PT  - JOURNAL ARTICLE
AU  - Venkata R. M. Chavali
AU  - Naqi Haider
AU  - Sonika Rathi
AU  - Vrathasha Vrathasha
AU  - Teja Alapati
AU  - Jie He
AU  - Kamaljot Gill
AU  - Roman Nikonov
AU  - Thu T. Duong
AU  - Devin S. McDougald
AU  - Sergei Nikonov
AU  - Joan O’ Brien
AU  - Jason A. Mills
TI  - Dual SMAD inhibition and Wnt inhibition enable efficient and reproducible differentiations of induced pluripotent stem cells into retinal ganglion cells
AID  - 10.1101/682666
DP  - 2020 Jan 01
TA  - bioRxiv
PG  - 682666
4099  - http://biorxiv.org/content/early/2020/05/30/682666.short
4100  - http://biorxiv.org/content/early/2020/05/30/682666.full
AB  - Glaucoma is a group of progressive optic neuropathies that share common biological and clinical characteristics including irreversible changes to the optic nerve and visual field loss caused by death of retinal ganglion cells (RGCs). The loss of RGCs manifests as characteristic cupping or optic nerve degeneration, resulting in visual field loss in patients with Glaucoma. Published studies on in vitro RGC differentiation from stem cells utilized classical RGC signaling pathways mimicking retinal development in vivo. Although many strategies allowed for the generation of RGCs, increased variability between experiments and lower yield hampered the cross comparison between individual lines and between experiments. To address this critical need, we developed a reproducible chemically defined in vitro methodology for generating retinal progenitor cell (RPC) populations from iPSCs, that are efficiently directed towards RGC lineage. Using this method, we reproducibly differentiated iPSCs into RGCs with greater than 80% purity, without any genetic modifications. We used small molecules and peptide modulators to inhibit BMP, TGF-β (SMAD), and canonical Wnt pathways that reduced variability between iPSC lines and yielded functional and mature iPSC-RGCs. Using CD90.2 antibody and Magnetic Activated Cell Sorter (MACS) technique, we successfully purified Thy-1 positive RGCs with nearly 95% purity.Competing Interest StatementThe authors have declared no competing interest.