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Hypoxic Induction of Inflammatory Responses in Chronic Sinusitis
Relationship Between Promoter Methylation of Multiple Genes and inflammatory Cell infiltration in Nasal Polyps c. li1, Y. Pang1, Q. Tao2, D. Wang1; 1Otolaryngology, National University of Singapore, Singapore, SINGAPORE, 2Chinese University of Hong Kong, Hong Kong, CHINA. RATIONALE: Since promoter hypermethylation of some genes was found in certain inflammatory diseases, we attempted to investigate whether these epigenetic alterations are related to the inflammatory cell infiltration (e.g., CD8+ and eosinophils) in nasal polyps (NPs). METHODS: NP tissues from 20 patients, and inferior turbinate (IT) biopsies of 11 subjects with no history of NP (controls) were obtained from the local population of Singapore. Hematoxylin and eosin staining and CD8 immunostaining were carried out in all samples. Methylation specific PCR (MSP) was applied to detect the methylation status of p16, RASSF1A, SOCS1, SHP1, TSLC1, E-cadherin and DAP in all samples. In addition, positive MSP results were confirmed by bisulfite genomic sequencing. RESULTS: For all samples, no methylation was found in p16, RASSF1A and SOCS1. In NP patients, promoter methylation in SHP1, TSLC1, E-cadherin and DAP-kinase was detected in 100%, 55%, 75% and 60%, respectively. In IT of the controls, methylation was detected in 100% of SHP1, 36% of TSLC1, 63% of both E-cadherin and DAP-kinase. NP patients with DAP-kinase methylation showed higher infiltration of CD8 T cells than those without (p<0.05), while methylation of E-cadherin and TSLC1 was not related to CD8 T cell percentage in NP or non-NP tissues. There was no significant difference between the methylation status of E-cadherin, DAP and TSLC1 and the infiltration of Eosinophils in NP or non-NP tissues. CONCLUSIONS: The methylation of SHP1 may be a tissue specific phenomenon, while methylation of the pro-apoptotic factor (DAP-kinase) may be involved in the CD8 T cell infiltration in NPs. Funding: National University of Singapore
284
Unique Genetic Expression in Nasal Mucosa and Blood of Patients with Nasal Polyposis and Inflammatory Nasal Conditions T. A. Miller1, J. Kukowska-Latallo2, A. M. Singer1, M. A. Pynnonen3, J. E. Terrell3, J. R. Baker, Jr.4; 1Department Of Internal Medicine, Division of Allergy & Clinical Immunology, University of Michigan Health System, Ann Arbor, MI, 2Michigan Nanotechnology Institute for Medicine & Biological Sciences, University of Michigan, Ann Arbor, MI, 3Department Of Otolaryngology, University of Michigan Health System, Ann Arbor, MI, 4Michigan Nanotechnology Institute for Medicine & Biological Sciences, University of Michigan Health System, Ann Arbor, MI. RATIONALE: Nasal Polyps (NP) represent a pathologic manifestation linked to a group of inflammatory nasal conditions. The exact cause of NP is unknown. We evaluated for a systemic genetic basis for NP by simultaneously analyzing nasal tissue and blood samples. METHODS: Patient with NP counseled to undergo surgery were recruited from the Allergy and Otolaryngology Clinics. Patients scheduled for nasal anatomic surgery were controls. All patients were given IRB-approved written information and gave informed consent to participate. A detailed questionnaire regarding medical history was administered to each patient. Medical records verified patient’s answers. Nasal tissue and whole blood was collected at the time of surgery. RNA was extracted by standard protocol. Clean-up was performed by RNeasy column. RNA was processed using Affymetrix HuGene133 Plus2.0 chips. RESULTS: Calculation of gene expression values were performed using robust multi-array average. Linear modeling revealed differentially expressed genes between NP and non-NP patients with a p-value of 0.05 or 2-fold change. The most upregulated gene was mammaglobin (SCGB2A), with a fold value of + 2.3. RT-PCR confirmed the presence of this gene in nasal tissue. Mammaglobin was not upregulated in blood. Interestingly, in subgroup analysis, Interferon alpha-Inducible Protein 27 (IFI27) was significantly downregulated (in nasal tissue and blood) in a patient with aspirin sensitivity (ASRD).
285
CONCLUSIONS: There is significant evidence to suggest a genetic cause for nasal polyposis. Our data confirm the differential expression of certain genes in the nasal tissue of patients with NP. The possibility of systemic genetic regulation among patients with NP warrants further investigation. Funding: Benz Research Grant Heterogeneity of Effects in the Appearance of Rhinitis and/or Asthma According to Gender and Atopy M. A. Tejedor, A. Rosado, C. Vila, M. Moro, M. D. Alonso, M. Mugica, M. Fernández-Rivas, T. Verdura; Unidad de alergia, FUNDACION HOSPITAL ALCORCON, ALCORCON, SPAIN. RATIONALE: Our aim has been to know how atopy and gender influence the asthma and rhinitis appearance simultaneously or in a solitary way. METHODS: Between 1998 and 2003 12,223 patients were diagnosed in our Unit of rhinitis or asthma. They were classified into patients with only rhinitis ®, rhinitis and asthma (R+A) and only asthma (A). An univariant analysis and a multinomial logistic regression were made to analyze the interaction of gender and atopy, controlling the age. RESULTS: In our non stratified population, among the patients with rhinitis, atopic patients had more risk to having asthma (OR 2.2), and among the patients with asthma, atopic patients had more risk to having rhinitis (OR 6.7). In both analysis the risk was higher in males (R+A against R, OR: males 2.6, females 1.9; R+A against A, OR: males 8, females 6.7). In the multivariant analysis, absence of atopy favoured the absence of asthma among patients with rhinitis, being the influence of gender very weak (OR: atopic males =1; non atopic females 0.53, atopic females 1.1, non atopic males 0.3). On the other hand, the absence of atopy was the most important factor which favoured the presence of A and not R, although the influence of gender being also weak (OR: atopic males =1, non atopic females 3.1, atopic females 1.3, non atopic males 2). CONCLUSIONS: In our population atopy favours the presence of both illnesses simultaneously (R+A), instead of only rhinitis or only asthma while the gender had only a weak influence.
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Hypoxic Induction of Inflammatory Responses in Chronic Sinusitis J. W. Steinke, S. B. Early, K. Hise, L. Borish; Asthma and Allergic Diseases Center, University of Virginia, Charlottesville, VA. RATIONALE: Chronic sinusitis is a noninfectious inflammatory disorder characterized by hyperplasia of immune cells and sinus tissue. Nasal polyps, which frequently complicate the disease, can occlude the opening to the sinus cavity decreasing the level of oxygen available to the sinus tissue. Hypoxia has been studied in many diseases and results in increased recruitment of inflammatory cells and release of cytokines. The role of hypoxia in chronic sinusitis is unknown. We hypothesize that hypoxia induces production of mediators that recruit cells into the sinus tissue and are involved in remodeling of the nasal mucosa. METHODS: We compared data from unstimulated nasal polyp-derived fibroblasts with cultured in hypoxic (10% O2) and anoxic (0% O2) environments. Changes in mRNA expression (semiquantitative PCR) and protein levels (ELISA) of cytokines and chemokines were measured along with changes in cellular proliferation. RESULTS: Hypoxic conditions did not change the proliferative capacity of fibroblasts, while anoxia led to a 40% decrease in cellular proliferation (p<0.05). Both hypoxia and anoxia led to increases in secretion of many cytokines including VEGF and IL-8. As a marker of remodeling, procollagen production was significantly increased under hypoxic conditions, but was decreased with anoxic conditions. CONCLUSIONS: Hypoxic conditions present in the sinus tissue increase production of proinflammatory and remodeling cytokines that contribute to the inflammation observed in sinusitis. Surgical intervention may temporarily help decrease inflammation by allowing reoxygenation of the sinus cavity and decrease the hypoxic induction of cytokines and remodeling factors. Funding: NIH
287
SATURDAY
Abstracts S73
J ALLERGY CLIN IMMUNOL VOLUME 117, NUMBER 2
284
Unique Genetic Expression in Nasal Mucosa and Blood of Patients with Nasal Polyposis and Inflammatory Nasal Conditions T. A. Miller1, J. Kukowska-Latallo2, A. M. Singer1, M. A. Pynnonen3, J. E. Terrell3, J. R. Baker, Jr.4; 1Department Of Internal Medicine, Division of Allergy & Clinical Immunology, University of Michigan Health System, Ann Arbor, MI, 2Michigan Nanotechnology Institute for Medicine & Biological Sciences, University of Michigan, Ann Arbor, MI, 3Department Of Otolaryngology, University of Michigan Health System, Ann Arbor, MI, 4Michigan Nanotechnology Institute for Medicine & Biological Sciences, University of Michigan Health System, Ann Arbor, MI. RATIONALE: Nasal Polyps (NP) represent a pathologic manifestation linked to a group of inflammatory nasal conditions. The exact cause of NP is unknown. We evaluated for a systemic genetic basis for NP by simultaneously analyzing nasal tissue and blood samples. METHODS: Patient with NP counseled to undergo surgery were recruited from the Allergy and Otolaryngology Clinics. Patients scheduled for nasal anatomic surgery were controls. All patients were given IRB-approved written information and gave informed consent to participate. A detailed questionnaire regarding medical history was administered to each patient. Medical records verified patient’s answers. Nasal tissue and whole blood was collected at the time of surgery. RNA was extracted by standard protocol. Clean-up was performed by RNeasy column. RNA was processed using Affymetrix HuGene133 Plus2.0 chips. RESULTS: Calculation of gene expression values were performed using robust multi-array average. Linear modeling revealed differentially expressed genes between NP and non-NP patients with a p-value of 0.05 or 2-fold change. The most upregulated gene was mammaglobin (SCGB2A), with a fold value of + 2.3. RT-PCR confirmed the presence of this gene in nasal tissue. Mammaglobin was not upregulated in blood. Interestingly, in subgroup analysis, Interferon alpha-Inducible Protein 27 (IFI27) was significantly downregulated (in nasal tissue and blood) in a patient with aspirin sensitivity (ASRD).
285
CONCLUSIONS: There is significant evidence to suggest a genetic cause for nasal polyposis. Our data confirm the differential expression of certain genes in the nasal tissue of patients with NP. The possibility of systemic genetic regulation among patients with NP warrants further investigation. Funding: Benz Research Grant Heterogeneity of Effects in the Appearance of Rhinitis and/or Asthma According to Gender and Atopy M. A. Tejedor, A. Rosado, C. Vila, M. Moro, M. D. Alonso, M. Mugica, M. Fernández-Rivas, T. Verdura; Unidad de alergia, FUNDACION HOSPITAL ALCORCON, ALCORCON, SPAIN. RATIONALE: Our aim has been to know how atopy and gender influence the asthma and rhinitis appearance simultaneously or in a solitary way. METHODS: Between 1998 and 2003 12,223 patients were diagnosed in our Unit of rhinitis or asthma. They were classified into patients with only rhinitis ®, rhinitis and asthma (R+A) and only asthma (A). An univariant analysis and a multinomial logistic regression were made to analyze the interaction of gender and atopy, controlling the age. RESULTS: In our non stratified population, among the patients with rhinitis, atopic patients had more risk to having asthma (OR 2.2), and among the patients with asthma, atopic patients had more risk to having rhinitis (OR 6.7). In both analysis the risk was higher in males (R+A against R, OR: males 2.6, females 1.9; R+A against A, OR: males 8, females 6.7). In the multivariant analysis, absence of atopy favoured the absence of asthma among patients with rhinitis, being the influence of gender very weak (OR: atopic males =1; non atopic females 0.53, atopic females 1.1, non atopic males 0.3). On the other hand, the absence of atopy was the most important factor which favoured the presence of A and not R, although the influence of gender being also weak (OR: atopic males =1, non atopic females 3.1, atopic females 1.3, non atopic males 2). CONCLUSIONS: In our population atopy favours the presence of both illnesses simultaneously (R+A), instead of only rhinitis or only asthma while the gender had only a weak influence.
286
Hypoxic Induction of Inflammatory Responses in Chronic Sinusitis J. W. Steinke, S. B. Early, K. Hise, L. Borish; Asthma and Allergic Diseases Center, University of Virginia, Charlottesville, VA. RATIONALE: Chronic sinusitis is a noninfectious inflammatory disorder characterized by hyperplasia of immune cells and sinus tissue. Nasal polyps, which frequently complicate the disease, can occlude the opening to the sinus cavity decreasing the level of oxygen available to the sinus tissue. Hypoxia has been studied in many diseases and results in increased recruitment of inflammatory cells and release of cytokines. The role of hypoxia in chronic sinusitis is unknown. We hypothesize that hypoxia induces production of mediators that recruit cells into the sinus tissue and are involved in remodeling of the nasal mucosa. METHODS: We compared data from unstimulated nasal polyp-derived fibroblasts with cultured in hypoxic (10% O2) and anoxic (0% O2) environments. Changes in mRNA expression (semiquantitative PCR) and protein levels (ELISA) of cytokines and chemokines were measured along with changes in cellular proliferation. RESULTS: Hypoxic conditions did not change the proliferative capacity of fibroblasts, while anoxia led to a 40% decrease in cellular proliferation (p<0.05). Both hypoxia and anoxia led to increases in secretion of many cytokines including VEGF and IL-8. As a marker of remodeling, procollagen production was significantly increased under hypoxic conditions, but was decreased with anoxic conditions. CONCLUSIONS: Hypoxic conditions present in the sinus tissue increase production of proinflammatory and remodeling cytokines that contribute to the inflammation observed in sinusitis. Surgical intervention may temporarily help decrease inflammation by allowing reoxygenation of the sinus cavity and decrease the hypoxic induction of cytokines and remodeling factors. Funding: NIH
287
SATURDAY
Abstracts S73
J ALLERGY CLIN IMMUNOL VOLUME 117, NUMBER 2