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S. J. Upadhyay, J. A. Hossain, M. P. Krebs, P. Baciu, S. Kaushal; Effect of HDAC Inhibitors on Oxidative Apoptosis of RPE Cells. Invest. Ophthalmol. Vis. Sci. 2010;51(13):496.
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© ARVO (1962-2015); The Authors (2016-present)
To determine the effect of the HDAC inhibitors valproic acid (VPA), 4-phenylbutyrate (PBA) and the VPA analog valpromide (VPD) on the survival and function of RPE cells in response to oxidative stress and complement attack both in vitro and in vivo.
ARPE-19 cells were cultured by standard methods. For hydroquinone (HQ)-induced death, HQ was used at concentrations previously established to induce RPE oxidative damage (250-450 µM). ARPE-19 cells were then pretreated for 24 hrs with various concentrations of VPA, VPD, or PBA, followed by re-treatment and challenged with HQ for 48 hrs. Apoptosis and necrosis were assayed by dual Annexin V and propidum iodide staining, as well as cleaved caspase-3 analysis via flow cytometry. For complement-induced death, pretreated ARPE-19 cells were primed with a complement-fixing antibody, followed by treatment with C1q-deficient serum in the presence or absence of conditioned media, and level of cell death was monitored by accumulation of a cell membrane impermeant nuclear stain. VPA effects in vivo were also investigated. Oxidative stress was induced by intravitreal paraquat injection in SOD1+/- mice, and VPA protection was determined by ERGs.
Dose-dependent ARPE-19 cell death was observed at all concentrations of HQ, with more than 80% of cells dead at concentrations of 350 µM and above. Both VPA and VPD significantly increased cell viability in response to HQ challenge, and VPA reduced apoptosis and downstream caspase-3 activation. VPA and VPD also protected against complement-mediated cell death, however conditioned media from pretreated cells was required for protection. VPA also protected against paraquat-induced oxidative stress in vivo.
VPA can reduce RPE cell death in in vitro models of oxidative injury and complement attack and in vivo against paraquat-induced oxidative stress. This protective and anti-apoptotic activity of VPA on the RPE and photoreceptors supports the use of VPA in clinical ARMD studies.
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