Super-Resolution Imaging Captures Mitophagy Dynamics: Tricolor Fluorescence Labels Mitochondria, Autophagosomes, and Lysosomes.

Anal Chem

Shandong Provincial Key Laboratory of Tumor Imaging Equipment Development and Integrated Diagnosis and Treatment Technology, Linyi University, Linyi 276000, China.

Published: September 2025


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Article Abstract

Mitophagy is a vital lysosome-dependent process in which damaged mitochondria exhibiting elevated HO production are selectively engulfed by autophagosomes and delivered to lysosomes for degradation, thereby maintaining intracellular homeostasis. Consequently, monitoring mitophagy holds significant potential for disease diagnosis and therapeutic development. In this study, HO-activated lysosome-targeted fluorescent probe, , was developed for the super-resolution imaging of the mitophagic process. The initiation of mitophagy induces the significant production of autophagosomes. Red-stained mitochondria are engulfed by blue-stained autophagosomes and subsequently fuse with lysosomes, leading to an increase in the HO concentration within the lysosomes. This activates the green fluorescence signal of . The entire mitophagic process can be visualized and monitored by observing changes in the fluorescence intensity of the autophagosomes and lysosomes as well as the overlap of fluorescence signals. Super-resolution imaging enables the triple-signal nanoscale visualization of the dynamic processes in three organelles. This probe was successfully applied to monitor rapamycin- and starvation-induced mitophagy and was further utilized to assess the role of mitophagy inhibition in enhancing antitumor activity.

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http://dx.doi.org/10.1021/acs.analchem.5c03247DOI Listing

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