Efficient Differentiation of Mouse Induced Pluripotent Stem Cells into Alveolar Epithelium Type II with a BRD4 Inhibitor
Regenerative medicine has continued to progress for lung biology and lung diseases. Efforts have focused on a variety of different applications for pluripotent stem cells. Several groups have reported successful methods for inducing differentiation of induced pluripotent stem cells (iPSCs) into the...
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| Main Authors: | , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Wiley
2019-01-01
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| Series: | Stem Cells International |
| Online Access: | http://dx.doi.org/10.1155/2019/1271682 |
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| author | Toru Momozane Eriko Fukui Soichiro Funaki Makoto Fujii Yuhei Kinehara Emiko Ito Shigeru Miyagawa Yuko Ohno Yoshiki Sawa Meinoshin Okumura Yasushi Shintani |
| author_facet | Toru Momozane Eriko Fukui Soichiro Funaki Makoto Fujii Yuhei Kinehara Emiko Ito Shigeru Miyagawa Yuko Ohno Yoshiki Sawa Meinoshin Okumura Yasushi Shintani |
| author_sort | Toru Momozane |
| collection | DOAJ |
| description | Regenerative medicine has continued to progress for lung biology and lung diseases. Efforts have focused on a variety of different applications for pluripotent stem cells. Several groups have reported successful methods for inducing differentiation of induced pluripotent stem cells (iPSCs) into the airway epithelium such as alveolar epithelium type II (ATII). However, differentiation efficiency varies among reports and improvements are needed. In the present paper, we propose a novel method for elimination of residual undifferentiated murine iPSCs using JQ1, a potent inhibitor of bromodomain (BRD) and extraterminal domain (BET) family proteins, for efficient differentiation into ATII. First, the murine iPSC line 20D-17 was induced to differentiate into ATII over a period of 26 days (days 0-26) using previously reported embryoid body seeding and stepwise differentiation methods. mRNA expressions of differentiation markers including surfactant protein C (Sftpc) were confirmed by real-time reverse transcription-polymerase chain reaction (RT-PCR) results, and 17% of the cells were shown positive for prosurfactant protein C (proSPC) in flow cytometry analysis. Next, those cells were cultured three-dimensionally in Matrigel for an additional 14 days (days 26-40), during which JQ1 was added for 4 days (days 28-32) to remove residual undifferentiated iPSCs. As a result, on day 40, the mRNA expression level of Sftpc in the three-dimensional culture was maintained at the same level as on day 26 and shown to be further increased by the addition of JQ1, with 39% of the cells found to express proSPC, showing that differentiation efficiency could be further increased. Three-dimensional culture with BRD4 inhibition by JQ1 improved the differentiation induction efficiency to ATII by removing residual undifferentiated murine iPSCs during the differentiation induction process. |
| format | Article |
| id | doaj-art-f81fdcd02a714ef7a38d6ff1615f69d9 |
| institution | Kabale University |
| issn | 1687-966X 1687-9678 |
| language | English |
| publishDate | 2019-01-01 |
| publisher | Wiley |
| record_format | Article |
| series | Stem Cells International |
| spelling | doaj-art-f81fdcd02a714ef7a38d6ff1615f69d92025-02-03T01:11:53ZengWileyStem Cells International1687-966X1687-96782019-01-01201910.1155/2019/12716821271682Efficient Differentiation of Mouse Induced Pluripotent Stem Cells into Alveolar Epithelium Type II with a BRD4 InhibitorToru Momozane0Eriko Fukui1Soichiro Funaki2Makoto Fujii3Yuhei Kinehara4Emiko Ito5Shigeru Miyagawa6Yuko Ohno7Yoshiki Sawa8Meinoshin Okumura9Yasushi Shintani10Department of General Thoracic Surgery, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of General Thoracic Surgery, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of General Thoracic Surgery, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Mathematical Health Science, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Respiratory Medicine and Clinical Immunology, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Cardiovascular Surgery, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Cardiovascular Surgery, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Mathematical Health Science, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of Cardiovascular Surgery, Osaka University Graduate School of Medicine, Osaka, JapanDepartment of General Thoracic Surgery, National Hospital Organization Toneyama Hospital, Osaka, JapanDepartment of General Thoracic Surgery, Osaka University Graduate School of Medicine, Osaka, JapanRegenerative medicine has continued to progress for lung biology and lung diseases. Efforts have focused on a variety of different applications for pluripotent stem cells. Several groups have reported successful methods for inducing differentiation of induced pluripotent stem cells (iPSCs) into the airway epithelium such as alveolar epithelium type II (ATII). However, differentiation efficiency varies among reports and improvements are needed. In the present paper, we propose a novel method for elimination of residual undifferentiated murine iPSCs using JQ1, a potent inhibitor of bromodomain (BRD) and extraterminal domain (BET) family proteins, for efficient differentiation into ATII. First, the murine iPSC line 20D-17 was induced to differentiate into ATII over a period of 26 days (days 0-26) using previously reported embryoid body seeding and stepwise differentiation methods. mRNA expressions of differentiation markers including surfactant protein C (Sftpc) were confirmed by real-time reverse transcription-polymerase chain reaction (RT-PCR) results, and 17% of the cells were shown positive for prosurfactant protein C (proSPC) in flow cytometry analysis. Next, those cells were cultured three-dimensionally in Matrigel for an additional 14 days (days 26-40), during which JQ1 was added for 4 days (days 28-32) to remove residual undifferentiated iPSCs. As a result, on day 40, the mRNA expression level of Sftpc in the three-dimensional culture was maintained at the same level as on day 26 and shown to be further increased by the addition of JQ1, with 39% of the cells found to express proSPC, showing that differentiation efficiency could be further increased. Three-dimensional culture with BRD4 inhibition by JQ1 improved the differentiation induction efficiency to ATII by removing residual undifferentiated murine iPSCs during the differentiation induction process.http://dx.doi.org/10.1155/2019/1271682 |
| spellingShingle | Toru Momozane Eriko Fukui Soichiro Funaki Makoto Fujii Yuhei Kinehara Emiko Ito Shigeru Miyagawa Yuko Ohno Yoshiki Sawa Meinoshin Okumura Yasushi Shintani Efficient Differentiation of Mouse Induced Pluripotent Stem Cells into Alveolar Epithelium Type II with a BRD4 Inhibitor Stem Cells International |
| title | Efficient Differentiation of Mouse Induced Pluripotent Stem Cells into Alveolar Epithelium Type II with a BRD4 Inhibitor |
| title_full | Efficient Differentiation of Mouse Induced Pluripotent Stem Cells into Alveolar Epithelium Type II with a BRD4 Inhibitor |
| title_fullStr | Efficient Differentiation of Mouse Induced Pluripotent Stem Cells into Alveolar Epithelium Type II with a BRD4 Inhibitor |
| title_full_unstemmed | Efficient Differentiation of Mouse Induced Pluripotent Stem Cells into Alveolar Epithelium Type II with a BRD4 Inhibitor |
| title_short | Efficient Differentiation of Mouse Induced Pluripotent Stem Cells into Alveolar Epithelium Type II with a BRD4 Inhibitor |
| title_sort | efficient differentiation of mouse induced pluripotent stem cells into alveolar epithelium type ii with a brd4 inhibitor |
| url | http://dx.doi.org/10.1155/2019/1271682 |
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