J Korean Ophthalmol Soc.  2016 Jan;57(1):106-112. 10.3341/jkos.2016.57.1.106.

Cytoprotective Effect of Polyphenolic Compounds against Oxidative Stress in Cultured Retinal Pigment Epithelial Cells

• Seo KH
• Yu SY
• Kwak HW

• KMID: 2213851
• DOI: http://doi.org/10.3341/jkos.2016.57.1.106

Abstract

PURPOSE
Grape seed-derived polyphenols (GSPs) provide a concentrated source of polyphenols having antioxidant capacity. In this study we investigated the cytoprotective effect of GSP against oxidative stress-induced cell damage in cultured human retinal pigment epithelial (RPE) cells.
METHODS
Cultured adult retinal pigment epithelium (ARPE)-19 cells were incubated with GSP from Vitis vinifera (0.1, 0.5, 1, 5 or 10 microg/mL) for 24 hours and treated with hydrogen peroxide (H2O2, 0.4 mM) for 24 hours to induce oxidative stress. Cell viability was measured using 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Intracellular reactive oxygen species (ROS) was quantified using 2',7'-dichlorofluorescein diacetate (DCF-DA) fluorescence.
RESULTS
The percentage of viable RPE cells was significantly lower in cultures treated with H2O2 0.4 mM than in control cultures. GSP significantly reduced H2O2-induced cell death in a dose dependent manner. GSP at 0.1, 0.5, 1, 5 and 10 microg/mL significantly reduced cell mortality due to the treatment with H2O2. Intracellular ROS production increased significantly in cultures treated with H2O2 0.4 mM compared with control. There was a significant dose-dependent decrease in intracellular ROS levels after treatment of RPE with GSP.
CONCLUSIONS
GSP, a natural polyphenolic compound, can protect RPE cells from H2O2-induced oxidative stress and reduce intracellular ROS production by scavenging free radicals. This suggests potential effects of polyphenolic compounds against retinal diseases associated with oxidative stress.

Keyword

Grape seed extract; Oxidative stress; Polyphenols; Proanthocyanidins; Retinal pigment epithelium

MeSH Terms

Adult
Cell Death
Cell Survival
Epithelial Cells*
Fluorescence
Free Radicals
Grape Seed Extract
Humans
Hydrogen Peroxide
Mortality
Oxidative Stress*
Polyphenols
Proanthocyanidins
Reactive Oxygen Species
Retinal Diseases
Retinal Pigment Epithelium
Retinaldehyde*
Vitis
Free Radicals
Grape Seed Extract
Hydrogen Peroxide
Polyphenols
Proanthocyanidins
Reactive Oxygen Species
Retinaldehyde

Figure

• 

  Figure 1. Cytotoxic effect of grape seed-derived polyphenols (GSP). The ARPE-19 cells were incubated with different concen-tration of GSP for 24 hours. (A) GSP showed cytotoxicity at 100 μ g/mL. (B) The asterisk (*) indicates p < 0.05 versus untreated control. ARPE = adult retinal pigment epithelium.

• 

  Figure 2. Grape seed-derived polyphenols (GSP) protected against H2 O2 cell death in ARPE-19 cells. (A) The ARPE-19 cells were treated with H2 O2 at dose of 200, 400, and 600 μ M for 24 hours. (B) The cells were pretreated with GSP for 24 hours then exposed to H2 O2 (400 μ M) for 24 hours. Data was expressed as a percentage of the untreated control. The aster-isk (*) indicates p < 0.05 versus untreated control (A), or H2 O2-induced cells without GSP pretreatment (B). ARPE = adult retinal pigment epithelium.

• 

  Figure 3. Grape seed-derived polyphenols (GSP) reduced H2 O2-induced intracellular reactive oxygen species (ROS) generation in ARPE-19 cells. (A) The ARPE-19 cells were treated with H2 O2 at dose of 200, 400, and 600 μ M for 24 hours. (B) The cell were pretreated with GSP for 24 hours then treated with 400 μ M for 4 hours. The intracellular ROS was examined by a fluorescence spectrophotometer using DCF- DA. Data was expressed as a percentage of the untreated control. The asterisk (*) indicates p < 0.05 versus untreated control (A), or H2 O2-induced cells without GSP pretreatment (B). ARPE = adult retinal pigment epithelium; DCF-DA = 2′,7′-dichlorofluorescein diacetate.

• 

  Figure 4. Morphology of ARPE-19 cells exposed to H2 O2 (400 μ M) with/without grape seed-derived polyphenols (GSP). (A) 400 μ M H2 O2 without GSP. After 24-hour exposure, (B) 400 μ M H2 O2 with GSP. ARPE = adult retinal pigment epithelium.

Reference

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