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Anand Giddabasappa, Matthew N. Bauler, Christina M. Barrett, Christopher C. Coss, Zhongzhi Wu, Duane D. Miller, James T. Dalton, Jeetendra R. Eswaraka; GTx-822, an ERβ-Selective Agonist, Protects Retinal Pigment Epithelium (ARPE-19) from Oxidative Stress by Activating MAPK and PI3-K Pathways. Invest. Ophthalmol. Vis. Sci. 2010;51(11):5934-5942. doi: 10.1167/iovs.10-5630.
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The goal of this study was to determine whether an estrogen receptor-β (ERβ)-selective agonist (GTx-822; GTx, Inc., Memphis, TN) could prevent hydrogen peroxide (H2O2)-induced oxidative stress in ARPE-19 cells and to elucidate the molecular pathways involved in this protection.
The selectivity of GTx-822 for ERβ was determined by receptor-binding assay (RBA) and transactivation assay. Cultured ARPE-19 cells were subjected to oxidative stress with t-butyl hydroxide (t-BH) or hydrogen peroxide (H2O2) in the presence and absence of GTx-822. Reactive oxygen species (ROS) was measured by using H2DCFDA fluorescence. Apoptosis was evaluated by cell death ELISA. Mitochondrial membrane potential was measured with the JC-1 assay. Gene expression and protein expression and activation were quantitated with qRT-PCR and Western blot analysis. Phospho-protein arrays elucidated the activation of protein kinases.
The RBA and transactivation assay revealed that GTx-822 is an ERβ-selective agonist (K i = 0.53 nM). GTx-822 prevented oxidative stress in ARPE-19 cells. It preserved mitochondrial function and prevented cellular apoptosis. Pretreatment with GTx-822 increased ERβ gene and protein expression during oxidative stress. Upregulation of the phase II antioxidant genes GPx-2 and HO-1 was also seen in an ERβ-dependent mechanism. GTx-822 pretreatment induced phosphorylation of ERK1/2, PI3-K, and Bad.
This is the first report to show that GTx-822, an ERβ agonist, can protect ARPE-19 cells from the cellular apoptosis induced by oxidative stress. GTx-822 mediated cytoprotection was mediated through induction of both genomic and nongenomic pathways. The results of this study open new avenues for the use of a selective ERβ agonist in treatment of ocular diseases like AMD where oxidative stress plays a major role in disease pathogenesis.
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