Lineage Commitment of Mesenchymal Stem Cell in Allergen-induced Airway Remodeling

描述（由申请人提供）：哮喘是一种常见的慢性疾病，影响世界各地的许多儿童和成人。哮喘的预防和治疗的一个主要障碍是其病因的多样性和我们对生物学机制的认识不足。最近的研究已经改变了我们对哮喘的理解，将其作为一种纯粹的炎症性疾病，转变为一种炎症和结构成分同等参与的疾病。过敏原引起的早期上皮损伤允许环境过敏原进入气道组织，并可能导致慢性阶段气道重塑的发展。因此，在早期阶段靶向修复受损上皮的治疗可以预防病理性气道重塑并改善哮喘控制。间充质干细胞（Mesenchymal stem cells，MSCs）是一种具有多向分化潜能和旁分泌免疫调节功能的成体结缔组织祖细胞。在过敏原激发后，肺中的MSC显著增加，并可能参与气道修复/重塑。转化生长因子1（TGF）1）在募集干/祖细胞用于各种器官中的组织修复、再生和重塑中起重要作用。我们先前的研究提供了初步证据，即在过敏原诱导的哮喘模型中，气道中的MSC和活性TGF？1信号传导增加。此外，来自蟑螂提取物的条件培养基（ECM）刺激上皮诱导MSC迁移，而TGF β 1中和抗体拮抗这种迁移。最近的体外研究表明，MSC可以在TGF β 1存在下分化为上皮细胞。因此，该阶段将严格评估哮喘中TGF β 1信号转导对MSC的功能影响。假设：活性TGF？1是一种过敏原激活的内源性信使，其将骨髓来源的MSC募集到受损的气道，其分化为上皮细胞以在早期阶段修复受损的上皮或分化为成纤维细胞/肌成纤维细胞，从而在哮喘的慢性阶段促进纤维化气道重塑。目的1提出实验以确定活性TGF？1在哮喘中MSC向肺募集中的作用。我们将使用我们的基于气液界面（ALI）-ECM的细胞迁移系统确定TGF β 1在MSC迁移中的作用。我们将使用Nes-GFP小鼠进一步检查在哮喘的不同阶段内源性巢蛋白+MSC向变应原激发的气道的募集，然后使用TGF-β 1中和抗体或TGF-β 1受体I型（T-RI）抑制剂检查TGF-β 1在MSC募集中的作用。[Aim 2提出了实验来追踪哮喘急性和慢性阶段通过TGF ² 1信号传导在肺部招募的MSC的谱系定型/分化]。我们已经建立了一个可诱导的MSC谱系追踪小鼠模型（nestin-CreERT 2; ROSA 26-EGFP），我们将使用它来追踪急性和慢性过敏原诱导的哮喘模型气道中招募的MSC的谱系定型。这将为TGF-β 1信号传导和MSC在过敏性哮喘中的作用提供新的见解，并为新的治疗提供机会。

DESCRIPTION (provided by applicant): Asthma is a common chronic illness that affects many children and adults worldwide. A major obstacle to prevention and treatment of asthma has been its diverse etiologies and our inadequate understanding of the biological mechanisms. Recent studies have changed our understanding of asthma as a purely inflammatory disease to a disease in which both inflammatory and structural components are equally involved. Allergen caused epithelial damage in early stages allows environmental allergens access to the airway tissue and may lead to the development of airway remodeling in chronic stages. Therefore, therapies that target the repair of the damaged epithelium in early stages could prevent the pathological airway remodeling and improve asthma control. Mesenchymal stem cells (MSCs) are adult connective tissue progenitor cells with multi-lineage differentiation potential and poten paracrine immunomodulatory properties. MSCs are significantly increased in the lungs after allergen challenge and may participate in airway repair/remodeling. Transforming growth factor ¿1 (TGF¿1) plays an important role in the recruitment of stem/progenitor cells for tissue repair, regeneration, and remodeling in various organs. Our previous studies have provided initial evidence that MSCs and active TGF¿1 signaling are increased in airway in an allergen induced asthma model. Furthermore, conditioned medium (ECM) from cockroach extract challenged epithelium induces the migration of MSCs, while TGF¿1 neutralizing antibody antagonized this migration. Recent studies in vitro have shown that MSCs can differentiate into epithelial cells in the presence of TGF¿1. The stage is thus set to critically evaluate the functional effect on MSCs of TGF¿1signaling in asthma. HYPOTHESIS: Active TGF¿1 is an allergen-activated endogenous messenger that recruits bone marrow-derived MSCs to the injured airways, which differentiate into epithelial cells to repair the damaged epithelium in early stages or into fibroblasts/myofibroblasts contributing to fibrotic airway remodeling in chronic stage of asthma. Aim 1 proposes experiments to determine the role of active TGF¿1 in the recruitment of MSCs to the lung in asthma. We will determine the role of TGF¿1 in migration of MSCs using our Air-Liquid Interface (ALI)-ECM-based cell migration system. We will further examine the recruitment of endogenous nestin+MSCs to the allergen-challenged airways during different stages of asthma using Nes-GFP mice, followed by the examination of the role of TGF¿1 in the recruitment of MSCs using a TGF¿1 neutralizing antibody or TGF-¿ receptor type I (T¿RI) inhibitor. [Aim 2 proposes experiments to track the lineage commitment/differentiation of MSC recruited in lungs through TGF¿1 signaling during acute and chronic stages of asthma]. We have established an inducible MSC lineage tracing mouse model (nestin-CreERT2; ROSA26-EGFP) that we will use to track the lineage commitment of recruited MSCs in airway of acute and chronic allergen-induced models of asthma. This will provide novel insights into the role of TGF-¿1 signaling and MSCs in allergic asthma and offer an opportunity for novel therapies.