Abstract
Purpose::
Development of immortalized mouse retinal pigmented epithelial cell lines that retain their in vivo phenotype, would aid in studies of ocular diseases including age related macular degeneration.
Methods::
RPE sheets and cells were isolated from 14 month old ERKOα and ERKOß mice and their C57Bl/6 wildtype littermates. We confirmed the presence of epithelial cell markers, ZO1, cytokeratin 8 and 18 by immunofluorescence staining. RPE65, an in vivo marker of RPE cells, was detected by real-time RT PCR, and western analysis. Cells were immortalized using human papilloma virus 16 (E6/E7). Following immortalization, the above markers as well as estrogen receptor (ER) subtypes ERα and ERß and extracellular matrix components were examined to determine if any changes occurred from immortalization.
Results::
RPE cells isolated from mice retain RPE-specific in vivo markers and morphology. There was no evidence of any alterations in the parameters that we examined including MMP-2, TIMP-2, collagen type IV, and ERα and ERß protein expression and ER copy number ratio.
Conclusions::
Immortalized mouse RPE cell lines can be isolated from either pharmacologically or genetically manipulated mice. Our cell lines retain their in vivo phenotype and will be useful to study mechanisms that may lead to ECM dysregulation in AMD.
Keywords: age-related macular degeneration • retinal pigment epithelium