Complete suspension culture of human induced pluripotent stem cells supplemented with suppressors of spontaneous differentiation
Human induced pluripotent stem cells (hiPSCs) are promising resources for producing various types of tissues in regenerative medicine; however, the improvement in a scalable culture system that can precisely control the cellular status of hiPSCs is needed. Utilizing suspension culture without microc...
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| Format: | Article |
| Language: | English |
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eLife Sciences Publications Ltd
2024-11-01
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| Series: | eLife |
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| Online Access: | https://elifesciences.org/articles/89724 |
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| author | Mami Matsuo-Takasaki Sho Kambayashi Yasuko Hemmi Tamami Wakabayashi Tomoya Shimizu Yuri An Hidenori Ito Kazuhiro Takeuchi Masato Ibuki Terasu Kawashima Rio Masayasu Manami Suzuki Yoshikazu Kawai Masafumi Umekage Tomoaki M Kato Michiya Noguchi Koji Nakade Yukio Nakamura Tomoyuki Nakaishi Naoki Nishishita Masayoshi Tsukahara Yohei Hayashi |
| author_facet | Mami Matsuo-Takasaki Sho Kambayashi Yasuko Hemmi Tamami Wakabayashi Tomoya Shimizu Yuri An Hidenori Ito Kazuhiro Takeuchi Masato Ibuki Terasu Kawashima Rio Masayasu Manami Suzuki Yoshikazu Kawai Masafumi Umekage Tomoaki M Kato Michiya Noguchi Koji Nakade Yukio Nakamura Tomoyuki Nakaishi Naoki Nishishita Masayoshi Tsukahara Yohei Hayashi |
| author_sort | Mami Matsuo-Takasaki |
| collection | DOAJ |
| description | Human induced pluripotent stem cells (hiPSCs) are promising resources for producing various types of tissues in regenerative medicine; however, the improvement in a scalable culture system that can precisely control the cellular status of hiPSCs is needed. Utilizing suspension culture without microcarriers or special materials allows for massive production, automation, cost-effectiveness, and safety assurance in industrialized regenerative medicine. Here, we found that hiPSCs cultured in suspension conditions with continuous agitation without microcarriers or extracellular matrix components were more prone to spontaneous differentiation than those cultured in conventional adherent conditions. Adding PKCβ and Wnt signaling pathway inhibitors in the suspension conditions suppressed the spontaneous differentiation of hiPSCs into ectoderm and mesendoderm, respectively. In these conditions, we successfully completed the culture processes of hiPSCs, including the generation of hiPSCs from peripheral blood mononuclear cells with the expansion of bulk population and single-cell sorted clones, long-term culture with robust self-renewal characteristics, single-cell cloning, direct cryopreservation from suspension culture and their successful recovery, and efficient mass production of a clinical-grade hiPSC line. Our results demonstrate that precise control of the cellular status in suspension culture conditions paves the way for their stable and automated clinical application. |
| format | Article |
| id | doaj-art-9741ffd7753d4338bddd028973796412 |
| institution | OA Journals |
| issn | 2050-084X |
| language | English |
| publishDate | 2024-11-01 |
| publisher | eLife Sciences Publications Ltd |
| record_format | Article |
| series | eLife |
| spelling | doaj-art-9741ffd7753d4338bddd0289737964122025-08-20T02:14:02ZengeLife Sciences Publications LtdeLife2050-084X2024-11-011210.7554/eLife.89724Complete suspension culture of human induced pluripotent stem cells supplemented with suppressors of spontaneous differentiationMami Matsuo-Takasaki0Sho Kambayashi1Yasuko Hemmi2Tamami Wakabayashi3Tomoya Shimizu4Yuri An5Hidenori Ito6Kazuhiro Takeuchi7Masato Ibuki8Terasu Kawashima9Rio Masayasu10Manami Suzuki11Yoshikazu Kawai12Masafumi Umekage13Tomoaki M Kato14Michiya Noguchi15Koji Nakade16Yukio Nakamura17Tomoyuki Nakaishi18Naoki Nishishita19Masayoshi Tsukahara20Yohei Hayashi21https://orcid.org/0000-0001-5490-7052iPS Cell Advanced Characterization and Development Team, RIKEN BioResource Research Center, Ibaraki, JapanRegenerative Medicine and Cell Therapy Laboratories, KANEKA CORPORATION, Kobe, JapaniPS Cell Advanced Characterization and Development Team, RIKEN BioResource Research Center, Ibaraki, JapaniPS Cell Advanced Characterization and Development Team, RIKEN BioResource Research Center, Ibaraki, JapaniPS Cell Advanced Characterization and Development Team, RIKEN BioResource Research Center, Ibaraki, JapaniPS Cell Advanced Characterization and Development Team, RIKEN BioResource Research Center, Ibaraki, JapaniPS Cell Advanced Characterization and Development Team, RIKEN BioResource Research Center, Ibaraki, JapanRegenerative Medicine and Cell Therapy Laboratories, KANEKA CORPORATION, Kobe, JapanRegenerative Medicine and Cell Therapy Laboratories, KANEKA CORPORATION, Kobe, JapanRegenerative Medicine and Cell Therapy Laboratories, KANEKA CORPORATION, Kobe, JapanRegenerative Medicine and Cell Therapy Laboratories, KANEKA CORPORATION, Kobe, JapanRegenerative Medicine and Cell Therapy Laboratories, KANEKA CORPORATION, Kobe, JapanRegenerative Medicine and Cell Therapy Laboratories, KANEKA CORPORATION, Kobe, JapanResearch and Development Center, CiRA Foundation, Kyoto, JapanResearch and Development Center, CiRA Foundation, Kyoto, JapanCell Engineering Division, RIKEN BioResource Research Center, Ibaraki, JapanGene Engineering Division, RIKEN BioResource Research Center, Ibaraki, JapanCell Engineering Division, RIKEN BioResource Research Center, Ibaraki, JapanRegenerative Medicine and Cell Therapy Laboratories, KANEKA CORPORATION, Kobe, JapanRegenerative Medicine and Cell Therapy Laboratories, KANEKA CORPORATION, Kobe, JapanResearch and Development Center, CiRA Foundation, Kyoto, JapaniPS Cell Advanced Characterization and Development Team, RIKEN BioResource Research Center, Ibaraki, Japan; Faculty of Medicine and School of Integrative and Global Majors, University of Tsukuba, Ibaraki, JapanHuman induced pluripotent stem cells (hiPSCs) are promising resources for producing various types of tissues in regenerative medicine; however, the improvement in a scalable culture system that can precisely control the cellular status of hiPSCs is needed. Utilizing suspension culture without microcarriers or special materials allows for massive production, automation, cost-effectiveness, and safety assurance in industrialized regenerative medicine. Here, we found that hiPSCs cultured in suspension conditions with continuous agitation without microcarriers or extracellular matrix components were more prone to spontaneous differentiation than those cultured in conventional adherent conditions. Adding PKCβ and Wnt signaling pathway inhibitors in the suspension conditions suppressed the spontaneous differentiation of hiPSCs into ectoderm and mesendoderm, respectively. In these conditions, we successfully completed the culture processes of hiPSCs, including the generation of hiPSCs from peripheral blood mononuclear cells with the expansion of bulk population and single-cell sorted clones, long-term culture with robust self-renewal characteristics, single-cell cloning, direct cryopreservation from suspension culture and their successful recovery, and efficient mass production of a clinical-grade hiPSC line. Our results demonstrate that precise control of the cellular status in suspension culture conditions paves the way for their stable and automated clinical application.https://elifesciences.org/articles/89724induced pluripotent stem cellssuspension cultureprotein kinase CWNTbioreactor |
| spellingShingle | Mami Matsuo-Takasaki Sho Kambayashi Yasuko Hemmi Tamami Wakabayashi Tomoya Shimizu Yuri An Hidenori Ito Kazuhiro Takeuchi Masato Ibuki Terasu Kawashima Rio Masayasu Manami Suzuki Yoshikazu Kawai Masafumi Umekage Tomoaki M Kato Michiya Noguchi Koji Nakade Yukio Nakamura Tomoyuki Nakaishi Naoki Nishishita Masayoshi Tsukahara Yohei Hayashi Complete suspension culture of human induced pluripotent stem cells supplemented with suppressors of spontaneous differentiation eLife induced pluripotent stem cells suspension culture protein kinase C WNT bioreactor |
| title | Complete suspension culture of human induced pluripotent stem cells supplemented with suppressors of spontaneous differentiation |
| title_full | Complete suspension culture of human induced pluripotent stem cells supplemented with suppressors of spontaneous differentiation |
| title_fullStr | Complete suspension culture of human induced pluripotent stem cells supplemented with suppressors of spontaneous differentiation |
| title_full_unstemmed | Complete suspension culture of human induced pluripotent stem cells supplemented with suppressors of spontaneous differentiation |
| title_short | Complete suspension culture of human induced pluripotent stem cells supplemented with suppressors of spontaneous differentiation |
| title_sort | complete suspension culture of human induced pluripotent stem cells supplemented with suppressors of spontaneous differentiation |
| topic | induced pluripotent stem cells suspension culture protein kinase C WNT bioreactor |
| url | https://elifesciences.org/articles/89724 |
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