Respiratory infections are a real threat for humans, and therefore the pig model is of interest for studies. As one of a case for studies, Actinobacillus pleuropneumoniae (APP) caused infections and still worries many pig breeders around the world. To better understand the influence of pathogenic effect of APP on a respiratory system-lungs and tracheobronchial lymph nodes (TBLN), we aimed to employ matrix-assisted laser desorption/ionization time-of-flight mass spectrometry imaging (MALDI-TOF MSI). In this study, six pigs were intranasally infected by APP and two were used as non-infected control, and 48 cryosections have been obtained. MALDI-TOF MSI and immunohistochemistry (IHC) were used to study spatial distribution of infectious markers, especially interleukins, in cryosections of porcine tissues of lungs (necrotic area, marginal zone) and tracheobronchial lymph nodes (TBLN) from pigs infected by APP. CD163, interleukin 1β (IL‑1β) and a protegrin-4 precursor were successfully detected based on their tryptic fragments. CD163 and IL‑1β were confirmed also by IHC. The protegrin-4 precursor was identified by MALDI-TOF/TOF directly on the tissue cryosections. CD163, IL‑1β and protegrin‑4 precursor were all significantly (p < 0.001) more expressed in necrotic areas of lungs infected by APP than in marginal zone, TBLN and in control lungs.
- MeSH
- Actinobacillus pleuropneumoniae patogenita MeSH
- antigeny diferenciační myelomonocytární metabolismus MeSH
- biologické markery metabolismus MeSH
- bronchy metabolismus MeSH
- CD antigeny metabolismus MeSH
- infekce bakteriemi rodu Actinobacillus metabolismus mikrobiologie MeSH
- infekce dýchací soustavy metabolismus mikrobiologie MeSH
- interleukin-1beta metabolismus MeSH
- kationické antimikrobiální peptidy metabolismus MeSH
- lymfatické uzliny metabolismus MeSH
- plíce metabolismus MeSH
- prasata MeSH
- receptory buněčného povrchu metabolismus MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice metody MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Occupational exposure to diesel exhaust may cause lung cancer in humans. Mechanisms include DNA-damage and inflammatory responses. Here, the potential of NIST SRM2975 diesel exhaust particles (DEP) to transform human bronchial epithelial cells (HBEC3) in vitro was investigated. Long-term exposure of HBEC3 to DEP led to increased colony growth in soft agar. Several DEP-transformed cell lines were established and based on the expression of epithelial-to-mesenchymal-transition (EMT) marker genes, one of them (T2-HBEC3) was further characterized. T2-HBEC3 showed a mesenchymal/fibroblast-like morphology, reduced expression of CDH1, and induction of CDH2 and VIM. T2-HBEC3 had reduced migration potential compared with HBEC3 and little invasion capacity. Gene expression profiling showed baseline differences between HBEC3 and T2-HBEC3 linked to lung carcinogenesis. Next, to assess differences in sensitivity to DEP between parental HBEC3 and T2-HBEC3, gene expression profiling was carried out after DEP short-term exposure. Results revealed changes in genes involved in metabolism of xenobiotics and lipids, as well as inflammation. HBEC3 displayed a higher steady state of IL1B gene expression and release of IL-1β compared with T2-HBEC3. HBEC3 and T2-HBEC3 showed similar susceptibility towards DEP-induced genotoxic effects. Liquid-chromatography-tandem-mass-spectrometry was used to measure secretion of eicosanoids. Generally, major prostaglandin species were released in higher concentrations from T2-HBEC3 than from HBEC3 and several analytes were altered after DEP-exposure. In conclusion, long-term exposure to DEP-transformed human bronchial epithelial cells in vitro. Differences between HBEC3 and T2-HBEC3 regarding baseline levels and DEP-induced changes of particularly CYP1A1, IL-1β, PGE2, and PGF2α may have implications for acute inflammation and carcinogenesis.
- MeSH
- bronchy účinky léků metabolismus ultrastruktura MeSH
- buněčné kultury MeSH
- epitelo-mezenchymální tranzice účinky léků genetika MeSH
- epitelové buňky účinky léků metabolismus ultrastruktura MeSH
- interleukin-1beta genetika MeSH
- látky znečišťující vzduch toxicita MeSH
- lidé MeSH
- pevné částice toxicita MeSH
- poškození DNA MeSH
- stanovení celkové genové exprese MeSH
- transformované buněčné linie MeSH
- transkriptom účinky léků MeSH
- výfukové emise vozidel toxicita MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The airway epithelium restricts the penetration of inhaled pathogens into the underlying tissue and plays a crucial role in the innate immune defense against respiratory infections. The whooping cough agent, Bordetella pertussis, adheres to ciliated cells of the human airway epithelium and subverts its defense functions through the action of secreted toxins and other virulence factors. We examined the impact of B. pertussis infection and of adenylate cyclase toxin-hemolysin (CyaA) action on the functional integrity of human bronchial epithelial cells cultured at the air-liquid interface (ALI). B. pertussis adhesion to the apical surface of polarized pseudostratified VA10 cell layers provoked a disruption of tight junctions and caused a drop in transepithelial electrical resistance (TEER). The reduction of TEER depended on the capacity of the secreted CyaA toxin to elicit cAMP signaling in epithelial cells through its adenylyl cyclase enzyme activity. Both purified CyaA and cAMP-signaling drugs triggered a decrease in the TEER of VA10 cell layers. Toxin-produced cAMP signaling caused actin cytoskeleton rearrangement and induced mucin 5AC production and interleukin-6 (IL-6) secretion, while it inhibited the IL-17A-induced secretion of the IL-8 chemokine and of the antimicrobial peptide beta-defensin 2. These results indicate that CyaA toxin activity compromises the barrier and innate immune functions of Bordetella-infected airway epithelia.
- MeSH
- adenylátcyklasový toxin genetika metabolismus toxicita MeSH
- AMP cyklický metabolismus MeSH
- Bordetella pertussis genetika metabolismus MeSH
- bronchy cytologie metabolismus mikrobiologie MeSH
- cytoskelet metabolismus MeSH
- epitelové buňky metabolismus mikrobiologie MeSH
- interleukin-6 metabolismus MeSH
- lidé MeSH
- mucin 5AC metabolismus MeSH
- pertuse genetika metabolismus mikrobiologie MeSH
- signální transdukce účinky léků MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
This study used toxicogenomics to identify the complex biological response of human lung BEAS-2B cells treated with organic components of particulate matter in the exhaust of a diesel engine. First, we characterized particles from standard diesel (B0), biodiesel (methylesters of rapeseed oil) in its neat form (B100) and 30% by volume blend with diesel fuel (B30), and neat hydrotreated vegetable oil (NEXBTL100). The concentration of polycyclic aromatic hydrocarbons (PAHs) and their derivatives in organic extracts was the lowest for NEXBTL100 and higher for biodiesel. We further analyzed global gene expression changes in BEAS-2B cells following 4 h and 24 h treatment with extracts. The concentrations of 50 µg extract/mL induced a similar molecular response. The common processes induced after 4 h treatment included antioxidant defense, metabolism of xenobiotics and lipids, suppression of pro-apoptotic stimuli, or induction of plasminogen activating cascade; 24 h treatment affected fewer processes, particularly those involved in detoxification of xenobiotics, including PAHs. The majority of distinctively deregulated genes detected after both 4 h and 24 h treatment were induced by NEXBTL100; the deregulated genes included, e.g., those involved in antioxidant defense and cell cycle regulation and proliferation. B100 extract, with the highest PAH concentrations, additionally affected several cell cycle regulatory genes and p38 signaling.
- MeSH
- anotace sekvence MeSH
- benzin analýza toxicita MeSH
- biopaliva analýza toxicita MeSH
- bronchy cytologie účinky léků metabolismus MeSH
- epitelové buňky cytologie účinky léků metabolismus MeSH
- látky znečišťující vzduch analýza toxicita MeSH
- lidé MeSH
- oleje rostlin chemie MeSH
- pevné částice analýza toxicita MeSH
- polycyklické aromatické uhlovodíky analýza toxicita MeSH
- regulace genové exprese u rostlin * MeSH
- rostlinné proteiny genetika metabolismus MeSH
- signální transdukce MeSH
- stanovení celkové genové exprese MeSH
- transformované buněčné linie MeSH
- výfukové emise vozidel analýza MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- srovnávací studie MeSH
Researchers have been studying aerosol transport in human lungs for some decades. The overall lung deposition can be predicted with sufficient precision nowadays. However, the prediction of local deposition remains an unsolved problem. Numerical modeling of aerosol transport can provide detailed data with such precision and spatial resolution which were unavailable in the past. Yet, the necessary validation of numerical results represents a difficult task, as the experimental data in a sufficient spatial resolution are hardly available. This article introduces a method based on positron emission tomography, which allows acquisition of detailed experimental data on local aerosol deposition in a realistic model of human lungs. The method utilizes the Condensation Monodisperse Aerosol Generator modified for a safe production of radioactive aerosol particles and a special measuring rig. The scanning of the model is performed on a positron emission tomography-computed tomography scanner. The evaluation of aerosol deposition is based on a volume radioactivity analysis in a specialized, yet publicly available software. The reliability of the method was tested and its first results are discussed in the article. The measurements performed using the presented method can serve for validation of numerical simulations, since the presented lung model digital geometry is available.
- MeSH
- aerosoly aplikace a dávkování farmakokinetika MeSH
- biologické modely * MeSH
- bronchy metabolismus MeSH
- lidé MeSH
- pozitronová emisní tomografie metody MeSH
- systémy cílené aplikace léků MeSH
- trachea metabolismus MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Chronic obstructive pulmonary disease (COPD) and bronchial asthma (BA) are 2 severe respiratory disorders with different predominated immunopathologies. There are several "novel molecules" from different families that are proposed as part of the etiopathogenesis of COPD and BA. Proteinase-activated receptor 2 (PAR-2), thymic stromal lymphoprotein (TSLP), interleukin-4 and its receptor (CD124), Yin-Yang 1 (YY1), and transforming growth factor beta (TGF-β) have been previously shown to be involved in the pathophysiology of both these diseases. We investigated PAR-2, TSLP, CD124 (interleukin-4R), TGF-β, and YY1 immunohistochemical expression in endobronchial and transbronchial biopsies from 22 BA patients and 20 COPD patients. Immunostaining for the above-mentioned antigens was quantified using a modified semiquantitative scoring system and statistically evaluated. The values of TGF-β in the epithelial cells (P=0.0007) and TGF-β in the submucosa (P=0.0075) were higher in the BA samples, whereas values of CD124 (P=0.0015) and TSLP (P=0.0106) were higher in the COPD samples. No statistically significant differences between the groups were recorded for PAR-2 and YY1. Airway inflammatory reaction diversity in BA and COPD seems to be disease specific; however, there are also shared mechanisms involved in the pathophysiology of both diseases.
- MeSH
- biopsie MeSH
- bronchiální astma * metabolismus patologie MeSH
- bronchy metabolismus patologie MeSH
- chronická obstrukční plicní nemoc * metabolismus patologie MeSH
- cytokiny biosyntéza MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- receptor interleukinu-4 - alfa-podjednotka biosyntéza MeSH
- receptor PAR-2 biosyntéza MeSH
- regulace genové exprese u nádorů * MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- transformující růstový faktor beta biosyntéza MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- klinické zkoušky MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
OBJECTIVE: Alveolar concentration (C(A)NO) and bronchial flux (J(aw)NO) of nitric oxide (NO) characterize the contributions of peripheral and proximal airways to exhaled NO. Both parameters can be estimated using a two-compartment model if the fraction of NO in orally exhaled air (FE(NO)) is measured at multiple constant expiratory flow rates (V). The aim of this study was to evaluate how departures from linearity influence the estimates of C(A)NO and J(aw)NO obtained with the help of linear regression analysis of the relationships between FE(NO) and 1/V (method P), and between the NO output (V(NO) = FE(NO) × V) and V (method T). Furthermore, differences between patients with atopic asthma (AA) and allergic rhinitis (AR) and between methods P and T were assessed. DESIGN: Measurements of FE(NO) were performed with a chemiluminiscence analyzer at five levels of V ranging from 50 to 250 ml/sec in school children and adolescents with mild to moderate-severe AA treated by inhaled corticosteroids (N = 42) and AR (N = 20). RESULTS: Violation of the linearity condition at V ≤ 100 ml/sec caused shifts between methods with regard to the partition of exhaled NO into alveolar (C(A)NO: P > T) and bronchial (J(aw)NO: T > P) components. Both methods gave similar results in the linear range of 150-250 ml/sec: The mean ratios P/T and limits of agreement calculated in AA and AR patients were 1.03 (0.49-1.56) and 1.07 (0.55-1.59) for C(A)NO and 1.03 (0.73-1.33) and 0.99 (0.90-1.10) for J(aw)NO, respectively. No significant differences between AA and AR were found in C(A)NO and J(aw)NO calculated in the linear range by the T method {medians (inter-quartile ranges): 1.7 ppb (0.9-3.9) vs. 2.3 ppb (0.8-3.7), P = 0.91; 1,800 pl/sec (950-3,560) vs. 1,180 pl/sec (639-1,950), P = 0.061}. However, the flow-dependency of the estimates was markedly higher in AA than in AR patients: C(A) NO was decreased 2.8-fold vs. 1.5-fold and J(aw) NO was increased 1.5-fold vs. 1.2-fold in the linear range as compared to the range of 50-250 ml/sec. In both groups, the median standard errors (SE) of the J(aw) NO estimates were similar for the metods P and T and small (<15%) regardless of the range for expiratory flows. The precision of C(A) NO estimates was less in all ranges. For both methods, the SE of the estimates obtained in the range of 150-250 ml/sec exceeded 50% in asthmatics and 30% in AR patients, respectively. The results show that FE(NO) has to be measured at several expiratory flows ≥100 ml/sec for the accurate estimation of C(A) NO and J(aw) NO using linear methods P and T in children and adolescents with AA and AR. A stepwise procedure for detecting nonlinearity and evaluating the quality of FE(NO) measurements is suggested.
- MeSH
- biologické modely * MeSH
- bronchiální astma metabolismus patofyziologie MeSH
- bronchy chemie metabolismus patofyziologie MeSH
- celoroční alergická rýma metabolismus patofyziologie MeSH
- dechové testy metody MeSH
- dítě MeSH
- lidé MeSH
- lineární modely MeSH
- mladiství MeSH
- oxid dusnatý analýza metabolismus MeSH
- plicní alveoly chemie metabolismus patofyziologie MeSH
- respirační funkční testy MeSH
- stupeň závažnosti nemoci MeSH
- Check Tag
- dítě MeSH
- lidé MeSH
- mladiství MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- hodnotící studie MeSH
- práce podpořená grantem MeSH
BACKGROUND: Lung carcinogenesis is a multistep process of accumulation of genetic changes, including loss of heterozygosity (LOH), and precedes phenotypic transformation of the bronchial mucosa. The activity of telomerase, correlating with the hTERT mRNA expression, is detectable in a majority of neoplasms. In this study, the frequency of LOH and hTERT expression in bronchial mucosa of heavy smokers in bronchoscopic biopsies was analyzed. METHODS: LOH was examined in 122 bronchial specimens from 81 smokers (67 normal mucosa/bronchitis, 12 squamous metaplasia, 28 dysplasia, 15 bronchogenic carcinoma specimens) by polymerase chain reaction (PCR) and capillary electrophoresis by using 7 fluorescence-labeled markers matching 5 chromosomal regions. hTERT expression was analyzed in 87 specimens (45 normal mucosa/bronchitis, 12 squamous metaplasia, 18 dysplasia, 12 bronchogenic carcinoma specimens) by real-time quantitative reverse-transcription PCR. RESULTS: LOH was detected in at least 1 chromosomal region in 51 of 122 (41.8%) specimens; the incidence in normal bronchial mucosa and preneoplastic lesions was similar (20%-40%); a substantial rise (87%) occurred in carcinomas. The median normalized hTERT(N) values were 6.67 in normal epithelium/chronic bronchitis, 18.38 in squamous metaplasia, 13.31 in epithelial dysplasia, and 75.46 in carcinomas. These results were significantly different (P=.0036). With an increasing number of LOH, the median value of hTERT(N) expression rose, but hTERT was expressed also in tissue samples without any LOH detection. CONCLUSIONS: Results indicated that hTERT expression, together with LOH, represent early events in lung carcinogenesis, as both were detected in precancerous lesions and in normal epithelium of heavy smokers.
- MeSH
- bronchogenní karcinom enzymologie genetika MeSH
- bronchy metabolismus patologie MeSH
- epitelové buňky metabolismus patologie MeSH
- financování organizované MeSH
- kouření genetika MeSH
- lidé MeSH
- messenger RNA metabolismus MeSH
- metaplazie MeSH
- nádory plic enzymologie genetika MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- prekancerózy metabolismus patologie MeSH
- sliznice metabolismus patologie MeSH
- telomerasa genetika metabolismus MeSH
- ztráta heterozygozity MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
