Toxicology study of emissions from a burn pit simulator

烧坑模拟器排放物的毒理学研究

• 批准号: 10339413
• 负责人: Nathan RAVI
• 金额: --
• 依托单位: ST. LOUIS VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-10-01 至 2023-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10339413
• 关键词: Acute; Address; Aerosols; Afghanistan; Air Pollution; Apoptosis; Area; Aromatic Polycyclic Hydrocarbons; Asphyxia; Binding Sites; Biomass; Cancer Etiology; Cancerous; Carbon; Carcinogens; Cell Culture Techniques; Cell Death; Cell Line; Cell Survival; Cell physiology; Cell-Matrix Junction; Cells; Cessation of life; Charge; Chemicals; Chemiluminescence assay; Chinese Hamster Ovary Cell; Chronic; Chronic Disease; Cohort Studies; Cornea; Culture Media; DNA; DNA Damage; Data; Deposition; Development; Dioxins; Disease; Electrodes; Endocrine; Endocrine system; Engineering; Ensure; Environment; Epithelial; Ethylenes; Event; Exhibits; Exposure to; Foundations; Functional disorder; Furans; Gases; Generations; Genes; Genome; Genomics; Goals; Gold; Gulf War; Gulf War veteran; Human; Human Cell Line; Hydrophobicity; Impairment; In Vitro; Institute of Medicine (U.S.); Iraq; Lead; Light; Mammalian Cell; Measurement; Measures; Medical; Medicine; Methodology; Military Personnel; Minerals; Molecular; Monitor; Nature; Nitrogen; Onset of illness; Outcome; Oxidative Stress; Oxygen; Particulate Matter; Pathway interactions; Persian Gulf Syndrome; Phase; Preventive; Proliferating; Property; Publishing; RNA; Reactive Oxygen Species; Recommendation; Reporting; Research Design; Retina; Retrospective Studies; Review Literature; Sampling; Sedimentation process; Shapes; Soldier; Soot; Stream; Structure; Surface; Syndrome; System; Technology; Temperature; Testing; Therapeutic; Time; Toxic effect; Toxicology; Toxin; United States National Academy of Sciences; Universities; Veterans; War; Washington; Wildfire; Work; World Health Organization; acute toxicity; airway inflammation; base; biomaterial compatibility; burn pit; carcinogenicity; cell type; chemical property; combustion product; cost; culture plates; cytotoxicity; design; electric impedance; experimental study; irritation; lens; lung injury; metallicity; nanoGold; nephelometry; non-genomic; operation; physical property; response; tissue culture; tool; toxicant; transcriptome sequencing; volatile organic compound; wasting; zeta potential

Carbonaceous (i.e., carbon-containing) particulate matter (PM) emitted by the combustion of waste in military burn pits is associated with post-deployment sudden illnesses, such as the Gulf War Illness (GWI). GWI is a syndrome where otherwise healthy soldiers return with debilitating conditions ranging from acute respiratory inflammations to chronic impaired endocrine function. The nature by which it develops is unknown. To shed light over the effect of military burn pit emissions contributing to GWI, we propose to evaluate the combustion envi- ronments within, using a simulator we built, and understand how these are conducive to toxicant formation. To identify underlying mechanisms, our immediate goal is to modify and calibrate the simulator. Then, we will syn- thesize carbonaceous PM representative of burn pit emissions and expose it to mammalian cell lines. The simulator operates on the bench scale and, to the best of our knowledge, is the only one of its kind within the VA system capable of reliably reproducing carbonaceous PM (and gases) emitted by military burn pits. The simulator’s design allows modulation of the feed (fuel, oxygen, and nitrogen) in the respective inlet streams. The ability to modulate these three independent variables offers tight control over the structure and temperature of the flame that precedes PM synthesis. As a result, the physical and chemical properties of the PM can be predicted. We will generate a statistically designed full-factorial experiments for mixtures. In this approach, we will characterize the PM for size, shape, charge, and surface area-to-volume ratio as well as polycyclic aromatic hydrocarbon (PAH) content, and plot these responses as a function of the independent variables. The response surface methodology (RSM) is a statistical tool, used commonly in engineering, which will allow us to explore the relationship between the three independent factors (that regulate the flame structure and temperature) and the properties of the PM using the least number of experiments. While several PAH molecules that exist in the gas-phase are toxic, the known carcinogens are larger hydrophobic molecules adsorbed on or deposited within the PM. To separate the toxicological effect of PM from the gaseous species, we will collect PM on filters. We will sterilize and disperse the PM samples in cell culture media. To ensure the dispersions are stable, we will record their sizes and zeta potentials using dynamic light scattering (DLS) measurements, and sedimen- tation rates. Cell viability will be monitored by electric cell-substrate impedance sensing (ECIS). ECIS measures the total electrical impedance across two gold electrodes at the bottom of the tissue culture plates, so cell viability is determined by the area of the gold electrodes covered by cell attachment. Biocompatibility will be monitored by apoptosis (cell death) and reactive oxygen species (ROS) generation using chemiluminescence measure- ments. ROS measurements are an indication of oxidative stress to the cells; an environment adverse for healthy cell function. Finally, to determine the chemical pathways initiated by the PM at the molecular level, we will perform RNA sequencing. Before the cells show signs of death or detachment from the electrode, we will extract the total RNA. The RNA will then be analyzed in the Genome Technology Access Center (GTAC) at Washington University. Through GTAC we will determine which genes are under- or over-expressed in each cell line com- pared to control cell lines. We anticipate this data will illustrate the effects of carbonaceous PM on the cells. By initially excluding other types of PM encountered by military personnel (e.g., metallic, halogenated, heterocyclic, or mineral) we will be able to pinpoint PAH-initiated pathways that we believe are significantly re- sponsible for GWI. This will serve as a foundation to direct efforts on how studies (using real material) are to be undertaken. After identifying how real carbonaceous PM causes disease, we will explore therapeutic options.

碳质（即，含碳）颗粒物（PM）的军事废物燃烧排放 燃烧坑与部署后的突发性疾病有关，例如海湾战争疾病（GWI）。GWI是一个 综合症，即健康的士兵返回时出现从急性呼吸道疾病 炎症到慢性内分泌功能受损。其发展的性质是未知的。揭示 考虑到军用燃烧坑排放对GWI的影响，我们建议对燃烧环境进行评估， 使用我们建立的模拟器，了解这些是如何有助于有毒物质的形成。到 识别潜在的机制，我们的直接目标是修改和校准模拟器。然后，我们将同步- 将燃烧坑排放物的代表性碳质PM的大小并将其暴露于哺乳动物细胞系。 该模拟器在实验室规模上运行，据我们所知，这是同类产品中唯一的 在VA系统中，能够可靠地再生军事燃烧坑排放的含碳PM（和气体）。 模拟器的设计允许调节各个入口流中的进料（燃料、氧气和氮气）。 调节这三个独立变量的能力提供了对结构和温度的严格控制 PM合成前的火焰因此，PM的物理和化学性质可以是 预测了我们将针对混合物生成统计设计的全因子实验。在这种方法中，我们 将表征PM的大小，形状，电荷，表面积与体积比以及多环芳烃 碳氢化合物（PAH）含量，并绘制这些响应作为自变量的函数。响应 表面方法（RSM）是一种统计工具，通常用于工程，这将使我们能够探索 三个独立因素（调节火焰结构和温度）之间的关系， 使用最少数量的实验的PM的属性。虽然存在于细胞中的几种PAH分子 气相是有毒的，已知的致癌物是吸附或沉积在气相中的较大的疏水分子。 首相为了从气态物质中分离PM的毒理学效应，我们将在过滤器上收集PM。 我们将对PM样品进行灭菌并将其分散在细胞培养基中。为了确保分散体稳定， 我们将使用动态光散射（DLS）测量和沉积来记录它们的尺寸和zeta电位- 税率。将通过电细胞基质阻抗传感（ECIS）监测细胞活力。ECIS措施 组织培养板底部两个金电极的总电阻抗， 由细胞附着覆盖的金电极面积决定。将监测生物相容性 通过细胞凋亡（细胞死亡）和活性氧（ROS）的产生，使用化学发光测量- 的部分。ROS测量是对细胞的氧化应激的指示;对健康不利的环境 细胞功能最后，为了在分子水平上确定PM引发的化学途径，我们将 进行RNA测序。在细胞显示出死亡或从电极上脱落的迹象之前，我们将提取 总RNA。然后，RNA将在华盛顿的基因组技术访问中心（GTAC）进行分析 大学通过GTAC，我们将确定哪些基因在每个细胞系中表达不足或过度， 对照细胞系。我们预计这些数据将说明碳质PM对细胞的影响。 通过最初排除军事人员遇到的其他类型的PM（例如，金属，卤化， 杂环，或矿物质），我们将能够查明多环芳烃引发的途径，我们认为是显着重新， 为GWI负责。这将作为指导研究（使用真实的材料）的基础。 进行。在确定了真实的碳质PM如何导致疾病后，我们将探索治疗方案。