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Meta-analysis of Transcriptome Data using Pathway-based Approach Reveals Biologically Relevant Asthma Genes
AB136 Abstracts
516
SUNDAY
Increased Platelet Adherence to Leukocytes Results in Cysteinyl Leukotriene (cysLT) Overproduction in Aspirin Exacerbated Respiratory Disease (AERD) T. M. Laidlaw1, M. S. Kidder1, N. Bhattacharyya1, G. L. Milne2, J. A. Boyce1; 1Brigham and Women’s Hospital, Boston, MA, 2Vanderbilt University Medical Center, Nashville, TN. RATIONALE: Overproduction of cysLTs is a hallmark of AERD, but the mechanisms for their production are incompletely defined. Because leukocytes can provide the precursor leukotriene (LT)A4 to adherent platelets for transcellular conversion to the parent cysLT, LTC4, we hypothesized that platelet-dependent leukotriene metabolism contributes to cysLT production in AERD. METHODS: Blood, urine, and nasal polyp tissues were obtained from subjects with AERD and from aspirin-tolerant asthmatic and nonasthmatic controls. Circulating platelet-leukocyte aggregates were quantified cytofluorographically, as were the effects of adherent platelets on leukocyte expression of adhesion markers. Platelet contribution to granulocytederived leukotrienes in vitro was quantified by HPLC, by measuring release of 5-lipoxygenase products from freshly isolated or platelet-stripped granulocytes. Urinary LTE4 was quantified by mass spectrometry. RESULTS: Percentages of circulating leukocytes with adherent platelets were markedly higher in subjects with AERD than in control groups, and platelet-adherent leukocytes were abundant in their nasal polyp tissue. Subsets of platelet-adherent monocytes and granulocytes expressed significantly higher levels of several adhesion markers than did platelet nonadherent subsets. Adherent platelets contributed more than half of the leukotriene C4 synthase (LTC4S) activity found in peripheral blood granulocytes from subjects with AERD and primed them for 5-lipoxygenase activity and conversion of endogenous LTA4 to LTC4. Urinary LTE4 levels correlated strongly with percentages of neutrophil-platelet (r50.78) and eosinophil-platelet (r50.73) aggregates. CONCLUSIONS: The increased frequency of leukocyte-platelet aggregates in AERD permits both adherence to endothelial cells and augmented transcellular conversion of leukotrienes, and may be partly responsible for the tissue inflammation and systemic overproduction of cysLTs that characterize this disease.
517
MiR150 Transgenic Mice Exhibit Altered T Cell Response, Cytokine Profile and Lung Inflammation J. Wang, K. Li, G. Hellermann, S. Mohapatra, R. Lockey, S. Mohapatra; University of South Florida, Tampa, FL. RATIONALE: Regulatory T (Treg) cells play a pivotal role in regulating T cell response and inflammation in asthmatic lungs; however the mechanism of Treg generation and function is poorly understood. A micro-RNA profile in Tregs from asthmatic mice showed a significant decrease in miR-150 expression. MiRNA150 may thus be useful as an antiinflammatory treatment for asthma. Our goal was to determine the effect of miR-150 overexpression on T cell response and lung pathology in a mouse model of asthma. METHODS: Total RNA was isolated from Treg and non-Treg cells and subjected to miRNA profiling. Transgenic mice overexpressing miR-150 were generated by pronuclear injection. T cell responses, inflammation and airway hyperreactivity were measured in asthmatic mice given miR-150 chitosan nanoparticles. RESULTS: MiR-150 was significantly down-regulated during asthmatic inflammation in Treg and non-Treg CD4+ T cells and in Th1 and Th2 cells. Transgenic mice overexpressing miR-150 showed lower airway hyperreactivity, inflammatory cytokine production and NKT cells than WT. Overexpression of miR-150 delivered by chitosan nanoparticles inhibited lung inflammation and cytokines in asthmatic mice and did not cause adverse side effects. MiR-150 suppressed expression of Akt3, Cbl1 and Elk1 but up-regulated p53, inhibited cell proliferation and promoted apoptosis of Jurkat T cells. CONCLUSIONS: MiR-150 decreases inflammation in asthma by inhibiting cytokine production, inducing apoptosis and repressing cell growth by regulating critical genes including Akt, Elk, Cbl1 and p53.
J ALLERGY CLIN IMMUNOL FEBRUARY 2012
Deregulation of miR-150 may be involved in the pathogenesis of asthma. Thus, overexpressing miR-150 may be useful as a safe anti-inflammatory therapeutic strategy for attenuating lung inflammation.
518
Meta-analysis of Transcriptome Data using Pathway-based Approach Reveals Biologically Relevant Asthma Genes T. M. Baye, S. Amirisetty, G. Khurana Hershey; Cincinnati Children’s Hospital Medical Center, Cincinnati, OH. RATIONAL: Asthma is a disease of chronic airway inflammation that affects over 300 million individuals worldwide including 20 million in the U.S. While the causes of asthma are not fully understood, genetic differences may be partly responsible for the variations in asthma susceptibility. Using microarray experiments, many independent groups have identified differentially expressed genes (DEGs) between healthy versus asthmatic patients. However, there is little overlap in the DEG lists identified among different experiments due to variation from random noise, biological and experimental differences, and differences in the extraction and handling of RNA samples. Therefore, a statistics-based meta-analysis is necessary to identify a set of genes that are consistently dysregulated among multiple independent microarray studies. METHODS: A recent search of asthma-related expression data in the GEO dataset yielded 1379 experiments of which 1190 were humanspecific (http://www.ncbi.nlm.nih.gov/, accessed on September 2, 2011). In this study, we focused our meta-analysis and network/pathway analysis on the 5 microarray human data sets, which had individual experiment sample sizes of over 100. RESULTS: Of the 5534 genes analyzed, 550 (12%) were up-regulated and 456 (8%) were down-regulated. Many key genes including SERPINB2, CLCA1 and P2RY14 were up-regulated and MUC5AC/MUC5B mucins and C6 were down-regulated. These genes were involved in inflammatory diseases, mucus over-production, airway obstructions, immunological diseases, and hypersensitivity responses. CONCLUSIONS: Meta-analytic dataset and network/pathway analysis of publicly available databases provide a starting point to identify biologically plausible potential candidate genes linked to asthma and to explore the mechanistic basis for the observed expression patterns in asthma.
516
SUNDAY
Increased Platelet Adherence to Leukocytes Results in Cysteinyl Leukotriene (cysLT) Overproduction in Aspirin Exacerbated Respiratory Disease (AERD) T. M. Laidlaw1, M. S. Kidder1, N. Bhattacharyya1, G. L. Milne2, J. A. Boyce1; 1Brigham and Women’s Hospital, Boston, MA, 2Vanderbilt University Medical Center, Nashville, TN. RATIONALE: Overproduction of cysLTs is a hallmark of AERD, but the mechanisms for their production are incompletely defined. Because leukocytes can provide the precursor leukotriene (LT)A4 to adherent platelets for transcellular conversion to the parent cysLT, LTC4, we hypothesized that platelet-dependent leukotriene metabolism contributes to cysLT production in AERD. METHODS: Blood, urine, and nasal polyp tissues were obtained from subjects with AERD and from aspirin-tolerant asthmatic and nonasthmatic controls. Circulating platelet-leukocyte aggregates were quantified cytofluorographically, as were the effects of adherent platelets on leukocyte expression of adhesion markers. Platelet contribution to granulocytederived leukotrienes in vitro was quantified by HPLC, by measuring release of 5-lipoxygenase products from freshly isolated or platelet-stripped granulocytes. Urinary LTE4 was quantified by mass spectrometry. RESULTS: Percentages of circulating leukocytes with adherent platelets were markedly higher in subjects with AERD than in control groups, and platelet-adherent leukocytes were abundant in their nasal polyp tissue. Subsets of platelet-adherent monocytes and granulocytes expressed significantly higher levels of several adhesion markers than did platelet nonadherent subsets. Adherent platelets contributed more than half of the leukotriene C4 synthase (LTC4S) activity found in peripheral blood granulocytes from subjects with AERD and primed them for 5-lipoxygenase activity and conversion of endogenous LTA4 to LTC4. Urinary LTE4 levels correlated strongly with percentages of neutrophil-platelet (r50.78) and eosinophil-platelet (r50.73) aggregates. CONCLUSIONS: The increased frequency of leukocyte-platelet aggregates in AERD permits both adherence to endothelial cells and augmented transcellular conversion of leukotrienes, and may be partly responsible for the tissue inflammation and systemic overproduction of cysLTs that characterize this disease.
517
MiR150 Transgenic Mice Exhibit Altered T Cell Response, Cytokine Profile and Lung Inflammation J. Wang, K. Li, G. Hellermann, S. Mohapatra, R. Lockey, S. Mohapatra; University of South Florida, Tampa, FL. RATIONALE: Regulatory T (Treg) cells play a pivotal role in regulating T cell response and inflammation in asthmatic lungs; however the mechanism of Treg generation and function is poorly understood. A micro-RNA profile in Tregs from asthmatic mice showed a significant decrease in miR-150 expression. MiRNA150 may thus be useful as an antiinflammatory treatment for asthma. Our goal was to determine the effect of miR-150 overexpression on T cell response and lung pathology in a mouse model of asthma. METHODS: Total RNA was isolated from Treg and non-Treg cells and subjected to miRNA profiling. Transgenic mice overexpressing miR-150 were generated by pronuclear injection. T cell responses, inflammation and airway hyperreactivity were measured in asthmatic mice given miR-150 chitosan nanoparticles. RESULTS: MiR-150 was significantly down-regulated during asthmatic inflammation in Treg and non-Treg CD4+ T cells and in Th1 and Th2 cells. Transgenic mice overexpressing miR-150 showed lower airway hyperreactivity, inflammatory cytokine production and NKT cells than WT. Overexpression of miR-150 delivered by chitosan nanoparticles inhibited lung inflammation and cytokines in asthmatic mice and did not cause adverse side effects. MiR-150 suppressed expression of Akt3, Cbl1 and Elk1 but up-regulated p53, inhibited cell proliferation and promoted apoptosis of Jurkat T cells. CONCLUSIONS: MiR-150 decreases inflammation in asthma by inhibiting cytokine production, inducing apoptosis and repressing cell growth by regulating critical genes including Akt, Elk, Cbl1 and p53.
J ALLERGY CLIN IMMUNOL FEBRUARY 2012
Deregulation of miR-150 may be involved in the pathogenesis of asthma. Thus, overexpressing miR-150 may be useful as a safe anti-inflammatory therapeutic strategy for attenuating lung inflammation.
518
Meta-analysis of Transcriptome Data using Pathway-based Approach Reveals Biologically Relevant Asthma Genes T. M. Baye, S. Amirisetty, G. Khurana Hershey; Cincinnati Children’s Hospital Medical Center, Cincinnati, OH. RATIONAL: Asthma is a disease of chronic airway inflammation that affects over 300 million individuals worldwide including 20 million in the U.S. While the causes of asthma are not fully understood, genetic differences may be partly responsible for the variations in asthma susceptibility. Using microarray experiments, many independent groups have identified differentially expressed genes (DEGs) between healthy versus asthmatic patients. However, there is little overlap in the DEG lists identified among different experiments due to variation from random noise, biological and experimental differences, and differences in the extraction and handling of RNA samples. Therefore, a statistics-based meta-analysis is necessary to identify a set of genes that are consistently dysregulated among multiple independent microarray studies. METHODS: A recent search of asthma-related expression data in the GEO dataset yielded 1379 experiments of which 1190 were humanspecific (http://www.ncbi.nlm.nih.gov/, accessed on September 2, 2011). In this study, we focused our meta-analysis and network/pathway analysis on the 5 microarray human data sets, which had individual experiment sample sizes of over 100. RESULTS: Of the 5534 genes analyzed, 550 (12%) were up-regulated and 456 (8%) were down-regulated. Many key genes including SERPINB2, CLCA1 and P2RY14 were up-regulated and MUC5AC/MUC5B mucins and C6 were down-regulated. These genes were involved in inflammatory diseases, mucus over-production, airway obstructions, immunological diseases, and hypersensitivity responses. CONCLUSIONS: Meta-analytic dataset and network/pathway analysis of publicly available databases provide a starting point to identify biologically plausible potential candidate genes linked to asthma and to explore the mechanistic basis for the observed expression patterns in asthma.