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确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，并且膜的完整性对细胞功能至关重要。然而，这些相互作用知之甚少，并没有被检查或连接到仿生膜表现出异质性或不对称性。本CAREER提案的研究目标是阐明模型细菌膜中的纳米颗粒-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将采用低温显微镜和微量热法研究局部纳米颗粒-膜相互作用如何产生膜结构和功能的全局变化。AIM 2将采用显微镜和光谱学技术专门研究纳米颗粒聚集在膜/水界面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查多糖（葡聚糖）包被的囊泡作为模型细胞壁/膜屏障，以将潜在的纳米颗粒-膜相互作用与现实的复合膜联系起来。PI的职业计划将研究与教育和宣传目标相结合，以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。建立在PI？她以前的经验，一个新的高中计划认为小/梦想大！将在大普罗维登斯，国际扶轮地区的城市学校的科学类开发。该计划将利用最近授予NSF MRI资助的透射电子显微镜的远程操作能力，并为学生提供？虚拟？暴露于使用最先进的仪器分析纳米材料。这项建议还将加强一个新的大一通识教育课程，旨在教育学生的社会，经济和环境影响的纳米技术，以及需要有效地沟通新兴技术，以广泛的受众。专业发展活动，包括研究和专门讲习班，将补充课程编制，并提供加强？软？和研究生学习所需的技术技能。总之，研究和教育计划将支持PI作为成功的学者和教育家的独立职业道路。智力优势。研究是由以下假设驱动的：（1）一个？最佳？存在尺寸和表面化学的组合以促进纳米颗粒-膜结合和膜重构，这通过净粘合强度和用于使膜变形的能量损失来平衡;（2）纳米颗粒在膜/水界面处的聚集导致尺寸依赖性膜孔形成和纳米颗粒内陷;以及（3）模仿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构。定量信息，这是目前缺乏的，涉及纳米粒子结合的程度，这如何影响膜结构，相行为，和渗透性将获得环境相关的细菌膜。这项工作是变革性的，因为纳米颗粒-膜相互作用和细胞壁的作用还没有被检查细菌膜。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，这种理解可以用来创建或修改基于纳米颗粒的治疗方法，设计混合有机无机胶体，并确定新的生物传感或药物输送策略。传播将通过出版物、介绍以及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宣传材料，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