Mutagenicity of tobacco smoke in human cell co-cultures

人类细胞共培养物中烟草烟雾的致突变性

• 批准号: 7146535
• 负责人: JOSEPH B GUTTENPLAN
• 金额: $ 14.54万
• 依托单位: NEW YORK UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-07-18 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7146535
• 关键词: culture; human; model; smoking; tissue /cell culture; tobacco; tobacco abuse

DESCRIPTION (provided by applicant): Tobacco smoking is a major cause of cancers at many sites, particularly the aerodigestive tract. For many years, filter and reduced tar cigarettes have been available, but have not resulted in reduced incidences of smoking-related cancers, because of compensatory smoking behavior. Recently, newer versions of "reduced-risk" cigarettes have been marketed, but their relative carcinogenicities are unknown. Cancer arises through multistage carcinogenesis, which involves the accumulation of genetic damage (such as mutations), over years, until a cell converts to cancer. Current assays for potential carcinogenesis in humans all have certain deficiencies. Few models employ normal human cells and mutagenesis is generally not an endpoint. To better understand risk from potential carcinogens, new model systems that can evaluate such risk are needed. In preliminary studies, we showed that in monolayer co-culture of normal human oral epithelial cells (NOE's) with a lacI rat reporter cell line (BB cells - that contain a lambda-based shuttle vector for detecting mutagenesis), increased mutagenesis induced by benzo(a)pyrene (BaP), relative to BB cells alone. This increased mutagenesis is most likely due to activation of BaP by the NOE's. We hypothesize that co-culturing NOE's with lacI reporter cells will allow detection of mutagenesis that is causally linked to metabolic activation of carcinogens by the normal human oral cells. To validate the model, mutagenesis induced by the two tobacco carcinogens (BaP and nitrosonornicotine) will be investigated. In Aim 1 we will establish and characterize a co- culture model composed of NOE's co-cultured with BB reporter cells. In Aim 2, this model will be used: 1) to compare mutagenesis induced by tobacco smoke condensate (TSC) from conventional cigarettes and recently-introduced "low-risk" cigarettes and 2) to determine inter-individual variation in the metabolism of these TSC's. Since human cells from a variety of organ sites can be grown in culture, the model has potential to become a flexible broad-based system to evaluate carcinogenicity risk of tobacco products in a number of human organs. A third aim will be a pilot study on the levels of cytochrome P- 450's in NOE's with and without the presence of TSC's, using gene expression arrays and confirmation by RT-PCR and western blotting. As NOE's are the very cells that are destined to develop into oral cancers, the model fills a void with the potential of being an organ specific technique to quantify mutagenesis, a major step in carcinogenesis. Clearly, it is preferable to determine their potential risk prospectively, rather than retrospectively.

描述（由申请人提供）：吸烟是许多部位癌症的主要原因，特别是呼吸消化道癌症。多年来，过滤嘴和低焦油香烟已经问世，但由于补偿性吸烟行为，并没有降低与吸烟相关的癌症的发病率。最近，新型“低风险”香烟已上市，但其相对致癌性尚不清楚。癌症是通过多阶段致癌作用产生的，其中涉及多年以来遗传损伤（例如突变）的积累，直到细胞转化为癌症。目前对人类潜在致癌作用的检测都存在一定的缺陷。很少有模型采用正常人类细胞，并且诱变通常不是终点。为了更好地了解潜在致癌物的风险，需要能够评估此类风险的新模型系统。在初步研究中，我们发现，在正常人口腔上皮细胞 (NOE) 与 lacI 大鼠报告细胞系（BB 细胞 - 含有用于检测诱变的基于 lambda 的穿梭载体）的单层共培养中，苯并 (a) 芘 (BaP) 诱导的诱变相对于单独的 BB 细胞增加。这种增加的诱变很可能是由于 NOE 激活 BaP 所致。我们假设 NOE 与 lacI 报告细胞共培养将能够检测与正常人口腔细胞致癌物代谢激活有因果关系的诱变。为了验证该模型，将研究两种烟草致癌物（BaP 和亚硝基降烟碱）诱导的诱变。在目标 1 中，我们将建立并表征由 NOE 与 BB 报告细胞共培养组成的共培养模型。在目标 2 中，该模型将用于：1) 比较传统卷烟和最近推出的“低风险”卷烟的烟草烟雾冷凝物 (TSC) 引起的诱变，2) 确定这些 TSC 代谢的个体间差异。由于来自不同器官部位的人体细胞可以在培养物中生长，因此该模型有可能成为一个灵活的基础广泛的系统，用于评估烟草产品在许多人体器官中的致癌风险。第三个目标是使用基因表达阵列并通过 RT-PCR 和蛋白质印迹进行确认，对存在和不存在 TSC 的 NOE 中细胞色素 P-450 的水平进行初步研究。由于 NOE 细胞注定会发展成口腔癌，因此该模型填补了一项空白，具有成为量化突变（致癌过程中的一个重要步骤）的器官特异性技术的潜力。显然，最好是前瞻性地确定其潜在风险，而不是回顾性地确定。