Unveiling the hypoxia-induced mitophagy process through two-channel real-time imaging of NTR and viscosity under the same excitation.

Mitophagy is an essential physiological process that eliminates damaged mitochondria via lysosomes. It is reported that hypoxia, inflammatory stimuli or other stress conditions could lead to mitochondrial damage and mitochondrial dysfunction, which induces the process of mitophagy. Herein, we report a novel fluorescent probe PC-NTR for imaging hypoxia-induced mitophagy by monitoring the change of nitroreductase and viscosity simultaneously. To our delight, PC-NTR could respond simultaneously to nitroreductase and viscosity at different fluorescence channels with no mutual interference under the same excitation wavelength. The fluorescence emission around 535 nm was enhanced dramatically after addition of nitroreductase while the fluorescence emission around 635 nm heightened as the viscosity increased. The probe would be able to selectively targeting of mitochondria in cells because of the positively charged pyridine salt structure of PC-NTR. The probe was successfully applied to assess the different levels of hypoxia and real-time imaging of mitochondrial autophagy in live cells. More importantly, using dual channel imaging, PC-NTR could be used to distinguish cancer cells from normal cells and was successfully applied to imaging experiments in HeLa-derived tumor-bearing nude mice. Therefore, PC-NTR would be an important molecular tool for hypoxia imaging and detecting solid tumors in vivo.