J Korean Ophthalmol Soc.  2009 Aug;50(8):1247-1253. 10.3341/jkos.2009.50.8.1247.

Effects of Oxidative Stress and Antioxidant on the Expression of Heme Oxygenase-1 in Human RPE

Affiliations
  • 1Department of Ophthalmology, Inha University School of Medicine, Incheon, Korea. hschin@inha.ac.kr

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 (H2O2), 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 H2O2. RESULTS: The expression of the HO-1 mRNA level increased after H2O2 exposure, and this level was proportional to the increased H2O2 concentration (p<0.05). Expression of the HO-1 mRNA level decreased when the RPE cells were preincubated with NAC (p<0.05). CONCLUSIONS: In human RPE cells, the HO-1 level increased in proportion to the degree of oxidative stress and decreased with exposure to antioxidants. Expression of HO-1 may be helpful in preventing retinal disease, which occurs due to oxidative stresses such as age-related macular degeneration.

Keyword

Age related macular degeneration; Antioxidant; Heme oxygenase-1; Oxidative stress; Retinal pigment epithelium

MeSH Terms

Antioxidants
Heme
Heme Oxygenase-1
Humans
Hydrogen Peroxide
Macular Degeneration
Oxidative Stress
Retinal Diseases
Retinal Pigment Epithelium
Retinaldehyde
RNA, Messenger
Antioxidants
Heme
Heme Oxygenase-1
Hydrogen Peroxide
RNA, Messenger
Retinaldehyde

Figure

  • 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.


Reference

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