Live-cell imaging with fluorogenic radical-trapping antioxidant probes reveals the onset and progression of...

Ferroptosis is a form of cell death involving the formation of lipid peroxyl radicals, with potential therapeutic applications. Sensitivity to ferroptosis is expected to vary in different organelles. To monitor in real time the onset and progression of lipid peroxidation in ferroptosis, here we report lipophilic fluorogenic radical-trapping antioxidants, embedding in endoplasmic reticulum, lysosomes, mitochondria and plasma membrane. We show that endoplasmic reticulum- and lysosome-embedding fluorogenic radical-trapping antioxidants are most effective in protecting from cell death. The onset of lipid peroxidation happens in the endoplasmic reticulum, with lipid hydroperoxide accumulating in Golgi-associated vesicles. Disintegration of these structures spreads lipid hydroperoxide intracellularly, acting as 'free radical embers'. Outwards migration of oxidized lipids to plasma membrane, the ultimate sink for oxidized lipids, was recorded. Our results underscore Golgi-associated structures as a site to regulate ferroptosis progression. The work further positions fluorogenic radical-trapping antioxidants as valuable tools for unravelling the dynamic subcellular progression of ferroptosis.