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Differentiation of Human Protein-Induced Pluripotent Stem Cells toward a Retinal Pigment Epithelial Cell Fate.
PLoS One. 2015; 10(11):e0143272.Plos

Abstract

Compared with many induced pluripotent stem cell (iPSC) lines generated using retrovirus and other non-integrating methods, the utilization of human protein-induced iPSC (piPSC) lines may provide a safer alternative for the generation of retinal pigment epithelial (RPE) cells for transplantation in retinal degenerative diseases. Here we assess the ability of piPSCs to differentiate into RPE cells, and to perform native RPE cell behavior. piPSCs were seeded in 6-well low-attachment plates to allow embryoid body formation, and then analyzed for pluripotent stem cell markers NANOG, SSEA4 and TRA-1-60 by immunofluorescence. Following colony formation, piPSCs were assessed for confirmation of RPE cell differentiation by staining for zonula occludens (ZO-1), bestrophin, microphthalmia-associated transcription factor (MITF) and retinal pigment epithelium specific protein-65 (RPE65). To evaluate piPSC-RPE cell phagocytic ability, adult bovine photoreceptor rod outer segments (ROS) were fed to piPSC-RPE cells, which were analyzed by fluorescent microscopy and flow cytometry. Undifferentiated piPSCs expressed all pluripotent markers assessed and formed embryoid body aggregates after 7 days. Differentiated piPSC-RPE cells expressed ZO-1, bestrophin, MITF and RPE65, typical RPE cell markers. Flow cytometry revealed robust ingestion of fluorescently-labeled ROS by piPSC-RPE cells, which was over four-times greater than that of undifferentiated piPSCs and comparable to that of an immortalized RPE cell line. Phagocytosis activity by piPSC-RPE cells was significantly reduced after the addition of anti-integrin αVβ5. In conclusion, piPSCs can be differentiated toward an RPE cell fate, expressing RPE cell markers and resembling native RPE cells in behavior. These results demonstrate that piPSCs can be differentiated into RPE-like cells using a method that has an increased safety profile, a critical consideration for the development of better treatments for retinal degenerative diseases such as age-related macular degeneration (AMD).

Authors+Show Affiliations

Department of Ophthalmology, Storm Eye Institute, Medical University of South Carolina, Charleston, SC, United States of America.Department of Ophthalmology, Storm Eye Institute, Medical University of South Carolina, Charleston, SC, United States of America. Department of Regenerative Medicine and Cell Biology, Medical University of South Carolina, Charleston, SC, United States of America.Department of Ophthalmology, Storm Eye Institute, Medical University of South Carolina, Charleston, SC, United States of America.Department of Ophthalmology, Storm Eye Institute, Medical University of South Carolina, Charleston, SC, United States of America.Department of Drug Discovery and Biomedical Sciences, Medical University of South Carolina, Charleston, SC, United States of America.Department of Ophthalmology, Storm Eye Institute, Medical University of South Carolina, Charleston, SC, United States of America.Department of Ophthalmology, Storm Eye Institute, Medical University of South Carolina, Charleston, SC, United States of America.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

26606685

Citation

Gong, Jie, et al. "Differentiation of Human Protein-Induced Pluripotent Stem Cells Toward a Retinal Pigment Epithelial Cell Fate." PloS One, vol. 10, no. 11, 2015, pp. e0143272.
Gong J, Fields MA, Moreira EF, et al. Differentiation of Human Protein-Induced Pluripotent Stem Cells toward a Retinal Pigment Epithelial Cell Fate. PLoS One. 2015;10(11):e0143272.
Gong, J., Fields, M. A., Moreira, E. F., Bowrey, H. E., Gooz, M., Ablonczy, Z., & Del Priore, L. V. (2015). Differentiation of Human Protein-Induced Pluripotent Stem Cells toward a Retinal Pigment Epithelial Cell Fate. PloS One, 10(11), e0143272. https://doi.org/10.1371/journal.pone.0143272
Gong J, et al. Differentiation of Human Protein-Induced Pluripotent Stem Cells Toward a Retinal Pigment Epithelial Cell Fate. PLoS One. 2015;10(11):e0143272. PubMed PMID: 26606685.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Differentiation of Human Protein-Induced Pluripotent Stem Cells toward a Retinal Pigment Epithelial Cell Fate. AU - Gong,Jie, AU - Fields,Mark A, AU - Moreira,Ernesto F, AU - Bowrey,Hannah E, AU - Gooz,Monika, AU - Ablonczy,Zsolt, AU - Del Priore,Lucian V, Y1 - 2015/11/25/ PY - 2015/08/17/received PY - 2015/11/03/accepted PY - 2015/11/26/entrez PY - 2015/11/26/pubmed PY - 2016/6/28/medline SP - e0143272 EP - e0143272 JF - PloS one JO - PLoS One VL - 10 IS - 11 N2 - Compared with many induced pluripotent stem cell (iPSC) lines generated using retrovirus and other non-integrating methods, the utilization of human protein-induced iPSC (piPSC) lines may provide a safer alternative for the generation of retinal pigment epithelial (RPE) cells for transplantation in retinal degenerative diseases. Here we assess the ability of piPSCs to differentiate into RPE cells, and to perform native RPE cell behavior. piPSCs were seeded in 6-well low-attachment plates to allow embryoid body formation, and then analyzed for pluripotent stem cell markers NANOG, SSEA4 and TRA-1-60 by immunofluorescence. Following colony formation, piPSCs were assessed for confirmation of RPE cell differentiation by staining for zonula occludens (ZO-1), bestrophin, microphthalmia-associated transcription factor (MITF) and retinal pigment epithelium specific protein-65 (RPE65). To evaluate piPSC-RPE cell phagocytic ability, adult bovine photoreceptor rod outer segments (ROS) were fed to piPSC-RPE cells, which were analyzed by fluorescent microscopy and flow cytometry. Undifferentiated piPSCs expressed all pluripotent markers assessed and formed embryoid body aggregates after 7 days. Differentiated piPSC-RPE cells expressed ZO-1, bestrophin, MITF and RPE65, typical RPE cell markers. Flow cytometry revealed robust ingestion of fluorescently-labeled ROS by piPSC-RPE cells, which was over four-times greater than that of undifferentiated piPSCs and comparable to that of an immortalized RPE cell line. Phagocytosis activity by piPSC-RPE cells was significantly reduced after the addition of anti-integrin αVβ5. In conclusion, piPSCs can be differentiated toward an RPE cell fate, expressing RPE cell markers and resembling native RPE cells in behavior. These results demonstrate that piPSCs can be differentiated into RPE-like cells using a method that has an increased safety profile, a critical consideration for the development of better treatments for retinal degenerative diseases such as age-related macular degeneration (AMD). SN - 1932-6203 UR - https://www.unboundmedicine.com/medline/citation/26606685/Differentiation_of_Human_Protein_Induced_Pluripotent_Stem_Cells_toward_a_Retinal_Pigment_Epithelial_Cell_Fate_ L2 - https://dx.plos.org/10.1371/journal.pone.0143272 DB - PRIME DP - Unbound Medicine ER -