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Differentiation of RPE cells from integration-free iPS cells and their cell biological characterization.
Stem Cell Res Ther. 2017 10 02; 8(1):217.SC

Abstract

BACKGROUND

Dysfunction of the retinal pigment epithelium (RPE) is implicated in numerous forms of retinal degeneration. The readily accessible environment of the eye makes it particularly suitable for the transplantation of RPE cells, which can now be derived from autologous induced pluripotent stem cells (iPSCs), to treat retinal degeneration. For RPE transplantation to become feasible in the clinic, patient-specific somatic cells should be reprogrammed to iPSCs without the introduction of reprogramming genes into the genome of the host cell, and then subsequently differentiated into RPE cells that are well characterized for safety and functionality prior to transplantation.

METHODS

We have reprogrammed human dermal fibroblasts to iPSCs using nonintegrating RNA, and differentiated the iPSCs toward an RPE fate (iPSC-RPE), under Good Manufacturing Practice (GMP)-compatible conditions.

RESULTS

Using highly sensitive assays for cell polarity, structure, organelle trafficking, and function, we found that iPSC-RPE cells in culture exhibited key characteristics of native RPE. Importantly, we demonstrate for the first time with any stem cell-derived RPE cell that live cells are able to support dynamic organelle transport. This highly sensitive test is critical for RPE cells intended for transplantation, since defects in intracellular motility have been shown to promote RPE pathogenesis akin to that found in macular degeneration. To test their capabilities for in-vivo transplantation, we injected the iPSC-RPE cells into the subretinal space of a mouse model of retinal degeneration, and demonstrated that the transplanted cells are capable of rescuing lost RPE function.

CONCLUSIONS

This report documents the successful generation, under GMP-compatible conditions, of human iPSC-RPE cells that possess specific characteristics of healthy RPE. The report adds to a growing literature on the utility of human iPSC-RPE cells for cell culture investigations on pathogenicity and for therapeutic transplantation, by corroborating findings of others, and providing important new information on essential RPE cell biological properties.

Authors+Show Affiliations

Stein Eye Institute and Department of Ophthalmology, David Geffen School of Medicine at UCLA, 100 Stein Plaza, Los Angeles, CA, 90095, USA.Department of Microbiology Immunology and Molecular Genetics, Los Angeles, CA, USA. Skarumbayaram@mednet.ucla.edu. Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA, Los Angeles, CA, USA. Skarumbayaram@mednet.ucla.edu. Jonsson Comprehensive Cancer Center, Los Angeles, CA, USA. Skarumbayaram@mednet.ucla.edu.Stein Eye Institute and Department of Ophthalmology, David Geffen School of Medicine at UCLA, 100 Stein Plaza, Los Angeles, CA, 90095, USA.Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA, Los Angeles, CA, USA.Stein Eye Institute and Department of Ophthalmology, David Geffen School of Medicine at UCLA, 100 Stein Plaza, Los Angeles, CA, 90095, USA.Stein Eye Institute and Department of Ophthalmology, David Geffen School of Medicine at UCLA, 100 Stein Plaza, Los Angeles, CA, 90095, USA. Department of Molecular Cell and Developmental Biology, Los Angeles, CA, USA.Stein Eye Institute and Department of Ophthalmology, David Geffen School of Medicine at UCLA, 100 Stein Plaza, Los Angeles, CA, 90095, USA.Department of Pediatrics, David Geffen School of Medicine, Los Angeles, CA, USA. Jonsson Comprehensive Cancer Center, Los Angeles, CA, USA.Department of Population Sciences, City of Hope National Medical Center, Duarte, CA, USA.Department of Microbiology Immunology and Molecular Genetics, Los Angeles, CA, USA. Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA, Los Angeles, CA, USA. Jonsson Comprehensive Cancer Center, Los Angeles, CA, USA. Department of Medicine, David Geffen School of Medicine, Los Angeles, CA, USA.Department of Microbiology Immunology and Molecular Genetics, Los Angeles, CA, USA. Department of Pediatrics, David Geffen School of Medicine, Los Angeles, CA, USA. Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA, Los Angeles, CA, USA. Jonsson Comprehensive Cancer Center, Los Angeles, CA, USA.Department of Pediatrics, David Geffen School of Medicine, Los Angeles, CA, USA. Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA, Los Angeles, CA, USA. Jonsson Comprehensive Cancer Center, Los Angeles, CA, USA.Department of Microbiology Immunology and Molecular Genetics, Los Angeles, CA, USA. Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA, Los Angeles, CA, USA. Jonsson Comprehensive Cancer Center, Los Angeles, CA, USA.Department of Molecular Cell and Developmental Biology, Los Angeles, CA, USA. Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research at UCLA, Los Angeles, CA, USA. Jonsson Comprehensive Cancer Center, Los Angeles, CA, USA.Stein Eye Institute and Department of Ophthalmology, David Geffen School of Medicine at UCLA, 100 Stein Plaza, Los Angeles, CA, 90095, USA. dswilliams@ucla.edu. Department of Neurobiology, David Geffen School of Medicine, Los Angeles, CA, USA. dswilliams@ucla.edu. Molecular Biology Institute, Los Angeles, CA, USA. dswilliams@ucla.edu. Brain Research Institute, University of California, Los Angeles, CA, USA. dswilliams@ucla.edu.

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

28969679

Citation

Hazim, Roni A., et al. "Differentiation of RPE Cells From Integration-free iPS Cells and Their Cell Biological Characterization." Stem Cell Research & Therapy, vol. 8, no. 1, 2017, p. 217.
Hazim RA, Karumbayaram S, Jiang M, et al. Differentiation of RPE cells from integration-free iPS cells and their cell biological characterization. Stem Cell Res Ther. 2017;8(1):217.
Hazim, R. A., Karumbayaram, S., Jiang, M., Dimashkie, A., Lopes, V. S., Li, D., Burgess, B. L., Vijayaraj, P., Alva-Ornelas, J. A., Zack, J. A., Kohn, D. B., Gomperts, B. N., Pyle, A. D., Lowry, W. E., & Williams, D. S. (2017). Differentiation of RPE cells from integration-free iPS cells and their cell biological characterization. Stem Cell Research & Therapy, 8(1), 217. https://doi.org/10.1186/s13287-017-0652-9
Hazim RA, et al. Differentiation of RPE Cells From Integration-free iPS Cells and Their Cell Biological Characterization. Stem Cell Res Ther. 2017 10 2;8(1):217. PubMed PMID: 28969679.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Differentiation of RPE cells from integration-free iPS cells and their cell biological characterization. AU - Hazim,Roni A, AU - Karumbayaram,Saravanan, AU - Jiang,Mei, AU - Dimashkie,Anupama, AU - Lopes,Vanda S, AU - Li,Douran, AU - Burgess,Barry L, AU - Vijayaraj,Preethi, AU - Alva-Ornelas,Jackelyn A, AU - Zack,Jerome A, AU - Kohn,Donald B, AU - Gomperts,Brigitte N, AU - Pyle,April D, AU - Lowry,William E, AU - Williams,David S, Y1 - 2017/10/02/ PY - 2017/05/06/received PY - 2017/08/29/accepted PY - 2017/08/16/revised PY - 2017/10/4/entrez PY - 2017/10/4/pubmed PY - 2018/5/31/medline KW - Induced pluripotent stem cells KW - Live-cell imaging KW - Phagocytosis KW - RPE cytoskeleton KW - Retinal pigment epithelium SP - 217 EP - 217 JF - Stem cell research & therapy JO - Stem Cell Res Ther VL - 8 IS - 1 N2 - BACKGROUND: Dysfunction of the retinal pigment epithelium (RPE) is implicated in numerous forms of retinal degeneration. The readily accessible environment of the eye makes it particularly suitable for the transplantation of RPE cells, which can now be derived from autologous induced pluripotent stem cells (iPSCs), to treat retinal degeneration. For RPE transplantation to become feasible in the clinic, patient-specific somatic cells should be reprogrammed to iPSCs without the introduction of reprogramming genes into the genome of the host cell, and then subsequently differentiated into RPE cells that are well characterized for safety and functionality prior to transplantation. METHODS: We have reprogrammed human dermal fibroblasts to iPSCs using nonintegrating RNA, and differentiated the iPSCs toward an RPE fate (iPSC-RPE), under Good Manufacturing Practice (GMP)-compatible conditions. RESULTS: Using highly sensitive assays for cell polarity, structure, organelle trafficking, and function, we found that iPSC-RPE cells in culture exhibited key characteristics of native RPE. Importantly, we demonstrate for the first time with any stem cell-derived RPE cell that live cells are able to support dynamic organelle transport. This highly sensitive test is critical for RPE cells intended for transplantation, since defects in intracellular motility have been shown to promote RPE pathogenesis akin to that found in macular degeneration. To test their capabilities for in-vivo transplantation, we injected the iPSC-RPE cells into the subretinal space of a mouse model of retinal degeneration, and demonstrated that the transplanted cells are capable of rescuing lost RPE function. CONCLUSIONS: This report documents the successful generation, under GMP-compatible conditions, of human iPSC-RPE cells that possess specific characteristics of healthy RPE. The report adds to a growing literature on the utility of human iPSC-RPE cells for cell culture investigations on pathogenicity and for therapeutic transplantation, by corroborating findings of others, and providing important new information on essential RPE cell biological properties. SN - 1757-6512 UR - https://www.unboundmedicine.com/medline/citation/28969679/Differentiation_of_RPE_cells_from_integration_free_iPS_cells_and_their_cell_biological_characterization_ DB - PRIME DP - Unbound Medicine ER -