Airway eosinophilia is an important marker of asthma severity, risk of exacerbation, and response to therapy. Based upon the current understanding of the function of eosinophils (EOS), the preliminary data and studies accomplished over the previous funding cycle, form the basis for the hypothesis that EOS play a pivotal role in asthma pathophysiology by enhancing airway inflammation and remodeling. The studies in this PPG renewal application are designed to answer clinically important questions formulated to take maximum advantage of our collective expertise and resources. They will allow us to address critically, comprehensively and interactively the role of EOS in allergic airway inflammation using ex vivo and in vivo experiments in human subjects complemented, as appropriate, by focused use of cell lines and animal models. Project 1 will investigate key EOS functions including promoting IL-17 responses and enhancing fibroblast (Fb) function via expression of SEMA7A, which leads to promoting myoFb differentiation and generation of extracellular matrix (ECM) components. The Th2 skewed environment in the asthmatic airway promotes the expression of periostin, an important marker of allergic inflammation. The proposed studies in Project 2 will determine how this ECM protein interacts with EOS to enhance their survival, mediator release, and response to EOS-active cytokines. Studies will define the requirements for periostin generation in the airway and recognition by the EOS integrin alphaMBeta2. Finally in Project 3, the molecular mechanisms regulating the pro-fibrotic cytokine, TGF-Beta1 signaling in EOS and Fb will be investigated focusing on the role of Pin1 interaction with proximal Smads as well as PI-3-K and Akt. Our group discovered that up-regulation of isomerase Pin1 is an essential step in the TGF-Beta1 signaling pathway and seek to determine how Pin1 influences Smad6 and Smad3 including the role of Akt and PI-3-Kgamma in this process. It can be argued that few other centers, if any, around the world are capable of doing the clinical studies needed to retrieve airway EOS and at the same time have the experience necessary to critically dissect EOS biological functions to the same level of sophistication that our group is capable of doing. The collaborative studies among the three laboratories and the rich clinical research experience will allow us to move back and forth from ex vivo to in vivo experiments to better understand the role of EOS in modulating allergic airway inflammation and remodeling. Given the prominence of eosinophilic inflammation in a significant proportion of severe asthma patients, these advances will have direct implications for the patients most affected by this very common illness.
From these collaborative and integrated approaches, we will unveil novel insights into the pathobiology of asthma, with implications for other illnesses where eosinophils are known to be an important participant These advances will be crucial to focusing the quest for the next generation of asthma therapeutics with important implications for other eosinophilic diseases such as hypereosinophilic syndrome, Churg-Strauss Syndrome and eosinophilic esophagitis/gastritis. (End of Abstract) Project 1 - Role of Eosinophils in Airway Inflammation and Remodeling (Jarjour, Nizar N) RESUME AND SUMMARY OF DISCUSSION: This is a very innovative proposal that focuses on newly described pathways for eosinophil function in the asthma airway. The investigator and environment are well suited to the experiments. This is an outstanding project with high potential to advance the field in understanding the role of eosinophils, production of IL-1b, activation of TH17 and semaphonin production and the relationship of eosinophils to airway response and fibrosis. (End of Reviewers' Comments) Description (as provided by applicant): There is a fundamental gap in understanding how eosinophils (EOS) participate in inflammation and remodeling in asthmatic subjects. The lack of knowledge in this field is problematic to develop appropriate drugs to better treat a heterogenic disease such as asthma. The long-term goal is to understand the mechanisms by which EOS contribute to the development of asthma, particularly the more severe phenotypes. The objective in this Project 1 is to identify how activated EOS produce pro-inflammatory and remodeling factors that are relevant in allergic asthma. The central hypothesis is that IL-3 activates EOS to produce 1) the pro-inflammatory cytokine IL-1B and 2) the pro-fibrotic membrane protein, semaphorin7A (SEMA7A). We propose that IL-1 B drives the increase of the highly inflammatory cytokine, IL-17 in lymphocytes in the context of an allergic response. SEMA7A induces fibroblasts (Fb) differentiation toward myoFb, amplifying airway remodeling. Our preliminary data will allow us to test this hypothesis by pursuing three specific aims: 1) Determine the expression of IL-17 in the airways after segmental allergen challenge in patients with asthma. Measure IL-1B release from blood and airway EOS, and connect the release of IL-1B with IL-17 expression in vivo and with EOS ability to increase IL-17 by CD4+ T lymphocytes in vitro. 2) Define the mechanisms responsible for IL-3-induced IL-1B. The mechanisms analyzed include mRNA stability and IL-1B maturation through the inflammasome. 3) Analyze the expression and regulation of SEMA7A on blood and airway EOS and determine the effect of SEMA7A on human bronchial Fb. The approach is innovative addressing new functions attributed to EOS. The release of bioactive IL-1B by EOS, as well as the expression of SEMA7A on EOS, are both novel. The implication of EOS in IL-17 production has never been shown and the function of SEMA7A on human Fb is unknown. These mechanisms will be analyzed in IL-3-activated EOS in vitro or in vivo in the context of a segmental allergen challenge. This latter in vivo approach, which mimics natural allergic asthma, arguably sets our group apart in our abilities to analyze airway EOS in humans. The proposed research is significant because it is expected to advance our understanding of the role of EOS in asthma, and of EOS potential function in the development of severe asthma. Ultimately, the knowledge acquired by these studies, will help to define new potential targets for drugs in needs for patients refractory to current treatments. PUBLIC HEALTH RELEVANCE: We propose that IL-3 contributes to eosinophil function in asthma by 1) influencing CD4+ T cell production of the pro-inflammatory cytokine IL-17, and 2) inducing expression of the pro-fibrotic molecule semaphorin 7 A. The focus on IL-3 is a paradigm shift from current IL-5-targeted therapies. Understanding the cellular and bio-molecular mechanisms down-stream of eosinophil activation by IL-3 will reveal novel pathways with therapeutic implications in asthma and other eosinophil related diseases. (End of Abstract)
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