单细胞水平下多环芳烃的实时原位跟踪、跨膜运移过程和生物富集机制

The environmental behavior of pollutants in mineral-water media and interfaces has been extensively and deeply studied, but the environmental behavior of pollutants in biological micro-interfaces is emergently required. The environmental behaviors of contaminants are usually obtained from observations in a large group of cells using the average strategy. These traditional methods cannot fully reveal the complex behaviors of typical pollutants and potentially ignore the heterogeneity and diversity of environmental behaviors of pollutants in a single cell. Using single molecule fluorescence spectroscopy (SMFM), in this proposal we aim to real-time visualize and in-situ quantify the environmental behaviors and processes of persistent pollutants (polycyclic aromatic hydrocarbons, PAHs) at single-cellular level with micro-second temporal resolution. We would reveal a series of micro-environmental behaviors and processes of PAHs in bacteria, alga, and model mice, respectively, including their environment-biological partitioning and distribution, trans-membrane transfer (absorption and excretion), bioaccumulation and bio-concentration, at single-cell level with high temporal and spatial resolution. The technical advantages of SMFM high-throughput screening and large-data statistics will enable us discover the randomness, heterogeneity and diversity of those environmental behaviors of PAHs in single cells. Finally, we would discuss the molecular and cellular mechanisms resulting in the epigenetic diversity of bio-environmental behaviors of pollutants in different cells, and its significant applications and implications for environmental sciences.

污染物在矿质-水环境介质和界面中的环境行为研究已得到广泛而深入的展开，但污染物在生物微界面内的环境行为研究相对较少。传统的“一锅烩”和“平均”方法所获得的污染物的环境行为信息，严重掩盖了污染物在单个细胞内环境行为的异质性和多样性。本课题拟用单分子荧光显微技术 (SMFM)，在毫秒级和单细胞分辨的时空模式下，对持久性污染物（多环芳烃为例）在细胞内的微观环境行为和动态过程进行实时跟踪和原位成像。本项目以成像模式和电影拍摄的视角，高灵敏、高时空分辨、单细胞水平深刻揭示多环芳烃分别在单细菌、单细胞藻类和高等模式生物中的环境生物分配、分布、跨膜运移（吸收和排出）、生物富集和蓄积等微观环境行为和界面过程。单分子荧光显微技术高通量检测和大数据统计的技术便利，能帮助我们揭示污染物在不同细胞内上述环境行为的随机性、异质性和多样性。我们将从多角度探讨引起这种表观异质性的细胞生物学机制和重要的环境

传统的“一锅烩”和“平均”方法所获得的污染物的环境行为信息，严重掩盖了污染物在单个细胞内环境行为的异质性和多样性。利用单分子荧光显微镜，我们不但在单细胞水平上探究了微生物对HOCs的富集和分配过程，还观察了HOCs的分子团簇现象及其裂殖分配过程。相较于传统环境分析技术，单分子荧光显微镜可以得到HOCs在环境介质中的微区分布动态过程及物理聚集状体，为HOCs的环境过程的研究提供了新的视角和信息。通过结合 GC-MS 和单分子荧光显微镜 (SMFM)，我们还揭示了四种不同多环芳烃，包括菲 (Phe)、芘 (Pyr)、苝 (Per) 和苯并[a]芘 (BaP)，在动脉粥样硬化模型小鼠（ApoE-KO 小鼠）中长期口服暴露后的宏观组织和微观区域水平分布特征。我们观察到许多微簇热区，显示出比其他常见区域高得多的Per荧光强度。我们观察到在动脉粥样硬化斑块区域，Per 热点与炎症反应最严重的微区域共定位。PAHs 富集非常高的热点区可能刺激局部炎症并导致主动脉过度损伤，导致动脉粥样硬化病变相对面积显著增加并加重动脉粥样硬化。