Project 1 Cellular uptake, clearance, and effects of C60 and MWCNTs in epithelial

摘要 项目1的总体目标是通过使用从肺、心血管和生殖系统分离的大鼠、小鼠和人类上皮和内皮细胞开发数据，用于项目3，以构建和验证基于生理学的药代动力学/药效学（PBPK/PD）模型，从而推断与纳米材料暴露相关的人类健康风险。人类接触纳米材料 可能通过职业暴露、环境暴露或有意用作治疗剂的递送载体而发生。迄今为止，大多数关于纳米材料相关毒性的动物研究都集中在雄性或非妊娠啮齿动物身上。正如本提案的总体主题中所讨论的，我们正在利用怀孕啮齿动物来模拟潜在易感人群，这些人群具有更大的不良健康影响风险 与纳米材料暴露有关。在项目1的具体目标1中，我们将生成，表征和放射性标记石墨烯类中的两种碳基纳米材料-富勒烯C60和多壁碳纳米管（MWCNT）形式，这将用于每个项目。具体而言，项目1将利用未标记的C60和三种形式的多壁碳纳米管：封端的多壁碳纳米管，氧化损伤的多壁碳纳米管（多壁碳纳米管（COOH）n）， 和含铁氧化损伤的MWCNT（FemMWCNT（COOH）n）。我们将使用术语“MWCNT的形式”来指这三种形式的MWCNT。项目1还将利用碳-14均匀标记的类似物[C（U）]C60、[rC（U）] MWCNT、[rC（U）]MWCNT（COOH）n和[rC（U）]FemMWCNT（COOH）n。 为了开始提供这些纳米材料的细胞命运数据，用于在项目3中建立PBPK/PD模型，我们将利用三种细胞类型代表潜在的靶器官：肺（肺泡1型和11型上皮细胞）、心血管系统（主动脉内皮细胞）和生殖系统（胎盘内皮细胞）。在项目1的前2年，纳米颗粒的分布和效果将在体外使用大鼠细胞进行评估，以补充项目2中获得的体内数据。在第3-4年期间，我们将开始通过测试C60和多壁碳纳米管形式对从小鼠分离的相同细胞群体的体外作用来验证大鼠模型。最后，在项目1的第5年，我们将研究C60和多壁碳纳米管形式对人类上皮细胞和内皮细胞的影响，以外推到人类风险评估。项目1共同负责人将与中心管理团队和国家环境健康科学研究所（NIEHS）项目官员密切合作，以确定数据开发方向的任何必要变化。项目1共同负责人将向项目2和项目3共同负责人提供体外结果，以便改进项目2和项目3的研究范围。C60和多壁碳纳米管形式在细胞和亚细胞水平上的作用将用下面概述的三个具体目标进行测试。

Abstract The overall aim of Project 1 is to develop data, through the use of rat, mouse and human epithelial and endothelial cells isolated from the pulmonary, cardiovascular, and reproductive system, for use in Project 3 to construct and validate a physiologically based pharmacokinetic/pharmacodynamic (PBPK/PD) model to extrapolate human health risks associated with exposure to nanomaterials. Human exposure to nanomaterials may occur through occupational exposure, environmental exposure, or intentional use as a delivery vehicle for therapeutic agents. To date, the majority of animal studies on the toxicity associated with nanomaterials have focused on male or non-pregnant rodents. As discussed in the overall theme of this proposal, we are utilizing the pregnant rodent to model potentially susceptible populations at greater risk for adverse health effects associated with nanomaterial exposure. In Specific Aim 1 of Project 1, we will generate, characterize, and radiolabel the two carbon-based nanomaterials in the graphene class¿ the fullerene C60 and forms of multiwalled carbon nanotube (MWCNT), that will be used in each project. Specifically, Project 1 will utilize unlabeled C60 and three forms of MWCNTs: end-capped MWCNTs, oxidation-damaged MWCNTs (MWCNT(COOH)n), and iron-containing oxidation-damaged MWCNTs (FemMWCNT(COOH)n). We will use the terms "forms of MWCNTs" to refer to these three forms of MWCNTs. Project 1 will also utilize the carbon-14 uniformly labeled analogs f C(U)]C60, rC(U)]MWCNTs, r''C(U)]MWCNT(COOH)n, and r^C(U)]FemMWCNT(COOH)n. To begin providing data on the cellular fate of these nanomaterials for use in establishing a PBPK/PD model in Project 3, we will utilize three cell types to represent potential target organs: lung (alveolar type 1 & 11 epithelial cells), the cardiovascular system (aortic endothelial cells), and the reproductive system (placental endothelial cells). In the first 2 years of Project 1, the distribution and effect ofthe nanoparticles will be evaluated in vitro using rat cells to complement the in vivo data obtained from Project 2. During Years 3-4, we will begin to validate the rat model by testing the in vitro effects of C60 and forms of MWCNTs on the same population of cells isolated from mice. Lastly, in Year 5 of Project 1, we will examine the effects of C60 and forms of MWCNTs on human epithelial and endothelial cells to extrapolate to human risk assessment. Project 1 co-Leaders will work closely with the Center Management Team and the National Institute of Environmental Health Sciences (NIEHS) Project Officer to determine any necessary changes in direction as data is developed. Project 1 co-Leaders will provide the in vitro results to Projects 2 and 3 co-Leaders as they become available to permit refinement of the scope of research in Projects 2 and 3. The effects of C60 and forms of MWCNTs at the cellular and sub-cellular level will be tested with the three Specific Aims outlined below.