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Patients with obstructive lung diseases commonly undergo bronchodilator reversibility testing during examination of their pulmonary function by spirometry. A positive response is defined by an increase in forced expiratory volume in 1 s (FEV1). FEV1 is a rather nonspecific criterion not allowing the regional effects of bronchodilator to be assessed. We employed the imaging technique of electrical impedance tomography (EIT) to visualize the spatial and temporal ventilation distribution in 35 patients with chronic obstructive pulmonary disease at baseline and 5, 10, and 20 min after bronchodilator inhalation. EIT scanning was performed during tidal breathing and forced full expiration maneuver in parallel with spirometry. Ventilation distribution was determined by EIT by calculating the image pixel values of FEV1, forced vital capacity (FVC), tidal volume, peak flow, and mean forced expiratory flow between 25 and 75% of FVC. The global inhomogeneity indexes of each measure and histograms of pixel FEV1/FVC values were then determined to assess the bronchodilator effect on spatial ventilation distribution. Temporal ventilation distribution was analyzed from pixel values of times needed to exhale 75 and 90% of pixel FVC. Based on spirometric FEV1, significant bronchodilator response was found in 17 patients. These patients exhibited higher postbronchodilator values of all regional EIT-derived lung function measures in contrast to nonresponders. Ventilation distribution was inhomogeneous in both groups. Significant improvements were noted for spatial distribution of pixel FEV1 and tidal volume and temporal distribution in responders. By providing regional data, EIT might increase the diagnostic and prognostic information derived from reversibility testing. [ABSTRACT FROM AUTHOR]

Background: Inhaled corticosteroids (ICS) are widely used in the treatment of obstructive lung diseases. Recent data suggest a higher pneumonia risk in chronic obstructive pulmonary disease (COPD) patients treated with ICS. Objective: Since non-typeable Haemophilus influenzae (NTHi) is the most common pathogen associated with acute exacerbations of COPD, we investigated the effects of budesonide (BUD) on NTHi-induced inflammation and invasive infection. Methods: The alveolar epithelial cell line A549 and specimens of human lung tissue (HLT) were used in our experiments. Intracellular infection was determined by a lysis/culture assay of infected cells. Activated p38 mitogen-associated protein kinase (MAPK) was assessed using Western blotting and immunohistochemistry, expression of toll-like receptor 2 (TLR2) was determined by PCR, and CXCL-8 levels were measured using ELISA. Immunohistochemistry was used for detection of CXCL-8, platelet-activating factor receptor (PAF-R) and NTHi. Results: BUD significantly reduced CXCL-8 secretion in A549 cells and lung tissue infected with NTHi. Furthermore, BUD decreased the expression of PAF-R in HLT and A549 cells. In A549 cells and HLT, BUD inhibited intracellular infection and - synergistically with NTHi - increased the expression of TLR2 (in A549 cells). TLR2 stimulation did not influence the intracellular infection of A549 cells, but p38 MAPK inhibition resulted in a significant reduction of infection. Conclusion: The present study adds new insights into the effects of glucocorticoids on pulmonary host defence after NTHi infection. Although the inflammatory response to infection is suppressed by BUD, interestingly, the intracellular infection is also inhibited. This effect seems to depend on the inhibition of p38 MAPK - a key enzyme in many pro-inflammatory pathways - as well as of PAF-R expression. © 2015 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background: Upper airway inflammation has been previously demonstrated in obstructive sleep apnea (OSA). However, investigation has been hampered by the necessity of invasive tissue biopsies. Objective: To evaluate the pharyngeal lavage (PHAL) as a new tool to analyze mucosal inflammation in the pharynx of patients with sleep-related disordered breathing. Patients and Methods: 36 patients with a diagnosis of OSA, 14 patients with heavy snorer syndrome (HS) or body position dependent OSA (bd-OSA), and 14 healthy volunteers underwent PHAL. Inflammatory cell counts were compared. Results: Neutrophils were the predominant cells in PHAL in all groups (94.3%±0.7%, 98.5%±0.6%, 94.3%±0.7%, and 96.2%±1.4%). OSA patients had significantly increased numbers of lymphocytes (3.2%±0.4%) compared to bd-OSA/HS and controls group (0.5%±0.1% and 0.6%±0.2%, respectively; P<0.05). Patients with moderate to severe OSA had significantly higher numbers of lymphocytes compared to patients with mild OSA (P<0.05). Conclusions: Data from this study suggest that PHAL is a feasible tool to investigate upper airway inflammation in OSA. In addition, PHAL demonstrates lymphocytic inflammation of the pharynx in OSA patients. Future studies are warranted to evaluate whether PHAL can be used to monitor disease and whether lymphocytic inflammation is affected by OSA treatment. [ABSTRACT FROM AUTHOR]

Intestinal edema remains a serious clinical problem, and novel approaches to study its pathophysiology are needed. It was our aim to develop a long-term stable isolated perfused rat small bowel preparation permitting analysis of vascular, luminal, interstitial, and lymphatic compartments and to demonstrate the utility of this model by studying the effects of the proinflammatory mediator platelet-activating factor (PAF). A temperature-controlled chamber with an integrated balance was designed to perfuse isolated intestines through the mesenteric artery and the gut lumen. Steroids or oxygen carriers were not needed. Functional and morphological integrity of the tissue was preserved for several hours as confirmed by oxygen consumption, venous lactate-to-pyruvate ratio, arterial and venous pH, lactose digestion and galactose uptake, intravascular and luminal pressures, maintained fluid homeostasis, gut motility, and quantitative light microscopic analysis. Administration of PAF caused typical effects such as vasoconstriction, gut atony, and loss of galactose uptake. PAF also elicited a transient loss of 20% of the perfusate liquid from the mesenteric vascular bed, two-thirds of which were transferred to the lumen. All these responses were entirely reversible. This new model provides detailed insights into the physiology of the small intestine and will allow to study fundamental processes such as fluid homeostasis, barrier functions, transport mechanisms, and immune responses in this organ. Using this model, here we show a dramatic and yet reversible response of the rat small bowel to PAF, suggesting luminal water clearance as a novel safety factor in the intestine that may be of clinical relevance. [ABSTRACT FROM AUTHOR]

Background: While lipopolysaccharide (LPS) from Gram-negative bacteria has been shown to augment inflammation in ventilated lungs information on the effect of Gram-positive bacteria is lacking. Therefore the effect of LPS and a lipopetide from Gram-positive bacteria, PAM3, on ventilated lungs were investigated. Methods: C57/Bl6 mice were mechanically ventilated. Sterile saline (sham) and different concentrations of LPS (1 μg and 5 μg) and PAM3 (50 nM and 200 nM) were applied intratracheally. Lung function parameters and expression of MIP-2 and TNFa as well as influx of neutrophils were measured. Results: Mechanical ventilation increased resistance and decreased compliance over time. PAM3 but not LPS significantly increased resistance compared to sham challenge (P < 0.05). Both LPS and PAM3 significantly increased MIP-2 and TNFα mRNA expression compared to sham challenge (P < 0.05). The numbers of neutrophils were significantly increased after LPS at a concentration of 5 μg compared to sham (P < 0.05). PAM3 significantly increased the numbers of neutrophils at both concentrations compared to sham (P < 0.05). Conclusions: These data suggest that PAM3 similar to LPS enhances ventilator-induced inflammation. Moreover, PAM3 but not LPS increases pulmonary resistance in ventilated lungs. Further studies are warranted to define the role of lipopetides in ventilator-associated lung injury. [ABSTRACT FROM AUTHOR]

Background: A large number of studies have investigated the effects of high tidal volume ventilation in mouse models. In contrast data on very short term effects of low tidal volume ventilation are sparse. Therefore we investigated the functional and structural effects of low tidal volume ventilation in mice. Methods: 38 Male C57/Bl6 mice were ventilated with different tidal volumes (Vt 5, 7, and 10 ml/kg) without or with application of PEEP (2 cm H2O). Four spontaneously breathing animals served as controls. Oxygen saturation and pulse rate were monitored. Lung function was measured every 5 min for at least 30 min. Afterwards lungs were removed and histological sections were stained for measurement of infiltration with polymorphonuclear leukocytes (PMN). Moreover, mRNA expression of macrophage inflammatory protein (MIP)-2 and tumor necrosis factor (TNF)a in the lungs was quantified using real time PCR. Results: Oxygen saturation did not change significantly over time of ventilation in all groups (P > 0.05). Pulse rate dropped in all groups without PEEP during mechanical ventilation. In contrast, in the groups with PEEP pulse rate increased over time. These effects were not statistically significant (P > 0.05). Tissue damping (G) and tissue elastance (H) were significantly increased in all groups after 30 min of ventilation (P < 0.05). Only the group with a Vt of 10 ml/kg and PEEP did not show a significant increase in H (P > 0.05). Mechanical ventilation significantly increased infiltration of the lungs with PMN (P < 0.05). Expression of MIP-2 was significantly induced by mechanical ventilation in all groups (P < 0.05). MIP-2 mRNA expression was lowest in the group with a Vt of 10 ml/kg + PEEP. Conclusions: Our data show that very short term mechanical ventilation with lower tidal volumes than 10 ml/kg did not reduce inflammation additionally. Formation of atelectasis and inadequate oxygenation with very low tidal volumes may be important factors. Application of PEEP attenuated inflammation. [ABSTRACT FROM AUTHOR]

Background: Nontypeable Haemophilus influenzae (NTHI) may play a role as an infectious trigger in the pathogenesis of chronic obstructive pulmonary disease (COPD). Few data are available regarding the influence of acute and persistent infection on tissue remodelling and repair factors such as transforming growth factor (TGF)-β. Methods: NTHI infection in lung tissues obtained from COPD patients and controls was studied in vivo and using an in vitro model. Infection experiments were performed with two different clinical isolates. Detection of NTHI was done using in situ hybridization (ISH) in unstimulated and in in vitro infected lung tissue. For characterization of TGF-β signaling molecules a transcriptome array was performed. Expression of the TGF-pseudoreceptor BMP and Activin Membrane-bound Inhibitor (BAMBI) was analyzed using immunohistochemistry (IHC), ISH and PCR. CXC chemokine ligand (CXCL)-8, tumor necrosis factor (TNF)-α and TGF-β expression were evaluated in lung tissue and cell culture using ELISA. Results: In 38% of COPD patients infection with NTHI was detected in vivo in contrast to 0% of controls (p < 0.05). Transcriptome arrays showed no significant changes of TGF-β receptors 1 and 2 and Smad-3 expression, whereas a strong expression of BAMBI with upregulation after in vitro infection of COPD lung tissue was demonstrated. BAMBI was expressed ubiquitously on alveolar macrophages (AM) and to a lesser degree on alveolar epithelial cells (AEC). Measurement of cytokine concentrations in lung tissue supernatants revealed a decreased expression of TGF-β (p < 0.05) in combination with a strong proinflammatory response (p < 0.01). Conclusions: We show for the first time the expression of the TGF pseudoreceptor BAMBI in the human lung, which is upregulated in response to NTHI infection in COPD lung tissue in vivo and in vitro. The combination of NTHI-mediated induction of proinflammatory cytokines and inhibition of TGF-β expression may influence inflammation induced tissue remodeling. [ABSTRACT FROM AUTHOR]

Bronchoalveolar lavage (BAL) is a useful diagnostic tool in interstitial lunge diseases (ILD). However, differential cell counts are often non specific and immunocytochemistry is time consuming. Staining of glyoproteins by periodic acid Schiff (PAS) reaction may help in discriminating different forms of ILD. In addition, PAS staining is easy to perform. BAL cells from patients with idiopathic pulmonary fibrosis (IPF) (n = 8), sarcoidosis (n = 9), and extrinsic allergic alveolitis (EAA) (n = 2) were investigated. Cytospins from BAL cells were made and cells were stained using Hemacolor quick stain and PAS staining. Lymphocytic alveolitis was found in sarcoidosis and EAA whereas in IPF both lymphocytes and neutrophils were increased. PAS positive cells were significantly decreased in EAA compared to IPF and sarcoidosis (25.5% ± 0.7% vs 59.8% ± 25.1% and 64.0% ± 19.7%, respectively) (P < 0.05). No significant correlation between PAS positive cells and inflammatory cells was observed. These results suggest that PAS staining of BAL cells may provide additional information in the differential diagnosis of ILD. Further studies ware warranted to evaluate PAS staining in larger numbers of BAL from patients with ILD. [ABSTRACT FROM AUTHOR]

Objectives/Hypothesis: Allergic inflammation of the upper airways is commonly associated with mucus hypersecretion. At present, there is no specific mucus regulating drug available. Our goal was to investigate the effect of glucocorticosteroids, acetyl-cysteine (ACC), and calcium-activated chloride channel (CLCA) inhibitors in a model of Th2 type cytokine induced mucin expression in human airway mucosa. Study Design: Prospective. Methods: Explanted tissue from human sinus mucosa was stimulated with interleukin (IL)-4, IL-9, or IL-13. Different concentrations of dexamethasone, ACC, or CLCA inhibitors [niflumic acid (NFA) or MSI-2216] were added to stimulated tissue. Epithelial mucin expression was quantified using periodic acid-Schiff staining. Results: IL-4, IL-9, and IL-13 significantly increased epithelial mucin expression ( P < .05). Dexamethasone reduced Th2 type cytokine induced mucin expression in a dose-dependant manner being statistically significant at concentrations ≥4.0 μmol/L (IL-4) and ≥40.0 μmol/L (IL-9 and IL-13) ( P < .05). ACC had no significant effect on IL-4 and IL-13 induced mucin expression, whereas IL-9 induced mucin expression was significantly decreased at concentrations ≥3.0 mmol/L ( P < .05). NFA and MSI-2216 decreased Th2 type cytokine induced mucin expression in a dose-dependant manner. This effect was statistically significant at concentrations ≥100 μmol/L (NFA) and ≥50 μmol/L (MSI-2216) ( P < .05). Conclusions: Th2 type cytokines can induce mucin expression in a model of explanted human airway mucosa. Th2 type cytokine induced mucin expression can be effectively reduced by either glucocorticosteroids or CLCA inhibitors ex vivo. Besides glucocorticosteroids CLCA inhibitors may offer an alternative therapeutic approach to treat allergic mucus hypersecretion. [ABSTRACT FROM AUTHOR]

In several tumors the transketolase activity, controlled inter alia by enzymes of the pentose phosphate pathway which is an alternative, energy generating reaction-cascade to glycolysis, has been correlated with proliferation. The increase of thiamine-dependant transketolase enzyme reactions is induced especially through upregulated transketolase-like enzyme 1 (TKTL1)-activity; that shows TKTL1 to be a causative enzyme for tumors enhanced, anaerobic glucose degradation. We investigated TKTL1-expression in 88 human, formalin-fixed non-small cell lung cancer tissues and 24 carcinomas of the breast by immunohistochemical stainings applying a 0 to 3 staining-score system (3 = strongest expression). For means of validation we additionally stained 40 NSCLC fixed and paraffin-embedded utilizing the HOPE-technique; showing comparable results to the formalin-fixed, paraffin-embedded specimens (not shown). Potential correlations with age, sex, TNM-classification parameters and tumor grading as well as tumor transcription factor 1 (TTF1) and surfactant protein A (SPA) expression were investigated. 40.9% of the analyzed lung tumors expressed TKTL1 weakly (Score 1), 38.6% moderately (score 2) and 17.1% strongly (score 3). 3 tumors were diagnosed TKTL1-negative (3.4%; score 0). All Breast cancer specimen stainings were positive and scored 1: 32%; scored 2: 36%; scored 3: 32%. Alveolar macrophages and Alveolar Epithelial Cells Type II were also found to be TKTL1-positive. None of the listed clinical parameters could be found to show a significant correlation to TKTL1 signal appearance. Although we describe the expression of TKTL1 in lung cancers, we need to state that up till now there is no scientific indication for any treatment regimens based upon these findings. [ABSTRACT FROM AUTHOR]

Rationale: Chlamydia pneumoniae (Cpn) infection may play a role in the pathogenesis of chronic obstructive pulmonary disease (COPD). Few data are available comparing persistent and acute infection of this pathogen in the human respiratory tract.Objectives: To study Cpn-induced innate immune responses in lung tissue from patients with COPD and control subjects ex vivo and in vitro.Methods: Cpn detection was done by nested polymerase chain reaction, in situ hybridization, and immunohistochemistry ex vivo in unstimulated tissue and in vitro using an acute Cpn infection model. As main endpoints for the assessment of early cellular responses, nuclear factor (NF)-kappaB activation and CXC chemokine ligand (CXCL)-8 expression were evaluated. The role of Toll-like receptors (TLRs) as recognition molecules in Cpn-induced innate responses was tested by blocking experiments.Measurements and Main Results: Fifteen percent of patients with COPD were chronically infected with Cpn in contrast to 0% of control subjects (p < 0.05). There were no differences in CXCL-8 and NF-kappaB expression between infected and noninfected COPD tissue ex vivo. In contrast, acute in vitro infection induced an intense innate immune response including up-regulation of TLR2. Blocking experiments demonstrated the predominant role of TLR2 in induction of the early immune response, whereas no influence on chlamydial infection rates was observed.Conclusions: Acute in vitro infection of human lung tissue with Cpn elicited a marked innate response via TLR2, whereas chronic chlamydial infection in patients with COPD was not associated with enhanced cellular activation. These findings suggest different roles of Cpn during acute and chronic stages of pulmonary infection. [ABSTRACT FROM AUTHOR]

Background: hCLCA1 is a member of the calcium-activated chloride channel family and is associated with disease-inducible mucus expression. Niflumic acid (NFA) and a closely related chemical structure are reported inhibitors of calcium-activated chloride channels and endotoxin-inducible mucus expression in the mouse. Therefore, we tested the hypothesis that hCLCA1 may be involved in lipopolysaccharide (LPS) induced mucin up-regulation in human airways. We also investigated the effect of NFA and MSI-2216 on LPS-induced mucin up-regulation.Materials and Methods : Explanted human airways and the muco-epidermoid cell line Calu-3 were stimulated with LPS. Different concentrations of NFA or MSI-2216 were added to LPS-stimulated airway mucosa and Calu-3 cells. Expression of hCLCA1 and MUC5AC mRNA and protein was quantified in human airways using real-time PCR and PAS staining. In addition, immunohistochemistry was performed for quantification of inflammatory cells (lymphocytes, monocytes, eosinophils, and neutrophils) in the submucosa of the airways. Expression of hCLCA1 protein in Calu-3 cells was analysed by FACS.Results: LPS significantly induced hCLCA1 and MUC5AC mRNA and protein expression in human airway mucosa (P < 0.05). NFA and MSI-2216 significantly decreased LPS-induced mucus expression in explanted airway mucosa in a dose-dependent manner (P < 0.05). In Calu-3 cells, LPS significantly increased hCLCA1 surface expression whereas intracellular expression was significantly decreased (P < 0.05). In Calu-3 cells, NFA and MSI-2216 also significantly reduced MUC5AC mRNA expression (P < 0.05).Conclusions: These data suggest that hCLCA1 may play a role in LPS-induced mucin expression in human airway mucosa. Calcium-activated chloride channel inhibitors significantly decreased LPS-induced mucus expression both ex vivo and in vitro . Therefore, blocking of hCLCA1 may offer a therapeutic approach to reduce bacterial-induced mucus hypersecretion. [ABSTRACT FROM PUBLISHER]

Background: hCLCA1 is a member of the calcium-activated chloride channel family and is associated with disease-inducible mucus expression. Niflumic acid (NFA) and a closely related chemical structure are reported inhibitors of calcium-activated chloride channels and endotoxininducible mucus expression in the mouse. Therefore, we tested the hypothesis that hCLCA1 may be involved in lipopolysaccharide (LPS) induced mucin up-regulation in human airways. We also investigated the effect of NFA and MSI-2216 on LPS-induced mucin up-regulation. Materials and Methods: Explanted human airways and the muco-epidermoid cell line Calu-3 were stimulated with LPS. Different concentrations of NFA or MSI-2216 were added to LPS-stimulated airway mucosa and Calu-3 cells. Expression of hCLCA1 and MUC5AC mRNA and protein was quantified in human airways using real-time PCR and PAS staining. In addition, immunohistochemistry was performed for quantification of inflammatory cells (lymphocytes, monocytes, eosinophils, and neutrophils) in the submucosa of the airways. Expression of hCLCA1 protein in Calu-3 cells was analysed by FACS. Results: LPS significantly induced hCLCA1 and MUC5AC mRNA and protein expression in human airway mucosa (P < 0.05). NFA and MSI-2216 significantly decreased LPS-induced mucus expression in explanted airway mucosa in a dose-dependent manner (P < 0.05). In Calu-3 cells, LPS significantly increased hCLCA1 surface expression whereas intracellular expression was significantly decreased (P < 0.05). In Calu-3 cells, NFA and MSI-2216 also significantly reduced MUC5AC mRNA expression (P < 0.05). Conclusions: These data suggest that hCLCA1 may play a role in LPS-induced mucin expression in human airway mucosa. Calcium-activated chloride channel inhibitors significantly decreased LPS-induced mucus expression both ex vivo and in vitro. Therefore, blocking of hCLCA1 may offer a therapeutic approach to reduce bacterial-induced mucus hypersecretion. [ABSTRACT FROM AUTHOR]

Background: Non-small cell lung cancer (NSCLC) causes most of cancer related deaths in humans and is characterized by poor prognosis regarding efficiency of chemotherapeutical treatment and long-term survival of the patients. The purpose of the present study was the development of a human ex vivo tissue culture model and the analysis of the effects of conventional chemotherapy, which then can serve as a tool to test new chemotherapeutical regimens in NSCLC. Methods: In a short-term tissue culture model designated STST (Short-Term Stimulation of Tissues) in combination with the novel *HOPE-fixation and paraffin embedding method we examined the responsiveness of 41 human NSCLC tissue specimens to the individual cytotoxic drugs carboplatin, vinorelbine or gemcitabine. Viability was analyzed by LIFE/ DEAD assay, TUNEL-staining and colorimetric MTT assay. Expression of Ki-67 protein and of BrdU (bromodeoxyuridine) uptake as markers for proliferation and of cleaved (activated) effector caspase-3 as indicator of late phase apoptosis were assessed by immunohistochemistry. Transcription of caspase-3 was analyzed by RT-PCR. Flow cytometry was utilized to determine caspase-3 in human cancer cell lines. Results: Viability, proliferation and apoptosis of the tissues were moderately affected by cultivation. In human breast cancer, small-cell lung cancer (SCLC) and human cell lines (CPC-N, HEK) proliferative capacity was clearly reduced by all 3 chemotherapeutic agents in a very similar manner. Cleavage of caspase-3 was induced in the chemo-sensitive types of cancer (breast cancer, SCLC). Drug-induced effects in human NSCLC tissues were less evident than in the chemosensitive tumors with more pronounced effects in adenocarcinomas as compared to squamous cell carcinomas. Conclusion: Although there was high heterogeneity among the individual tumor tissue responses as expected, we clearly demonstrate specific multiple drug-induced effects simultaneously. Thus, STST provides a useful human model to study numerous aspects of mechanisms underlying tumor responsiveness towards improved anticancer treatment. The results presented here shall serve as a base for multiple functional tests of novel chemotherapeutic approaches to NSCLC in the future. [ABSTRACT FROM AUTHOR]

As part of an investigation aimed at illuminating the possibilities and limits of the HOPE-fixation and paraffin-embedding technique we here describe a novel procedure which was developed in order to combine the benefits of the HOPE-technique with the capabilities of laser microdissection. The presented procedure avoids the need for amplification of template-RNA thus facilitating reliable and reproducible results. The excellent preservation of nucleic acids, proteins, and morphology in HOPE-fixed, paraffin-embedded tissues enhances the molecular applications available to date with materials acquired by laser microdissection when compared to formalin fixed, paraffin-embedded tissues, thus substantially extending the methodological panel in tissue based research. [ABSTRACT FROM AUTHOR]

Severe acute respiratory syndrome (SARS) is a new infectious disease caused by a novel coronavirus that leads to deleterious pulmonary pathological features. Due to its high morbidity and mortality and widespread occurrence, SARS has evolved as an important respiratory disease which may be encountered everywhere in the world. The virus was identified as the causative agent of SARS due to the efforts of a WHO-led laboratory network. The potential mutability of the SARS-CoV genome may lead to new SARS outbreaks and several regions of the viral genomes open reading frames have been identified which may contribute to the severe virulence of the virus. With regard to the pathogenesis of SARS, several mechanisms involving both direct effects on target cells and indirect effects via the immune system may exist. Vaccination would offer the most attractive approach to prevent new epidemics of SARS, but the development of vaccines is difficult due to missing data on the role of immune system-virus interactions and the potential mutability of the virus. Even in a situation of no new infections, SARS remains a major health hazard, as new epidemics may arise. Therefore, further experimental and clinical research is required to control the disease. [ABSTRACT FROM AUTHOR]

Backround: Cigarette smoke exposure including biologically active lipopolysaccharide (LPS) in the particulate phase of cigarette smoke induces activation of alveolar macrophages (AM) and alveolar epithelial cells leading to production of inflammatory mediators. This represents a crucial mechanism in the pathogenesis of chronic obstructive pulmonary disease (COPD). Respiratory pathogens are a major cause of exacerbations leading to recurrent cycles of injury and repair. The interaction between pathogen-associated molecular patterns and the host is mediated by pattern recognition receptors (PRR's). In the present study we characterized the expression of Toll-like receptor (TLR)-2, TLR4 and CD14 on human AM compared to autologous monocytes obtained from patients with COPD, healthy smokers and non-smokers. Methods: The study population consisted of 14 COPD patients without evidence for acute exacerbation, 10 healthy smokers and 17 healthy non-smokers stratified according to age. The expression of TLR2, TLR4 and CD14 surface molecules on human AM compared to autologous monocytes was assessed ex vivo using FACS analysis. In situ hybridization was performed on bronchoalveolar lavage (BAL) cells by application of the new developed HOPE-fixative. Results: The expression of TLR2, TLR4 and CD14 on AM from COPD patients, smokers and nonsmokers was reduced as compared to autologous monocytes. Comparing AM we detected a reduced expression of TLR2 in COPD patients and smokers. In addition TLR2 mRNA and protein expression was increased after LPS stimulation on non-smokers AM in contrast to smokers and COPD patients. Conclusion: Our data suggest a smoke related change in the phenotype of AM's and the cellular response to microbial stimulation which may be associated with impairment of host defenses in the lower respiratory tract. [ABSTRACT FROM AUTHOR]

Background: CpG-oligonucleotides (CpG-ODN), which induce signaling through Toll-like receptor 9 (TLR9), are currently under investigation as adjuvants in therapy against infections and cancer. CpG-ODN function as Th-1 adjuvants and are able to activate dendritic cells. In humans TLR9 has been described to be strongly expressed in B-lymphocytes, monocytes, plasmacytoid dendritic cells and at low levels in human respiratory cells. We determined whether a direct interaction of bacterial DNA with the tumor cells themselves is possible and investigated the expression and function of TLR9 in human malignant solid tumors and cell lines. TLR9 expression by malignant tumor cells, would affect treatment approaches using CpG-ODN on the one hand, and, on the other hand, provide additional novel information about the role of tumor cells in tumor-immunology. Methods: The expression of TLR9 in HOPE-fixed non-small lung cancer, non-malignant tissue and tumor cell lines was assessed using immunohistochemistry, confocal microscopy, in situ hybridization, RT-PCR and DNA-sequencing. Apoptosis and chemokine expression was detected by FACS analysis and the Bio-Plex system. Results: We found high TLR9 signal intensities in the cytoplasm of tumor cells in the majority of lung cancer specimens as well as in all tested tumor cell lines. In contrast to this non-malignant lung tissues showed only sporadically weak expression. Stimulation of HeLa and A549 cells with CpG-ODN induced secretion of monocyte chemoattractant protein-1 and reduction of spontaneous and tumor necrosis factor-alpha induced apoptosis. Conclusions: Here we show that TLR9 is expressed in a selection of human lung cancer tissues and various tumor cell lines. The expression of functionally active TLR9 in human malignant tumors might affect treatment approaches using CpG-ODN and shows that malignant cells can be regarded as active players in tumor-immunology. [ABSTRACT FROM AUTHOR]

Pulmonary presence of Chlamydia pneumoniae is associated with acute and chronic infections. We show that unapparent chlamydial infection in four out of 31 chronic obstructive pulmonary disease (COPD) patients (12.9%) is characterized by a significant increase in infected alveolar epithelial cells type II (18.2 ± 3.5% vs. 2.3 ± 0.9; IHC/ISH) compared to a newly established model of acute chlamydial infection (ACIM) in vital lung specimens from pulmonary lobectomy. Expression of cHSP60 demonstrated pathogen viability and virulence in the ACIM. We conclude that target cells differ in acute and chronic chlamydial infection and suggest the ACIM as a novel tool to analyze the host–pathogen-interactions in acute respiratory infections. [Copyright &y& Elsevier]

The ability of the host to recognize pulmonary invasion by pathogenic organisms and establish an appropriate host response to infection requires innate immune defense mechanisms. Early bacterial clearance in the lung is mediated by alveolar macrophages (AM) and polymorphonuclear neutrophils. Additionally alveolar epithelial cells type II (AEC-II) may act as immunoregulatory cells. The toll-like receptors (TLR) are part of this innate immune defense, recognizing conserved patterns on microorganisms. Toll-like receptor 2 (TLR2) is crucial in detecting components of gram-positive bacteria and mycobacteria. Signals initiated by the interaction of TLR2 with bacterial components direct the subsequent inflammatory response. The detection of TLR2 mRNA in human lung tissue prompted us to localize the expression of mRNA and protein at the cellular level using a novel method for tissue fixation. We utilized HOPE-fixed lung specimen sections for targeting mRNA by in situ hybridization and protein by immunohistochemistry using the monoclonal antibody TL2.1. In normal lung areas the expression of TLR2 mRNA and protein was found to be located in cells resembling AEC-II and AM. Expression of mRNA was verified by RT-PCR and DNA sequencing. These results indicate a potential mechanism of increased immunosurveillance at the alveolar level controlling the localized infection. [ABSTRACT FROM AUTHOR]

Background: In animal studies, gender differences were related to hormonal and immunologic changes that were associated with an increased susceptibility to sepsis in males. Objective: In a prospective study, gender differences in patients with surgical sepsis were evaluated in terms of survival, sex hormones, and proinflammatory as well as anti-inflammatory mediators. Setting: Surgical intensive care unit of a university hospital. Patients: Fifty-two patients (19 women and 33 men) with surgical sepsis. Measurements and Main Results: In a prospective study, tumor necrosis factor α and interleukin 6 bioactivity and plasma levels of interleukin 10 (using enzyme-linked immunosorbent assay), total testosterone, and 17-β estradiol (using radioimmunoassay) were determined on days 1, 3, 5, 7, 10, and 14 after diagnosis of sepsis. There were no differences in characteristics of patients in age (mean age, 55.4 years for women and 53.1 years for men) or cause and severity of sepsis (Acute Physiology and Chronic Health Evaluation II score, 17.3 for women and 18.5 for men; multiple organ dysfunction score, 9.9 vs 10.8, respectively). Although no difference could be found in the multiple organ dysfunction score from day 1 to day 28, the prognosis of sepsis was significantly different in women compared with men. Hospital mortality rate was 70% (23 of 33 patients) in male and 26% (5 of 19) in female patients (P<.008, log-rank test). Bioactivity of tumor necrosis factor continuously increased in men after diagnosis of sepsis, with significantly elevated levels on day 10 (P<.05, Mann-Whitney U test with Bonferroni correction), whereas no difference was found for interleukin 6 bioactivity. Women displayed enhanced interleukin 10 levels compared with men from day 1 to day 10 that reached a significant difference on days 3 and 5 (P<.05). Total testosterone levels were below the normal range for men, and estradiol levels were initially increased in both men and post... [ABSTRACT FROM AUTHOR]

Objective: To evaluate the effect of pentoxifylline on organ dysfunction, survival, and mediator response in patients with severe sepsis. Design: Randomized, double-blind, placebo-controlled study. Setting: Surgical intensive care units at 2 university hospitals. Patients: Fifty-one surgical patients with severe sepsis were randomized to receive pentoxifylline continuously (27 patients) or saline infusion as placebo (24 patients). Interventions: Patients received pentoxifylline (1 mg/kg of body weight per hour; maximum, 1800 mg/d) during 28 days or until they were discharged from the intensive care unit or died. Measurements and Main Results: Vital signs and organ function were determined at diagnosis; daily from day 1 to 7; on days 10, 14, 17, 21, and 24; and 28 days after diagnosis of sepsis. There were no differences in characteristics of patients at diagnosis in the Acute Physiology and Chronic Health Evaluation II (APACHE II) score (mean±SEM, 17±4 points for the pentoxifylline group and 18±5 points for the placebo group), the multiple organ dysfunction score (mean±SEM, 11.0±0.8 vs 11.8±1.0 points), tumor necrosis factor α and interleukin 6 bioactivity, serum endotoxin levels, or organ dysfunction. At study entrance, 23 of 27 patients in the pentoxifylline group and 21 of 24 patients in the placebo group experienced septic shock. No adverse effects of pentoxifylline treatment were observed. The 28-day mortality rate was 30% (8/27) in pentoxifylline-treated patients and 33% (8/24) in the placebo group. Hospital mortality was 41% (11/27) in the pentoxifylline group and 54% (13/24) in the placebo group. The multiple organ dysfunction score decreased in patients receiving pentoxifylline 4 days after diagnosis of sepsis compared with placebo-treated patients; a significant difference was reached on day 14 (P<.05; Student t test, Bonferoni correction). The PaO[sub 2]/FIO[sub 2] (fraction of inspired oxygen) ratio was signi... [ABSTRACT FROM AUTHOR]

No abstract available Copyright © 2012 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background: While lipopolysaccharide (LPS) from Gram-negative bacteria has been shown to augment inflammation in ventilated lungs information on the effect of Gram-positive bacteria is lacking. Therefore the effect of LPS and a lipopetide from Gram-positive bacteria, PAM3, on ventilated lungs were investigated.Methods: C57/Bl6 mice were mechanically ventilated. Sterile saline (sham) and different concentrations of LPS (1 microg and 5 microg) and PAM3 (50 nM and 200 nM) were applied intratracheally. Lung function parameters and expression of MIP-2 and TNFalpha as well as influx of neutrophils were measured.Results: Mechanical ventilation increased resistance and decreased compliance over time. PAM3 but not LPS significantly increased resistance compared to sham challenge (P < 0.05). Both LPS and PAM3 significantly increased MIP-2 and TNFalpha mRNA expression compared to sham challenge (P < 0.05). The numbers of neutrophils were significantly increased after LPS at a concentration of 5 microg compared to sham (P < 0.05). PAM3 significantly increased the numbers of neutrophils at both concentrations compared to sham (P < 0.05).Conclusions: These data suggest that PAM3 similar to LPS enhances ventilator-induced inflammation. Moreover, PAM3 but not LPS increases pulmonary resistance in ventilated lungs. Further studies are warranted to define the role of lipopetides in ventilator-associated lung injury. [ABSTRACT FROM AUTHOR]