Real-Time Assessment of Extracellular Vesicles by Intravital Microscopy Imaging

摘要

Extracellular vesicles (EVs), such as exosomes, are essential players in cell-cell communication, which are being considered for developing novel drug delivery applications. Thus, there is much interest in developing reliable methods for tracing EVs in biological settings in real-time. Current standard methods are based on lipophilic fluorescent dyes, though this labeling has not been validated and may result in biased understanding of EVs. Herein, we developed labeling methods directed to faithfully follow the EVs for assessing the fate in biological settings both in vitro and in vivo. Our results showed that by installing fluorescent probes on the surface of EVs by conjugation to the EVs' amino groups, it is possible to preserve the EVs concentration and stability. Moreover, we found that the EVs labeled by our method were stable in the bloodstream, while the EVs labeled with the conventional lipophilic dyes were immediately removed from circulation, highly accumulating in the lungs. Finally, by using our new labeling method for imaging EVs, we found that EVs from murine breast cancer 4T1 accumulated along the brain capillaries, which correlate with the incidence of brain metastasis from breast tumors.