Abstract
The study was aimed at evaluation of the role of secondary oxidative stress in the stress-induced premature senescence (SIPS) of human fibroblasts induced by H2O2. Two fibroblast lines were used: lung MRC-5 and ear H8F2p25LM fibroblasts. The lines differed considerably in sensitivity to H2O2 (IC50 of 528 and 33.5 μM, respectively). The cells were exposed to H2O2 concentrations corresponding to IC50 and after 24 h supplemented with a range of antioxidants. Most of antioxidants studied slightly augmented the survival of fibroblasts at single concentrations or in a narrow concentration range, but the results were not consistent among the cell lines. Chosen antioxidants (4-amino-TEMPO, curcumin, caffeic acid and p-coumaric acid) did not restore the level of glutathione decreased by H2O2. Hydrogen peroxide treatment did not induce secondary production of H2O2 and even decreased it, decreased mitochondrial potential in both cell lines and induced changes in the mitochondrial mass inconsistent between the lines. Antioxidant protected mitochondrial potential only in H8F2p25LM cells, but attenuated changes in mitochondrial mass. These results speak against the intermediacy of secondary oxidative stress in the SIPS induced by H2O2 and suggest that the small protective action of antioxidants is due to their effects on mitochondria.
Highlights
Aging is an irreversible process affecting all higher organisms, characterized by progressive deterioration leading to a loss of function of cells, tissues, organs and death
Hydrogen peroxide showed a dose-dependent cytotoxicity against normal human fibroblast line [MRC-5 (CCL-171)] obtained from lung and primary human fibroblast line [H8F2p25LM] obtained from ear skin of an adult donor (Figure 1A, 1B)
24 hours after H2O2 treatment, antioxidants were added to the cells to study their effects on the processes dependent on secondary oxidant-dependent signaling leading to decrease in cell survival
Summary
Aging is an irreversible process affecting all higher organisms, characterized by progressive deterioration leading to a loss of function of cells, tissues, organs and death. Cellular aging can be accelerated by using non-lethal stresses; this phenomenon is referred to as stress induced premature senescence (SIPS). Premature aging of cultured cells is usually associated with the exposure of cells to environmental stress factors. Stress induced premature senescence can be defined as the long-term effect of sub-cytotoxic stress on proliferative cell types including appearance of many features of replicative senescence. Various genotoxic agents, such as hydrogen peroxide (H2O2), tert-butyl hydroperoxide, copper sulfate, diperoxovanadate, ethanol, mitomycin C, other cytostatic drugs, heat shock or UV radiation are wellestablished inducers of SIPS. It has been suggested that SIPS can be used in toxicology to identify xenobiotics that may induce premature senescence. Oxidative stress is believed to be the major cause of SIPS program activation in normal cells [2,3,4,5,6]
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