Abstract: Glaucoma is characterized by chronic progressive optic nerve damage and retinal ganglion cell death. Although extensive research has been conducted on neuroprotection for retinal ganglion cells, there is still no treatment for clinical use. Recent evidence shows that extracellular vesicles isolated from a variety of stem cells are efficacious in retinal ganglion cell neuroprotection. In this study, we tested the novel extracellular vesicle source of the retinal progenitor R-28 cell line in vitro and in vivo. We isolated and characterized extracellular vesicles from R-28 cells and tested their therapeutic efficacy in terms of retinal ganglion cell survival in vitro and in an in vivo glaucoma model, measuring retinal ganglion cell survival and preservation of their axons. Additionally, we tested extracellular vesicles for their neuroprotective capacity in retinal ganglion cells differentiated from human embryonic stem cells. Finally, we investigated miRNA changes in retinal ganglion cells with R-28 extracellular vesicle treatment, and predicted possible pathways that may be modulated. R-28 extracellular vesicles improved retinal ganglion cell survival but failed to preserve axons significantly. Moreover, the results also illustrated the neuroprotection of R-28 extracellular vesicles on human retinal ganglion cells. Finally, we also showed changes in hsa-miRNA-4443, hsa-miRNA-216a-5p, hsa-let-7e-5p, hsa-miRNA-374b-5p, hsa-miRNA-331-3p, and hsa-miRNA-421 expressions, which may have neuroprotective potential on retinal ganglion cell degeneration. This study will pave the way for miRNA and extracellular vesicle-based neuroprotective therapies for glaucoma.

Key words: extracellular vesicles, glaucoma, miRNA, neuroprotection, R-28 cell line, retinal ganglion cells