Abstract
Purpose :
Age-related macular degeneration (AMD) leads to blindness in millions of individuals globally, primarily due to the degeneration and demise of the retinal pigment epithelium (RPE). At present, there are no effective treatments available to regenerate lost RPEs as a result of AMD. Pluripotent stem cells emerged as an attractive method to replace lost RPEs and restore vision in these patients. However, these methods are cumbersome, time consuming and outcome with variable quality of RPEs making it challenging for clinical applications. Developing therapies that specifically target the stimulation of endogenous RPE regeneration offers a compelling approach to treat AMD and other RPE degenerative diseases. In this study, we present a straightforward chemical-based approach to stimulate RPE regeneration from Muller cells both in vitro and in vivo.
Methods :
A combination of a six-chemical cocktail plus two factors was administered as eye drops in both normal and sodium iodate-injured mice, allowing observation of subsequent responses in Muller cells, including proliferation, migration, and differentiation using immunofluorescence. To track cell lineages, an AAV2-GLAST-GFP reporter and GLAST-CreERT;Ai9 mice were employed following the administration of the eye drops. In addition, human primary Muller cells were subjected to the chemical cocktail for in vitro reprogramming and were analyzed using immunofluorescence, alongside a Lenti-BEST1-GFP promoter reporter.
Results :
We observed chemical cocktail eyedrop application can effectively regenerate RPEs from Müller cells, both uninjured and sodium iodate injured retina. These RPEs displayed a cobblestone-like morphology, express bestrophin1 (BEST1) and were highly pigmented, indicating their resemblance to bonafide RPEs. Moreover, the same chemical cocktail was able to reprogram human primary Muller cells into hexagonal, pigmented, and RPE65+ and BEST1-GFP+ RPE cells. Lineage tracing experiments utilizing AAV2 GLAST-GFP and GLAST-CRE-Tdtomato mice provided confirmation of the Müller cell origin of the newly generated RPEs
Conclusions :
We successfully developed an endogenous RPE regeneration protocol from Müller cells using a chemical cocktail both in vivo and in vitro.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.