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interleukin (IL)-5 generation in the airway
Abstracts $299
J ALLERGY CUN IMMUNOL VOLUME 111, NUMBER 2
chemokine receptors by bradykinin in human airway tissue in subjects with AR. Our findings indicate that bradykinin plays a potential role in focusing the al]ergic response in the airway in part through the upregulation of chemokine receptor expression. Funding: National Institutes of Health
lrl14 •
The Effect of a Rhinovirus(RV)16 Infection on Interferon (IFN)g/Inlerleukin (IL)-5 Generation in the Airway
W, W. Busse, R. F. Vrtis, K. A. Schwantes, J. E. Gern; Medicine, University of Wisconsin, Madison, Wl. RATIONALE: Although RV infections are major causes of asthma exacerbations, the airway sites and patterns (Thl vs Th2) of cytokine production to this respiratory virus are not established. M E T H O D S : To determine the pattern of cytokine response to a rhinovirus infection, six individuals were experimentally infected with RVI6. Nasal lavage, sputum, bronchioalveolar lavage (BAL) cells, and mucosal biopsies were obtained prior to and during the acute infections. Total RNA was extracted from these samples and real-time PCR was used to measure mRNA for [FN-yand IL-5, which were expressed on an IFNy/IL-5 ratio (high values = Thl, low values = Th2). RESULTS: RVI6 infections caused an increase in the IFN-7/IL-5 ratio which was measured at the time of acute symptoms and virus shedding. The pattern of this response was variable and dependent upon airway site of the samples. The mean IFN-y/IL-5 ratio in nasal cells was 874, in sputum cells 94, BAL cells 195, and biopsy tissue 126. CONCLUSIONS: These data indicate a predominant Thl response to the RVI6 infection, which was most dramatic in the upper airway, the primary site of the infection. These data also suggest that the cytokine response, eg, protective vs. inflammatory, may relate to the site of infection (upper vs. lower airway) or the constitutive composition of site-specific inflammatory ceils. Funding: Supported by NIH grant No. 1 POI A150500 and GCRC grant No. MOI RR03186
21 tions Oifferential T-bet and STAT4Expressionin NK Cell Subpopulawith Disparate Capacities for IFN-gamma Production L. A. Rosentha] 1, L. D. Mikus j, R. L. Sorkness z,2, R. F. Lemanske. Jr J; IUniversity of Wisconsin Medical School. Madison, WI, 2University of Wisconsin School of Pharmacy, Madison, Wl. RATIONALE: BN, but not F344, rats develop a post-bronchiolitis asthma-like phenotype after Sendal virus infection as weanlings, and reduced IFN-y production by BN, compared with F344, weanlings is important for this difference. Because Sendal virus-induced IFN-7 production by splenocytes is NK cell and IL-12 dependent, and BN, compared with F344, weanling NK cells have a reduced capacity for IFN-y secretion, we investigated mechanisms regulating NK cell IFN-y production, M E T H O D S : Two subpopulations of splenic NK cells (NKRP 1A+brightCD3- or NKR-PI A+aimCD3- cells), purified by flow cytometry from uninfected weanlings, were cultured +_-IL- 12 tbr 24 h, and IFN-y was measured by ELISA. T-bet and STAT4 levels were determined by immunoblot analysis. RESULTS: BN, compared with F344, weanlings had a reduced proportion of NKR-PI A+bright cells. NKR-P1A+bright cells produced significantly more IL-12-induced IFN-"/than did NKR-PIA+ dim cells (p < 0.01), with no observed strain differences. F344 NKR-PIA+ dim cells produced significantly more IL-12-induced IFN-~/than did BN NKR-PIA+ dim cells (p < 0.05), although these amounts were relatively small. NKR-PIA+bngh~ and NKR-PIA+ dim cells expressed comparable levels of T-bet (a transcription factor mediating IFN-y expression), whereas NKR-P 1A+ dim cells expressed markedly less STAT4 (a transcription factor mediating IL-12 signaling) than did NKRPIA+bnght ceils, with no observed strain differences. CONCLUSIONS: Capacity for IFN-yproduction by NK cell subpopulations was directly and selectively associated with the level of STAT4 expression. Skewing of NK cell population dynamics toward an NK cell subpopulation with a diminished capacity for 1FN-y production may be a developmentally regulated mechanism underlying reduced IFN-y responses in BN weanlings. Funding: NIH/NIAID
920 Identification afa Novel Metalloproteinase ADAM-8 in the 922 Modulation of Eosinophilia and Cytokinesof Bronchnalveolar Pathophysiologyof Experimental Asthma Lavage Fluid (BALF) by CpG ODN in a Mouse Model of EstabN. E. King, N. Zimmermann, S. M. Pope, A. Mishra, D. P. Witte, M. E. Rothenberg; Cincinnati Children's Hospital Medical Center, Cincinnati, OH. RATIONALE: We utilized an empirical approach involving transcript profiling of asthmatic lung RNA to elucidate the molecular mechanisms of asthma. We were struck by the over-expression of a novel membranebound metalloproteinase (MP) with disintegrin activity, ADAM-8, in the asthmatic lung. Notably, a recent human genome search identified ADAM-33 as an asthma-associated gene. Collectively, these findings provided the rationale/'or analyzing the expression and regulation of ADAM genes in asthma. METHODS: RNA transcript profiles from mice with experimental asthma were initially analyzed by microarray analysis using the Affymetrix U74Av2 DNA chip. Northern blot and in situ hybridization were used to analyze gene expression in wild type and STAT-6 deficient mice lbllowing allergen or Th2-cytokine treatment. RESULTS: Microarray analysis revealed upregulated expression of ADAM-8 (but not 22 other ADAMs) following ovalbumin (OVA) or Aspergillus antigen delivery to the lungs. There was a dose and time dependent induction of ADAM-8 following OVA challenge, with peak expression seen 18 hours after the second allergen challenge. Following transgenic or pharmacological delivery of IL-4 or 1L-13 to the lungs, ADAM-8 expression was strongly increased. Interestingly, ADAM-8 induction was only partially dependent on STAT-6 tbllowing OVA or IL4 treatment. In situ hybridization revealed tow expression of ADAM-8 mRNA in multiple lung parenchymal cells and induction of a stronger signal in peribronchial and perivascular cells following allergen challenge. CONCLUSIONS: ADAM-8 is an allergen-, IL-4-, and lL-13-induced gene in the asthmatic lung. These results suggest that allergic lung responses involve the interplay of diverse members of the ADAM family. Funding: NIH, ILSI. BWE HFSP, AlIA
lished Airway Inflammation S. Choir, Z, Peng2; I Department of Pediatrics, Dongguk University, Kyung-ju, REPUBLIC OF KOREA, 2Department of Pediatrics and Child Health, University of Manitoba, Winnipeg, MB, CANADA. RATIONALE: Oligodeoxynucleotides containing a CpG motif (CpG ODN), as potent inducers ofTh I immunity, are considered promising candidates for immune modulation in asthma. In this study we wanted to investigate the effect of CpG ODN on eosinophilia and cytokines of BALF in a mouse model established airway inflammation and the optimal route (systemic vs mucosal) of CpG ODN. We examined the difference of immunologic responses between CpG ODN and corticosteroids. METHODS: Female BALB/c mice, induced pulmonary allergic inflammation, were treated intranasally or intraperitoneally with CpG ODN and Dexamethasone. Allergen-specific antibody responses, cytokines (IL-4, IL-5, IL-12), and eosinophilic inflammation of the airways were investigated on BALF and splenocyte. RESULTS: CpG ODN effectively induced IL-12 (p<0.05) and inhibited IL-4 and IL-5 as well as eosinophilic inflammation (p<0.05) when CpG ODN was administered intranasally or intraperitoneally with allergen challenge. Therapy with corticosteroides, while effective inhibiting IL-5 generation, did not induced IL-12 in BALF (p<0.05). CONCLUSIONS: Systemic or mucosal administration of CpG ODN effectively stimulated the production of Thl cytokines and suppressed eosinophilic airway inflammation in contrast of corticosteroids and control ODN~ Thus, CpG ODN vaccination is a potentially useful approach for immunomodulation of established airway inflammation in a mouse model of asthma. Funding: Dongguk University Foundation
J ALLERGY CUN IMMUNOL VOLUME 111, NUMBER 2
chemokine receptors by bradykinin in human airway tissue in subjects with AR. Our findings indicate that bradykinin plays a potential role in focusing the al]ergic response in the airway in part through the upregulation of chemokine receptor expression. Funding: National Institutes of Health
lrl14 •
The Effect of a Rhinovirus(RV)16 Infection on Interferon (IFN)g/Inlerleukin (IL)-5 Generation in the Airway
W, W. Busse, R. F. Vrtis, K. A. Schwantes, J. E. Gern; Medicine, University of Wisconsin, Madison, Wl. RATIONALE: Although RV infections are major causes of asthma exacerbations, the airway sites and patterns (Thl vs Th2) of cytokine production to this respiratory virus are not established. M E T H O D S : To determine the pattern of cytokine response to a rhinovirus infection, six individuals were experimentally infected with RVI6. Nasal lavage, sputum, bronchioalveolar lavage (BAL) cells, and mucosal biopsies were obtained prior to and during the acute infections. Total RNA was extracted from these samples and real-time PCR was used to measure mRNA for [FN-yand IL-5, which were expressed on an IFNy/IL-5 ratio (high values = Thl, low values = Th2). RESULTS: RVI6 infections caused an increase in the IFN-7/IL-5 ratio which was measured at the time of acute symptoms and virus shedding. The pattern of this response was variable and dependent upon airway site of the samples. The mean IFN-y/IL-5 ratio in nasal cells was 874, in sputum cells 94, BAL cells 195, and biopsy tissue 126. CONCLUSIONS: These data indicate a predominant Thl response to the RVI6 infection, which was most dramatic in the upper airway, the primary site of the infection. These data also suggest that the cytokine response, eg, protective vs. inflammatory, may relate to the site of infection (upper vs. lower airway) or the constitutive composition of site-specific inflammatory ceils. Funding: Supported by NIH grant No. 1 POI A150500 and GCRC grant No. MOI RR03186
21 tions Oifferential T-bet and STAT4Expressionin NK Cell Subpopulawith Disparate Capacities for IFN-gamma Production L. A. Rosentha] 1, L. D. Mikus j, R. L. Sorkness z,2, R. F. Lemanske. Jr J; IUniversity of Wisconsin Medical School. Madison, WI, 2University of Wisconsin School of Pharmacy, Madison, Wl. RATIONALE: BN, but not F344, rats develop a post-bronchiolitis asthma-like phenotype after Sendal virus infection as weanlings, and reduced IFN-y production by BN, compared with F344, weanlings is important for this difference. Because Sendal virus-induced IFN-7 production by splenocytes is NK cell and IL-12 dependent, and BN, compared with F344, weanling NK cells have a reduced capacity for IFN-y secretion, we investigated mechanisms regulating NK cell IFN-y production, M E T H O D S : Two subpopulations of splenic NK cells (NKRP 1A+brightCD3- or NKR-PI A+aimCD3- cells), purified by flow cytometry from uninfected weanlings, were cultured +_-IL- 12 tbr 24 h, and IFN-y was measured by ELISA. T-bet and STAT4 levels were determined by immunoblot analysis. RESULTS: BN, compared with F344, weanlings had a reduced proportion of NKR-PI A+bright cells. NKR-P1A+bright cells produced significantly more IL-12-induced IFN-"/than did NKR-PIA+ dim cells (p < 0.01), with no observed strain differences. F344 NKR-PIA+ dim cells produced significantly more IL-12-induced IFN-~/than did BN NKR-PIA+ dim cells (p < 0.05), although these amounts were relatively small. NKR-PIA+bngh~ and NKR-PIA+ dim cells expressed comparable levels of T-bet (a transcription factor mediating IFN-y expression), whereas NKR-P 1A+ dim cells expressed markedly less STAT4 (a transcription factor mediating IL-12 signaling) than did NKRPIA+bnght ceils, with no observed strain differences. CONCLUSIONS: Capacity for IFN-yproduction by NK cell subpopulations was directly and selectively associated with the level of STAT4 expression. Skewing of NK cell population dynamics toward an NK cell subpopulation with a diminished capacity for 1FN-y production may be a developmentally regulated mechanism underlying reduced IFN-y responses in BN weanlings. Funding: NIH/NIAID
920 Identification afa Novel Metalloproteinase ADAM-8 in the 922 Modulation of Eosinophilia and Cytokinesof Bronchnalveolar Pathophysiologyof Experimental Asthma Lavage Fluid (BALF) by CpG ODN in a Mouse Model of EstabN. E. King, N. Zimmermann, S. M. Pope, A. Mishra, D. P. Witte, M. E. Rothenberg; Cincinnati Children's Hospital Medical Center, Cincinnati, OH. RATIONALE: We utilized an empirical approach involving transcript profiling of asthmatic lung RNA to elucidate the molecular mechanisms of asthma. We were struck by the over-expression of a novel membranebound metalloproteinase (MP) with disintegrin activity, ADAM-8, in the asthmatic lung. Notably, a recent human genome search identified ADAM-33 as an asthma-associated gene. Collectively, these findings provided the rationale/'or analyzing the expression and regulation of ADAM genes in asthma. METHODS: RNA transcript profiles from mice with experimental asthma were initially analyzed by microarray analysis using the Affymetrix U74Av2 DNA chip. Northern blot and in situ hybridization were used to analyze gene expression in wild type and STAT-6 deficient mice lbllowing allergen or Th2-cytokine treatment. RESULTS: Microarray analysis revealed upregulated expression of ADAM-8 (but not 22 other ADAMs) following ovalbumin (OVA) or Aspergillus antigen delivery to the lungs. There was a dose and time dependent induction of ADAM-8 following OVA challenge, with peak expression seen 18 hours after the second allergen challenge. Following transgenic or pharmacological delivery of IL-4 or 1L-13 to the lungs, ADAM-8 expression was strongly increased. Interestingly, ADAM-8 induction was only partially dependent on STAT-6 tbllowing OVA or IL4 treatment. In situ hybridization revealed tow expression of ADAM-8 mRNA in multiple lung parenchymal cells and induction of a stronger signal in peribronchial and perivascular cells following allergen challenge. CONCLUSIONS: ADAM-8 is an allergen-, IL-4-, and lL-13-induced gene in the asthmatic lung. These results suggest that allergic lung responses involve the interplay of diverse members of the ADAM family. Funding: NIH, ILSI. BWE HFSP, AlIA
lished Airway Inflammation S. Choir, Z, Peng2; I Department of Pediatrics, Dongguk University, Kyung-ju, REPUBLIC OF KOREA, 2Department of Pediatrics and Child Health, University of Manitoba, Winnipeg, MB, CANADA. RATIONALE: Oligodeoxynucleotides containing a CpG motif (CpG ODN), as potent inducers ofTh I immunity, are considered promising candidates for immune modulation in asthma. In this study we wanted to investigate the effect of CpG ODN on eosinophilia and cytokines of BALF in a mouse model established airway inflammation and the optimal route (systemic vs mucosal) of CpG ODN. We examined the difference of immunologic responses between CpG ODN and corticosteroids. METHODS: Female BALB/c mice, induced pulmonary allergic inflammation, were treated intranasally or intraperitoneally with CpG ODN and Dexamethasone. Allergen-specific antibody responses, cytokines (IL-4, IL-5, IL-12), and eosinophilic inflammation of the airways were investigated on BALF and splenocyte. RESULTS: CpG ODN effectively induced IL-12 (p<0.05) and inhibited IL-4 and IL-5 as well as eosinophilic inflammation (p<0.05) when CpG ODN was administered intranasally or intraperitoneally with allergen challenge. Therapy with corticosteroides, while effective inhibiting IL-5 generation, did not induced IL-12 in BALF (p<0.05). CONCLUSIONS: Systemic or mucosal administration of CpG ODN effectively stimulated the production of Thl cytokines and suppressed eosinophilic airway inflammation in contrast of corticosteroids and control ODN~ Thus, CpG ODN vaccination is a potentially useful approach for immunomodulation of established airway inflammation in a mouse model of asthma. Funding: Dongguk University Foundation