Epigallocatechin Gallate (EGCG) Prevents H₂O₂-Induced Oxidative Stress in Primary Rat Retinal Pigment Epithelial Cells

Abstract

Purpose: To determine whether the green tea polyphenol epigallocatechin gallate (EGCG) could prevent H₂O₂-induced oxidative stress in primary rat retinal pigment epithelial cells.

Methods: Primary cultures of retinal pigment epithelium (RPE) cells were established from Long–Evans newborn rats. RPE cells were pretreated with various concentrations of EGCG for 24 h before being exposed to hydrogen peroxide (H₂O₂) for 2 h to induce oxidative stress. Cell metabolic activity was measured using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell death was quantified by flow cytometry using propidium iodide (PI).

Results: Treatment of RPE cells with EGCG alone does not affect the cell viability up to 50 µM. Exposure of RPE cells to 600 µM H₂O₂ caused a significant decrease in cell viability; whereas pretreatment with 10, 25, and 50 µM EGCG significantly reduced this decrease in a dose-dependent manner. The proportion of PI-positive cells increased significantly in cultures treated with H₂O₂ alone; whereas pretreatment of RPE cells with 50 µM EGCG significantly reduced H₂O₂-induced RPE cell death.

Conclusions: Our study shows that EGCG pretreatment can protect primary rat RPE cells from H₂O₂-induced death. This suggests potential effect of EGCG in the prevention of retinal diseases associated with H₂O₂-induced oxidative stress.

• Age-related macular degeneration
• epigallocatechin gallate
• hydrogen peroxide
• oxidative stress
• primary retinal pigment epithelial cell

Acknowledgements

We thank Arlette Tridon (Laboratoire d’Immunologie UMR 1019, UFR Pharmacie, Clermont Université, France) for allowing us to use their microplate reader, and Marie-Paule Vasson (Laboratoire de Biochimie Biologie Moléculaire et Nutrition UMR 1019, UFR Pharmacie, Clermont Université, France) for allowing us to use their flow cytometer.

This article was presented in part at the ARVO annual meeting, Fort-Lauderdale, FL, USA, 1 May–5 May 2011.