How nicotine delivered by Electronic Nicotine Delivery Systems (ENDS) affects the lung and its recovery from cigarette smoking

自从电子尼古丁输送系统（ENDS）进入市场以来， 经历了迅速增长的普及作为一种手段，提供尼古丁的用户在前所未有的高 浓度和纯度。几乎没有证据表明尼古丁的这种自我途径会产生什么后果- 给药对初次使用者或使用本产品帮助吸烟（CS）的人的健康的影响 停止此Merit应用程序旨在填补我们对这些问题的理解中的空白。这里的重点 将是如何肺形态和正常基因表达，包括一个明确的反应，吸入 屋尘螨过敏原受ENDS：nic暴露的影响，特别是在以前暴露于 香烟烟雾。有3个互动的目的来解决项目假设：尼古丁的使用作为交付 通过电子尼古丁给药系统（ENDS：nic），无论是在幼稚肺还是在停止使用 吸烟（CS）影响暴露肺的形态和免疫反应性， 使CS停止后的正常恢复过程复杂化。具体目标1。是否结束：nic暴露 影响肺形态学、粘液产生和初治肺和既往肺纤维化 暴露在CS中会有什么损害？ENDS的测量：对肺的影响将包括形态学变化 在肺泡腔中，粘蛋白产生（例如，Muc 5 b）和沉积，以及胶原沉积的变化（纤维化）。 结果将确定尼古丁的形式（盐与游离碱）及其相对浓度如何影响 这些参数。我们进一步假设ENDS：nic暴露（可能单独携带）会改变肺功能， 并且先前暴露于CS的小鼠将不会恢复到与允许的小鼠相同的程度。 在没有END：NIC使用和可能的载体的情况下恢复。具体目标2。ENDS：nic影响 未接触过香烟的小鼠的免疫功能以及从香烟烟雾到ENDS的过渡：nic暴露 阻碍免疫功能恢复？Nic递送强烈抑制肺嗜酸性粒细胞反应 吸入常见的屋尘螨过敏原。这种可靠的实验测量提供了 一个终点，以量化不同离子形式的尼古丁，其浓度和相关的电子 液体载体化合物。重点也将是对HDM的AM和特异性转录反应 在ENDS背景下从先前CS相关损伤恢复期间的肺细胞信号传导反应：nic 相对于没有产品使用以及ENDS：nic本身的影响。具体目标3。alpha 7是如何被调节的 受ENDS影响的肺泡巨噬细胞（AM）中的细胞信号传导机制：nic暴露？这些 实验将集中在肺泡巨噬细胞（AM），它构成了免疫细胞中的大多数。 支气管肺泡肺液（BALF）。新开发的方法，联合收割机AM富集从BALF的 ENDS：nic和CS暴露的小鼠和对用定义的细胞因子（IL-4+IL-10）处理的体外应答将被描述。 在细胞信号传导中间体的特异性抑制剂和细胞信号传导的正变构调节剂的存在下进行。 AM尼古丁的靶点，烟碱受体α 7。AM转录反应机制将是 使用RT-PCR以及RNA-Seq测量。 总的来说，这项研究将提供第一个全面的观点AM反应后，ENDS：nic 在从先前吸烟中恢复期间，ENDS：nic使用如何影响肺部。

Since their introduction to the marketplace, Electronic Nicotine Delivery Systems (ENDS) have experienced a rapid growth in popularity as a means to deliver nicotine to the user in unprecedented high concentrations and purity. There is little evidence pertaining to the consequences this route of nicotine self- administration has on the health of the naïve user or those who use this product to aid in cigarette smoking (CS) cessation. This Merit application is designed to fill gaps in our understanding of these issues. Here the focus will be on how the lung morphology and normal gene expression including a defined response to the inhaled house dust mite allergen is impacted by ENDS:nic exposure, especially in the lung previously exposed to cigarette smoke. There are 3 interactive Aims to address the Project Hypothesis: The use of nicotine as delivered by electronic nicotine delivery systems (ENDS:nic), both in the naïve lung and subsequent to cessation of cigarette smoking (CS), impacts the morphology and immune responsiveness of the exposed lung and complicates normal recovery processes following CS cessation. Specific Aim 1. Does ENDS:nic exposure impact lung morphology, mucin production and fibrosis in the naïve lung and the lung previously damaged by CS exposure? Measurement of ENDS:nic effects on the lung will include morphologic changes in alveolar spaces, mucin production (e.g., Muc5b) and deposition, and changes to collagen deposition (fibrosis). Results will determine how the form of nicotine (salt versus free-base) and its relative concentrations impact these parameters. We further hypothesize that ENDS:nic exposure (and possibly carrier alone) will alter lung morphology and that mice previously exposed to CS will not recover to the same extent as mice that are allowed to recover in the absence of END:nic use and possibly carrier. Specific Aim 2. Does ENDS:nic impact immune function in naïve mice and does the transition from cigarette smoke to ENDS:nic exposure impede immune functional recovery? ENDS:Nic delivery strongly suppresses the lung eosinophil response to inhalation of the common house dust mite (HDM) allergen. This reliable experimental measurement provides an end-point to quantify the impact by different ionic forms of nicotine, their concentration and the associated e- liquid carrier compounds. The focus will also be on transcriptional responsiveness to HDM by AM and specific lung cellular signaling responses during recovery from prior CS-associated damage in the context of ENDS:nic relative to no product use as well as the impact by ENDS:nic itself. Specific Aim 3. How are alpha7-modulated cell-signaling mechanisms in the alveolar macrophage (AM) affected by ENDS:nic exposure? These experiments will focus on alveolar macrophages (AM) which constitute the majority of immune cells in the bronchial alveolar lung fluids (BALF). Newly developed methods that combine AM enrichment from the BALF of ENDS:nic and CS exposed mice and in vitro response to treatment with defined cytokines (IL-4+IL-10) will be done in the presence of specific inhibitors of cell signaling intermediates and positive allosteric modulators of the AM target of nicotine, the nicotinic receptor alpha7. The AM transcriptional response mechanisms will be measured using RT-PCR as well as RNA-Seq. Collectively, this study will provide the first comprehensive view of AM responsiveness after ENDS:nic exposure and how ENDS:nic use impacts the lung during recovery from previous cigarette smoking.