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Anti-Interleukin (IL)-9 Antibody Increases Induction of Oral Tolerance in Murine Allergic Rhinitis
Abstracts AB69
224
Anti-Interleukin (IL)-9 Antibody Increases Induction of Oral Tolerance in Murine Allergic Rhinitis
226
Autolysosomal Formation Is Required for Autophagy-Dependent IL-18 Release from Airway Epithelial Cells
Soo Whan Kim; The Catholic University Of Korea and Jihyun SHIN, The Catholic University Of Korea, Seoul, Korea. RATIONALE: The purpose of this study is to investigate the effects of anti-IL-9 antibody on oral tolerance in a mouse model of allergic rhinitis. METHODS: BALB/c mice were divided into 4 groups: control, allergic rhinitis (AR), oral tolerance (OT) , and OT with anti-IL-9 antibody (OT + IL9AB) group. All mice except control group were sensitized with ovalbumin (OVA) three times for two weeks consecutively. After two weeks, mice in OT and OT + IL9AB group underwent immunotherapy by feeding of OVA. During the immunotherapy, mice in OT + IL9AB group were injected with purified anti-mouse IL-9 antibody. All sensitized mice were challenged intranasally with OVA. Allergic symptoms, eosinophil counts in nasal mucosa and serum OVA-specific IgE were measured. Interferone-g, IL-4, IL-17, IL-10, TGF-ß, T-bet, GATA-3, ROR- g t, PU.1 and Foxp3 mRNA expression in nasal mucosa determined by real-time polymerase chain reaction. Flow cytometry of CD4+CD25+Foxp3+ T cells in spleen was analyzed. RESULTS: In OT and OT + IL9AB group, symptom score, serum OVAspecific IgE, eosinophils, IL-4, PU.1 and GATA-3 mRNA were decreased (p<0.05), and IL-10, Foxp3 mRNA, and CD4+CD25+Foxp3+ T cells were increased compared with those in AR group (p<0.05). In OT + IL9AB group, symptom score and serum OVA-specific IgE were lower than those in OT group (p<0.05). Foxp3 mRNA and CD4+CD25+Foxp3+ T cells were higher than those in OT group (p<0.05). CONCLUSIONS: Administration of anti-IL-9 antibody increased the induction of tolerance in a mouse model of allergic rhinitis. These results indicate that anti-IL-9 antibody have immunomodulatory effect on immune tolerance in allergic rhinitis model.
Hiroki Murai, MD, PhD1, Shintaro Okazaki, MD1, Hisako Hayashi, MD, PhD1, Akiko Kawakita, MD1, Motoko Yasutomi, MD, PhD1, Sanjiv Sur, MD2, Yusei Ohshima, MD, PhD1; 1University of Fukui, Fukui, Japan, 2 University of Texas Medical Branch, Galveston, TX. RATIONALE: We have previously reported that IL-18 release from airway epithelial cells in response to Alternaria extract (ALT-E) is depend on autophagy activation but not on caspase 1 activation. LPS has been shown to activate caspase 1 and autophagy and to induce oxidative stress in macrophages through TLR4. Here, we compared LPS- and ALT-E-stimuli, and analyzed the precise roles of activating processes of autophagy in IL18 release from airway epithelial cells. METHODS: After A549 or NHBE cells were stimulated with ALT-E or LPS, IL-18 release was measured by ELISA. Autophagosome and autolysosome formations were measured by western blotting analysis using LC3 and p62 antibodies, respectively. To assess the involvement of oxidative stress in the IL-18 release, the cells were pre-treated with antioxidants, N-acetyl cysteine (NAC) and MG132 for 1 h prior to stimulation with ALT-E. RESULTS: Stimulation with ALT-E but not with LPS induced IL-18 release from AECs. NAC and MG132 ameliorated the ALT-E-induced IL18 release. Both ALT-E and LPS provoked the formation of LC3-II, which occurred in early phase of autophagy activation. ALT-E stimulation facilitated the degradation of p62, which arose in the late phase of autophagy activation, whereas LPS did not affects p62 degradation. CONCLUSIONS: Stimulation with ALT-E induced oxidative stress, which was involved in IL-18 release as well as autophagy activation. The autolysosomal formation during autophagy activation seems essential for the IL-18 release from airway epithelial cells.
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Reduced Nasal Brain Derived Neurotrophic Factor in Aspirin Exacerbated Respiratory Disease
Michele Pham, MD1, Rachel Baum, BS1, David Broide, MB, ChB, FAAAAI1, Andrew White, MD, FAAAAI2, Taylor Doherty, MD, FAAAAI1; 1University of California San Diego, La Jolla, CA, 2Scripps Clinic, Division of Allergy, Asthma and Immunology, San Diego, CA. RATIONALE: Aspirin-exacerbated respiratory disease (AERD) is a complex syndrome of eosinophilic sinus inflammation and asthma characterized by hypersensitivity reactions to COX-1 inhibition. Neurotrophins may contribute to inflammatory responses as previous studies have revealed an elevation in brain derived neutrotrophic factor (BDNF) levels in allergic airway disease. Further, TSLP and IL-33 have known roles in promoting eosinophilic tissue inflammation. However, levels of IL-33, TSLP, and BDNF in AERD have not been reported. METHODS: Nasal lavage (17 patients), blood (32 patients), and urine (45 patients) samples were obtained from AERD patients. Samples from normal controls (11 patients) were also obtained. For AERD patients, samples were collected at the initial visit, after aspirin desensitization, and greater than 30 days after aspirin desensitization. ELISAs were performed with urine, serum, and nasal lavage for BDNF, IL33, and TSLP. RESULTS: Only BDNF levels in nasal lavage samples were different and showed reduced levels (p50.0005) in AERD patients compared with normal controls. There were no differences in BDNF, IL33, or TSLP levels pre and post aspirin desensitization. CONCLUSIONS: Recent data suggests upregulation of BDNF in severe asthma and previous studies have demonstrated elevated BDNF in chronic sinusitis. In AERD, our data demonstrates lower BDNF levels in nasal secretions. There are a variety of factors that may affect BDNF which include concomitant medications and the unique inflammatory nature of AERD. It is also possible that the reduction in BDNF is a result of counterregulatory factors in the disease process or related to anosmia found in these patients.
MD2 Facilitates Pollen and Cat Dander-Induced Innate and Allergic Airway Inflammation
Koa Hosoki, MD, PhD, Toshiko Itazawa, MD, PhD, Istvan Boldogh, PhD, Sanjiv Sur, MD; University of Texas Medical Branch, Galveston, TX. RATIONALE: The NHANES study identified several pollens and cat dander as the most common allergens that induce allergic sensitization and allergic diseases. We recently reported that ragweed pollen extract (RWPE) requires TLR4 to stimulate CXCL-mediated innate neutrophilic inflammation that facilitates allergic sensitization and airway inflammation. Myeloid differentiation protein-2 (MD2) is a TLR4 coreceptor, but its role in pollen and cat dander-induced innate and allergic inflammation has not been critically evaluated. To elucidate the role of MD2 in inducing pollen and cat dander-induced innate and allergic airway inflammation. METHODS: TCMNull (TLR4Null, CD14Null, MD2Null), TLR4Hi, TCMHicells and human bronchial epithelial cells with siRNA-induced downregulation of MD2 were stimulated with RWPE, other pollen allergic extracts, or cat dander extract (CDE), and activation of NF-kB and/or secretion of the NF-kB-dependent CXCL8 were quantified. Wild type (WT) mice or mice with siRNA knockdown of lung MD2 were challenged intranasally with RWPE or CDE, and innate and allergic inflammation were quantified. RESULTS: RWPE stimulated MD2-dependent NF-kB activation and CXCL secretion. Likewise, Bermuda, rye, timothy, pigweed, Russian thistle, cottonwood, walnut and CDE stimulated MD2-dependent CXCL secretion. RWPE and CDE challenge induced MD2-dependent, CD14independent innate neutrophil recruitment. RWPE induced MD2-dependent allergic sensitization and airway inflammation. CONCLUSIONS: MD2 plays an important role in induction of allergic sensitization to cat dander and common pollens relevant to human allergic diseases.
SATURDAY
J ALLERGY CLIN IMMUNOL VOLUME 137, NUMBER 2
224
Anti-Interleukin (IL)-9 Antibody Increases Induction of Oral Tolerance in Murine Allergic Rhinitis
226
Autolysosomal Formation Is Required for Autophagy-Dependent IL-18 Release from Airway Epithelial Cells
Soo Whan Kim; The Catholic University Of Korea and Jihyun SHIN, The Catholic University Of Korea, Seoul, Korea. RATIONALE: The purpose of this study is to investigate the effects of anti-IL-9 antibody on oral tolerance in a mouse model of allergic rhinitis. METHODS: BALB/c mice were divided into 4 groups: control, allergic rhinitis (AR), oral tolerance (OT) , and OT with anti-IL-9 antibody (OT + IL9AB) group. All mice except control group were sensitized with ovalbumin (OVA) three times for two weeks consecutively. After two weeks, mice in OT and OT + IL9AB group underwent immunotherapy by feeding of OVA. During the immunotherapy, mice in OT + IL9AB group were injected with purified anti-mouse IL-9 antibody. All sensitized mice were challenged intranasally with OVA. Allergic symptoms, eosinophil counts in nasal mucosa and serum OVA-specific IgE were measured. Interferone-g, IL-4, IL-17, IL-10, TGF-ß, T-bet, GATA-3, ROR- g t, PU.1 and Foxp3 mRNA expression in nasal mucosa determined by real-time polymerase chain reaction. Flow cytometry of CD4+CD25+Foxp3+ T cells in spleen was analyzed. RESULTS: In OT and OT + IL9AB group, symptom score, serum OVAspecific IgE, eosinophils, IL-4, PU.1 and GATA-3 mRNA were decreased (p<0.05), and IL-10, Foxp3 mRNA, and CD4+CD25+Foxp3+ T cells were increased compared with those in AR group (p<0.05). In OT + IL9AB group, symptom score and serum OVA-specific IgE were lower than those in OT group (p<0.05). Foxp3 mRNA and CD4+CD25+Foxp3+ T cells were higher than those in OT group (p<0.05). CONCLUSIONS: Administration of anti-IL-9 antibody increased the induction of tolerance in a mouse model of allergic rhinitis. These results indicate that anti-IL-9 antibody have immunomodulatory effect on immune tolerance in allergic rhinitis model.
Hiroki Murai, MD, PhD1, Shintaro Okazaki, MD1, Hisako Hayashi, MD, PhD1, Akiko Kawakita, MD1, Motoko Yasutomi, MD, PhD1, Sanjiv Sur, MD2, Yusei Ohshima, MD, PhD1; 1University of Fukui, Fukui, Japan, 2 University of Texas Medical Branch, Galveston, TX. RATIONALE: We have previously reported that IL-18 release from airway epithelial cells in response to Alternaria extract (ALT-E) is depend on autophagy activation but not on caspase 1 activation. LPS has been shown to activate caspase 1 and autophagy and to induce oxidative stress in macrophages through TLR4. Here, we compared LPS- and ALT-E-stimuli, and analyzed the precise roles of activating processes of autophagy in IL18 release from airway epithelial cells. METHODS: After A549 or NHBE cells were stimulated with ALT-E or LPS, IL-18 release was measured by ELISA. Autophagosome and autolysosome formations were measured by western blotting analysis using LC3 and p62 antibodies, respectively. To assess the involvement of oxidative stress in the IL-18 release, the cells were pre-treated with antioxidants, N-acetyl cysteine (NAC) and MG132 for 1 h prior to stimulation with ALT-E. RESULTS: Stimulation with ALT-E but not with LPS induced IL-18 release from AECs. NAC and MG132 ameliorated the ALT-E-induced IL18 release. Both ALT-E and LPS provoked the formation of LC3-II, which occurred in early phase of autophagy activation. ALT-E stimulation facilitated the degradation of p62, which arose in the late phase of autophagy activation, whereas LPS did not affects p62 degradation. CONCLUSIONS: Stimulation with ALT-E induced oxidative stress, which was involved in IL-18 release as well as autophagy activation. The autolysosomal formation during autophagy activation seems essential for the IL-18 release from airway epithelial cells.
225
227
Reduced Nasal Brain Derived Neurotrophic Factor in Aspirin Exacerbated Respiratory Disease
Michele Pham, MD1, Rachel Baum, BS1, David Broide, MB, ChB, FAAAAI1, Andrew White, MD, FAAAAI2, Taylor Doherty, MD, FAAAAI1; 1University of California San Diego, La Jolla, CA, 2Scripps Clinic, Division of Allergy, Asthma and Immunology, San Diego, CA. RATIONALE: Aspirin-exacerbated respiratory disease (AERD) is a complex syndrome of eosinophilic sinus inflammation and asthma characterized by hypersensitivity reactions to COX-1 inhibition. Neurotrophins may contribute to inflammatory responses as previous studies have revealed an elevation in brain derived neutrotrophic factor (BDNF) levels in allergic airway disease. Further, TSLP and IL-33 have known roles in promoting eosinophilic tissue inflammation. However, levels of IL-33, TSLP, and BDNF in AERD have not been reported. METHODS: Nasal lavage (17 patients), blood (32 patients), and urine (45 patients) samples were obtained from AERD patients. Samples from normal controls (11 patients) were also obtained. For AERD patients, samples were collected at the initial visit, after aspirin desensitization, and greater than 30 days after aspirin desensitization. ELISAs were performed with urine, serum, and nasal lavage for BDNF, IL33, and TSLP. RESULTS: Only BDNF levels in nasal lavage samples were different and showed reduced levels (p50.0005) in AERD patients compared with normal controls. There were no differences in BDNF, IL33, or TSLP levels pre and post aspirin desensitization. CONCLUSIONS: Recent data suggests upregulation of BDNF in severe asthma and previous studies have demonstrated elevated BDNF in chronic sinusitis. In AERD, our data demonstrates lower BDNF levels in nasal secretions. There are a variety of factors that may affect BDNF which include concomitant medications and the unique inflammatory nature of AERD. It is also possible that the reduction in BDNF is a result of counterregulatory factors in the disease process or related to anosmia found in these patients.
MD2 Facilitates Pollen and Cat Dander-Induced Innate and Allergic Airway Inflammation
Koa Hosoki, MD, PhD, Toshiko Itazawa, MD, PhD, Istvan Boldogh, PhD, Sanjiv Sur, MD; University of Texas Medical Branch, Galveston, TX. RATIONALE: The NHANES study identified several pollens and cat dander as the most common allergens that induce allergic sensitization and allergic diseases. We recently reported that ragweed pollen extract (RWPE) requires TLR4 to stimulate CXCL-mediated innate neutrophilic inflammation that facilitates allergic sensitization and airway inflammation. Myeloid differentiation protein-2 (MD2) is a TLR4 coreceptor, but its role in pollen and cat dander-induced innate and allergic inflammation has not been critically evaluated. To elucidate the role of MD2 in inducing pollen and cat dander-induced innate and allergic airway inflammation. METHODS: TCMNull (TLR4Null, CD14Null, MD2Null), TLR4Hi, TCMHicells and human bronchial epithelial cells with siRNA-induced downregulation of MD2 were stimulated with RWPE, other pollen allergic extracts, or cat dander extract (CDE), and activation of NF-kB and/or secretion of the NF-kB-dependent CXCL8 were quantified. Wild type (WT) mice or mice with siRNA knockdown of lung MD2 were challenged intranasally with RWPE or CDE, and innate and allergic inflammation were quantified. RESULTS: RWPE stimulated MD2-dependent NF-kB activation and CXCL secretion. Likewise, Bermuda, rye, timothy, pigweed, Russian thistle, cottonwood, walnut and CDE stimulated MD2-dependent CXCL secretion. RWPE and CDE challenge induced MD2-dependent, CD14independent innate neutrophil recruitment. RWPE induced MD2-dependent allergic sensitization and airway inflammation. CONCLUSIONS: MD2 plays an important role in induction of allergic sensitization to cat dander and common pollens relevant to human allergic diseases.
SATURDAY
J ALLERGY CLIN IMMUNOL VOLUME 137, NUMBER 2