Health effects of manufactured nanoparticles (NP): molecular and cellular biology and toxicology (NanoBioTox)

该项目旨在促进我们对纳米颗粒（NPs）与呼吸道蛋白质，组织和细胞相互作用的理解。在上一个资助期间，我们开发了一套具有明确定义的特定属性的NP工具包。我们已经分析了模型蛋白与我们的NP的结合动力学和平衡。在体外研究中，我们已经证明，NP的表面性质决定了它们的吸收和它们在细胞内的最终目的地，以及细胞毒性作用。我们进一步提供的证据表明，在体内，NP表面特性，包括蛋白质吸附，同样影响其在体内的分布。基于这些结果，我们计划继续我们的研究，包括旨在阐明NPs被细胞摄取后的命运的实验，特别是关于包裹NPs的“蛋白质冠”。我们的工作分为三个主要领域。第一个（“NP工具箱”）的目的是合成和表征的水溶性功能纳米粒子。修饰纳米颗粒周围的蛋白质冠将发挥重要作用。第二个主题（“NP-蛋白质复合物”）侧重于NP与蛋白质和体液中其他生物分子的相互作用。特别注意将致力于上皮衬里液，一个含水的液体层，含有糖蛋白和肺表面活性物质成分，并在内部肺表面的气液界面的连续的肺表面活性物质膜。我们将量化的动力学和平衡行为的关键蛋白质结合到纳米粒子作为我们的纳米粒子的特定物理和化学性质的函数。此外，我们将详细研究蛋白质冠的结构。蛋白质是以明确的方向吸附到纳米粒子上还是随机吸附？蛋白质结构在吸附后是保守的，还是被吸附的蛋白质向变性构象发展？在第三个领域（“NP-细胞相互作用和细胞反应”），我们研究了NP与离体肺组织细胞、体外肺细胞及其细胞器的相互作用。此外，在这里，蛋白质冠的影响将进行调查。细胞摄取NP后，蛋白质冠的结构是否会保留下来？最后，我们将研究吸附的蛋白质是否以及如何影响纳米粒子引起的毒性作用。

This project aims to advance our understanding of the interactions of nanoparticles (NPs) with proteins, tissue and cells of the respiratory tract. In the previous funding period, we have developed a toolkit of NPs with well-defined, specific properties. We have analyzed the binding of model proteins to our NPs with respect to kinetics and equilibrium. With in-vitro studies, we have demonstrated that NP surface properties govern their uptake by and their final destination inside cells, and also cytotoxic effects. We further provided evidence that, in vivo, NP surface properties, including protein adsorption, likewise affect their distribution within the body. Based on these results, we plan to continue our studies by including experiments aimed at elucidating the fate of NPs after their uptake by cells, specifically, with regard to the “protein corona” enshrouding the NPs. Our work is structured in three major areas. The first one ("NP toolbox") aims at the synthesis and characterization of water-soluble functional NPs. Modification of the protein corona around NPs will play a major role. The second topic ("NP-protein complexes") focuses on NP interactions with proteins and other biomolecules of body fluids. Special attention will be devoted to the epithelial lining fluid, an aqueous liquid layer containing glycoproteins and pulmonary surfactant components, and to the continuous pulmonary surfactant film at the air-liquid interface of the internal lung surface. We will quantify kinetic and equilibrium behavior of key proteins binding to NPs as a function of the specific physical and chemical properties of our NPs. Moreover, we will examine the structure of the protein corona in detail. Do proteins adsorb to NPs in well defined orientations or randomly? Is the protein structure conserved upon adsorption, or do adsorbed proteins develop toward denatured conformations? In the third area ("NP-cell interactions and cellular responses") we study the interaction of NPs with cells from lung tissue ex vivo, lung cells in vitro and their organelles. Also here, the influence of the protein corona will be investigated. Will the structure of the protein corona be preserved after NP uptake by the cell? Finally, we will investigate if and how adsorbed proteins affect toxic effects elicited by NPs.