Integrins in the Developing Lung

项目总结 人和小鼠的肺发育需要肺上皮细胞和 围绕细胞外基质(ECM)，但ECM指导的控制上皮细胞行为的机制 在肺中的作用仍不明确。在上皮组织中，整合素作为基底膜的受体。 组成胶原和层粘连蛋白(LMS)，其中LMS是对肺最重要的ECM蛋白 器官发生。肺上皮细胞通过整合素α3β1、α6β1和α6β4与肺泡巨噬细胞结合 报道称，这些整合素的突变会导致肺发育不良或新生儿肺气肿并发 异常的呼吸道，表明它们在人类发育性肺部疾病中发挥着重要作用。 为了研究LM结合整合素的作用，我们产生了肺上皮细胞特异性整合素缺失。 β1和α6亚基的缺失会导致明显的分枝缺陷和早期死亡。删除α3 仅造成轻微的呼吸道分支中断。β4缺陷小鼠表现出正常的分支，但令人惊讶的是， 对围产期也是致命的。β-4组织学检查显示有蛋白质物质充填于 呼吸道和纤毛缺乏，类似于α6缺陷小鼠，表明它们的死亡是由于呼吸道功能障碍造成的。 对α6缺陷肺测序数据的路径分析显示，骨形态发生蛋白信号转导中断，这是一个关键的 呼吸道分支的通路。在α6缺失的上皮细胞中，骨形态发生蛋白受体的表达增加，但骨形态发生蛋白 靶基因表达仍然显著降低，暗示含有α6的整合素参与了对 胎肺中的骨形态发生蛋白途径。与β4缺陷小鼠纤毛丧失一致，β4缺失的上皮细胞 与MCC末端分化相关的转录因子表达减少。作为一个关键组件， 半桥粒，α6β4控制与基底膜的紧密黏附，并与 细胞内角蛋白中间丝。角蛋白也为纤毛形成了顶端的支持网络，这表明 α-6-β-4对角蛋白组织的调控对肺内巨噬细胞的终末分化至关重要。加在一起， 这些发现表明：1)α6β1是可能通过骨形态发生蛋白的气道分支所需的主要整合素 2)α-6-β-4调节MCC的终末分化。根据初步数据，我们建议 假设含有α6的整合素是胎儿肺发育的关键整合素 BMP信号在多纤毛动物气道分支和终末分化过程中的调节 上皮细胞。 目的1：确定α6整合素调节肺分支的机制 形态发生。 目的2：明确α-6整合素在胚胎发育过程中调控骨形态发生蛋白信号的机制 肺部发育。 目的3：探讨α6β4整合素在呼吸道上皮细胞分化中的作用。

PROJECT SUMMARY Human and murine lung development requires the coordinated efforts of the lung epithelium with the surrounding extracellular matrix (ECM), but the ECM-directed mechanisms that govern epithelial cell behavior in the lung remain undefined. In epithelial tissues, integrins serve as receptors for the basement membrane components collagen and laminins (LMs), with LMs being the most important ECM protein for lung organogenesis. Lung epithelial cells bind to LMs through integrins α3β1, α6β1, and α6β4. Recent studies reported that mutations in these integrins cause pulmonary hypoplasia or neonatal emphysema complicated by abnormal airways, suggesting they play a major role in human developmental lung diseases. To investigate the role of LM-binding integrins, we generated lung epithelial specific integrin deletions. Deletion of both the β1 and α6 subunits resulted in marked branching defects and early death. Deletion of α3 caused only minor airway branching disruption. β4 deficient mice exhibited normal branching but, surprisingly, were also perinatal lethal. The β4 histological examination was notable for proteinaceous material filling the airways and lack of cilia, similar to α6 deficient mice, suggesting their demise resulted from airway dysfunction. Pathway analysis of α6 deficient lung sequencing data revealed disruptions in BMP signaling, a critical pathway for airway branching. BMP receptor expression was increased in α6-null epithelial cells, but BMP target gene expression remained markedly reduced, implicating α6-containing integrins in regulation of the BMP pathway in the fetal lung. Consistent with loss of cilia in β4 deficient mice, β4-null epithelial cells exhibited reduced expression of transcription factors linked to MCC terminal differentiation. As a critical component of hemidesmosomes, α6β4 controls tight adhesion to the basement membrane and connects with the intracellular keratin intermediate filaments. Keratin also forms a support network apically for cilia, suggesting that α6β4 regulates keratin organization critical for terminal differentiation of MCCs in the lung. Taken together, these findings indicate that: 1) α6β1 is the principal integrin required for airway branching likely through BMP signaling and 2) α6β4 regulates terminal differentiation of MCCs. Based on preliminary data, we propose the hypothesis that α6-containing integrins are critical integrins for fetal lung development through regulation of BMP signaling during airway branching and terminal differentiation of multi-ciliated epithelial cells. AIM 1: Determine the mechanisms whereby α6-containing integrins regulate lung branching morphogenesis. AIM 2: Define the mechanisms whereby α6-containing integrins regulate BMP signaling during fetal lung development. AIM 3: Identify the role of α6β4 integrin in airway epithelial cell differentiation.