Mechanotransduction and Lung Alveolar Differentiation

力传导和肺泡分化

• 批准号: 7372106
• 负责人: JUAN R SANCHEZ-ESTEBAN
• 金额: $ 24.75万
• 依托单位: WOMEN AND INFANTS HOSPITAL-RHODE ISLAND
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-05-01 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7372106
• 关键词: Adult; Alveolar; Area; Biology; Breathing; Bronchopulmonary Dysplasia; Cell Differentiation process; Cell Maturation; Cells; Cleaved cell; Clinical; Condition; Conditioned Culture Media; Development; Differentiation and Growth; Endopeptidases; Epidermal Growth Factor Receptor; Epithelial; Epithelial Cells; Extremely Low Birth Weight Infant; Fetal Lung; Goals; Growth; Growth Factor; In Vitro; Integrins; Investigation; Knockout Mice; Ligand Binding Domain; Ligands; Ligation; Liquid substance; Lung; Mechanical Stimulation; Mechanical ventilation; Mechanics; Mediating; Membrane; Mesenchymal; Modeling; Modification; Morphogenesis; Movement; Pathway interactions; Peptide Hydrolases; Phenotype; Physiological; Play; Pneumonectomy; Process; Protein Kinase; Pulmonary Surfactants; Rattus; Regulatory Pathway; Role; Role playing therapy; Signal Transduction; Signaling Protein; Simulate; Source; Stretching; Techniques; Transgenic Mice; Type II Epithelial Receptor Cell; airway remodeling; autocrine; base; extracellular; fetal; in utero; lung development; lung injury; lung maturation; neutralizing antibody; novel strategies; postnatal; pressure; receptor; research study; response

DESCRIPTION (provided by applicant): Mechanical forces generated in utero by repetitive breathing movements and by fluid distension are essential to mammalian lung development. However, the mechanisms by which pulmonary cells sense and transduce mechanical signals are largely unknown. The long-term goals are to define how mechanical forces promote lung maturation. Epidermal growth factor receptor (EGFR) is critical for fetal lung development. Although past studies indicate that EGFR is important for differentiation of type II cells and stretch-mediated compensatory growth after pneumonectomy, the mechanisms by which EGFR is activated are not known. Our investigations have identified potential roles for the EGFR and specific integrins in stretch-induced fetal type II cell differentiation. Further studies have shown that this process may be mediated by force-induced release of EGFR ligands, since strain-induced type II cell differentiation was markedly inhibited when the ligand-binding domain of the EGFR was blocked with neutralizing antibodies. We also showed that conditioned medium from stretched cells promoted type II cell differentiation when added to unstretched cells. This hypothesis is further supported by experiments performed in fetal lambs, which demonstrate that lung fluid composition after tracheal ligation, and not just increase in intrapulmonary pressure, is critical to accelerate lung growth and differentiation. Therefore, the specific hypothesis of this application is that strain-induced differentiation of fetal type II cells is mediated via autocrine release of membrane-anchored EGFR ligands. Our Specific Aims are: 1) To identify EGFR ligands released by lung epithelial cells in response to mechanical strain that promote type II cell differentiation. 2) To analyze the signaling mechanisms by which mechanical stretch induces EGFR ligand release. 3) To demonstrate the physiological role of force-induced release of soluble growth factors in organotypic models of epithelial strain. Using the Flexercell Strain Unit apparatus, cell signaling techniques, modification of the proposed key signaling proteins by inducible or knockdown expression and EGFR and ADAM17 knockout mice, we will examine EGFR ligands released by force in fetal type II cells that promote lung differentiation. We will analyze the role of ADAM17 as the protease that cleaves membrane-anchored EGFR ligands after mechanical stimulation of integrin receptors. Finally, we will validate our in vitro findings using ex vivo fetal lung explant models. Based on the critical role played by mechanical forces during normal fetal lung development, the identification of key regulatory pathways activated by strain in fetal lungs may provide a unique opportunity to rescue the phenotype and to facilitate development of new approaches to accelerate lung maturation in clinical conditions where lung development is impaired, including pulmonary hypoplasia, bronchopulmonary dysplasia and postnatal lung growth in extremely-low-birth-weight infants.

描述（由申请人提供）：子宫内通过重复呼吸运动和液体膨胀产生的机械力对于哺乳动物肺部发育至关重要。然而，肺细胞感知和转导机械信号的机制在很大程度上尚不清楚。长期目标是确定机械力如何促进肺成熟。表皮生长因子受体（EGFR）对于胎儿肺部发育至关重要。尽管过去的研究表明 EGFR 对于 II 型细胞的分化和肺切除术后拉伸介导的代偿性生长很重要，但 EGFR 激活的机制尚不清楚。我们的研究已经确定了 EGFR 和特定整合素在拉伸诱导的胎儿 II 型细胞分化中的潜在作用。进一步的研究表明，这一过程可能是通过强制诱导 EGFR 配体释放来介导的，因为当 EGFR 的配体结合域被中和抗体阻断时，菌株诱导的 II 型细胞分化受到显着抑制。我们还表明，当将来自拉伸细胞的条件培养基添加到未拉伸细胞中时，可促进 II 型细胞分化。在胎羔中进行的实验进一步支持了这一假设，这些实验表明，气管结扎后的肺液成分，而不仅仅是肺内压的增加，对于加速肺生长和分化至关重要。因此，本申请的具体假设是，菌株诱导的胎儿II型细胞的分化是通过膜锚定的EGFR配体的自分泌释放来介导的。我们的具体目标是： 1) 鉴定肺上皮细胞响应促进 II 型细胞分化的机械应变而释放的 EGFR 配体。 2) 分析机械拉伸诱导EGFR配体释放的信号机制。 3）证明力诱导释放可溶性生长因子在上皮应变的器官模型中的生理作用。使用 Flexercell 菌株装置、细胞信号传导技术、通过诱导或敲低表达对拟议的关键信号传导蛋白进行修饰以及 EGFR 和 ADAM17 敲除小鼠，我们将检查胎儿 II 型细胞中强制释放的 EGFR 配体，以促进肺分化。我们将分析 ADAM17 作为蛋白酶在整联蛋白受体机械刺激后裂解膜锚定 EGFR 配体的作用。最后，我们将使用离体胎儿肺外植体模型验证我们的体外研究结果。基于机械力在正常胎儿肺部发育过程中发挥的关键作用，识别胎儿肺部应变激活的关键调节途径可能提供独特的机会来挽救表型，并促进开发新方法，以在肺发育受损的临床条件下加速肺成熟，包括肺发育不全、支气管肺发育不良和出生后肺生长。 极低出生体重婴儿。