Molecular mechanism of alveolar injury caused by cigarette smoke

描述（由申请人提供）：我们之前对肺气肿中肺细胞凋亡失调的分子机制的研究使我们发现肺气肿患者肺部和暴露于香烟烟雾（CS）的细胞中鞘脂神经酰胺种类显着增加。我们的发现暗示神经酰胺是肺氧化应激和细胞凋亡的放大器。我们假设神经酰胺是cs诱导的氧化应激和肺泡细胞损伤的重要近端介质，因此可能是肺气肿的治疗靶点。因此，了解CS调节肺神经酰胺的机制是至关重要的。我们假设CS通过失调细胞膜内脂质相互作用刺激神经酰胺合成。此外，我们假设神经酰胺是香烟引起的早期肺泡肺损伤的介质，其特征是肺泡屏障功能的改变。我们将以以下具体目的来调查这些假设：为了确定CS上调神经酰胺的生物物理机制，我们将在膜脂双分子层模型中进行生物物理研究，并辅以细胞损伤模型；2.具体目标为了确定神经酰胺是cs诱导肺泡细胞屏障功能损伤的介质，我们将实时研究原代肺泡细胞单层的电容，并首次开发和利用完整大鼠肺双光子发射显微镜（TPEM）的活体应用。这项工作完成后，将阐明脂质双分子层膜的生物力学变化与导致神经酰胺信号传导的生化事件之间的联系，并通过首次实现肺活体TPEM成像，将其广泛应用于肺部健康和疾病的多种机制，从而加速肺生物医学研究。

DESCRIPTION (provided by applicant): Our previous investigations of the molecular mechanisms of dysregulated lung cell apoptosis in emphysema led us to identify a marked increase in sphingolipid ceramide species in the lungs of patients with emphysema and in cells exposed to cigarette smoke (CS). Our findings implicated ceramide as an amplifier of oxidative stress and apoptosis in the lung. We postulate that ceramide is an important proximal mediator of CS-induced oxidative stress and injury in lung alveolar cells and therefore a putative therapeutic target in emphysema. Therefore, understanding the mechanisms by which CS upregulates ceramides in the lung is of utmost importance. We hypothesize that CS stimulates ceramide synthesis by dysregulating lipid interactions within the cellular membranes. Furthermore, we hypothesize that ceramide is a mediator of early alveolar lung injury induced by cigarette smoke characterized by alterations of the alveolar barrier function. We will investigate these hypotheses with the following specific aims: Specific Aim 1. To determine the biophysical mechanism by which CS upregulates ceramides, we will conduct biophysical investigations in membrane lipid bilayer models complemented with cellular injury models; and Specific Aim 2. To establish that ceramides are mediators of the CS-induced injury of the alveolar cell barrier function, we will study the capacitance of primary alveolar cell monolayers in real-time and we will develop and utilize for the first time an intravital application of two-photon emission microscopy (TPEM) of the intact rat lung. When completed, this work will elucidate the connection between bio-mechanical changes in the lipid bilayer membrane and biochemical events leading to ceramide signaling and, by implementing intravital TPEM imaging of the lung for the first time, it will accelerate biomedical lung research by its broad application to multiple mechanisms of lung health and disease. PUBLIC HEALTH RELEVANCE: We will investigate how cigarette smoke triggers the increase in ceramides in lipid membranes which are lining the cells and important organelles within the cells and are the sites of ceramides synthesis. Besides using classical methods, we propose a novel technique of real time recording of barrier function in the intact lung utilizing an advanced imaging modality to study if the increase in ceramide is a necessary step in the earliest lung injury induced by cigarette smoke. Our work is expected to provide the rationale for a therapeutic strategy that targets ceramide in patients with COPD and will develop a new intravital molecular imaging modality of the whole lung which can be applied to the studies of all lung diseases.