title: Polarized human embryonic stem cell-derived retinal pigment epithelial cell monolayers have higher resistance to oxidative stress-induced cell death than nonpolarized cultures.
abstract: Oxidative stress-mediated injury to the retinal pigment epithelium (RPE) is a major factor involved in the pathogenesis of age-related macular degeneration (AMD), the leading cause of blindness in the elderly. Human embryonic stem cell (hESC)-derived RPE cells are currently being evaluated for their potential for cell therapy in AMD patients through subretinal injection of cells in suspension and subretinal placement as a polarized monolayer. To gain an understanding of how transplanted RPE cells will respond to the highly oxidatively stressed environment of an AMD patient eye, we compared the survival of polarized and nonpolarized RPE cultures following oxidative stress treatment. Polarized, nonpolarized/confluent, nonpolarized/subconfluent hESC-RPE cells were treated with H2O2. Terminal deoxynucleotidyl transferase dUTP nick end labeling stains revealed the highest amount of cell death in subconfluent hESC-RPE cells and little cell death in polarized hESC-RPE cells with H2O2 treatment. There were higher levels of proapoptotic factors (phosphorylated p38, phosphorylated c-Jun NH2-terminal kinase, Bax, and cleaved caspase 3 fragments) in treated nonpolarized RPE-particularly subconfluent cells-relative to polarized cells. On the other hand, polarized RPE cells had constitutively higher levels of cell survival and antiapoptotic signaling factors such as p-Akt and Bcl-2, as well as antioxidants superoxide dismutase 1 and catalase relative to nonpolarized cells, that possibly contributed to polarized cells' higher tolerance to oxidative stress compared with nonpolarized RPE cells. Subconfluent cells were particularly sensitive to oxidative stress-induced apoptosis. These results suggest that implantation of polarized hESC-RPE monolayers for treating AMD patients with geographic atrophy should have better survival than injections of hESC-RPE cells in suspension.
authors.affiliation: Departments of Pathology and Ophthalmology, Keck School of Medicine of the University of Southern California, Los Angeles, California, USA.
authors.forename: Jamie
authors.initials: J
authors.lastname: Hsiung
authors.affiliation: Departments of Pathology and Ophthalmology, Keck School of Medicine of the University of Southern California, Los Angeles, California, USA.
authors.forename: Danhong
authors.initials: D
authors.lastname: Zhu
authors.affiliation: Departments of Pathology and Ophthalmology, Keck School of Medicine of the University of Southern California, Los Angeles, California, USA dhinton@usc.edu.
authors.forename: David R
authors.initials: DR
authors.lastname: Hinton
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D016764:Cell Polarity; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects; D006801:Humans; D006861:Hydrogen Peroxide / Q000633:toxicity; D020287:In Situ Nick-End Labeling; D016877:Oxidants / Q000633:toxicity; D018384:Oxidative Stress / Q000502:physiology*; D060888:Real-Time Polymerase Chain Reaction; D055213:Retinal Pigment Epithelium / Q000166:cytology* / Q000187:drug effects; D023822:Tissue Engineering / Q000379:methods
chemical_list: D016877:Oxidants; D006861:Hydrogen Peroxide
keywords: Age-related macular degeneration; Apoptosis; Embryonic stem cells; Oxidative stress; Polarized retinal pigment epithelial cell; Retinal pigment epithelial cell