Biology of the Oral Epithelium of E-Cigarette Smokers

 描述（由申请人提供）：电子烟（EC）是电池供电的尼古丁输送装置，可雾化尼古丁。尽管EC的使用越来越多，但很少有人注意到其对人类健康可能产生的不利影响。理论上，风险与尼古丁本身和/或EC气雾剂中的推进剂或污染物有关。我们申请的假设是，慢性EC吸烟会扰乱口腔上皮细胞的生物学，口腔上皮细胞是暴露于吸入EC蒸气的第一个细胞群。有令人信服的证据支持这一假设：（1）EC蒸汽含有尼古丁，口腔上皮表达尼古丁受体，上皮暴露于尼古丁激活尼古丁通路;（2）EC蒸汽还含有可能影响口腔上皮生物学的污染物;和（3）体外研究表明EC蒸气改变上皮生物学，并且我们实验室产生的数据表明即使是短暂的，健康的非吸烟者急性暴露于EC蒸气诱导气道上皮转录组的显著变化，包括尼古丁和p53途径中的基因表达。使用横截面，队列比较EC吸烟者相比，年龄，性别和种族匹配的从不吸烟者，我们建议评估口腔上皮活检获得200 EC吸烟者和50名非吸烟者。EC研究队列将限于年轻人（年龄21-35岁），既往无吸烟史，但吸烟时间>6个月。由于细胞生物学的紊乱早在临床疾病之前就发生了，我们将在表观基因组（DNA甲基化）和转录组（mRNA和miRNA）水平上评估口腔上皮（特异性目的1）。随着共同专业人员的补充专业知识（R。Crystal，上皮生物学，J. Mezey，计算生物学），通过在表观基因组和转录组水平上整合分析，我们将鉴定被EC蒸气扰乱的基因/途径/群落（具体目标2）。基于我们的初步数据，我们将首先集中在尼古丁，p53和蛋白质和翻译后水平的相关途径。由于对EC吸烟对口腔上皮的影响知之甚少，我们无法预测哪些其他生物过程会受到影响，但EC蒸汽可能会影响与宿主防御和/或早期致癌相关的生物学。基于这些考虑，我们提出了两个具体目标。具体目标1：为了评估EC吸烟改变口腔上皮生物学的假设，将从慢性EC吸烟者和健康非吸烟者对照的颊粘膜收集活检，这些人都没有吸烟史。将生成全基因组数据集，以评估DNA甲基化以及编码和非编码RNA转录组。具体目标二：使用计算和实验方法，将评估特定目标1中生成的活检和全基因组数据集，以确定EC吸烟者口腔上皮中改变的基因，途径和基因群落，包括分析我们的试点研究中确定的两种途径（尼古丁，p53）。

 DESCRIPTION (provided by applicant): Electronic cigarettes (EC) are battery powered nicotine delivery devices that aerosolize nicotine. Despite the increasing use of EC, little attention has been paid to their possible adverse effects on human health. Theoretically, the risk relates to nicotine per se and/or the propellants or contaminants in the EC aerosol. The hypothesis underlying our application is that chronic EC smoking disorders the biology of the oral epithelium, the first cell population exposed to inhaled EC vapors. There is compelling evidence to support this hypothesis: (1) EC vapors contain nicotine, the oral epithelium expresses nicotine receptors, and exposure of epithelia to nicotine activates the nicotine pathway; (2) EC vapors also contain contaminants that potentially can affect oral epithelial biology; and (3) in vitro studies suggest that EC vapors modify epithelial biology and data generated by our laboratory demonstrates that even a brief, acute exposure of healthy nonsmokers to EC vapors induces significant changes in the airway epithelial transcriptome, including the expression of genes in the nicotine and p53 pathways. Using a cross-sectional, cohort-comparison of EC smokers compared to age, gender and ethnicity-matched never smokers, we propose to assess the oral epithelium obtained by biopsy from 200 EC smokers and 50 nonsmokers. The EC study cohort will be restricted to young adults (age 21-35 yr) with no prior history of tobacco smoking, but who have smoked EC for >6 months. With the knowledge that disordering of cell biology occurs long before clinical disease, we will evaluate the oral epithelium at the epigenome (DNA methylation) and transcriptome (mRNA and miRNA) levels (Specific Aim 1). With the complementary expertise of the co-PIs (R. Crystal, biology of the epithelium, J. Mezey, computational biology), by integrating analysis at the epigenome and transcriptome levels, we will identify gene(s)/pathways/communities disordered by EC vapors (Specific Aim 2). Based on our preliminary data, we will initially focus on the nicotine, p53 and related pathways at the protein and post-translational levels. Because there is so little known about the effects of EC smoking on the oral epithelium, we cannot predict which other biologic processes will be affected, but it is likely that EC vapors influence biology linked to host defens and/or early carcinogenesis. Based on these considerations, we propose 2 Specific Aims. Specific Aim 1: To evaluate the hypothesis that EC smoking alters oral epithelial biology, biopsies will be collected from the buccal mucosa of chronic EC smokers and healthy nonsmoker controls, none of whom have a history of smoking any tobacco products. Genome-wide datasets will be generated to assess DNA methylation and both coding and non-coding RNA transcriptomes. Specific Aim 2: Using both computational and experimental approaches, the biopsies and genome-wide datasets generated in Specific Aim 1 will be assessed to identify genes, pathways and gene communities altered in the oral epithelium of EC smokers, including analyses of two pathways (nicotine, p53) identified in our pilot study.