Original ContributionLipofuscin-bound iron is a major intracellular source of oxidants: Role in senescent cells
Section snippets
Materials
If not indicated differently all chemicals were obtained from Sigma (Deisenhofen, Germany) and the cell culture materials were purchased from Biochrom (Berlin, Germany). The caspase-3 assay kit from BD Bioscience (Pharmingen, San Diego, CA, USA) was used.
Cell culture
Experiments were performed using human dermal fibroblasts obtained from skin tissue. Cells were cultured in Dulbecco's MEM supplemented with 10% fetal calf medium, 1% penicillin/streptomycin, and 1% Glutamax at 37°C in a humidified 5% CO2
Formation of free radicals in young and SIPS cells with and without inhibition of the respiratory chain
Cellular senescence is often considered to be accompanied by an enhanced formation of oxidants. Therefore, we tested whether our SIPS fibroblasts also have this phenotype. As shown in Fig. 1A, the amount of free radicals in SIPS cells, quantified via DCF fluorescence, is significantly increased (up to 310%) compared to unexposed young human fibroblasts. After inhibition of the respiratory chain (with antimycin A and rotenone for 24 h) the amount of DCF fluorescence is still significantly
Discussion
Keeping the cytosol clear of the consequences of oxidative damage is one main function of the lysosomal and proteasomal systems. The capacity of these systems decreases over time in postmitotic cells, in several pathologies, and during oxidative stress [12]. The mitochondria are the main intracellular source of oxidative stress [29], contributing about 50%, whereas other main enzymes are xanthine- and NADPH-oxidase [43]. The main primary radical generated by the three systems mentioned is the
Acknowledgment
TG was supported by the DFG.
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