To test whether the possibly enhanced sensitivity of aged cells to oxidative stress may depend on their content of ceroid/lipofuscin, AG-1518 human fibroblasts with various amounts of the pigment accumulated due to prolonged cultivation under normobaric hyperoxia were exposed to acute oxidative stress (2.5 microM naphthazarin, 15 min) and then returned to standard culture conditions. Twenty-four hours after the naphthazarin treatment, 37% of the cells were still vital, whereas others had undergone oxidative stress-induced apoptosis with ensuing postapoptotic necrosis. The average amount of ceroid/lipofuscin within the surviving cells was only about half of that of the initial population of cells, as measured before the naphthazarin exposure. This finding suggests that ceroid/lipofuscin-rich cells have an increased sensitivity to oxidative stress. The ceroid/lipofuscin quantity strongly positively correlated with the size of the acidic compartment (as evaluated by uptake of the weakly basic lysosomotropic fluorochrome acridine orange) and with its content of the lysosomal protease cathepsin D, as assayed by immunocytochemistry. We hypothesize that the enhanced sensitivity of ceroid/lipofuscin-loaded cells to oxidative stress may be caused by the increased amounts of lysosomal enzymes, known as mediators of oxidative damage, and/or by catalysis of intralysosomal oxidative reactions by lipofuscin-associated iron.