Imaging HOCl Generation during the Mitochondria Peripheral Fission with a Tailor-Made Fluorescent Probe

Mitochondrial fission is a highly regulated process that can affect metabolism, proliferation, and Apoptosis. Division at the periphery enables damaged material to be shed into smaller mitochondria destined for Mitophagy, which is found preceded by increased Ca²⁺ and Reactive Oxygen Species, as well as reduced membrane potential and pH. However, the variation of hypochlorous acid (HOCl) during the peripheral fission has not been well studied, and the existing fluorescent probes are unsuitable for detecting mitochondrial HOCl because of the 0.8-fold decreased pH during this process. Herein, we design a novel CCS (changeable π-conjugation system)-based probe (ON-mito) with a dibenzo[1,4]oxazepine core, which can selectively react with HOCl at pH 6.4, generating an oxazine-containing product that emits at 660 nm. The capability of ON-mito for imaging the HOCl generation in HeLa cells during Mitophagy is demonstrated under weakly acidic condition. Further, with ON-mito, we find for the first time a burst increase of the mitochondrial HOCl in COS-7 cells during peripheral fission, which may serve as an important indicator of this process. Probe ON-mito may be useful for studying mitochondrial damage under diverse conditions.