Project 1: Mechanisms of Nanomaterial-Cell Interactions

项目1的首要目标是了解工程纳米颗粒（ENPs）的物理化学组成和结构如何决定它们与细胞的生物相互作用，最终影响信号传导和生理学。我们假设清道夫受体通路在决定 ENPs在巨噬细胞中的细胞内剂量和ENPs在体内的生物分布。我们进一步假设ENPs“劫持”清道夫受体摄取途径可能会损害巨噬细胞吞噬和/或杀死常见细菌性肺病原体的能力。为了检验这些假设，将使用显微镜、流式细胞术和磁性颗粒检测（MPD）来定量确定ENPs（氧化铈、二氧化硅、氧化铁）的大小、表面化学、电荷和聚集状态的变化如何影响其在来自野生型小鼠和A类清道夫受体（Aim 1）缺陷小鼠的巨噬细胞中的摄取和运输途径的速率。我们将确定如何物理化学性质的 ENPs影响溶酶体稳定性和它们激活炎性体信号传导的能力（目的2）。将进行体外巨噬细胞感染研究以确定通过清道夫受体途径内化的ENPs是否损害巨噬细胞识别、吞噬和杀死病原体的能力，使用 肺炎链球菌作为人肺病原体模型（目的3）。最后，我们将应用基因组微阵列和生物信息学分析来确定由清道夫受体调节的基因调控途径和ENPs可能影响巨噬细胞先天免疫功能的expore机制（目的4）。本项目中的剂量-反应研究旨在直接为项目3中的定量构效关系分析和剂量测定和风险模型的开发提供数据。这些结果还将为项目2中进行的平行体内生物分布和病原体感染研究的实验设计和解释提供分子基础。我们设想所提出的方法可以提供 用于危害和风险分析的定量数据，重点关注可能受到ENP低水平暴露影响的巨噬细胞功能的临床相关指标。

The overarching goal of Project 1 is to understand how the physicochemical composition and structure of engineered nanoparticles (ENPs) dictate their biological interaction with cells to ultimately affect signaling and physiology. We hypothesize that scavenger receptor pathways plays a prominent role in determining the intracellular dose of ENPs in macrophages and biodistribution of ENPs in vivo. We further hypothesize that ENPs that "hijack' scavenger receptor uptake pathways may compromise macrophages ability to phagocytose and/or kill common bacterial lung pathogens. To test these hypotheses, microscopy, flow cytometry, and magnetic particle detection (MPD) will be used to quantitatively determine how varying the size, surface chemistry, charge and agglomeration state of ENPs (cerium oxides, silica oxides, iron oxides) affects their rate of uptake and trafficking pathways in macrophages derived from wildtype mice and mice deficient in class A scavenger receptors (Aim1). We will determine how the physicochemical properties of the ENPs affect stability of lysosomes, and their ability to activation inflammasome signaling (Aim 2). In vitro macrophage infection studies will be conducted to determine whether ENPs internalized by scavenger receptor pathways compromise the macrophages ability to recognize, phagocytosis and kill pathogens, using streptococcus pneumoniae as a model human lung pathogen (Aim 3). Finally, we will apply genomic microarray and bioinformatic analyses to identify gene regulatory pathways modulated by scavenger receptors and expore mechanisms by which ENPs may affect innate immune functions of macrophages (Aim 4). The dose-response studies in this project are designed to feed data directly for quantitative structure activity relationship analysis and development of dosimetry and risk models in Project 3. The results will also provide a molecular basis for experimental design and interpretation of parallel in vivo biodistribution and pathogen infection studies to be conducted in Project 2. We envision the proposed approach can provide quantitative data for hazard and risk analysis that is focused on clinically relevant measures of macrophage function that are potentially impacted by low level exposures to ENPs.