Dual SMAD inhibition and Wnt inhibition enable efficient and reproducible differentiations of induced pluripotent stem cells into retinal ganglion cells

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 the death of retinal ganglion cells (RGCs). The loss of RGCs manifests as characteristic cupping or optic...

Full description

Saved in:
Bibliographic Details
Published in:Scientific reports 2020-07, Vol.10 (1), p.11828-11828, Article 11828
Main Authors: Chavali, Venkata R M, Haider, Naqi, Rathi, Sonika, Vrathasha, Vrathasha, Alapati, Teja, He, Jie, Gill, Kamaljot, Nikonov, Roman, Duong, Thu T, McDougald, Devin S, Nikonov, Sergei, O'Brien, Joan, Mills, Jason A
Format: Article
Language:English
Subjects:
CD90 antigen
Cell death
Cell differentiation
Cell Differentiation - drug effects
Computational Biology
Degeneration
Gene Expression Profiling
Glaucoma
Humans
Immunohistochemistry
Immunophenotyping
Induced Pluripotent Stem Cells - cytology
Induced Pluripotent Stem Cells - metabolism
Inhibitory postsynaptic potentials
Mimicry
Neurogenesis
Neuromodulation
Optic nerve
Pluripotency
Progenitor cells
Retina
Retina - cytology
Retinal ganglion cells
Retinal Ganglion Cells - cytology
Retinal Ganglion Cells - metabolism
Signal Transduction
Smad protein
Smad Proteins - antagonists & inhibitors
Stem cells
Visual field
Wnt protein
Wnt Proteins - antagonists & inhibitors
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary: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 the 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.
ISSN:2045-2322
2045-2322
DOI:10.1038/s41598-020-68811-8