Molecular sensing and imaging of human disease cells and their responses to biochemical stimuli.

摘要

Advancement in microscopic and spectroscopic techniques could significantly improve our ability in the study and diagnosis of diseases. Especially, being able to image and detect human diseases at the cellular and molecular level allows people to diagnose diseases at early stages and to study the molecular mechanisms behind various diseases. Currently, histopathological techniques are most widely used for prognosis and diagnosis of human diseases. However, conventional histopathology requires a complex process of sample preparation, which limits the diagnostic efficiency of this technique. More importantly, it requires fixation of tissue or cell sample, making it unsuitable for the study of dynamic cellular activities in the progress of diseases. This dissertation mainly discusses the progress in development of noninvasive imaging techniques that can be applied to study human diseases at the cellular level.;One approach is to use atomic force microscopy (AFM) and Raman spectroscopy to quantitatively measure the biomechanical and biochemical properties of cells, and then use these properties to differentiate between different cell types, or cells at different states. Here we have utilized our tandem AFM-Raman spectroscopy system to differentiate between cancerous and healthy human lung epithelial cells, and monitor their different responses to anticancer drug treatments. Generally, this technique (AFM-Raman) can serve as a complementary approach to study various diseased cells, providing additional information to help doctors identify diseases at an early stage and investigate the progress of diseases.;Another approach is specifically target and image disease marker molecules using advanced microscopic and spectroscopic techniques. Epidermal growth factor receptor (EGFR), as a cancer marker molecule, has been used as a model to develop noninvasive imaging methods. A nanoparticle-based imagine probe has been synthesized for specific imaging of EGFR at a single cell surface using surface-enhance Raman spectroscopy (SERS). Due to the noninvasive feature of SERS, it can monitor the receptor-mediated endocytosis of a nanoparticle in real time. Furthermore, an AFM-based simultaneous Topography and RECognition (TREC) imaging technique has been developed to localize EGFR subcellular distribution with nanoscale resolution. This TREC technique exhibits potential to monitor the binding between EGFR and its ligands at single molecule level.;A multimodal imaging nanoprobe, which integrates different imaging modalities into one single nanoparticle, can incorporate advantages and compensate for weaknesses of respective imaging techniques. In this dissertation, we have functionalized a previously reported nanoprobe for magnetic resonance imaging (MRI), trying to incorporate SERS function into this probe to realize MRI-SERS bimodal imaging. We have tested the SERS performance of the probe by using it to detect EGFR in three human cancer cell lines. This nanoprobe demonstrates the potential for in vivo MRI-SERS bimodal imaging with improved sensitivity from SERS. In addition, we have synthesized another composite nanoprobe for SERS-fluorescence bimodal imaging of a fat-responsive G protein-coupled receptor 120 (GPR120). Fluorescence is used as a fast indicator while SERS is for accurate localization of GPR120. Using this probe, we can also quantitatively measure the changes of GPR120 activities in response to fatty acid binding, showing the potential to study the molecular mechanism of fatty acid chemoreception.