The influence of particulate matter properties on the biophysical entry mechanisms into lung cells

环境空气中颗粒物(PM)的污染是一个日益严重的问题，对数百万人的健康构成了严重威胁。暴露于高水平的PM会显著增加患心血管疾病、肺部疾病和癌症的风险。直径<lt；1微米的细小至超细颗粒可进入肺细胞并引发炎症反应。尽管有大量的流行病学研究，但控制颗粒物和单细胞之间直接相互作用的机制还很难理解。特别是，涉及结合和摄取的细胞机械概念是未知的。因此，需要研究光学概念，它可以获得纳米和毫秒级的细胞响应。为此，我们将使用新的光学技术和生物物理概念。特别是，我们将使用光镊子、三维热噪声跟踪和快速、无标记的超分辨率显微镜来实现对相关进入细胞机制的新见解。通过这些工具，我们将回答以下问题：什么生物物理原理支配着颗粒物进入细胞的过程，颗粒特性如何影响与细胞接触的颗粒的命运，以及不同类型的肺细胞如何处理颗粒物。利用我们的光子力显微镜，我们将以一种可重复的方式将颗粒物带到肺细胞附近，以测量与细胞接触的颗粒结合强度和摩擦力的变化。这些相互作用参数可以从粒子的热位置涨落中提取出来。这些实验还得到了使用旋转相干散射(ROCS)显微镜的活细胞超分辨率显微镜的补充，这使我们能够观察到细胞骨架在100赫兹对颗粒物暴露的反应。通过使用不同的荧光细胞毒性分析，细胞反应模式将与促炎细胞因子的存在相关联。这项研究是为两名在不同方面密切合作的博士生设计的。一名博士生将使用光学力显微镜研究颗粒物进入肺细胞的情况，并通过热噪声跟踪来表征控制不同颗粒物吞噬的力学原理。另一名博士生将专注于使用ROCS显微镜研究不同颗粒结合和进入肺上皮细胞时细胞骨架的反应。通过这项研究项目，我们希望提供重要的补充知识，以更好地评估颗粒物特性对肺部疾病的影响，这些疾病与环境毒理学和肺病学领域最相关。

The pollution of the ambient air with particulate matter (PM), microscopic suspended particles, is an aggravating problem that poses a serious health risk to millions of people. The exposure to elevated levels of PM significantly increases the risk for cardiovascular diseases, lung diseases and cancer. Fine to ultrafine particles with diameters < 1µm can enter lung cells and trigger inflammatory responses. Despite a large number of epidemiological studies, the mechanisms governing the direct interaction between particulate matter and single cells is hardly understood. In particular, the cell mechanical concepts involving binding and uptake are unknown. Hence, investigative optical concepts are needed, which can access cellular responses on scales of nanometers and milliseconds.For this, we will use novel optical technologies and biophysical concepts. In particular, we will use optical tweezers, three-dimensional thermal noise tracking and fast, label-free super resolution microscopy to achieve new insights into the relevant entry mechanisms into cells. Through these tools, we will answer questions about what biophysical principles govern the entry process of particulate matter into cells and how particle properties influence the fate of the particle in contact with the cell, but also how different lung cell types handle particulate matter. With our Photonic Force Microscopy, we will bring particulate matter in the vicinity of lung cells in a reproducible way to measure the change in binding strength and friction of the particle in contact with the cell. These interaction parameters can be extracted from thermal position fluctuations of the particles. The experiments are complemented by live-cell super-resolution microscopy using rotating coherent scattering (ROCS) microscopy, which enables us to observe cytoskeleton reorganization in 100 Hz in response to particulate matter exposure. By using various fluorescence cytotoxicity assays, the cellular response pattern will be correlated with the presence of pro-inflammatory cytokines.The research is devised for two PhD students working closely together on different aspects. The one PhD-student shall investigate the entry of particulate matter into lung cells using photonic force microscopy and characterize the mechanical principles governing the engulfment of different particles by thermal noise tracking. The other PhD student will focus on the investigation of the cytoskeleton response during binding and entry of different particles into lung epithelial cells using ROCS microscopy. With this research project, we expect to provide important additional knowledge to better assess the influence of particulate matter properties on lung diseases, most relevant for the fields of environmental toxicology and pulmonology.