BACKGROUND: ? Normal human epidermal keratinocytes (NHEKs) and HaCaT cells are the most common models used to study the effects of various factors on skin cells. These cell lines share some common characteristics, but little is known about their differences in handling hydrogen peroxide (H(2) O(2) )-induced oxidative stress. AIM: To investigate the differential response of NHEKs and HaCaT cells to H(2) O(2) -induced oxidative stress. METHODS: We examined differences in NHEKs and HaCaT cells after H(2) O(2) treatment, assessing changes in cell viability; levels of intracellular reactive oxygen species (ROS), superoxide dismutase (SOD) and caspase-3/7; percentage of cells arrested in G1 phase; number of senescence-associated β-galactosidase (SA-β-Gal)-positive cells and; expression of senescence-related protein Klotho. RESULTS: The viability of NHEKs and HaCaT cells decreased in a concentration-dependent and time-dependent manner after exposure to H(2) O(2) . The inhibitory effect of 150 μmol/L H(2) O(2) on cell viability was greater in HaCaT cells than in NHEKs (P<0.05). Levels of ROS and caspase-3/7, and the percentage of cells arrested in G1 phase, were higher in HaCaT cells than in NHEKs, whereas intracellular SOD was higher in NHEKs than in HaCaT cells after exposure to 150 μmol/L H(2) O(2) (P < 0.05). SA-β-Gal positive cells increased significantly in NHEKs after treatment with H(2) O(2) (P < 0.05). Klotho was significantly downregulated in both NHEKs and HaCaT cells after H(2) O(2) treatment, but no SA-β-Gal-positive HaCaT cells were seen, even after treatment with H(2) O(2) . CONCLUSIONS: Normal human epidermal keratinocytes are more resistant than HaCaT cells to H(2) O(2) -induced oxidative stress. HaCaT cells have senescence phenotypes, but do not express β-Gal.

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