Evaluation of Retinal Pigment Epithelial Cell Cytotoxicity of Recombinant Tissue Plasminogen Activator Using Human-Induced Pluripotent Stem Cells
Purpose. The evaluation of drug-induced cytotoxicity is of great importance for the clinical application of pharmaceutical products, and human-induced pluripotent stem cells (hiPSCs) have received considerable scrutiny as a cell source for in vitro cytotoxicity testing. The aim of this study is to v...
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| Format: | Article |
| Language: | English |
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Wiley
2019-01-01
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| Series: | Journal of Ophthalmology |
| Online Access: | http://dx.doi.org/10.1155/2019/7189241 |
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| _version_ | 1832561221043224576 |
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| author | Hiroyuki Kamao Atsushi Miki Junichi Kiryu |
| author_facet | Hiroyuki Kamao Atsushi Miki Junichi Kiryu |
| author_sort | Hiroyuki Kamao |
| collection | DOAJ |
| description | Purpose. The evaluation of drug-induced cytotoxicity is of great importance for the clinical application of pharmaceutical products, and human-induced pluripotent stem cells (hiPSCs) have received considerable scrutiny as a cell source for in vitro cytotoxicity testing. The aim of this study is to validate the concept of cytotoxicity testing using hiPSC-derived retinal pigment epithelium (hiPSC-RPE) by comparing the responsiveness of human fetal RPE (hfRPE) and human RPE cell line (ARPE19) to recombinant tissue plasminogen activator (rtPA). Methods. HfRPE, two types of hiPSC-RPE, and ARPE19 were cultured in media with or without rtPA. A lactate dehydrogenase release assay was performed to investigate the dose- and time-dependent effects of rtPA on cell death. RPE function was evaluated by measuring the secretion of pigment epithelium-derived factor (PEDF) and vascular endothelial growth factor (VEGF) and RPE-specific gene expression. Results. Rates of cell damage in hfRPE and both hiPS-RPE were increased by rtPA supplementation (2000 and 4000 μg/ml) for 1 hour, whereas ARPE19 cell damage was increased by supplementation with rtPA at concentrations higher than 50 μg/ml. Although 100 μg/ml rtPA for 24 hours did not affect RPE cell function, sustained rtPA exposure induced prolonged cytotoxic effects in hfRPE and two hiPSC-RPE, but not ARPE19. Conclusion. The responsiveness of hiPSC-RPE to rtPA is similar to that of hfRPE in terms of cell death and cell function. Thus, hiPSC-RPE is a valuable cell source for in vitro cytotoxicity testing. |
| format | Article |
| id | doaj-art-668e398884704ae3935daaadfb00bf3d |
| institution | Kabale University |
| issn | 2090-004X 2090-0058 |
| language | English |
| publishDate | 2019-01-01 |
| publisher | Wiley |
| record_format | Article |
| series | Journal of Ophthalmology |
| spelling | doaj-art-668e398884704ae3935daaadfb00bf3d2025-02-03T01:25:39ZengWileyJournal of Ophthalmology2090-004X2090-00582019-01-01201910.1155/2019/71892417189241Evaluation of Retinal Pigment Epithelial Cell Cytotoxicity of Recombinant Tissue Plasminogen Activator Using Human-Induced Pluripotent Stem CellsHiroyuki Kamao0Atsushi Miki1Junichi Kiryu2Department of Ophthalmology, Kawasaki Medical School, 577 Matsushima, Kurashiki, Okayama 701-0114, JapanDepartment of Ophthalmology, Kawasaki Medical School, 577 Matsushima, Kurashiki, Okayama 701-0114, JapanDepartment of Ophthalmology, Kawasaki Medical School, 577 Matsushima, Kurashiki, Okayama 701-0114, JapanPurpose. The evaluation of drug-induced cytotoxicity is of great importance for the clinical application of pharmaceutical products, and human-induced pluripotent stem cells (hiPSCs) have received considerable scrutiny as a cell source for in vitro cytotoxicity testing. The aim of this study is to validate the concept of cytotoxicity testing using hiPSC-derived retinal pigment epithelium (hiPSC-RPE) by comparing the responsiveness of human fetal RPE (hfRPE) and human RPE cell line (ARPE19) to recombinant tissue plasminogen activator (rtPA). Methods. HfRPE, two types of hiPSC-RPE, and ARPE19 were cultured in media with or without rtPA. A lactate dehydrogenase release assay was performed to investigate the dose- and time-dependent effects of rtPA on cell death. RPE function was evaluated by measuring the secretion of pigment epithelium-derived factor (PEDF) and vascular endothelial growth factor (VEGF) and RPE-specific gene expression. Results. Rates of cell damage in hfRPE and both hiPS-RPE were increased by rtPA supplementation (2000 and 4000 μg/ml) for 1 hour, whereas ARPE19 cell damage was increased by supplementation with rtPA at concentrations higher than 50 μg/ml. Although 100 μg/ml rtPA for 24 hours did not affect RPE cell function, sustained rtPA exposure induced prolonged cytotoxic effects in hfRPE and two hiPSC-RPE, but not ARPE19. Conclusion. The responsiveness of hiPSC-RPE to rtPA is similar to that of hfRPE in terms of cell death and cell function. Thus, hiPSC-RPE is a valuable cell source for in vitro cytotoxicity testing.http://dx.doi.org/10.1155/2019/7189241 |
| spellingShingle | Hiroyuki Kamao Atsushi Miki Junichi Kiryu Evaluation of Retinal Pigment Epithelial Cell Cytotoxicity of Recombinant Tissue Plasminogen Activator Using Human-Induced Pluripotent Stem Cells Journal of Ophthalmology |
| title | Evaluation of Retinal Pigment Epithelial Cell Cytotoxicity of Recombinant Tissue Plasminogen Activator Using Human-Induced Pluripotent Stem Cells |
| title_full | Evaluation of Retinal Pigment Epithelial Cell Cytotoxicity of Recombinant Tissue Plasminogen Activator Using Human-Induced Pluripotent Stem Cells |
| title_fullStr | Evaluation of Retinal Pigment Epithelial Cell Cytotoxicity of Recombinant Tissue Plasminogen Activator Using Human-Induced Pluripotent Stem Cells |
| title_full_unstemmed | Evaluation of Retinal Pigment Epithelial Cell Cytotoxicity of Recombinant Tissue Plasminogen Activator Using Human-Induced Pluripotent Stem Cells |
| title_short | Evaluation of Retinal Pigment Epithelial Cell Cytotoxicity of Recombinant Tissue Plasminogen Activator Using Human-Induced Pluripotent Stem Cells |
| title_sort | evaluation of retinal pigment epithelial cell cytotoxicity of recombinant tissue plasminogen activator using human induced pluripotent stem cells |
| url | http://dx.doi.org/10.1155/2019/7189241 |
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