CAREER: Nanoparticle-Bacterial Membrane Interactions and their Role in Nanotoxicology

细胞反应最终决定了有意或无意释放到环境中的纳米颗粒的毒理学影响。物理纳米颗粒-细胞膜相互作用正在成为决定细胞摄取和细胞毒性的重要因素，因为膜是纳米颗粒-细胞相互作用的接触点，而膜的完整性对细胞功能至关重要。然而，人们对这些相互作用知之甚少，也没有在表现出异质性或不对称性的仿生膜中进行研究或与之相关。本CAREER提案的研究目的是阐明模型细菌膜中纳米颗粒-膜相互作用作为纳米颗粒尺寸和表面化学以及盐浓度的函数。AIM 1将使用低温显微镜和微量热法检查局部纳米颗粒-膜相互作用如何产生膜结构和功能的全局变化。AIM 2将使用显微镜和光谱学技术专门研究纳米颗粒在膜/水界面聚集的作用及其对诱导膜内陷（即吞噬纳米颗粒）和穿孔的影响。最后，AIM 3将采用上述实验技术来检测多糖（葡聚糖）包被的囊泡作为细胞壁/膜屏障的模型，将潜在的纳米颗粒-膜相互作用与现实的复合膜联系起来。PI的职业规划将研究与教育和推广目标相结合，吸引代表性不足的学生和当地社区，加强新兴技术的职业准备，特别是在纳米技术/环境/生物界面，并推进课程。以PI为基础？年代以前的经验,新的高中项目认为小/大梦想!将会在国际扶轮大普罗维登斯地区的城市学校发展科学课程。该项目将利用最近获得的NSF核磁共振资助的透射电子显微镜的远程操作能力，为学生提供虚拟的电子显微镜。接触使用最先进的仪器分析纳米材料。该提案还将加强URI的一门新的新生通识教育课程，该课程旨在教育学生纳米技术的社会、经济和环境影响，以及向广大受众有效传播新兴技术的必要性。专业发展活动，包括研究和专门讲习班，将补充课程发展，并提供增强的“软？以及从事研究工作和研究生学习所需的技术技能。总的来说，研究和教育计划将支持PI作为一个成功的学者和教育家的独立职业道路。知识价值。研究是由以下假设驱动的：(1)最优？尺寸和表面化学的结合促进了纳米颗粒与膜的结合和膜的重组，这是由净粘附强度和扭曲膜的能量惩罚来平衡的；(2)纳米颗粒在膜/水界面聚集导致膜孔形成和纳米颗粒内陷；(3)模拟细胞壁的多糖涂层将减少，但不能消除纳米颗粒诱导的膜重组。目前缺乏的与纳米颗粒结合程度有关的定量信息，以及它如何影响膜结构、相行为和通透性，将获得与环境相关的细菌膜。这项工作是变革性的，因为纳米颗粒-膜的相互作用和细胞壁的作用还没有被研究过。更广泛的影响。对纳米颗粒-膜相互作用的定量理解将为细胞毒性机制提供新的见解，并有助于制定预测纳米颗粒-细胞结合和设计生物相容性纳米材料的一般指南。除了细胞毒性之外，这种理解还可以用于创建或修改基于纳米颗粒的治疗方法，设计有机无机混合胶体，并确定新的生物传感或药物递送策略。传播将通过出版物、简报和纳入课程和外联活动来实现。该提案的关键在于，研究概念可以用作教育和推广材料，以激励代表性不足的学生实现STEM职业，参与并告知社区组织

Cellular responses ultimately determine the toxicological impacts of nanoparticles released intentionally or unintentionally into the environment. Physical nanoparticle-cell membrane interactions are emerging as an important factor in determining cell uptake and cytotoxicity as membranes are the point-of-contact for nanoparticle-cell interactions and membranes integrity is vital to cell function. However, these interactions are poorly understood and have not been examined in or connected to biomimetic membranes that exhibit heterogeneity or asymmetry. The research objective of this CAREER proposal is to elucidate nanoparticle-membrane interactions in model bacterial membranes as a function of nanoparticle size and surface chemistry, and salt concentration. AIM 1 will employ cryogenic microscopy and microcalorimetry examine how local nanoparticle-membrane interactions can yield global changes in membrane structure and function. AIM 2 will employ microscopy and spectroscopy techniques to specifically examine the role of nanoparticle aggregation at membrane/water interfaces and its effects on inducing membrane invagination (i.e. engulfing nanoparticles) and poration. Finally, AIM 3 will employ the aforementioned experimental techniques to examine polysaccharide (dextran)-coated vesicles as model cell wall/membrane barriers to link underlying nanoparticle-membrane interactions with a realistic composite membrane. The CAREER plan of the PI integrates research with the educational and outreach objective of engaging underrepresented students and local communities, enhancing career preparedness in emerging technologies, specifically at the nanotechnology/environmental/biological interface, and advancing curriculum. Building on the PI?s previous experiences, a new high school program Think Small/Dream Big! will be developed for science classes in urban schools in the greater Providence, RI area. This program will leverage the remote operating capabilities of the recently awarded NSF MRI-funded transmission electron microscope and provide students ?virtual? exposure to analyzing nanomaterials using state-of-the-art instrumentation. This proposal will also enhance a new freshman general education course at URI aimed at educating students about the social, economic, and environmental impacts of nanotechnology, as well as the need to effectively communicate emerging technologies to broad audiences. Professional development activities, including research and specialized workshops, will supplement curriculum development and provide enhanced ?soft? and technical skills needed for research careers and graduate studies. Taken together, the research and education plans will support the independent career path of the PI as a successful scholar and educator. Intellectual Merit. Research is driven by the hypotheses that (1) an ?optimal? combination of size and surface chemistry exists to facilitate nanoparticle-membrane binding and membrane restructuring, which is balanced by the net adhesive strength and energetic penalty for distorting a membrane; (2) nanoparticle aggregation at membrane/water interfaces leads to size-dependent membrane pore formation and nanoparticle invagination; and that (3) polysaccharide-coatings mimicking cell walls will reduce, but not eliminate nanoparticle-induced membrane restructuring. Quantitative information, which is currently lacking, relating to the degree of nanoparticle binding, how this affects membrane structure, phase behavior, and permeability will be gained for environmentally-relevant bacterial membranes. This work is transformative because nanoparticle-membrane interactions and the role of the cell wall have not been examined for bacterial membrane. Broader Impacts. A quantitative understanding of nanoparticle-membrane interactions will provide new insight into cytotoxicity mechanisms, and could help develop general guidelines for predicting nanoparticle-cell association and designing biocompatible nanomaterials. Beyond cytotoxicity, this understanding could be used to create or modify nanoparticle-based therapeutics, design hybrid organicinorganic colloids, and identify new biosensing or drug delivery strategies. Dissemination will be achieved through publications, presentations, and integration into courses and outreach activities. The crux of this proposal is that the research concepts can be used as educational and outreach material to inspire underrepresented students to achieve STEM careers, engage and inform community organizations