Interactions of engineered nanomaterials with lung alveolar epithelium

描述(申请人提供)：尽管由于纳米生物医学/技术的科学和应用的扩展，工程纳米材料(ENM)的使用有望显著增加，但ENM损伤和/或转运到/穿过肺泡上皮的机制尚不清楚。吸入环境中的超细颗粒(其大小范围与当前纳米材料的定义重叠)已被证明会导致不利的心血管、肺部和血液影响。如果任何ENM被意外吸入，它们最有可能进入体循环的途径是穿过肺泡上皮。根据我们正在进行的几类具有明确物理化学特征的纳米颗粒(例如聚苯乙烯、二氧化硅和金属(氧化物))的肺损伤和转运的研究，以及最近关于吸入超细空气污染物颗粒和其他纳米材料(特别是富勒烯及其衍生物)对健康影响的报道，我们假设不同形式的ENM之间的相互作用(例如，负富勒烯和正富勒烯；原始(疏水)富勒烯和衍生(亲水性)富勒烯；不同质量的富勒烯(例如，C60、C70、C80和聚合物富勒烯)和肺泡上皮细胞I可以以ENM特异性的方式破坏正常的肺泡上皮细胞动态平衡并诱导细胞特性和肺泡上皮屏障功能的改变，II)可以为ENM提供进入体循环的主要门户(例如，富勒烯可以通过跨皮转位途径转移)，以及III)高度依赖于ENM的物理化学性质(例如，富勒烯)。利用表面特征经过适当修饰的富勒烯在体外模型(包括我们建立的原代培养的大鼠肺泡上皮细胞单层)和体内大鼠肺组织中，我们将通过研究下列四个目标来验证这些假说：1)顶部暴露富勒烯对体外肺泡上皮主动和被动屏障特性的影响；2)富勒烯在体外肺泡上皮细胞中的内化、去向和作用；3)富勒烯在体外跨肺泡上皮的转运；以及4)富勒烯在体内大鼠肺内的内化和转运，珊瑚在体内与体外相比与远端呼吸上皮的损伤和转运有关。本文提出的研究结果将为具有明确理化性质的富勒烯内化/跨肺泡上皮的细胞毒性和内化/转运机制提供见解。我们的主要目标是获得有关富勒烯(和其他ENM)与肺泡上皮相互作用的新信息，以帮助了解与肺的相互作用，并帮助改进未来的纳米生物医学应用(例如，肺部药物/基因输送)。 与公共健康相关：吸入人造纳米材料(直径100纳米)，包括环境中的超细污染物颗粒和经工程/制造的纳米材料(ENM：例如富勒烯(由60个碳原子组成)、碳纳米管(CNT)、量子点和金属/金属氧化物纳米颗粒)，可能与各种与心脏、血液和肺相关的健康影响有关。看来，这些影响可能会增加易感人群的发病率和死亡率。在这些纳米材料中，富勒烯已经以惊人的数量被制造出来，用于广泛的应用，然而这种特殊的ENM(例如，具有正负表面电荷的富勒烯和/或具有正负表面电荷的富勒烯和/或原始的(喜脂的)富勒烯与衍生的(喜水的)富勒烯)损伤和/或移位到肺泡上皮内和/或跨肺泡上皮(其中气体交换和离子传输发生在肺的远端部分)的机制尚不清楚。我们将确定与体外和体内大鼠肺远端气血屏障(即肺泡上皮)模型的相互作用，以帮助了解吸入具有明确表面特性(例如，电荷和疏水性)的富勒烯可能造成的损害。有了这些知识，使用具有特定表面属性(不会造成伤害)的富勒烯的改进的纳米医学和生物应用(例如，药物/基因输送)可以设计用于未来的应用。

DESCRIPTION (provided by applicant): Although utilization of engineered nanomaterials (ENM) due to expansion of the science and application of nanobiomedicine/technology is expected to markedly increase, the mechanisms by which ENM injure and/or are transported into/across lung alveolar epithelium are not well known. Inhalation of ambient ultrafine particulates (whose size range overlaps the current definition of nanomaterials) has been shown to result in adverse cardiovascular, pulmonary and hematologic effects. If any ENM are accidently inhaled, their most likely route of entry into the systemic circulation is across the alveolar epithelium of the lung. Based on our ongoing research on lung injury and trafficking of several (e.g., polystyrene, silica and metal (oxides)) classes of nanoparticles with defined physicochemical characteristics, and recent reports on health effects of inhaled ultrafine air pollutant particulates and other nanomaterials (especially fullerenes and their derivatives), we hypothesize that interactions between various forms of ENM (e.g., negative vs. positive fullerenes; pristine (hydrophobic) vs. derivatized (hydrophilic) fullerenes; fullerenes of different mass (e.g., C60, C70, vs. C80 vs. polymeric fullerenes) and alveolar epithelial cells i) can disrupt normal alveolar epithelial cell homeostasis and induce changes in cellular properties and alveolar epithelial barrier function in an ENM-specific manner, ii) can provide the primary portal of entry for ENM into the systemic circulation (e.g., fullerenes may be translocated via transepithelial translocation pathways), and iii) are highly dependent on physicochemical properties of ENM (e.g., fullerenes). Utilizing fullerenes of appropriately modified surface characteristics in in vitro models (including our well-established primary cultured monolayers of rat alveolar epithelial cells) and rat lungs in vivo, we will test these hypotheses by investigating the following four aims: 1) effects of apically exposing fullerenes on active and passive barrier properties of alveolar epithelium in vitro; 2) internalization, fate and effects of fullerenes in alveolar epithelial cells in vitro; 3) trafficking of fullerenes across alveolar epithelium in vitro; and 4) fullerene internalization and trafficking in rat lungs in vivo, correlating injury to and trafficking across distal respiratory epithelium in vivo vs. in vitro. Findings from the investigations proposed herein will provide insights into cytotoxicity and mechanisms of internalization/trafficking of fullerenes with defined physicochemical properties into/across lung alveolar epithelium. Our major objective is to obtain new information on fullerene (and other ENM) interactions with alveolar epithelium in order to help understand interactions with the lung, and help improve future nanobiomedical applications (e.g., pulmonary drug/gene delivery). PUBLIC HEALTH RELEVANCE: Inhalation of man-made nanomaterials (<100 nm in diameter), including ultrafine ambient pollutant particles and engineered/manufactured nanomaterials (ENM: examples are fullerenes (composed of >60 carbon atoms), carbon nanotubes (CNT), quantum dots and metal /metal oxide nanoparticles), may be associated with various heart-, blood- and lung-related health effects. It appears that these effects may increase morbidity and mortality in susceptible populations. Of these nanomaterials, fullerenes have been manufactured in an astonishingly large quantity for wide applications, yet the mechanisms by which this particular ENM (e.g., fullerenes with positive vs. negative surface charges and/or pristine (lipid-loving) vs. derivatized (water-loving)) injure and/or are translocated into and/or across alveolar epithelium (where gas exchange and ion transport occur in the distal portion of the lung) are not well understood. We will determine interactions with both in vitro and in vivo rat models of the distal air-blood barrier of the lung (i.e., alveolar epithelium) in order to help understand possible injury from inhaled fullerenes with defined surface properties (e.g., charge and hydrophobicity). With this knowledge, improved nanomedical and biological applications (e.g., drug/gene delivery) using fullerenes bearing specific surface properties (that do not cause injury) can be designed for future applications.