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Abstract
Purpose:
To evaluate the effects of oxidative stress and antioxidantson heme oxygenase-1 (HO-1) in cultured human retinal pigment epithelial (RPE) cells.
Methods:
Cultured RPE cells were challenged with different concentrations of hydrogen peroxide (H2 O2), and the HO-1 mRNA level was determined by RT-PCR after 24 hours and 48 hours of incubation independently. Additionally, the HO-1 mRNA level was measured after preincubating RPE cells with N-acetylcystein (NAC) as the antioxidant for 30 minutes and then challenging the cells with H2 O2.
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Figure 1.
Human retinal pigment epithelial (RPE) cells were cultured for 24 hours at various concentrations of H2 O2. RPE cell survival showed progressive decrease with increasing concentrations of H2 O2. The cell viability was determined using MTT assay. The experiment was performed three times independently.
Figure 2.
RT-PCR results of HO-1 mRNA expression in 24 hours culture. Human retinal pigment epithelial cells were cultured at various concentrations of H2 O2 with or without NAC for 24 hours. HO-1 mRNA expression increased with H2 O2 concentration increase. And after NAC treatment, HO-1 mRNA expression decreased in comparison to the cells without NAC treatment. The mRNA expression of HO-1 was normalized to GAPDH (house keeping gene). Data are shown as mean± SD of five experiments. ∗ p<0.05 versus control (H2 O2 0 μM, NAC 0 μM). † p<0.05 versus the cells treated with H2 O2 without NAC.
Figure 3.
RT-PCR results of HO-1 mRNA expression in 48 hours culture. Human retinal pigment epithelial cells were cultured at various concentrations of H2 O2 with or without NAC for 48 hours. HO-1 mRNA expression increased with H2 O2 concentration increase. And after NAC treatment, HO-1 mRNA expression decreased in comparison to the cells without NAC treatment. The mRNA expression of HO-1 was normalized to GAPDH (house keeping gene). Data are shown as mean± SD of five experiments. ∗ p<0.05 versus control (H2 O2 0 μM, NAC 0 μM). † p<0.05 versus the cells treated with H2 O2 without NAC.
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