mRNA-transfection of Retinal Pigmented Epithelial cells 1 Efficient delivery and functional expression of transfected modified mRNA in human Embryonic Stem Cell-derived Retinal Pigmented Epithelial cells

Geneand cell-based therapies are promising strategies for the treatment of degenerative retinal diseases such as age-related macular degeneration (AMD), Stargardt disease and Retinitis Pigmentosa. Cellular engineering before transplantation may allow the delivery of cellular factors that can promote functional improvements such as increased engraftment or survival of transplanted cells. A current challenge in traditional DNA-based vector transfection is to find a delivery system that is both safe and efficient, but using mRNA as an alternative to DNA can circumvent these major roadblocks. In this study, we show that both unmodified and modified mRNA can be delivered to Retinal Pigmented Epithelial (RPE) cells with a high efficiency, when compared with conventional plasmid delivery systems. On the other hand, administration of unmodified mRNA induced a strong innate immune response, which was almost absent when using modified mRNA. Importantly, transfection of mRNA encoding a key regulator of RPE gene expression, Microphthalmia-associated transcription factor (MITF), confirmed the functionality of the delivered mRNA. Immunostaining showed that transfection with either type of mRNA led to the expression of roughly equal levels of MITF, primarily localized in the http://www.jbc.org/cgi/doi/10.1074/jbc.M114.618835 The latest version is at JBC Papers in Press. Published on January 2, 2015 as Manuscript M114.618835 Copyright 2015 by The American Society for Biochemistry and Molecular Biology, Inc. by gest on A uust 1, 2017 hp://w w w .jb.org/ D ow nladed from mRNA-transfection of Retinal Pigmented Epithelial cells