Network analysis of quantitative proteomics on asthmatic bronchi: effects of inhaled glucocorticoid treatment.

BACKGROUND: Proteomic studies of respiratory disorders have the potential to identify protein biomarkers for diagnosis and disease monitoring. Utilisation of sensitive quantitative proteomic methods creates opportunities to determine individual patient proteomes. The aim of the current study was to determine if quantitative proteomics of bronchial biopsies from asthmatics can distinguish relevant biological functions and whether inhaled glucocorticoid treatment affects these functions. METHODS: Endobronchial biopsies were taken from untreated asthmatic patients (n = 12) and healthy controls (n = 3). Asthmatic patients were randomised to double blind treatment with either placebo or budesonide (800 µg daily for 3 months) and new biopsies were obtained. Proteins extracted from the biopsies were digested and analysed using isobaric tags for relative and absolute quantitation combined with a nanoLC-LTQ Orbitrap mass spectrometer. Spectra obtained were used to identify and quantify proteins. Pathways analysis was performed using Ingenuity Pathway Analysis to identify significant biological pathways in asthma and determine how the expression of these pathways was changed by treatment. RESULTS: More than 1800 proteins were identified and quantified in the bronchial biopsies of subjects. The pathway analysis revealed acute phase response signalling, cell-to-cell signalling and tissue development associations with proteins expressed in asthmatics compared to controls. The functions and pathways associated with placebo and budesonide treatment showed distinct differences, including the decreased association with acute phase proteins as a result of budesonide treatment compared to placebo. CONCLUSIONS: Proteomic analysis of bronchial biopsy material can be used to identify and quantify proteins using highly sensitive technologies, without the need for pooling of samples from several patients. Distinct pathophysiological features of asthma can be identified using this approach and the expression of these features is changed by inhaled glucocorticoid treatment. Quantitative proteomics may be applied to identify mechanisms of disease that may assist in the accurate and timely diagnosis of asthma. TRIAL REGISTRATION: ClinicalTrials.gov registration NCT01378039.

Smoking affects eotaxin levels in asthma patients.

BACKGROUND: Chronic airway inflammation is most important pathological finding in asthma. Cigarette smoking may modify type of inflammation as well as may influence disease severity and response to the treatment. OBJECTIVE: Thus the aim of this study was to investigate whether cigarette smoking may have an influence on the levels of eotaxin-1, eotaxin-2, eotaxin-3 and IL-5 in patients with stable mild/moderate asthma. METHODS: 45 steroid naive asthmatics (mean age: 55.2 +/- 2.2 yrs) and 23 "healthy" smokers and non-smokers control subjects (mean age: 54.4 +/- 9.7 yrs) were investigated. Asthmatics were divided into two subgroups according to their smoking histories: asthmatic smokers (n = 19) who currently smoke and have a history of > 10 pack-years and asthmatic never-smokers (n = 26). BAL and induced sputum were performed. Cytospins of induced sputum and BAL were stained with May-Grunwald-Giemsa for differential cell counts. Eotaxin-1, eotaxin-2, eotaxin-3 and IL-5 concentrations in serum, sputum and BAL supernatant was measured using a commercial ELISA kit. RESULTS: In sputum supernatant from asthma smokers was significantly higher concentration of eotaxin-1 than in non-smokers asthmatics (203.4 +/- 10.0 vs. 140.2 +/- 9.5 respectively, p < 0.05). In non-smokers asthma patients levels of BAL eotaxin-1 strongly related to percent and absolute numbers of BAL eosinophils and neutrophils (Rs = 0.737 and Rs = 0.514 respectively, p < 0.05). The number and percent of sputum neutrophils and eosinophils, obtained from smokers asthmatics, significantly correlated with eotaxin-2 concentration in sputum supernatant (Rs = 0.58 and Rs = 0.75 respectively, p < 0.05). IL-5 levels in the serum and sputum from asthmatic never-smokers were significantly higher than they were from asthmatic smokers and "healthy" smokers. Asthmatic never-smokers showed a significantly higher amount of IL-5 in serum and sputum than the asthmatic smokers showed. CONCLUSIONS: This study showed the elevated levels of sputum eotaxin-1 as well as serum, sputum and BAL eotaxin-2 in asthmatic smokers without a significant increase of eosinophils compared to asthmatic never-smokers. The eotaxin concentrations were related not only with number of eosinophils but also with the number of neutrophils in all the studied tissue compartments. The data herein permits a suggestion that smoking may influence change in asthmatic airway inflammation by stimulating the production of eotaxins.

[Peculiarities of induced sputum inflammatory cell counts in allergic versus non-allergic asthma]. / Indukuotu skrepliu lastelines sudeties savybes sergant alergine ir nealergine astma.

OBJECTIVE: According to the Global Initiative for Asthma (GINA, updated 2004), although non-allergic asthma has a different clinical profile than allergic asthma, it is not a distinct immunopathological entity. We aimed to investigate the peculiarities of inflammatory cell counts in induced sputum from patients with allergic and non-allergic asthma. METHODS: We investigated 41 steroid naive patients (mean age 51.8+/-1.5 years) with mild persistent asthma. Depending on the results of allergic anamnesis and skin prick test, patients were divided into the two groups: 21-with allergic asthma, 20-with non-allergic asthma. Spirometry, bronchial provocation test with methacholine (PD20) and sputum induction with hypertonic saline were performed. Cytospins of induced sputum were stained with May-Grunwald-Giemsa for differential cell counts. RESULTS. In samples from non-allergic asthmatics we detected a significantly higher number of total cells (1.97+/-0.33 vs 1.23+/-0.11x10(6)/ml, p<0.01), neutrophils (1.34+/-0.18 vs 0.41+/-0.06x10(6)/ml, p<0.01) and lymphocytes (0.36+/-0.04 vs 0.15+/-0.03x10(6)/ml, p<0.01), than in samples from allergic asthmatics. Number of eosinophils and macrophages did not significantly differ between two groups. In non-allergic asthmatics, number of lymphocytes correlated with PD20 (Rs=0.41, p<0.05); in allergic asthmatics, number of eosinophils in induced sputum correlated with PD20 (Rs=0.71, p<0.05). CONCLUSIONS: Our data suggest that lymphocytes might play a more important role in pathogenesis of non-allergic asthma, while eosinophils -- in allergic asthma.

[Wiskott-Aldrich syndrome: the possibilities of diagnosis and treatment]. / Wiskott-Aldrich sindromas: diagnozavimo bei gydymo galimybes.

Wiskott-Aldrich syndrome is congenital X-linked immunodeficiency characterized by frequent infections, thrombocytopenia with small platelets, eczema and increased risk of autoimmune disorders and malignancies. This article is review of Wiskott-Aldrich syndrome actual diagnostics and treatment problems. Diagnostics problems exist due to clinical heterogenity of this syndrome, which is caused by mutations of the responsible gene. Recent 15-year studies showed, that bone marrow transplantations or use of cord blood as a source of stem cells prolonged median survival from 6.5 to 11 years. However, widespread use of splenectomy, intravenous immune globulin and prophylactic antibiotics did not change survival or appearance of infections, bleeding and autoimmune diseases. An attractive option for Wiskott-Aldrich syndrome is gene therapy, which leads to complete cure.