Mitophagy, as a crucial metabolic process, plays an essential role in maintaining cellular and tissue homeostasis. Various stresses especially hypoxia could improve intracellular reactive oxygen species (ROS) level to induce mitophagy. However, high-specific fluorescence imaging of mitophagy in living cells under hypoxia is still a challenge. Based on this, we report an azoreductase-responsive nanoprobe (termed Micelle@Mito-rHP@TATp, MCM@TATp) by encapsulating cationic spiropyrane derivative (Mito-rHP) to realize specific imaging of mitophagy in living cells under hypoxia. An azoreductase-responsive amphiphilic polymer, 1,2-distearoyl- sn-glycero-3-phosphoethanolamine-azo- N-[maleimide(polyethylene glycol-2000) (Mal-PEG2000-Azo-DSPE), was first self-assembled into a micelle in aqueous solution. Meanwhile, the synthetic Mito-rHP encapsulated into this formed micelle to construct MCM. By modifying the surface of MCM with cell-penetrating peptide (TATp) to form MCM@TATp, the nanoprobe could avoid endolysosomal trapping. Under hypoxic conditions, the azobenzene moiety-contained MCM@TATp would be disrupted by the highly expressed azoreductase, then the encapsulated Mito-rHP would be released. Since Mito-rHP is a mitochondria-targeted and pH-sensitive probe, thus it could target into mitochondria and displayed a desirable "off-on" fluorescence response to mitophagy during which mitochondria were regarded to undergo acidification. The results indicated that the MCM@TATp in our design could image mitophagy under hypoxia in high-specificity. As further application, we have also demonstrated that this MCM@TATp can perform well to realize mitophagy imaging under the photodynamic therapy (PDT) which can induce hypoxia in treatment of cancer. We expect this new strategy would be a powerful tool for hypoxia-related fundamental and clinical research.