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Lipids and their mediators have important regulatory functions in many cellular processes, including the innate antiviral response. The aim of this study was to compare the lipid membrane composition of in vitro differentiated primary bronchial epithelial cells (PBECs) with ex vivo bronchial brushings and to establish whether any changes in the lipid membrane composition affect antiviral defense of cells from donors without and with severe asthma. Using mass spectrometry, we showed that the lipid membrane of in vitro differentiated PBECs was deprived of polyunsaturated fatty acids (PUFAs) compared to ex vivo bronchial brushings. Supplementation of the culture medium with arachidonic acid (AA) increased the PUFA-content to more closely match the ex vivo membrane profile. Rhinovirus (RV16) infection of AA-supplemented cultures from healthy donors resulted in significantly reduced viral replication while release of inflammatory mediators and prostaglandin E2 (PGE2) was significantly increased. Indomethacin, an inhibitor of prostaglandin-endoperoxide synthases, suppressed RV16-induced PGE2 release and significantly reduced CXCL-8/IL-8 release from AA-supplemented cultures indicating a link between PGE2 and CXCL8/IL-8 release. In contrast, in AA-supplemented cultures from severe asthmatic donors, viral replication was enhanced whereas PTGS2 expression and PGE2 release were unchanged and CXCL8/IL-8 was significantly reduced in response to RV16 infection. While the PTGS2/COX-2 pathway is initially pro-inflammatory, its downstream products can promote symptom resolution. Thus, reduced PGE2 release during an RV-induced severe asthma exacerbation may lead to prolonged symptoms and slower recovery. Our data highlight the importance of reflecting the in vivo lipid profile in in vitro cell cultures for mechanistic studies.
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PURPOSE OF REVIEW: Chronic obstructive pulmonary disease (COPD) is a condition principally defined by airflow limitation that is not fully reversible. The main trigger, inhalation of noxious gases or particles (usually smoke) leads to complex pathology, including inflammation of the large and small airways, and destruction of the lung parenchyma. Overlap in pathophysiology with other chronic airways diseases leads to challenges in differential diagnosis, and furthermore, periodic exacerbations of disease symptoms also increase the complexity of the disease diagnosis and prediction of outcome. There is recognized need for biomarkers to aid in the determination of disease diagnosis, progression and response to intervention. This review describes the current status of biomarker identification in COPD. RECENT FINDINGS: Biomarkers of disease can take many forms other than the classical protein in serum, and their utility is dependent upon the clinical question to be addressed. No single protein marker has been adopted for routine clinical use to date. This review addresses the key issues around biomarker identification and utility in both stable and exacerbating COPD. SUMMARY: Biomarker identification in COPD is still a developing field, with increasing interest in patient phenotyping probably reflecting the challenges of biomarker development in a complex disease.
Asunto(s)
Biomarcadores/análisis , Enfermedad Pulmonar Obstructiva Crónica/diagnóstico , Progresión de la Enfermedad , Humanos , Enfermedad Pulmonar Obstructiva Crónica/fisiopatologíaRESUMEN
RATIONALE: Much effort is being made to discover noninvasive biomarkers of chronic airway disease that might enable better management, predict prognosis, and provide new therapeutic targets. OBJECTIVES: To undertake a comprehensive, unbiased proteomic analysis of induced sputum and identify novel noninvasive biomarkers for chronic obstructive pulmonary disease (COPD). METHODS: Induced sputum was obtained from patients with COPD with a spectrum of disease severity and from control subjects. Two-dimensional gel electrophoresis and mass spectrometric identification of differentially expressed proteins were first applied to induced sputum from patients with GOLD stage 2 COPD and healthy smoker control subjects. Initial results thus obtained were validated by a combination of immunoassays (Western blotting and ELISA) applied to a large subject cohort. The biomarkers were localized to bronchial mucosa by immunohistochemistry. MEASUREMENTS AND MAIN RESULTS: Of 1,325 individual protein spots identified, 37 were quantitatively and 3 qualitatively different between the two groups (P < 0.05%). Forty protein spots were subjected to tandem mass spectrometry, which identified 15 separate protein species. Seven of these were further quantified in induced sputum from 97 individuals. Using this sequential approach, two of these potential biomarkers (apolipoprotein A1 and lipocalin-1) were found to be significantly reduced in patients with COPD when compared with healthy smokers. Their levels correlated with FEV(1)/FVC, indicating their relationship to disease severity. CONCLUSIONS: A potential role for apolipoprotein A1 and lipocalin-1 in innate defense has been postulated previously; our discovery of their reduction in COPD indicates a deficient innate defense system in airway disease that could explain increased susceptibility to infectious exacerbations.
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Apolipoproteína A-I/metabolismo , Lipocalinas/metabolismo , Enfermedad Pulmonar Obstructiva Crónica/metabolismo , Adulto , Anciano , Apolipoproteína A-I/análisis , Asma/metabolismo , Biomarcadores/análisis , Biomarcadores/metabolismo , Western Blotting , Bronquitis Crónica/metabolismo , Estudios de Casos y Controles , Ensayo de Inmunoadsorción Enzimática , Femenino , Flujo Espiratorio Forzado , Humanos , Lipocalinas/análisis , Masculino , Persona de Mediana Edad , Mucosa Respiratoria/química , Índice de Severidad de la Enfermedad , Fumar/metabolismo , Esputo/química , Esputo/metabolismoRESUMEN
Biomarkers that describe the severity and progression of COPD and the responses of patients to treatment are a desirable addition to clinical measures of disease. In this review, we describe the current state of knowledge of biomarkers used for the diagnosis, staging and therapeutic response of COPD patients. The nature of these biomarkers is considered in relation to their intended use, and the desirable qualities of such entities are examined. Examples of biased and unbiased discovery platforms for COPD biomarker discovery are given, and the major findings of these studies are discussed. Cutting edge technology used for biomarker discovery, quantitation in biofluids and imaging biomarkers in whole body systems is reviewed.
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Biomarcadores/metabolismo , Proteómica/métodos , Enfermedad Pulmonar Obstructiva Crónica/metabolismo , Diagnóstico por Imagen , Ensayo de Inmunoadsorción Enzimática , Genómica/métodos , Humanos , Enfermedad Pulmonar Obstructiva Crónica/diagnóstico , Enfermedad Pulmonar Obstructiva Crónica/patología , Enfermedad Pulmonar Obstructiva Crónica/terapia , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
NIH mice were vaccinated subcutaneously or intranasally with chimaeric cow pea mosaic virus (CPMV) constructs expressing a 17-mer peptide sequence from canine parvovirus (CPV) as monomers or dimers on the small or large protein surface subunits. Responses to the chimaeric virus particles (CVPs) were compared with those of mice immunized with the native virus or with parvovirus peptide conjugated to keyhole limpet haemocyanin (KLH). The characteristics of the immune response to vaccination were examined by measuring serum and mucosal antibody responses in ELISA, in vitro antigen-induced spleen cell proliferation and cytokine responses. Mice made strong antibody responses to the native plant virus and peptide-specific responses to two of the four CVP constructs tested which were approximately 10-fold lower than responses to native plant virus. The immune response generated by the CVP constructs showed a marked TH1 bias, as determined by a predominantly IgG(2a) isotype peptide-specific antibody response and the release of IFN-gamma but not IL-4 or IL-5 from lymphocytes exposed to antigen in vitro. In comparison, parvovirus peptide conjugated to KLH generated an IgG(1)-biased (TH2) response. These data indicate that the presentation of peptides on viral particles could be used to bias the immune response in favor of a TH1 response.Anti-viral and anti-peptide IgA were detected in intestinal and bronchial lavage fluid of immunized mice, demonstrating that a mucosal immune response to CPV can be generated by systemic and mucosal immunization with CVP vaccines. Serum antibody from both subcutaneously-vaccinated and intranasally-vaccinated mice showed neutralizing activity against CPV in vitro.