Number Of Neutrophils In Bronchoalveolar Lavage Research Articles

PET in the diagnostic management of infectious/inflammatory pulmonary pathologies: a revisit in the era of COVID-19.

PET in the diagnostic management of infectious/inflammatory pulmonary pathologies: a revisit in the era of COVID-19.

Neutrophilic inflammation in asthma and defective epithelial translational control.

Neutrophilic inflammation in asthma is associated with interleukin (IL)-17A, corticosteroid-insensitivity and bronchodilator-induced forced expiratory volume in 1 s (FEV1) reversibility. IL-17A synergises with tumour necrosis factor (TNF)-α in the production of the neutrophil chemokine CXCL-8 by primary bronchial epithelial cells (PBECs).We hypothesised that local neutrophilic inflammation in asthma correlates with IL-17A and TNF-α-induced CXCL-8 production by PBECs from asthma patients.PBECs from most asthma patients displayed an exaggerated CXCL-8 production in response to TNF-α and IL-17A, but not to TNF-α alone, and which was also insensitive to corticosteroids. This hyperresponsiveness of PBECs strongly correlated with CXCL-8 levels and neutrophil numbers in bronchoalveolar lavage from the corresponding patients, but not with that of eosinophils. In addition, this hyperresponsiveness also correlated with bronchodilator-induced FEV1 % reversibility. At the molecular level, epithelial hyperresponsiveness was associated with failure of the translational repressor T-cell internal antigen-1 related protein (TiAR) to translocate to the cytoplasm to halt CXCL-8 production, as confirmed by TiAR knockdown. This is in line with the finding that hyperresponsive PBECs also produced enhanced levels of other inflammatory mediators.Hyperresponsive PBECs in asthma patients may underlie neutrophilic and corticosteroid-insensitive inflammation and a reduced FEV1, irrespective of eosinophilic inflammation. Normalising cytoplasmic translocation of TiAR is a potential therapeutic target in neutrophilic, corticosteroid-insensitive asthma.

Acute respiratory changes and pulmonary inflammation involving a pathway of TGF-β1 induction in a rat model of chlorine-induced lung injury

We investigated acute and delayed respiratory changes after inhalation exposure to chlorine (Cl2) with the aim to understand the pathogenesis of the long-term sequelae of Cl2-induced lung-injury.In a rat model of nose-only exposure we analyzed changes in airway hyperresponsiveness (AHR), inflammatory responses in airways, expression of pro-inflammatory markers and development of lung fibrosis during a time-course from 5h up to 90days after a single inhalation of Cl2. A single dose of dexamethasone (10mg/kg) was administered 1h following Cl2-exposure.A 15-min inhalation of 200ppm Cl2 was non-lethal in Sprague-Dawley rats. At 24h post exposure, Cl2-exposed rats displayed elevated numbers of leukocytes with an increase of neutrophils and eosinophils in bronchoalveolar lavage (BAL) and edema was shown both in lung tissue and the heart. At 24h, the inflammasome-associated cytokines IL-1β and IL-18 were detected in BAL. Concomitant with the acute inflammation a significant AHR was detected. At the later time-points, a delayed inflammatory response was observed together with signs of lung fibrosis as indicated by increased pulmonary macrophages, elevated TGF-β expression in BAL and collagen deposition around airways. Dexamethasone reduced the numbers of neutrophils in BAL at 24h but did not influence the AHR.Inhalation of Cl2 in rats leads to acute respiratory and cardiac changes as well as pulmonary inflammation involving induction of TGF-β1. The acute inflammatory response was followed by sustained macrophage response and lack of tissue repair. It was also found that pathways apart from the acute inflammatory response contribute to the Cl2-induced respiratory dysfunction.

Regulation of IL-17A expression in mice following subacute ozone exposure

Exposure to subacute ozone (O3) causes pulmonary neutrophil recruitment. In mice, this recruitment requires IL-17A. Ozone also causes expression of IL-23 and IL-1, which can induce IL-17A. The purpose of this study was to examine the hypothesis that IL-23 and IL-1 contribute to IL-17A expression and subsequent neutrophil recruitment after subacute O3 exposure. Wild-type, IL-23−/−, and Flt3l−/− mice were exposed to air or 0.3 ppm O3 for 72 h. Flt3l−/− mice lack conventional dendritic cells (cDC) that can express IL-23 and IL-1. Other wild-type mice were pre-treated with saline or the IL-1R1 antagonist anakinra prior to O3 exposure. After exposure, bronchoalveolar lavage (BAL) was performed and lung tissue harvested. The results indicated that pulmonary Il17a mRNA abundance and IL-17A+ F4/80+ cells were significantly reduced in O3-exposed IL-23−/− vs in wild-type mice. In contrast, anakinra had no effect on Il23a or Il17a pulmonary mRNA abundance or on BAL concentrations of the neutrophil survival factor G-CSF, but anakinra did reduce BAL neutrophil numbers, likely because anakinra also reduced BAL IL-6. Compared to air, O3 caused a significant increase in DC numbers in wild-type, but not in Flt3−/− mice. However, there was no significant difference in Il23a or Il17a mRNA abundance or in BAL neutrophil count in O3-exposed Flt3−/− vs in wild-type mice. From these results, it was concluded that IL-23 but not IL-1 contributes to the IL-17A expression induced by subacute O3 exposure. Induction of IL-23 by O3 does not appear to require cDC.

Effect of fluticasone propionate on virus-induced airways inflammation and anti-viral immune responses in mice

BackgroundInhaled corticosteroids are commonly used in the treatment of asthma and chronic obstructive pulmonary disease, but their effects on viral loads and anti-viral responses are poorly characterised. The aim of this study was to assess effects of inhaled fluticasone propionate on rhinovirus infection in vivo, in a mouse model. We tested the hypothesis that this treatment would reduce virus-induced airways inflammation but that the effect would be confounded by interference with anti-viral immune responses, leading to delayed viral clearance. MethodsC57BL/6 mice were intranasally dosed with fluticasone propionate (1 mg/kg) or vehicle (dimethyl sulfoxide, control), 1 h before infection with rhinovirus 1B. We assessed bronchoalveolar lavage (BAL) inflammatory cell numbers, and measured gene expression, protein production of innate mediators, or both by quantitative RT-PCR or ELISA. We compared mice treated with fluticasone with controls at various timepoints after infection. In additional experiments, recombinant interferon (IFN) beta was administered with fluticasone and rhinovirus 1B in both groups of mice. FindingsAt 24 h post infection, fluticasone treatment suppressed rhinovirus induction of type I and III IFNs in the airways (for the fluticasone-treated group compared with controls: mean IFNβ BAL protein 20·2 pg/mL [SD 16·7] vs 103·0 [30·9]; mean IFNλ BAL protein 102·6 pg/mL [17·4] vs 217 [44·6], p<0·001); it also impaired viral clearance, with increased lung tissue viral RNA copy numbers (4·7 × 105 copies [SD 1·3] vs 2·6 × 105 [0·8], p<0·001). Despite increasing viral loads, fluticasone inhibited rhinovirus-induced airway inflammation as evidenced by suppressed BAL neutrophil numbers in fluticasone compared with control mice (0·021 × 105 [0·012] vs 0·59 × 5 [0·39], p<0·001) and by suppressed lymphocyte numbers (0·092 × 105 [0·044] vs 0·45 × 105 [0·11], p<0·001). By contrast, fluticasone increased MUC5AC proteins (158·2 arbitrary units [29·9] vs 107·6 [7·1], p=0·0165) and MUC5B proteins (623·8 arbitrary units [231·9] vs 413·5 [70·5], p=0·0476) in BAL at day 7 post infection. Administration of intranasal recombinant IFN beta (104 units) with fluticasone and rhinovirus 1B led to upregulation of interferon-inducible cytokines OAS and CXCL10/IP-10 compared with control mice treated with fluticasone and rhinovirus alone and improved viral clearance without having any effect on suppression of inflammation by fluticasone. InterpretationOur findings suggest that fluticasone treatment suppresses rhinovirus-induced airways inflammation in vivo but also impairs anti-viral immune responses and increases viral titres, leading to mucus hypersecretion. Since asthma and chronic obstructive pulmonary disease are both associated with inherent deficient IFN responses to rhinovirus, inhaled corticosteroids might interact synergistically with disease to inhibit IFN and thus lead to increased severity of exacerbation. The clinical applicability of these findings requires confirmation in human models of disease. FundingWellcome Trust.

Abstract 1673: Alternative NF-κB signaling promotes inflammatory cell recruitment and lung tumor formation

Abstract Nuclear factor-κB (NF-κB) is a transcription factor that can be activated through either the classical or alternative signaling pathways. In lung tumors, high levels of classical and alternative pathway components have been observed. While many studies have demonstrated that classical NF-κB signaling is a central pathway regulating inflammation and tumorigenesis, the role of alternative NF-κB signaling remains undefined. We hypothesize that alternative NF-κB signaling in the lung epithelium plays a key role in inflammation and lung tumor formation through paracrine signaling to the inflammatory microenvironment. To investigate alternative NF-κB signaling in the lung epithelium, we generated a transgenic mouse model with doxycycline (dox)-inducible expression of the alternative pathway component p52 in airway epithelial cells (designated ALTA for alternative pathway trans-activated). When placed on dox, nuclear p52 is increased in the lungs of ALTA mice with no effect on activation of other NF-κB components. Surprisingly, alternative pathway activation alone did not cause a significant increase in inflammatory cell recruitment. However, in conjunction with the inflammatory stimulus lipopolysaccharide (LPS), alternative pathway activation caused increased mortality, which was associated with a significant increase in inflammation and increased lung permeability. To determine the effect of alternative pathway activation on lung tumor formation, mice were injected with the lung carcinogen urethane. Six weeks after urethane administration, ALTA mice demonstrated an increased number of bronchoalveolar lavage neutrophils, and by 6 months post-urethane, ALTA mice had an increased number of tumors, increased tumor size, and more advanced lesions. Together, these data suggest that alternative NF-κB signaling plays an important role in stimulus-induced inflammatory cell recruitment and that alternative pathway activation can promote lung tumor formation and progression. By defining the role of alternative NF-κB signaling in tumor formation, we hope to gain insight into the role of inflammation in lung carcinogenesis and to identify novel pathways and signaling targets that could potentially serve as therapeutic targets. Citation Format: Jamie Alicyn Ausborn, Dong-Sheng Cheng, Vasiliy Polosukhin, Wei Han, Fiona Yull, Timothy Blackwell. Alternative NF-κB signaling promotes inflammatory cell recruitment and lung tumor formation. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1673. doi:10.1158/1538-7445.AM2014-1673

CXC chemokines and antimicrobial peptides in rhinovirus‐induced experimental asthma exacerbations

SummaryRationaleRhinoviruses (RVs) are the major triggers of asthma exacerbations. We have shown previously that lower respiratory tract symptoms, airflow obstruction, and neutrophilic airway inflammation were increased in experimental RV‐induced asthma exacerbations.ObjectivesWe hypothesized that neutrophil‐related CXC chemokines and antimicrobial peptides are increased and related to clinical, virologic, and pathologic outcomes in RV‐induced exacerbations of asthma.MethodsProtein levels of antimicrobial peptides (SLPI, HNP 1–3, elafin, and LL‐37) and neutrophil chemokines (CXCL1/GRO‐α, CXCL2/GRO‐β, CXCL5/ENA‐78, CXCL6/GCP‐2, CXCL7/NAP‐2, and CXCL8/IL‐8) were determined in bronchoalveolar lavage (BAL) fluid of 10 asthmatics and 15 normal controls taken before, at day four during and 6 weeks post‐experimental infection.Results BAL HNP 1–3 and Elafin were higher, CXCL7/NAP‐2 was lower in asthmatics compared with controls at day 4 (P = 0.035, P = 0.048, and P = 0.025, respectively). BAL HNP 1–3 and CXCL8/IL‐8 were increased during infection (P = 0.003 and P = 0.011, respectively). There was a trend to increased BAL neutrophils at day 4 compared with baseline (P = 0.076). BAL HNP 1–3 was positively correlated with BAL neutrophil numbers at day 4. There were no correlations between clinical parameters and HNP1–3 or IL‐8 levels.ConclusionsWe propose that RV infection in asthma leads to increased release of CXCL8/IL‐8, attracting neutrophils into the airways where they release HNP 1–3, which further enhances airway neutrophilia. Strategies to inhibit CXCL8/IL‐8 may be useful in treatment of virus‐induced asthma exacerbations.

Inhibition of LPS-induced acute lung inflammation in mice by adenovirus mediated Foxp3 expression (HYP6P.275)

Abstract Forkhead transcription factor 3 (Foxp3) is critical transcription factor in CD4(+)CD25(+) regulatory T cells. There are increasing evidences that the functions of Foxp3 proteins are not restricted to regulatory T cells. The aim of this study is to evaluate the functions of Foxp3 proteins in LPS induced lung inflammation. In this study, LPS was administrated to Balb/c mice to induce a disease that resembles COPD. Two days prior to LPS administration, mice were infected once with the Foxp3-GFP or control GFP expression adenovirus to intra-trachea. The total cells and neutrophil numbers in bronchoalveolar lavage (BAL) fluid were counted and pro-inflammatory cytokine levels in BAL fluid were measured after 3 days later from LPS administration. Lung tissues were stained with hematoxylin and eosin (H&amp;E) for histologic evaluation. Total cells, including eosinophil, macrophage and lymphocytes was significantly reduced in the foxp3 expression adenovirus infected mouse BAL fluid compared with the control adenovirus infected mouse BAL fluid. Foxp3-GFP or GFP was only detected in the lung epithelial cells by confocal microscopy. Using a mouse lung epithelial cell line, LA-4, Foxp3 adenovirus infected cells reduced the expression of chemokines such as CCL3, CXCL2 and CXCL5 by LPS stimulation. These results suggest that Foxp3 expression in lung epithelial cells suppresses inflammation via inhibition of chemokines secretion in lung inflammation.

Interleukin-6 neutralization alleviates pulmonary inflammation in mice exposed to cigarette smoke and poly(I:C)

Increased systemic and pulmonary levels of IL-6 (interleukin-6) are associated with the severity of exacerbations and decline of lung function in patients with COPD (chronic obstructive pulmonary disease). Whether IL-6 is directly involved or plays a bystander role in the pathophysiology of COPD remains unclear. Here we hypothesized that neutralizing circulating levels of IL-6 would modulate episodes of acute pulmonary inflammation following CS (cigarette smoke) exposure and virus-like challenges. For this purpose, we used a model where C57BL/6 mice were exposed to CS twice daily via a nose-only system, and concomitant periodic intranasal challenge with poly(I:C), a synthetic ligand for TLR3 (Toll-like receptor 3) that mimics the encounter with double stranded RNA that is carried by influenza-like viruses. This protocol recapitulates several aspects of acute pulmonary inflammation associated with COPD, including prominent airway neutrophilia, insensitivity to steroid treatment and increased levels of several inflammatory cytokines in BAL (bronchoalveolar lavage) samples. Although IL-6-deficient mice exposed to CS/poly(I:C) developed pulmonary inflammation similar to WT (wild-type) controls, WT mice exposed to CS/poly(I:C) and treated intraperitoneally with IL-6-neutralizing antibodies showed significantly lower blood counts of lymphocytes and monocytes, lower BAL levels of IL-6 and CXCL1 (CXC chemokine ligand 1)/KC (keratinocyte chemoattractant), as well as reduced numbers of BAL neutrophils, lymphocytes and macrophages. Our results thus indicate that the systemic neutralization of IL-6 significantly reduces CS/poly(I:C)-induced pulmonary inflammation, which may be a relevant approach to the treatment of episodes of acute pulmonary inflammation associated with COPD.

Leukocyte‐Specific Protein 1 (LSP1) regulates neutrophil migration in acute lung inflammation

LSP1, an F‐actin binding protein, plays a role in neutrophil recruitment in peritoneum. Because mechanisms of excessive migration of activated neutrophils, credited with tissue damage, are not fully understood, we explored the hitherto unknown expression and role of LSP1 in neutrophil migration in acute lung inflammation (ALI). We induced ALI through intranasal E. coli LPS (80μg) in wild type 129/SVJ (WT) and LSP1 deficient (LSP−/− ) mice. WT (n=10) and LSP−/− (n=11) mice showed significant neutrophilia and more neutrophils in broncho‐alveolar lavage (BAL) at 9 hr post‐LPS challenge compared to respective saline‐treated controls (WT=7; LSP−/−=10). BAL neutrophil numbers were higher in LPS‐treated WT mice compared to LSP−/− mice (P&lt;0.001). Lung myeloperoxidase and Gr1+ were higher in LPS‐treated WT compared to the LSP−/− mice (P&lt;0.05). Lung tissue and BAL fluid KC, MCP1, MIP1α and MIP1β concentration and vascular permeability were not different between LPS‐treated WT and LSP−/− mice but TNFα concentration was higher in LPS‐treated WT mice. LSP1 expression was increased in inflamed lungs from LPS‐treated mice, and autopsied lungs from septic humans, compared to their respective controls. H&amp;E staining showed more septal congestion in LPS‐treated WT mice compared to LSP−/− mice. These data show that LSP1 expression is modulated in ALI and that LSP1 deficiency reduces neutrophil migration into ALI. Funding: NSERC

Elimination of Aspergillus fumigatus conidia from the airways of mice with allergic airway inflammation

BackgroundAspergillus fumigatus conidia can exacerbate asthma symptoms. Phagocytosis of conidia is a principal component of the host antifungal defense. We investigated whether allergic airway inflammation (AAI) affects the ability of phagocytic cells in the airways to internalize the resting fungal spores.MethodsUsing BALB/c mice with experimentally induced AAI, we tested the ability of neutrophils, macrophages, and dendritic cells to internalize A. fumigatus conidia at various anatomical locations. We used light microscopy and differential cell and conidium counts to determine the ingestion potential of neutrophils and macrophages present in bronchoalveolar lavage (BAL). To identify phagocyte-conidia interactions in conducting airways, conidia labeled with tetramethylrhodamine-(5-(and-6))-isothiocyanate were administered to the oropharyngeal cavity of mice. Confocal microscopy was used to quantify the ingestion potential of Ly-6G+ neutrophils and MHC II+ antigen-presenting cells located in the intraepithelial and subepithelial areas of conducting airways.ResultsAllergen challenge induced transient neutrophil recruitment to the airways. Application of A. fumigatus conidia at the acute phase of AAI provoked recurrent neutrophil infiltration, and consequently increased the number and the ingestion potential of the airway neutrophils. In the absence of recurrent allergen or conidia provocation, both the ingestion potential and the number of BAL neutrophils decreased. As a result, conidia were primarily internalized by alveolar macrophages in both AAI and control mice at 24 hours post-inhalation. Transient influx of neutrophils to conducting airways shortly after conidial application was observed in mice with AAI. In addition, the ingestion potential of conducting airway neutrophils in mice with induced asthma exceeded that of control mice. Although the number of neutrophils subsequently decreased, the ingestion capacity remained elevated in AAI mice, even at 24 hours post-conidia application.ConclusionsAspiration of allergen to sensitized mice enhanced the ingestion potential of conducting airway neutrophils. Such activation primes neutrophils so that they are sufficient to control dissemination of non-germinating A. fumigatus conidia. At the same time, it can be a reason for the development of sensitivity to fungi and subsequent asthma exacerbation.

Integrin β3 is not critical for neutrophil recruitment in a mouse model of pneumococcal pneumonia

Streptococcus pneumoniae is one of the most common causes of bacterial pneumonias in humans. Neutrophil migration into lungs infected with S. pneumoniae is central to the host defense but the mechanisms of neutrophil recruitment, as mediated by S. pneumoniae, into lungs are incompletely understood. Therefore, we have assessed the role of integrin αvβ3 by evaluating its subunit β3 in a mouse model of lung inflammation induced by S. pneumonia. Integrin subunit β3 knockout (β3(-/-)) and wild-type (WT) mice were intratracheally instilled with either S. pneumoniae or saline. Other groups of WT mice were treated intraperitoneally with 25μg or 50μg of antibody against integrin β3 or with isotype-matched antibody at 1h before instillation of S. pneumoniae. Mice were killed 24h after infection. Flow cytometry confirmed the absence or presence of integrin subunit β3 on peripheral blood neutrophils in β3(-/-) or WT mice, respectively. Neutrophil numbers in bronchoalveolar lavage (BAL) from infected β3(-/-) and WT mice showed no differences. Neutrophil numbers in BAL of infected WT mice treated with β3 antibody were lower compared with those without antibody but similar to those of mice administered isotype-matched antibody. Many neutrophils were present in the perivascular spaces of the lungs in β3(-/-) mice. Lungs from infected β3(-/-) mice had negligible mitogen-activated protein kinase expression compared with those of infected WT mice. Thus, integrin β3 or its heterodimer αvβ3 is not critical for neutrophil migration into lungs infected with S. pneumoniae.

Small macrophages are present in early childhood respiratory disease

BackgroundRecently, an established “small macrophage” phenotype has been observed in the sputum of patients with CF and COPD. However, little is known about the prevalence of this phenotype in the airways of young children. Since respiratory inflammation begins early in CF, we hypothesised that these small macrophages would be increased in paediatric CF bronchoalveolar lavage (BAL). MethodsMacrophage populations in CF and disease control BAL were assessed by multicolour flow cytometry. BAL inflammatory indices were collected as part of the AREST-CF programme. ResultsSmall macrophages were present in CF (n=35, mean 36±12% of BAL macrophages) but not significantly different to the respiratory disease controls (n=7, mean 40±21%). Number of small macrophages correlated significantly with number of BAL neutrophils (r=0.44, p<0.01) but not infection or IL-8. ConclusionsIn paediatric patients small macrophages are not unique to CF, but their establishment as the dominant phenotype in adults may be due to chronicity of inflammation and infection.

Effect of fermented Lonicera japonica on LPS-induced acute lung inflammation

Lonicera japonica that had been fermented with Lactobacillus casei (KCCM 10766P) was evaluated to determine if it exerted a protective effect against lipopolysaccaride (LPS) induced lung inflammation. In this study, LPS was administrated to Balb/c mice to induce a disease that resembles COPD. Two hours prior to LPS administration, mice were treated once with Fermented LJ (FLJ) via oral gavage. The total cell and neutrophil numbers in bronchoalveolar lavage (BAL) fluid were counted and pro-inflammatory cytokine levels in BAL fluid were measured. Lung tissues were stained with hematoxylin and eosin (H&E) for histologic evaluation. Neutrophil elastase (NE) and proliferating cell nuclear antigen (PCNA) were assessed by immunohistochemistry. On days 3 and 7 after LPS stimulation, an influx of neutrophils was noted, but the total cell number was significantly decreased in the FLJ group compared to the LPS group. In addition, TNF-α and IL-6 levels in BAL fluid were decreased in the FLJ group. Histologic results also demonstrated the attenuation effect of FLJ on LPS-induced lung inflammations. These data suggest that FLJ has a protective effect on LPS-induced lung inflammation. Therefore, fermented herbal medicine may represent a novel therapeutic agent for lung inflammation and in particular for COPD.

A Study on the Mechanism of Immunomodulating Effects of Moxifloxacin in Oleic Acid-Induced Acute Lung Injury

Background: It was hypothesized that the immunomodulating effects of moxifloxacin contribute to ameliorate oleic acid (OA)-induced acute lung injury (ALI) by suppression of cytosolic phospholipase A2 (cPLA2). This was based on observations from experiments on rats associated with neutrophilic respiratory burst, cPLA2 activity, and expressions of cPLA2, , and COX-II in the lung. Methods: ALI was induced by intravenous injection of OA in male Sprague-Dawley rats. Five hours after OA injection, protein content in bronchoalveolar lavage (BAL), lung myeloperoxidase (MPO) activity, and numbers of BAL neutrophils were measured. As an index of oxidative stress-induced lung injury, the content of malondialdehyde (MDA) in lung tissues was also determined. Lung histology, immunohistochemistry and determination of activity of cPLA2 in lung tissues were carried out. In addition, Western blotting of and COX-II in lung tissues was performed. Results: The accumulation of neutrophils in the lungs was observed after OA injection. BAL protein was increased along with neutrophilic infiltration and migration by OA. Moxifloxacin decreased all of these parameters of ALI and ameliorated ALI histologically. The increased malondialdehyde (MDA) in the lung by OA was also decreased by moxifloxacin. Moxifloxacin not only suppressed cPLA2 expression in the lungs and neutrophils but also decreased cPLA2 activity in lung tissues of rats given OA. The enhanced expressions of and COX-2 in the lung tissues of rats given OA were also suppressed by moxifloxacin. Conclusion: Moxifloxacin inhibited cPLA2 and down-regulated and COX-2 in the lungs of rats given OA, which resulted in the attenuation of inflammatory lung injury.

Changes in bronchoalveolar cell profiles of asthma patients induced by long-term glucocorticoid therapy

Effects of long-term glucocorticoid therapy on airway inflammatory cells were examined in 40 patients with bronchial asthma. 1. The dose of glucocorticoids required by asthmatic patients tended to increase and the level of serum cortisol tended to decrease with prolongation of the period of glucocorticoid therapy. 2. The proportion and number of lymphocytes in bronchoalveolar lavage (BAL) fluid decreased with prolongation of the period of glucocorticoid therapy. The proportion of BAL lymphocytes in asthmatic patients treated with glucocorticoids for more than 5 years was significantly lower than that in those treated for less than 4.9 years. 3. The proportion and number of BAL neutrophils increased with prolongation of the period of glucocorticoid therapy and the proportion of BAL neutrophils in patients treated with glucocorticoids for more than 10 years was significantly higher compared with those treated for less than 9.9 years. These findings indicate that long-term glucocorticoid therapy induces changes in airway inflammatory cell profiles, with a decrease in the number of lymphocytes and increase in the number of neutrophils.

Ventilation during Cardiopulmonary Bypass: Impact on Neutrophil Activation and Pulmonary Sequestration

Background: Cardiopulmonary bypass (CPB) is associated with neutrophil activation, pulmonary sequestration, and release of inflammatory mediators leading to pulmonary dysfunction. We investigate the effect of continuous ventilation during cardiopulmonary bypass on neutrophil activation and pulmonary sequestration. Methods: Forty-six patients undergoing coronary artery bypass grafting with cardiopulmonary bypass were prospectively randomized to continuous ventilation and nonventilation groups. Blood samples were collected, and bronchoalveolar lavage (BAL) was performed following induction of anesthesia and at 4 hr after aortic declamping. Differential white cell count was measured, and flow cytometry to determine cell count numbers and quantify CD45 and CD11b leukocyte cell surface adhesion molecule expression was performed on the blood and BAL samples. Results: Twenty-three patients were randomized to standard nonventilated CPB and 23 patients to ventilation throughout CPB. Significant increases in blood and BAL neutrophil numbers were detected at 4 hr following aortic declamping in both groups (Blood: NV p <. 0001, V p <. 0001; BAL: NV p =. 017, V p =. 0007). No significant inter-group differences in BAL and blood neutrophil numbers were found. Significantly higher blood neutrophil CD11b mean fluorescent intensity levels were present 4 hr following declamping compared with baseline in both groups (NV Blood, p =. 021; V Blood p <. 0001). No significant inter- or intragroup differences in BAL neutrophil CD11b mean fluorescent intensity levels were found. There was no death or major complication. Conclusions: Cardiopulmonary bypass during coronary artery bypass grafting is associated with increased neutrophil pulmonary sequestration, and blood neutrophil CD11b activation. Continuous ventilation during cardiopulmonary bypass does not significantly reduce neutrophil pulmonary sequestration or activation.

Role of p38 mitogen-activated protein kinase in ozone-induced airway hyperresponsiveness and inflammation

Ozone is a potent oxidant and causes airway hyperresponsiveness and neutrophilia. To determine the role of p38 mitogen-activated protein kinase (MAPK) activation, we studied the effect of a p38α inhibitor SD-282 (Scios Inc, Fremont, CA USA) on ozone-induced airway hyperresponsiveness and neutrophilia. Balb/c mice received SD-282 (30 or 90 mg/kg i.p) or vehicle 1 h before exposure to either ozone (3 ppm, 3 h) or air. Three hours after exposure, lungs were analysed for cytokine levels and bronchoalveolar lavage was performed. Another set of mice were dosed 6 h after exposure and 1 h before assessing airway hyperresponsiveness. SD-282 (90 mg/kg) significantly inhibited ozone-induced airway hyperresponsiveness (− LogPC 150: SD-282: − 1.73 ± 0.14 vs. vehicle: − 0.99 ± 0.15, P < 0.05). Bronchoalveolar lavage neutrophil numbers were time-dependently increased in vehicle-dosed, ozone-exposed mice, greatest at 20–24 h after exposure. SD-282 (30 and 90 mg/kg) significantly inhibited ozone induced neutrophil numbers at 3 h and 20–24 h after ozone SD-282 significantly inhibited ozone-induced increases in phosphorylated p38 MAPK expression, and in cyclooxygenase-2 (COX-2), interleukin-6 (IL-6) and IL-1β but not MIP-1α gene expression. We conclude that p38 MAPK is involved in ozone-induced airway hyperresponsiveness and lung neutrophilia. Inhibition of p38 MAPK with small molecule kinase inhibitors may be a means of reducing ozone-induced inflammation and airway hyperresponsiveness.

Effect of Pressure Stress Applied to the Airway on Cough-Reflex Sensitivity in Guinea Pigs

We hypothesized that cough stress of the airway wall results in a self-perpetuating cough-reflex cycle in which antigen-induced increase in cough-reflex sensitivity results in pathologic cough, and the cough in turn further amplifies cough-reflex sensitivity. To examine cough-reflex sensitivity in an experimental animal model. We developed an experimental guinea pig model in which airway collapse similar to that in cough was induced by rapid negative pressure applied to the airway of artificially ventilated animals. We examined the influence of this stimulus on cough-reflex sensitivity to inhaled capsaicin and bronchoalveolar lavage (BAL) cell components. After the termination of artificial ventilation, the number of coughs due to capsaicin was measured, and BAL was performed. Capsaicin cough-reflex sensitivity and the number of BAL neutrophils were increased 6 hours after stimulus application, decreasing to control levels by 24 hours. Cough-reflex sensitivity or BAL cell components were not changed in the absence of stimulus application. The number of BAL neutrophils correlated significantly with the number of coughs. Hydroxyurea inhibited the stimulus-induced increase in the number of coughs and airway neutrophil accumulation. Our findings suggest that cough itself is a traumatic mechanical stress to the airway wall that induces neutrophilic airway inflammation and cough-reflex hypersensitivity. Cough stress to the airway wall results in a self-perpetuating cough-reflex cycle.

Involvement of MMP-12 and phosphodiesterase type 4 in cigarette smoke-induced inflammation in mice

The aim of the present study was to characterise a mouse model of airways inflammation induced by cigarette smoke and to compare it with a lipopolysaccharide (LPS) model with regards to the efficacy of a PDE4 inhibitor (cilomilast), a corticosteroid (dexamethasone) and macrophage metalloelastase (MMP)-12 gene deletion. Cigarette smoke exposure for 3 days induced a time-dependent airway neutrophilia associated with an increased level of keratinocyte-derived chemokine (KC), macrophage inflammatory protein (MIP)-2, MIP-1alpha and MMP-9 in the bronchoalveolar lavage (BAL). LPS exposure also induced an increase in the number of neutrophils in BAL. Studies in MMP-12-/- mice showed that in contrast to the smoking model, MMP-12 did not have a critical role in LPS-induced inflammation. Both cilomilast and dexamethasone blocked LPS-induced neutrophilia in a dose-dependent manner. Cilomilast inhibited cigarette smoke-induced neutrophilia and MIP-1alpha, but only 10 mg.kg(-1) of dexamethasone was effective. Both anti-inflammatory treatments had no effect on the levels of KC and MIP-2 in the BAL. Although the inflammatory response was very similar in the smoking model and LPS, the pharmacological modulation and the MMP-12 gene deletion highlighted the differences in the mechanisms involved. Furthermore, the cigarette smoke model seemed to better represent the situation described in chronic obstructive pulmonary disease patients. In conclusion, these differences underline the importance of using an acute smoke-exposure model to investigate potential new treatments for chronic obstructive pulmonary disease.