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Shu-Zhen Wang, Run-Tao Yan; RPE for photoreceptor regeneration in mouse eye. Invest. Ophthalmol. Vis. Sci. 2013;54(15):1764. doi: https://doi.org/.
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Current research on producing mammalian photoreceptors, as regenerative medicine in general, focuses on ES cells and iPS cells. This project studies a different approach - tapping the naturally occurring wound healing mechanism of RPE for photoreceptor regeneration. Previous experiments found extra photoreceptor cells in transgenic mice generated with a DNA cassette expressing ngn1 from RPE promoter PVMD2. This study aims to determine RPE as a source of the extra photoreceptor cells.
Transgenic mice were generated with a DNA construct, in which RPE bestrophin-1 promoter (PVMD2) would drive the expression of ngn1, a regulatory gene with pro-photoreceptor activity. Eyes were analyzed for the presence of cells in “RPE-to-photoreceptor” transition with histology and immunohistology.
Cells in “RPE-to-photoreceptor” transition were detected in transgenic eyes. Some darkly pigmented cells, as well as discrete cells in the RPE, were recoverin+. Morphologically, these recoverin+ cells were more similar to RPE cells than to photoreceptor cells. In addition to their pigment granules, which otherwise are typically present in RPE cells, these recoverin+ cells displayed a nucleus that appeared (a) round (as in RPE cells) vs. elongated (as in photoreceptors), (b) large vs. compact, and (c) contained several discrete, brightly fluorescent spots of DAPI staining. Notably, in a 9-month-old PVMD2-ngn1 animal, anti-recoverin immunoreactivity was detected in essentially all cells within a particular region of the RPE. Nearby, a layer, reminiscent of an emerging extra photoreceptor layer, was present in the subretinal space. Cells in this layer showed pigment granules, as seen in RPE cells. Yet, they were recoverin+ and displayed an elongated cell body. At the tip of the emerging layer, a well defined extra photoreceptor layer was apparent. Immunostaining showed that the RPE in the region became negative for Otx2, a homeodomain factor involved in maintaining RPE vs. retina identity and no longer expressed in RPE-to-retina transdifferentiation in newt.
These results suggest that RPE-to-photoreceptor reprogramming was occurring in transgenic mouse and, thus, RPE was a source of the extra photoreceptor cells present in the transgenic mouse. The presence of cells in “RPE-to-photoreceptor” transition in a 9-month-old transgenic animal suggests that the reprogramming could occur in aged eyes.
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