Understanding the impact of engineered nanoparticles on the lysosome-autophagy system

CBET - 1336053过去十年见证了纳米技术行业的快速增长：目前市场上有1,000多种含有纳米颗粒（NPs）的消费品，每天都有各种新的工程纳米材料被开发出来。因此，纳米技术行业的工人、消费者以及最终的环境本身都不可避免地与工程纳米粒子接触。由于其独特的性质，包括其纳米级尺寸，大表面积，化学成分和反应性，纳米粒子与生物成分和系统相互作用，其中许多也在纳米级水平上运行。毫不奇怪，人们越来越担心这些相互作用可能对人类产生有害影响。目前，毒理学研究通常用于评估与接触新设计的纳米材料相关的风险。然而，即使当基于标准体内和体外活力测定认为NP没有毒性时，它们仍可能显著改变细胞生理学。关于NP的细胞摄取的大量信息是可用的，并且细胞内化NP的速率可以相对于NP大小、电荷和表面性质来控制。然而，这些物理化学性质对其他细胞过程的影响还没有被系统地研究。例如，已知在内化后，大多数NP引起细胞清除机制的反应。事实上，该项目团队最近发现，许多NP激活自噬，这是消除细胞废物的主要分解代谢途径。虽然自噬的激活可能导致废物的清除增强，但它也可以诱导细胞死亡程序的激活。此外，初步研究表明，特定电荷和组成的NP可以激活自噬反应，但也会损害介导降解的细胞组分，最终阻断自噬通量。该项目团队最近开发了一套分析工具来监测自噬通量和细胞模型系统，以测量自噬底物的积累，包括蛋白脂质和蛋白质聚集体。因此，该团队处于独特的位置，以研究NP对自噬系统的影响。拟议项目的总体目标是映射NP的物理化学性质及其诱导的自噬反应的性质，最终生成设计规则，以在与自噬系统的界面处设计具有所需性质的NP。为了实现这一目标，我们建议追求三个研究目标：1）确定激活自噬的NP的物理化学性质; 2）链接NP的物理化学性质与自噬激活相关的生物相容性和生物不良细胞反应;和3）确定调节自噬反应NPs的细胞网络。智力优点：本研究的结果将提供一个详细的表征NP对自噬系统的影响。该项目将确定NP理化性质（即几何参数、电荷和组成）对与自噬激活相关的下游效应（即NP和其他自噬底物的清除或自噬相关细胞死亡）的影响。该项目还将确定调节对NP的自噬反应的基因网络和NP诱导的自噬的细胞标志物，从而提供重要的预测工具来评估NP对细胞生理学的影响。更广泛的影响：这项研究将产生一个实验框架，用于调查NP对生物系统的影响。将开发和验证专门设计的报告系统，以将NP的理化性质与细胞反应及其潜在的分子机制联系起来。反过来，了解纳米粒子对细胞生理学的影响将有助于开发安全的纳米材料，并将使第三代纳米粒子的设计具有可调的特性，这些纳米粒子与生物系统的接口。最后，拟议的项目将为细胞和分子生物学研究提供新的工具。该研究计划还将提供广泛的教育和培训机会，总体目标是增加STEM中代表性不足的群体的数量和多样性。具体来说，这项研究计划将被利用1）为高中生提供研究和指导的机会，2）改善大学生的教学和学习。

CBET - 1336053 The last decade has witnessed rapid growth in the nanotechnology industry: more than 1,000 consumer products containing nanoparticles (NPs) are currently in the market place, and a variety of newly engineered nanomaterials are developed every day. As a result, workers in the nanotechnology industry, consumers, and ultimately the environment itself inevitably come in contact with engineered NPs. Because of their unique properties, including their nanoscale size, large surface area, chemical composition, and reactivity, NPs interact with biological components and systems, many of which also operate at the nanoscale level. Not surprisingly, growing concerns have been raised over the possibility that these interactions could have deleterious effects on humans. Currently, toxicology studies are typically conducted to assess the risk associated with exposure to newly engineered nanomaterials. However, even when NPs are not deemed toxic based on standard in vivo and in vitro viability assays, they might still significantly alter cell physiology. A wealth of information is available about the cellular uptake of NPs, and the rate at which cells internalize NPs can be controlled with respect to NP size, charge, and surface properties. However, the impact of these physicochemical properties on other cellular processes has not been investigated in a systematic fashion. It is known, for instance, that upon internalization, most NPs elicit the reaction of cellular clearance mechanisms. Indeed, the project team recently found that a number of NPs activate autophagy, the main catabolic pathway that eliminates cellular waste. While activation of autophagy may lead to enhanced clearance of waste material, it can also induce activation of cell death programs. Moreover, preliminary studies indicate that NPs of specific charege and composition can activate the autophagic response but also impair cellular components that mediate degradation, ultimately blocking autophagic flux.The project team recently developed a set of analytical tools to monitor the autophagic flux and cell model systems to measure the accumulation of autophagic substrates, including proteolipid and proteinaceous aggregates. The team is thus uniquely positioned to investigate the impact of NPs on the autophagic system. The overarching goal of the proposed project is to map the physicochemical properties of NPs with the nature of the autophagic response that they induce, ultimately generating the design rules to engineer NPs with the desired properties at the interface with the autophagy system. To achieve this goal, we propose to pursue three research objectives: 1) Identify the physicochemical properties of NPs that activate autophagy; 2) link the physicochemical properties of NPs to biocompatible and bioadverse cellular responses associated with autophagy activation; and 3) identify the cellular network that regulates the autophagic response to NPs.Intellectual Merit :Results from this study will provide a detailed characterization of the impact of NPs on the autophagy system. The project will establish the effect of NP physicochemical properties, namely geometric parameters, charge, and composition, on downstream effects associated with activation of autophagy, namely clearance of NPs and other autophagic substrates or autophagy-associated cell death. The project will also identify the gene network that regulates the autophagic response to NPs and cellular markers of NP-induced autophagy, thus providing important predictive tools to assess the impact of NPs on cell physiology.Broader Impacts :This study will generate an experimental framework for investigating the impact of NPs on biological systems. Specially designed reporter systems will be developed and validated to link the physicochemical properties of NPs to cellular responses and their underlying molecular mechanisms. Understanding the impact of NPs on cell physiology, in turn, will contribute to the development of safe nanomaterials and will enable the design of third-generation NPs with tunable properties where these NPs interface with biological systems. Finally, the proposed project will provide novel tools for cellular and molecular biology studies. This research program will also provide broadly reaching educational and training opportunities with the overall goal of increasing the number and diversity of underrepresented groups in STEM. Specifically, this research program will be leveraged 1) to provide research and mentoring opportunities for high school students, and 2) to improve teaching and learning for college students.