Abstract
Purpose:
Regenerating neurons and neural connection has been selected as NEI Audacious Goal. Our laboratory investigates the possibility of using a regulatory gene, ngn1 or ngn3, to reprogram the RPE for retinal regeneration in the mouse eye. In previous studies we found ectopic retinal tissue/cells in some PVMD2-ngn1 and PRPE65-ngn3 transgenic mice. This study examines whether (1) the RPE in adult mouse could undergo the gene-directed reprogramming, (2) the RPE, which plays a crucial role in the function and health of the retina, was preserved in those transgenic animals, and (3) Müller glia was involved.
Methods:
Cryosections of eyes from adult transgenic mice with photoreceptor-like cells in the subretinal space were immunostained with antibodies that recognize RPE proteins, photoreceptor protein Recoverin (Rcv), or Müller glia. The presence of a hybrid or mixed cell type, i.e., a cell co-expressing markers that otherwise are exclusive for either RPE or photoreceptor cells, was used as an indication of the cell amid RPE-to-photoreceptor reprogramming process.
Results:
At places with Rcv+ cells in the subretinal space, the highly melanized RPE was present and was immuno-positive for RPE proteins: CRALBP, Cytokeratin-18, Ezrin, and RPE65. Ezrin and RPE65 were also detected in some of the ectopic Rcv+ cells, especially in those containing some pigment granules, which otherwise are abundantly present in normal RPE cells. Otx2, a transcriptional factor known to be important for RPE differentiation and maintenance of RPE properties and to be down-regulated during the RPE-to-retina transdifferentiation observed in amphibians and chicken, was detected in the RPE at most places and not in regions where RPE was seemingly undergoing RPE-to-photoreceptor reprogramming. Some cells within the territory occupied by the subretinal Rcv+ cells were immuno-positive for Müller glia makers GS and CRALBP, and a few Rcv+/GS+ double-labeled cells were observed.
Conclusions:
The results suggest that the RPE in adult mice can be guided by ngn1 or ngn3 to produce retinal cells, including Müller glia, and can regenerate itself afterward. This raises a possibility using the RPE as stem-like cells for retinal regeneration in adult mammals.