An Endoplasmic Reticulum-targeting Hydroxyl Radical Fluorescence Probe for Imaging of Ferroptosis and Screening of Natural Protectants

The real-time and in situ detection of hydroxyl radicals (˙OH) in the endoplasmic reticulum (ER) is helpful to understand ferroptosis at its very early stage due to the crucial role of ˙OH in the ER in ferroptosis initiation. Herein, an ER-targeting ˙OH fluorescent probe (ER-OH) has been developed, which showed a large fluorescence increase at 645 nm in response to ˙OH. ER-OH was applied to monitor ferroptosis by fluorescence imaging, revealing a significant increase of the ˙OH level in the ER during this process. With the imaging of ER-OH, a high-throughput screening method was developed to evaluate the anti-ferroptosis activity of a series of natural protectants. Through this screening, the natural flavonoid derivative icariside I was found for the first time to be highly effective in inhibiting ferroptosis by direct scavenging of the excess cytotoxic oxides (e.g. ˙OH and lipid peroxides) and restoring the level of GPX4. ER-OH could also be used for in vivo imaging of ˙OH in a mouse tumor model. This work provides not only a new tool for ferroptosis monitoring but also a direct insight into the regulation mechanism of ferroptosis and development of new drugs for ferroptosis-related diseases.