2. Academic Validation
  3. Transcription factor overexpression drives reliable differentiation of retinal pigment epithelium from human induced pluripotent stem cells

Transcription factor overexpression drives reliable differentiation of retinal pigment epithelium from human induced pluripotent stem cells

  • Stem Cell Res. 2021 May:53:102368. doi: 10.1016/j.scr.2021.102368.
Tessa E Dewell 1,  Ketrin Gjoni 1,  Angela Z Liu 1,  Ashley R G Libby 2,  Anthony T Moore 3,  Po-Lin So 4,  Bruce R Conklin 5
Affiliations

Affiliations

  • 1 Gladstone Institutes, 1650 Owens Street, San Francisco, CA 94158, USA.
  • 2 Gladstone Institutes, 1650 Owens Street, San Francisco, CA 94158, USA; Developmental and Stem Cell Biology Program, University of California, 1675 Owens St, San Francisco, CA 94158, USA.
  • 3 UCSF Department of Ophthalmology, 10 Koret Way, San Francisco, CA 94143-0730, USA.
  • 4 Gladstone Institutes, 1650 Owens Street, San Francisco, CA 94158, USA; Innovative Genomics Institute, 2151 Berkeley Way, Berkeley, CA 94720, USA; Gladstone Institutes Stem Cell Core, 1650 Owens Street, San Francisco, CA 94158, USA.
  • 5 Gladstone Institutes, 1650 Owens Street, San Francisco, CA 94158, USA; UCSF Department of Ophthalmology, 10 Koret Way, San Francisco, CA 94143-0730, USA; Innovative Genomics Institute, 2151 Berkeley Way, Berkeley, CA 94720, USA; UCSF Department of Medicine, 535 Mission Bay Blvd South, San Francisco, CA 94158, USA. Electronic address: [email protected].
PMID: 34087997 DOI: 10.1016/j.scr.2021.102368
Abstract

Age-related macular degeneration and genetic forms of blindness such as Best Disease and Retinitis Pigmentosa can be caused by degeneration of the Retinal Pigment Epithelium (RPE). RPE generated from patient-derived induced pluripotent stem cells (iPSCs) is valuable for both the study of disease mechanisms and development of therapeutic strategies. However, protocols to produce iPSC-derived RPE in vitro are often inefficient, labor-intensive, low-throughput, and highly variable between cell lines and within batches. Here, we report a robust, scalable method to generate iPSC-RPE using doxycycline-inducible expression of eye field transcription factors OTX2, PAX6 and MITF paired with RPE-permissive culture media. Doxycycline addition induces exogenous expression of these transcription factors in Best Disease patient- and wildtype iPSCs to efficiently produce monolayers of RPE with characteristic morphology and gene expression. Further, these RPE monolayers display functionality features including light absorption via pigmentation, polarity-driven fluid transport, and phagocytosis. With this method, we achieve a highly efficient and easily scalable differentiation without the need for mechanical isolation or enrichment methods, generating RPE cultures applicable for in vitro studies.

Keywords

Cell Differentiation; Induced Pluripotent Stem Cells; Retinal Diseases; Retinal Pigment Epithelium; Transcription Factors; iPSC.

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