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Acute (LPS) and chronic airways inflammation blunts phagocytosis in vivo: Evidence of phenotype switching to a dendritic cell phenotype
Abstracts S205
J ALLERGY CLIN IMMUNOL VOLUME 115, NUMBER 2
Acute (LPS) and Chronic Airways Inflammation Blunts Phagocytosis In Vivo: Evidence of Phenotype Switching to a Dendritic Cell Phenotype N. Alexis; Center Environmental Med & Lung Biology, University NC, Chapel Hill, NC. RATIONALE: We recently reported that phagocytosis of airway cells is blunted in healthy volunteers following LPS inhalation (5ug, ~30,000 EU), and that blunting is related to the magnitude of the neutrophil influx. We investigated whether phagocytosis is altered in disease states marked by “chronic” neutrophilic inflammation (COPD N=7, CF N=18) as compared to “acute” inflammation following LPS inhalation in normals (N=9), and if phagocytes undergo phenotype switching to a non-phagocytic dendritic cell (DC) phenotype. METHODS: Phagocytosis (IgG-opsonized zymosan particles) and DC surface phenotypes (HLA-DR, CD80, CD86) were analyzed on induced sputum and BALF (CF) cells by flow cytometry. RESULTS: Neutrophilia was present 6h following LPS challenge and in COPD and CF airways. Phagocytosis (Mac, Mono, PMNs) was significantly (p<0.05) blunted following LPS (20,000 EU) challenge (vs. pre-exposure) and constitutively decreased (p<0.05) in COPD (vs. healthy) and CF (vs. non-CF). HLA-DR and CD86 expression (MFI) were significantly elevated (p<0.05) on Mac and Mono following LPS, and the proportion of Mono co-expressing CD80/CD86 was significantly increased (p<0.05) following both LPS challenge and in COPD subjects. GM-CSF, a potent DC maturation agent, was significantly elevated (p<0.05) following LPS challenge. In vitro, sputum phagocytes incubated with supernatants from stimulated (PMA) blood PMNs showed decreased phagocytosis compared to un-stimulated supernatants. CONCLUSION: Both acute (LPS) and chronic (COPD, CF) airways inflammation blunt phagocytosis, and activated neutrophils likely contribute to this effect. Airway phagocytes appear to undergo phenotype switching to a less phagocytic, antigen presenting DC phenotype. This may enhance T-cell responses and ultimately sensitize the airway to subsequently inhaled allergic stimuli. Funding: NIH 1RO1 HL66559; NIH 2RO1 HL62624; US EPA: CR824915; “Supp
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Inflammation and Remodelling of Distal Airways in a Murine Model of Chronic Experimental Asthma M. Wegmann1, H. Fehrenbach2, A. Ferhenbach2, T. Held1, C. Schramm3, H. Garn1, H. Renz1; 1Department of clinical chemistry and molecular
819
diagnostics, hospital of the philipps-university marburg, Marburg, GERMANY, 2Clinic of pulmonary medicine, hospital of the philipps-university marburg, Marburg, GERMANY, 3First poly clinic, hospital of the johannes gutenberg-university mainz, Mainz, GERMANY. RATIONALE: This study was aimed to analyze whether chronic inhalative allergen challenges lead to morphological and physiological changes in distal airways comparable to human bronchial asthma. METHODS: BALB/c mice were systemically sensitized to Ovalbumin / Al(OH)3 followed by OVA aerosol challenges on two consecutive days per week for 12 weeks. Broncho alveolar lavage fluid, lung histology and lung function were analyzed. RESULTS: In chronically challenged mice, progressive tissue inflammation was observed in proximal as well as distal airways. The cellular influx was characterized by lymphocytes and eosinophils. However, in comparision to acute allergic inflammation lymphocytes predominated airway inflammation. In contrast, intraluminal inflammation decreased over time. These changes were accompanied by markedly increased levels of transforming growth factor- (TGF-) in BAL fluid. Goblet cell hyperplasia and subepithelial fibrosis occurred throughout the airway tree. An increased number of myofibroblasts occurred both in proximal and in distal airways. In terms of lung function, these mice developed persistent bronchial hyperresponsiveness and progressive airflow limitation. CONCLUSION: Our data clearly demonstrate that chronic aerosol allergen challenges in mice lead to a phenotype representative to asthma pathology in distal as well as in proximal airways. Funding: BMBF Infection With Litosomoides Sigmodontis Suppresses Allergen-Induced Sensitization And Pulmonary Inflammation in a Murine Asthma Model A. Dittrich1, A. Erbacher2, F. Diesner1, B. Ahrens1, D. Quarcoo1, W. H. Hoffmann2, E. H. Hamelmann1; 1Pediatric Pneumology and Immunology, Charite University Hospital, Berlin, Germany, Berlin, GERMANY, 2Institute for Tropical Medicine, University of Tuebingen, Tuebingen, GERMANY. RATIONALE: Numerous epidemiological studies have shown an inverse relationship between helminth infections and the manifestation of atopic diseases. Yet, the immunological mechanisms governing this phenomenon remain unclear. To further delineate this question, we analyzed the effects of infection with a filarial parasite on allergeninduced sensitization and airway disease in a murine model of bronchial asthma. METHODS: Balb/c mice received an intraperitoneal (ip) implantation of female Litosomoides sigmodontis (Lsigmo) parasites (5/mouse) on day 1. Mice were sensitized via two ip injections (day 12/26) and intranasally challenged (day 40/41) with ovalbumin. Airway reactivity (body plethymography, day 43), allergen sensitization (antigen-specific serum immunoglobulins, antigen-specific proliferation and cytokine production) and airway inflammation (bronchoalveolar lavage cell differentiation, histology) were analyzed on day 44 of the protocol. RESULTS: Infection with Lsigmo significantly suppressed all aspects of the asthmatic phenotype: airway reactivity was reduced by appoximately 30%, antigen-specific IgG and IgE production was suppressed 4-20 fold, and airway eosinophilia was reduced to levels of negative controls. Antigen-specific recall proliferation and cytokine production (IL-4, IL-5 and IL-10) by spleen cells also showed reduction to background levels. Moreover, suppression of mitogen-induced proliferative responses pointed towards a generalized suppression rather than an allergen-specific mechanism. CONCLUSIONS: Parasitic infection prior to allergen sensitization and airway challenges nearly completely prevented the induction of allergen sensitization, airway inflammation and hyperreactivity by a mechanism independent of IL-10. The model will allow to further dissect the immunological mechanisms underlying the inverse correlation between helminth infections and atopic diseases and may point towards novel therapeutic approaches. Funding: Charite, University Medicine of Berlin
820
MONDAY
IL-10 Transfected Lactoccocus Lactis Prevent Food Allergy in a Mouse Model of Food-Induced Anaphylaxis P. A. Eigenmann, C. P. Frossard; Department of Pediatrics, University Hospital, Geneva, SWITZERLAND. RATIONALE: Sensitization to foods early in life is common in atopic children. It predisposes to food allergy as well as to other allergic diseases. Our aim was to explore the effect of Lactococcus Lactis in order to prevent food-induced anaphylaxis in a mouse model of food allergy. METHODS: Four to six-weeks old C3H/HeJ mice were gavaged with 109PFU of Lactococcus lactis (LL), or with an mIL-10 transfected Lactococcus Lactis strain (mIL10LL) for 3 days prior to four oral sensitizations with beta-lactoglobulin (BLG). BLG-specific IgE, IgG1, and IgA in the feces were measured, and one week after the fourth sensitization, the mice were challenged with BLG. RESULTS: Less mice pre-treated with LL and mIL10LL prior to BLG sensitization than sensitized mice without pre-treatment had symptoms of anaphylaxis upon antigen challenge (7/14 vs. 6/7). In addition, BLG-specific serum IgE titers were close to baseline in mIL-10LL (9600 +/- 1400 AU), and significantly decreased in LL treated mice (42.900 +/- 5100 AU) when compared to sensitized mice (62.900 +/- 4100 AU). IgG1 titers were similarly decrased in mIL10LL and LL-treated mice. BLG-specific titers in the feces were significantly increased in mIL10LL treated mice (1.281 +/- 596 AU vs. 235 +/-79 AU in sensitized mice). CONCLUSIONS: Pre-treatment with LL and mIL10LL prevents foodinduced anaphylaxis, reduces antigen-specific Th2-type Abs and generates an increased secretion of secretory IgA. This strategy might provide an interesting opportunity to prevent food allergy in young infants. Funding: Swiss National Scientific Fundation
817
J ALLERGY CLIN IMMUNOL VOLUME 115, NUMBER 2
Acute (LPS) and Chronic Airways Inflammation Blunts Phagocytosis In Vivo: Evidence of Phenotype Switching to a Dendritic Cell Phenotype N. Alexis; Center Environmental Med & Lung Biology, University NC, Chapel Hill, NC. RATIONALE: We recently reported that phagocytosis of airway cells is blunted in healthy volunteers following LPS inhalation (5ug, ~30,000 EU), and that blunting is related to the magnitude of the neutrophil influx. We investigated whether phagocytosis is altered in disease states marked by “chronic” neutrophilic inflammation (COPD N=7, CF N=18) as compared to “acute” inflammation following LPS inhalation in normals (N=9), and if phagocytes undergo phenotype switching to a non-phagocytic dendritic cell (DC) phenotype. METHODS: Phagocytosis (IgG-opsonized zymosan particles) and DC surface phenotypes (HLA-DR, CD80, CD86) were analyzed on induced sputum and BALF (CF) cells by flow cytometry. RESULTS: Neutrophilia was present 6h following LPS challenge and in COPD and CF airways. Phagocytosis (Mac, Mono, PMNs) was significantly (p<0.05) blunted following LPS (20,000 EU) challenge (vs. pre-exposure) and constitutively decreased (p<0.05) in COPD (vs. healthy) and CF (vs. non-CF). HLA-DR and CD86 expression (MFI) were significantly elevated (p<0.05) on Mac and Mono following LPS, and the proportion of Mono co-expressing CD80/CD86 was significantly increased (p<0.05) following both LPS challenge and in COPD subjects. GM-CSF, a potent DC maturation agent, was significantly elevated (p<0.05) following LPS challenge. In vitro, sputum phagocytes incubated with supernatants from stimulated (PMA) blood PMNs showed decreased phagocytosis compared to un-stimulated supernatants. CONCLUSION: Both acute (LPS) and chronic (COPD, CF) airways inflammation blunt phagocytosis, and activated neutrophils likely contribute to this effect. Airway phagocytes appear to undergo phenotype switching to a less phagocytic, antigen presenting DC phenotype. This may enhance T-cell responses and ultimately sensitize the airway to subsequently inhaled allergic stimuli. Funding: NIH 1RO1 HL66559; NIH 2RO1 HL62624; US EPA: CR824915; “Supp
818
Inflammation and Remodelling of Distal Airways in a Murine Model of Chronic Experimental Asthma M. Wegmann1, H. Fehrenbach2, A. Ferhenbach2, T. Held1, C. Schramm3, H. Garn1, H. Renz1; 1Department of clinical chemistry and molecular
819
diagnostics, hospital of the philipps-university marburg, Marburg, GERMANY, 2Clinic of pulmonary medicine, hospital of the philipps-university marburg, Marburg, GERMANY, 3First poly clinic, hospital of the johannes gutenberg-university mainz, Mainz, GERMANY. RATIONALE: This study was aimed to analyze whether chronic inhalative allergen challenges lead to morphological and physiological changes in distal airways comparable to human bronchial asthma. METHODS: BALB/c mice were systemically sensitized to Ovalbumin / Al(OH)3 followed by OVA aerosol challenges on two consecutive days per week for 12 weeks. Broncho alveolar lavage fluid, lung histology and lung function were analyzed. RESULTS: In chronically challenged mice, progressive tissue inflammation was observed in proximal as well as distal airways. The cellular influx was characterized by lymphocytes and eosinophils. However, in comparision to acute allergic inflammation lymphocytes predominated airway inflammation. In contrast, intraluminal inflammation decreased over time. These changes were accompanied by markedly increased levels of transforming growth factor- (TGF-) in BAL fluid. Goblet cell hyperplasia and subepithelial fibrosis occurred throughout the airway tree. An increased number of myofibroblasts occurred both in proximal and in distal airways. In terms of lung function, these mice developed persistent bronchial hyperresponsiveness and progressive airflow limitation. CONCLUSION: Our data clearly demonstrate that chronic aerosol allergen challenges in mice lead to a phenotype representative to asthma pathology in distal as well as in proximal airways. Funding: BMBF Infection With Litosomoides Sigmodontis Suppresses Allergen-Induced Sensitization And Pulmonary Inflammation in a Murine Asthma Model A. Dittrich1, A. Erbacher2, F. Diesner1, B. Ahrens1, D. Quarcoo1, W. H. Hoffmann2, E. H. Hamelmann1; 1Pediatric Pneumology and Immunology, Charite University Hospital, Berlin, Germany, Berlin, GERMANY, 2Institute for Tropical Medicine, University of Tuebingen, Tuebingen, GERMANY. RATIONALE: Numerous epidemiological studies have shown an inverse relationship between helminth infections and the manifestation of atopic diseases. Yet, the immunological mechanisms governing this phenomenon remain unclear. To further delineate this question, we analyzed the effects of infection with a filarial parasite on allergeninduced sensitization and airway disease in a murine model of bronchial asthma. METHODS: Balb/c mice received an intraperitoneal (ip) implantation of female Litosomoides sigmodontis (Lsigmo) parasites (5/mouse) on day 1. Mice were sensitized via two ip injections (day 12/26) and intranasally challenged (day 40/41) with ovalbumin. Airway reactivity (body plethymography, day 43), allergen sensitization (antigen-specific serum immunoglobulins, antigen-specific proliferation and cytokine production) and airway inflammation (bronchoalveolar lavage cell differentiation, histology) were analyzed on day 44 of the protocol. RESULTS: Infection with Lsigmo significantly suppressed all aspects of the asthmatic phenotype: airway reactivity was reduced by appoximately 30%, antigen-specific IgG and IgE production was suppressed 4-20 fold, and airway eosinophilia was reduced to levels of negative controls. Antigen-specific recall proliferation and cytokine production (IL-4, IL-5 and IL-10) by spleen cells also showed reduction to background levels. Moreover, suppression of mitogen-induced proliferative responses pointed towards a generalized suppression rather than an allergen-specific mechanism. CONCLUSIONS: Parasitic infection prior to allergen sensitization and airway challenges nearly completely prevented the induction of allergen sensitization, airway inflammation and hyperreactivity by a mechanism independent of IL-10. The model will allow to further dissect the immunological mechanisms underlying the inverse correlation between helminth infections and atopic diseases and may point towards novel therapeutic approaches. Funding: Charite, University Medicine of Berlin
820
MONDAY
IL-10 Transfected Lactoccocus Lactis Prevent Food Allergy in a Mouse Model of Food-Induced Anaphylaxis P. A. Eigenmann, C. P. Frossard; Department of Pediatrics, University Hospital, Geneva, SWITZERLAND. RATIONALE: Sensitization to foods early in life is common in atopic children. It predisposes to food allergy as well as to other allergic diseases. Our aim was to explore the effect of Lactococcus Lactis in order to prevent food-induced anaphylaxis in a mouse model of food allergy. METHODS: Four to six-weeks old C3H/HeJ mice were gavaged with 109PFU of Lactococcus lactis (LL), or with an mIL-10 transfected Lactococcus Lactis strain (mIL10LL) for 3 days prior to four oral sensitizations with beta-lactoglobulin (BLG). BLG-specific IgE, IgG1, and IgA in the feces were measured, and one week after the fourth sensitization, the mice were challenged with BLG. RESULTS: Less mice pre-treated with LL and mIL10LL prior to BLG sensitization than sensitized mice without pre-treatment had symptoms of anaphylaxis upon antigen challenge (7/14 vs. 6/7). In addition, BLG-specific serum IgE titers were close to baseline in mIL-10LL (9600 +/- 1400 AU), and significantly decreased in LL treated mice (42.900 +/- 5100 AU) when compared to sensitized mice (62.900 +/- 4100 AU). IgG1 titers were similarly decrased in mIL10LL and LL-treated mice. BLG-specific titers in the feces were significantly increased in mIL10LL treated mice (1.281 +/- 596 AU vs. 235 +/-79 AU in sensitized mice). CONCLUSIONS: Pre-treatment with LL and mIL10LL prevents foodinduced anaphylaxis, reduces antigen-specific Th2-type Abs and generates an increased secretion of secretory IgA. This strategy might provide an interesting opportunity to prevent food allergy in young infants. Funding: Swiss National Scientific Fundation
817