Bronchoalveolar Cell Number Research Articles

Attenuating immune pathology using a microbial-based intervention in a mouse model of cigarette smoke-induced lung inflammation

BackgroundCigarette smoke exposure is the major risk factor for developing COPD. Presently, available COPD treatments focus on suppressing inflammation and providing bronchodilation. However, these options have varying efficacy in controlling symptoms and do not reverse or limit the progression of COPD. Treatments strategies using bacterial-derived products have shown promise in diseases characterized by inflammation and immune dysfunction. This study investigated for the first time whether a novel immunotherapy produced from inactivated Klebsiella (hereafter referred to as KB) containing all the major Klebsiella macromolecules, could attenuate cigarette smoke exposure-induced immune responses. We hypothesized that KB, by re-directing damaging immune responses, would attenuate cigarette smoke-induced lung inflammation and bronchoalveolar (BAL) cytokine and chemokine production.MethodsKB was administered via a subcutaneous injection prophylactically before initiating a 3-week acute nose-only cigarette smoke exposure protocol. Control mice received placebo injection and room air. Total BAL and differential cell numbers were enumerated. BAL and serum were analysed for 31 cytokines, chemokines, and growth factors. Lung tissue and blood were analysed for Ly6CHI monocytes/macrophages and neutrophils. Body weight and clinical scores were recorded throughout the experiment.ResultsWe demonstrate that KB treatment attenuated cigarette smoke-induced lung inflammation as shown by reductions in levels of BAL IFNγ, CXCL9, CXCL10, CCL5, IL-6, G-CSF, and IL-17. KB additionally attenuated the quantity of BAL lymphocytes and macrophages. In parallel to the attenuation of lung inflammation, KB induced a systemic immune activation with increases in Ly6CHI monocytes/macrophages and neutrophils.ConclusionsThis is the first demonstration that subcutaneous administration of a microbial-based immunotherapy can attenuate cigarette smoke-induced lung inflammation, and modulate BAL lymphocyte and macrophage levels, while inducing a systemic immune activation and mobilization. These data provide a foundation for future studies exploring how KB may be used to either reverse or prevent progression of established emphysema and small airways disease associated with chronic cigarette smoke exposure. The data suggest the intriguing possibility that KB, which stimulates rather than suppresses systemic immune responses, might be a novel means by which the course of COPD pathogenesis may be altered.

Airway hyperresponsiveness in FVB/N delta F508 cystic fibrosis transmembrane conductance regulator mice

BackgroundAirway hyperresponsiveness is a feature of clinical CF lung disease. In this study, we investigated whether the FVB/N ΔF508 CFTR mouse model has altered airway mechanics. MethodsMechanics were measured in 12–14week old FVB/N Cftrtm1Eur (ΔF508) mice and wildtype littermates using the FlexiVent small animal ventilator. Lung disease was assayed by immunohistochemistry, histology and bronchoalveolar lavage analysis. ResultsCftrtm1Eur mice presented with increased airway resistance, compared to wildtype littermates, in response to methacholine challenge. No differences in bronchoalveolar cell number or differential, or in tissue lymphocyte, goblet cell or smooth muscle actin levels were evident in mice grouped by Cftr genotype. The bronchoalveolar lavage of Cftrtm1Eur mice included significantly increased levels of interleukin 12(p40) and CXCL1 compared to controls. ConclusionWe conclude that the pulmonary phenotype of Cftrtm1Eur mice includes airway hyperresponsiveness in the absence of overt lung inflammation or airway remodeling.

Radiation-Induced Lung Response of AcB/BcA Recombinant Congenic Mice

The genetic factors that influence the development of radiotherapy-induced lung disease are largely unknown. Herein we identified a strain difference in lung response to radiation wherein A/J mice developed alveolitis with increased levels of pulmonary mast cells and cells in bronchoalveolar lavage while the phenotype in C57BL/6J mice was fibrosis with fewer inflammatory cells. To identify genomic loci that may influence these phenotypes, we assessed recombinant congenic (RC) mice derived from the A/J and C57BL/6J strains for their propensity to develop alveolitis or fibrosis after 18 Gy whole-thorax irradiation. Mouse survival, lung histopathology and bronchoalveolar lavage cell types were recorded. Informative strains for each of mast cell influx, bronchoalveolar cell numbers, alveolitis and fibrosis were identified. In mice with the A/J strain background, the severity of alveolitis correlated with increased mast cell numbers while in C57BL/6J background strain mice fibrosis was correlated with the percentage of neutrophils in lavage. The data for RC mice support the association of specific inflammatory cells with the development of radiation-induced lung disease and provide informative strains with which to dissect the genetic basis of these complex traits.

Route of nutritional supply influences local, systemic, and remote organ responses to intraperitoneal bacterial challenge.

The authors' aim was to investigate whether antecedent nutritional routes influence immune responses after surgical insult. Total parenteral nutrition (TPN) may influence host responses to infection. To the best of the authors' knowledge, however, no study has focused on the mechanisms underlying the influence of nutritional route on local, systemic, and remote organ (lung) responses after surgical insult. Sixty-eight rats were divided into TPN and total enteral nutrition (TEN) groups. The two groups received identical nutrients for 7 days and were then challenged intraperitoneally with 3 x 10(8) Escherichia coli. In the first experiment, the rats were observed for survival. In the second experiment, the rats were killed before (0 hours) challenge or 2 or 6 hours after challenge. Peritoneal exudative cells (PEC) and bronchoalveolar cells (BALC) were harvested and cultured in vitro. Colony-forming units of bacteria in the peritoneal lavage fluid (PLF) were determined. Tumor necrosis factor (TNF), interleukin-1 alpha (IL-1 alpha), interferon-gamma (IFN-gamma) levels in serum, PLF, bronchoalveolar lavage fluid (BALF), and cell culture supernatants were measured. The 48-hour survival rate was higher in TEN than in TPN rats. Local immunity was depressed in the TPN group. Bacterial colony counts in PLF were significantly higher in the TPN group than in the TEN group after challenge. The number of PECs was significantly lower, and at 2 hours, local cytokine (TNF and IL-1 alpha) responses were diminished in the TPN group compared with the TEN group at 2 hours. The number of PECs showed a significant positive correlation with levels of local cytokines in the TEN group but not in the TPN group. Elevation of local IFN-gamma was significant from 0 to 6 hours in the TEN group but not in the TPN group. In vitro production of TNF by PEC was impaired in the TPN rats before challenge. Remote organ (lung) responses were suppressed in the TPN group. The number of BALCs and the TNF levels in BALF declined significantly between 0 and 2 hours in the TEN group but not in the TPN group. Interferon-gamma levels in BALF were higher in the TEN group than in the TPN group at 2 hours. Systemic cytokine responses were disturbed in the TPN group. Production of systemic TNF was greater, but the IFN-gamma response was diminished in the TPN group compared with the TEN group after intraperitoneal bacterial challenge. Local, systemic, and remote organ (lung) immune responses to intraperitoneal bacterial challenge are suppressed in TPN-treated animals, leading to poor survival after challenge. Enteral nutrition before surgical insult may enhance host immune responses after the insult as compared to parenteral nutrition.

Antioxidant therapy partially blocks immune-induced lung fibrosis.

A mouse model of hypersensitivity pneumonitis was generated by challenge with a thermophilic actinomycete. Oxygen radical scavengers were administered to challenged mice: vitamin E at 1000 units daily, polyethylene glycol-superoxide dismutase (SOD) at 500 units daily, polyethylene glycol-catalase at 10,000 units daily, 1,3,dimethyl-2-thiourea (DMTU) at 2 mg daily, and the biomimetic SOD, copper(II) [diisopropyl salicylate]2 (CuDIPS) at 1 mg daily. At three weeks after actinomycete challenge, a 10-fold increase in bronchoalveolar (BAL) cell number was observed. Treatments with catalase or DMTU were without effect on the BAL cell number in challenged mice. However, infusion of vitamin E was associated with an increased BAL cell influx (15-fold increase at two and three weeks). Similarly, treatment with PEG-SOD and CuDIPS resulted in an increase in cell number at two and three weeks. PEG-SOD or CuDIPS treatment resulted in a strong neutrophilia, whereas control challenged mice had a cellular influx mostly of macrophages and lymphocytes. Vitamin E treatment of challenged mice led to an increased T lymphocyte recruitment at two and three weeks. In vitro studies showed that actinomycete challenge was associated with an enhancement of alveolar macrophage O2- release, which was blocked by PEG-SOD, vitamin E, or DSC treatment but was unaffected by catalase or DMTU treatment. In control challenged mice, there was a 25-fold increase in the BAL albumin concentration at two weeks. PEG-SOD, vitamin E, or CuDIPS treatment all decreased the albumin concentration; the three modulators also diminished lung fibrosis at two or three weeks, as seen by a decrease in lung hydroxyproline and collagen synthesis by lung fibroblasts. Examination of sections from lungs of challenged animals showed evidence of cellular infiltrates around the bronchi and the blood vessels. Challenged mice given continuous infusions of vitamin E, SOD, or CuDIPS had lung histological scores that were significantly lower than control challenged mice or challenged mice treated with catalase or DMTU. Thus, therapies based on O2- scavenging or treatment with a general antioxidant such as vitamin E may hold some promise in the treatment of hypersensitivity pneumonitis.

Virus-induced airway hyperresponsiveness in the guinea-pig: possible involvement of histamine and inflammatory cells.

1. Guinea-pig tracheal contractions by histamine and by the cholinoceptor agonist, arecoline, are significantly enhanced (30% and 20%, respectively), 96 h after intra-tracheal inoculation with Parainfluenza-3 (PI-3) virus. 2. The airway hyperresponsiveness in animals inoculated with virus coincides with a significant increase in the number of broncho-alveolar cells (82%), and in the albumin concentration (121%) in lung lavage fluid, relative to values obtained in guinea-pigs challenged with control solution. 3. The chemiluminescence production by isolated broncho-alveolar cells, obtained from virus-infected guinea-pigs 96 h after inoculation stimulated with PI-3 virus in vitro, is significantly reduced by 42% relative to broncho-alveolar cells obtained from animals inoculated with control solution. This diminution was not specific for stimulation by PI-3 virus since the chemiluminescence production was also significantly reduced by 30% in response to zymosan. 4. Pretreatment of the guinea-pigs with the anti-allergic drugs, oxatomide (2.5 mg kg-1) or nedocromil (2.5 mg kg-1), or the specific H1-histamine receptor antagonist, levocabastine (0.25 mg kg-1), administered intra-peritoneally twice a day for five successive days, inhibits the virus-induced airway hyperresponsiveness, suppresses the influx of broncho-alveolar cells and increase in albumin content, and corrects the reduced chemiluminescence production by broncho-alveolar cells in response to zymosan. 5. In contrast, the cyclo-oxygenase inhibitor, suprofen (5.0 mg kg-1), the 5-HT2 receptor antagonist, ketanserin (0.63 mg kg-1), or the Ca2+ overload blocker, flunarizine (2.5 mg kg-1) do not modify the above mentioned processes. 6. The platelet-activating factor receptor antagonist, WEB 2170 (10 mg kg-1), reduces virus-induced airway hyperresponsiveness and influx of broncho-alveolar cells into the lungs but does not attenuate the increase of albumin in the bronchial lavage fluid. 7. Guinea-pigs nebulized with histamine, twice a day (30 min) during 4 successive days, do not demonstrate an increased airway responsiveness, but instead show tachyphylaxis in response to histamine in vitro. In addition, no influx of inflammatory cells is found in these animals. 8. These results suggest that histamine does not directly increase the responsiveness of the guinea-pig trachea; however, histamine may be involved in a cascade of events leading to airway hyperresponsiveness after a viral infection, a process that could be related to an influx and/or an activation of broncho-alveolar cells after PI-3 virus stimulation.

Murine hypersensitivity pneumonitis: interleukin-4 administration partially abrogates the disease process

C57BL-6 inbred mice were given intranasal instillations of Faeni rectivirgula (150 μg/day, 3 days a week for 3 weeks) to produce a lung inflammatory reaction which mimics Farmers' lung in humans. Challenged mice developed a strong inflammatory response in their lungs, based on various markers (lung index, bronchoalveolar cell number, fibrosis). The effect of interleukin-4 (IL-4) was studied by infusing mice intraperitoneally with 100, 1000 or 10000 units of IL-4 weekly during the challenge period. It was shown that IL-4 infusion decreased the inflammatory response, as seen by a decreased lung index (1.7 in mice given F. rectivirgula and 10 3 U IL-4 and 1.31 in mice given antigen and 10 4 U IL-4 weekly versus 2.3 in mice instilled with F. rectivirgula). Interleukin-4 infusion also partially abrogated the F. rectivirgula-induced alveolitis, as seen by a decrease in cell numbers in the broncho-alveolar lavage (BAL) (1.3 sx 10 5 cells in saline-instilled mice; 8.3 × 10 5 cells in mice given 10 3 U IL-4 and F. rectivirgula; 3.2 × 10 5 cells in mice given antigen and 10 4 U IL-4; 1.8 × 10 6 cells in mice given F. rectivirgula only). Also, it was apparent that IL-4 administration could partially block the appearance of the fibrosis induced by F. rectivirgula (220 μg of hydroxyproline/lung in challenged mice; 170 μg/lung in challenged mice given 10 3 U IL-4; 131 μg/lung in mice given antigen and 10 4 U IL-4 and c. 100 μg/lung in control animals). Infusion of 10 2 U IL-4 weekly had no statistical effect on any marker of inflammation. Blocking of IL-4 activity with 11B11 hybridoma supernatant blocked the anti-inflammatory effect of IL-4. Bronchoalveolar macrophages from mice given F. rectivirgula and excipient, released substantial amounts of tumour necrosis factor alpha (TNFα) upon lipopolysaccharide (LPS) stimulation, and the release of TNFα was significantly reduced by IL-4 treatment. Overall, these results suggest that IL-4 may have anti-inflammatory properties which may have therapeutic values in the treatment of lung inflammatory diseases.

Usefulness of bronchoalveolar cell profile in early detection of canine lung allograft rejection

To evaluate the value of a bronchoalveolar cell profile in the early detection of canine lung allograft rejection, bronchoalveolar lavages were done serially in mongrel dogs before and after single lung transplantation. The dogs were divided into 3 groups. In group 1, neither donor nor recipient dogs were treated with cyclosporine. In group 2, only donors were treated with cyclosporine, orally at a single dose of 20 mg/kg/day for 3 days prior to single lung transplantation. In group 3, only recipients were treated with cyclosporine (20 mg/kg/day) for 9 days after single lung transplantation. A marked increase in the number of bronchoalveolar cells and their cell differentials, and of major histocompatibility complex (MHC) class II-positive cells obtained from the grafted lungs after lung transplantation, was seen in groups 1 and 2. The changes in bronchoalveolar cell profile obtained from the rejecting grafted lungs were significantly different from those obtained from the normal and native lungs (P less than 0.05). In group 3, the bronchoalveolar cell profile obtained from the grafted lungs did not significantly differ from those present in normal and native lungs during the period of cyclosporine treatment after lung transplantation. On various days after withdrawal of cyclosporine treatment, bronchoalveolar cell profiles obtained from the grafted lungs showed similar changes to those observed in groups 1 and 2. Abnormal changes in bronchoalveolar cell profiles obtained from the grafted lungs heralded the appearance of abnormalities detected by chest X-ray films. Our results indicate that serial measurements of bronchoalveolar cell profile may serve as a useful means for early detection of canine lung allograft rejection.

Parainfluenza‐3 induced hyperreactivity of the guinea pig trachea coincides with an increased number of bronchoalveolar cells.

Guinea pigs were inoculated intra-tracheally with bovine parainfluenza type 3 virus (PI-3) to investigate whether airway hyperreactivity was associated with an increase in the number of broncho-alveolar cells. Two days after saline or PI-3 inoculation no difference in histamine reactivity of the isolated tracheal spirals was observed. The number of broncho-alveolar cells was also not changed. However, 4 days after PI-3 inoculation the histamine-induced contraction of the tracheal spirals was increased by 45% and the number of inflammatory cells in the airways by 150% as compared with the control group. In conclusion, PI-3 infection of the guinea pig respiratory airways induces tracheal hyperreactivity which is associated with an increase in broncho-alveolar cells.

Decreased suppressor cell activity of alveolar macrophages in bronchial asthma.

In 8 allergic asthmatic patients and 12 healthy volunteers, we investigated the effects of alveolar macrophages (AM) on lymphoproliferative responses to polyclonal T-cell mitogens of autologous peripheral blood mononuclear cells (PBMC). The AM were obtained by bronchoalveolar lavage. Peripheral blood mononuclear cells (PBMC) and mitomycin-treated autologous bronchoalveolar cells (BAC) were cocultured at various AM-to-PBMC ratios, and were stimulated or not by phytohemagglutinin and concanavalin A. Half of AM expressed HLA-DR antigens in both the asthmatic and the control group. The BAC from normal subjects were able to modify the lymphoproliferative responses of autologous PBMC to cell mitogens. A dose-dependent effect was observed related to the number of BAC added to PBMC--suppressive at high ratios but enhancing at low ratios. In allergic bronchial asthma, there was a decreased suppressor cell activity of BAC. Among BAC the adherent cells were responsible for this effect. This abnormality could be a part of more general decreased functional activity of AM in allergic bronchial asthma.