Search

Glaucoma is the leading cause of irreversible blindness and is a major health and economic burden. Current treatments do not address the neurodegenerative component of glaucoma. In animal models of glaucoma, the capacity to maintain retinal nicotinamide adenine dinucleotide (NAD) pools declines early during disease pathogenesis. Treatment with nicotinamide, an NAD precursor through the NAD salvage pathway, robustly protects against neurodegeneration in a number of glaucoma models and improves vision in existing glaucoma patients. However, it remains unknown in humans what retinal cell types are able to process nicotinamide to NAD and how these are affected in glaucoma. To address this, we utilized publicly available RNA-sequencing data (bulk, single cell, and single nucleus) and antibody labelling in highly preserved enucleated human eyes to identify expression of NAD synthesizing enzyme machinery. This identifies that the neural retina favors expression of the NAD salvage pathway, and that retinal ganglion cells are particularly enriched for these enzymes. NMNAT2, a key terminal enzyme in the salvage pathway, is predominantly expressed in retinal ganglion cell relevant layers of the retina and declines in glaucoma. These findings suggest that human retinal ganglion cells can directly utilize nicotinamide and could maintain a capacity to do so in glaucoma, showing promise for ongoing clinical trials.

In vitro differentiation of human pluripotent stem cells into functional retinal pigment epithelial (RPE) cells provides a potentially unlimited source for cell based reparative therapy of age-related macular degeneration. Although the inherent pigmentation of the RPE cells have been useful to grossly evaluate differentiation efficiency and allowed manual isolation of pigmented structures, accurate quantification and automated isolation has been challenging. To address this issue, here we perform a comprehensive antibody screening and identify cell surface markers for RPE cells. We show that these markers can be used to isolate RPE cells during in vitro differentiation and to track, quantify and improve differentiation efficiency. Finally, these surface markers aided to develop a robust, direct and scalable monolayer differentiation protocol on human recombinant laminin-111 and −521 without the need for manual isolation., Whilst pigmentation has been used to identify retinal pigment epithelial (RPE) cells, surface markers for these cells remain unclear. Here, the authors define surface markers for the RPE including CD140b, which help produce hPSC-derived RPE cells at a large scale following a robust, direct and scalable monolayer differentiation protocol.