asthma severity.
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Asma/diagnóstico , Asma/etiología , Biomarcadores , Eosinófilos/inmunología , Eosinófilos/metabolismo , MicroARNs/genética , Asma/sangre , Estudios de Casos y Controles , Biología Computacional/métodos , Femenino , Perfilación de la Expresión Génica , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Masculino , Curva ROC , Reacción en Cadena en Tiempo Real de la Polimerasa , Reproducibilidad de los Resultados , Índice de Severidad de la EnfermedadRESUMEN
BACKGROUND: Eosinophils secrete several granules that are involved in the propagation of inflammatory responses in patients with pathologies such as asthma. OBJECTIVE: We hypothesized that some of these granules are exosomes, which, when transferred to the recipient cells, could modulate asthma progression. METHODS: Eosinophils were purified from peripheral blood and cultured with or without IFN-γ or eotaxin. Multivesicular bodies (MVBs) in eosinophils were studied by using fluorescence microscopy, transmission electron microscopy (TEM), and flow cytometry. Exosome secretion was measured and exosome characterization was performed with TEM, Western blotting, and NanoSight analysis. RESULTS: Generation of MVBs in eosinophils was confirmed by using fluorescence microscopy and flow cytometry and corroborated by means of TEM. Having established that eosinophils contain MVBs, our aim was to demonstrate that eosinophils secrete exosomes. To do this, we purified exosomes from culture medium of eosinophils and characterized them. Using Western blot analysis, we demonstrated that eosinophils secreted exosomes and that the discharge of exosomes to extracellular media increases after IFN-γ stimulation. We measured exosome size and quantified exosome production from healthy and asthmatic subjects using nanotracking analysis. We found that exosome production was augmented in asthmatic patients. CONCLUSION: Our findings are the first to demonstrate that eosinophils contain functional MVBs and secrete exosomes and that their secretion is increased in asthmatic patients. Thus exosomes might play an important role in the progression of asthma and eventually be considered a biomarker.
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Asma/diagnóstico , Eosinófilos/metabolismo , Exosomas/metabolismo , Cuerpos Multivesiculares/metabolismo , Asma/inmunología , Asma/metabolismo , Asma/patología , Biomarcadores/análisis , Biomarcadores/metabolismo , Estudios de Casos y Controles , Fraccionamiento Celular , Separación Celular , Quimiocina CCL11/farmacología , Eosinófilos/efectos de los fármacos , Eosinófilos/inmunología , Eosinófilos/ultraestructura , Exosomas/inmunología , Exosomas/ultraestructura , Humanos , Interferón gamma/farmacología , Microscopía Electrónica de Transmisión , Cuerpos Multivesiculares/inmunología , Cuerpos Multivesiculares/ultraestructura , Tamaño de los Orgánulos , Cultivo Primario de CélulasRESUMEN
Eosinophils are one of the key inflammatory cells in asthma. Eosinophils can exert a wide variety of actions through expression and secretion of multiple molecules. Previously, we have demonstrated that eosinophils purified from peripheral blood from asthma patients express high levels of suppressor of cytokine signaling 3 (SOCS3). In this article, SOCS3 gene silencing in eosinophils from asthmatics has been carried out to achieve a better understanding of the suppressor function in eosinophils. SOCS3 siRNA treatment drastically reduced SOCS3 expression in eosinophils, leading to an inhibition of the regulatory transcription factors GATA-3 and FoxP3, also interleukin (IL)-10; in turn, an increased STAT3 phosphorilation was observed. Moreover, SOCS3 abrogation in eosinophils produced impaired migration, adhesion and degranulation. Therefore, SOCS3 might be regarded as an important regulator implicated in eosinophil mobilization from the bone marrow to the lungs during the asthmatic process.