17-β estradiol protects ARPE-19 cells from oxidative stress through estrogen receptor-β.

Giddabasappa A, Bauler M, Yepuru M, Chaum E, Dalton JT, Eswaraka J
Invest Ophthalmol Vis Sci. 2010 51 (10): 5278-87

PMID: 20463317 · DOI:10.1167/iovs.10-5316

PURPOSE - To elucidate the mechanism of 17-β estradiol (17β-E(2))-mediated protection of retinal pigment epithelium (RPE) from oxidative stress.

METHODS - Cultured ARPE-19 cells were subjected to oxidative stress with t-butyl hydroxide or hydrogen peroxide in the presence or absence of 17β-E(2). Reactive oxygen species (ROS) were measured using H(2)DCFDA fluorescence. Apoptosis was evaluated by cell-death ELISA kit and Hoechst-3486 staining. Mitochondrial membrane potential was measured using the JC-1 assay. Cellular localization of estrogen receptor (ER) was evaluated by confocal microscopy. Gene expression and protein expression was quantified using qRT-PCR and western blotting. Superoxide dismutase and ATP levels were measured using commercial kits.

RESULTS - ARPE-19 cells expressed significant amounts of ERα and ERβ. Pretreatment with 17β-E2 protected ARPE-19 cells from oxidative stress and apoptosis. 17β-E(2) reduced the ROS levels and mitochondrial depolarization. The 17β-E(2)-mediated cytoprotection was inhibited by ER antagonists ICI (ERα and ERβ) and THC (ERβ) but not by tamoxifen (ERα). Knockdown of ERβ expression by siRNA abolished the protective effects of 17β-E(2). Further, qRT-PCR analysis revealed that 17β-E(2) pretreatment upregulated the expression of ERβ and phase II cellular antioxidant genes.

CONCLUSIONS - These results indicate that 17β-E(2) protects ARPE-19 cells from oxidative stress through an ERβ-dependent mechanism. 17β-E(2)-mediated cytoprotection occurred through the preservation of mitochondrial function, reduction of ROS production, and induction of cellular antioxidant genes.

MeSH Terms (21)

Adenosine Triphosphate Apoptosis Blotting, Western Cells, Cultured Cytoprotection Enzyme-Linked Immunosorbent Assay Estradiol Estrogen Receptor beta Estrogens Humans Hydrogen Peroxide Membrane Potential, Mitochondrial Microscopy, Confocal Mitochondria Oxidative Stress Reactive Oxygen Species Retinal Pigment Epithelium Reverse Transcriptase Polymerase Chain Reaction RNA, Small Interfering Superoxide Dismutase tert-Butylhydroperoxide

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