Efficient Large-Scale 2D Culture System for Human Induced Pluripotent Stem Cells and Differentiated Cardiomyocytes

Shugo Tohyama, Jun Fujita, Chihana Fujita, Miho Yamaguchi, Sayaka Kanaami, Rei Ohno, Kazuho Sakamoto, Masami Kodama, Junko Kurokawa, Hideaki Kanazawa, Tomohisa Seki, Yoshikazu Kishino, Marina Okada, Kazuaki Nakajima, Sho Tanosaki, Shota Someya, Akinori Hirano, Shinji Kawaguchi, Eiji Kobayashi, Keiichi Fukuda

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

Cardiac regenerative therapies utilizing human induced pluripotent stem cells (hiPSCs) are hampered by ineffective large-scale culture. hiPSCs were cultured in multilayer culture plates (CPs) with active gas ventilation (AGV), resulting in stable proliferation and pluripotency. Seeding of 1 × 106 hiPSCs per layer yielded 7.2 × 108 hiPSCs in 4-layer CPs and 1.7 × 109 hiPSCs in 10-layer CPs with pluripotency. hiPSCs were sequentially differentiated into cardiomyocytes (CMs) in a two-dimensional (2D) differentiation protocol. The efficiency of cardiac differentiation using 10-layer CPs with AGV was 66%-87%. Approximately 6.2-7.0 × 108 cells (4-layer) and 1.5-2.8 × 109 cells (10-layer) were obtained with AGV. After metabolic purification with glucose- and glutamine-depleted and lactate-supplemented media, a massive amount of purified CMs was prepared. Here, we present a scalable 2D culture system using multilayer CPs with AGV for hiPSC-derived CMs, which will facilitate clinical applications for severe heart failure in the near future. In this article, Fujita and colleagues show that human induced pluripotent stem cells (hiPSCs) and hiPSC-derived cardiomyocytes (hiPSC-CMs) are stably and efficiently produced using multilayer culture plates with an active gas ventilation system because of stable CO2 and O2 concentration in culture media. Large numbers of obtained hiPSC-CMs are metabolically purified under glucose- and glutamine-depleted and lactate-supplemented conditions.

Original languageEnglish
JournalStem Cell Reports
DOIs
Publication statusAccepted/In press - 2017

Fingerprint

Induced Pluripotent Stem Cells
Stem cells
Cell culture
Cardiac Myocytes
Ventilation
Gases
Multilayers
Glutamine
Lactic Acid
Glucose
Purification
Culture Media
Heart Failure

Keywords

  • 2D culture
  • Cardiomyocytes
  • IPSCs
  • Large scale
  • Purification
  • Regenerative medicine

ASJC Scopus subject areas

  • Biochemistry
  • Genetics
  • Developmental Biology
  • Cell Biology

Cite this

Efficient Large-Scale 2D Culture System for Human Induced Pluripotent Stem Cells and Differentiated Cardiomyocytes. / Tohyama, Shugo; Fujita, Jun; Fujita, Chihana; Yamaguchi, Miho; Kanaami, Sayaka; Ohno, Rei; Sakamoto, Kazuho; Kodama, Masami; Kurokawa, Junko; Kanazawa, Hideaki; Seki, Tomohisa; Kishino, Yoshikazu; Okada, Marina; Nakajima, Kazuaki; Tanosaki, Sho; Someya, Shota; Hirano, Akinori; Kawaguchi, Shinji; Kobayashi, Eiji; Fukuda, Keiichi.

In: Stem Cell Reports, 2017.

Research output: Contribution to journalArticle

Tohyama, S, Fujita, J, Fujita, C, Yamaguchi, M, Kanaami, S, Ohno, R, Sakamoto, K, Kodama, M, Kurokawa, J, Kanazawa, H, Seki, T, Kishino, Y, Okada, M, Nakajima, K, Tanosaki, S, Someya, S, Hirano, A, Kawaguchi, S, Kobayashi, E & Fukuda, K 2017, 'Efficient Large-Scale 2D Culture System for Human Induced Pluripotent Stem Cells and Differentiated Cardiomyocytes', Stem Cell Reports. https://doi.org/10.1016/j.stemcr.2017.08.025
Tohyama, Shugo ; Fujita, Jun ; Fujita, Chihana ; Yamaguchi, Miho ; Kanaami, Sayaka ; Ohno, Rei ; Sakamoto, Kazuho ; Kodama, Masami ; Kurokawa, Junko ; Kanazawa, Hideaki ; Seki, Tomohisa ; Kishino, Yoshikazu ; Okada, Marina ; Nakajima, Kazuaki ; Tanosaki, Sho ; Someya, Shota ; Hirano, Akinori ; Kawaguchi, Shinji ; Kobayashi, Eiji ; Fukuda, Keiichi. / Efficient Large-Scale 2D Culture System for Human Induced Pluripotent Stem Cells and Differentiated Cardiomyocytes. In: Stem Cell Reports. 2017.
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abstract = "Cardiac regenerative therapies utilizing human induced pluripotent stem cells (hiPSCs) are hampered by ineffective large-scale culture. hiPSCs were cultured in multilayer culture plates (CPs) with active gas ventilation (AGV), resulting in stable proliferation and pluripotency. Seeding of 1 × 106 hiPSCs per layer yielded 7.2 × 108 hiPSCs in 4-layer CPs and 1.7 × 109 hiPSCs in 10-layer CPs with pluripotency. hiPSCs were sequentially differentiated into cardiomyocytes (CMs) in a two-dimensional (2D) differentiation protocol. The efficiency of cardiac differentiation using 10-layer CPs with AGV was 66{\%}-87{\%}. Approximately 6.2-7.0 × 108 cells (4-layer) and 1.5-2.8 × 109 cells (10-layer) were obtained with AGV. After metabolic purification with glucose- and glutamine-depleted and lactate-supplemented media, a massive amount of purified CMs was prepared. Here, we present a scalable 2D culture system using multilayer CPs with AGV for hiPSC-derived CMs, which will facilitate clinical applications for severe heart failure in the near future. In this article, Fujita and colleagues show that human induced pluripotent stem cells (hiPSCs) and hiPSC-derived cardiomyocytes (hiPSC-CMs) are stably and efficiently produced using multilayer culture plates with an active gas ventilation system because of stable CO2 and O2 concentration in culture media. Large numbers of obtained hiPSC-CMs are metabolically purified under glucose- and glutamine-depleted and lactate-supplemented conditions.",
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AU - Tohyama, Shugo

AU - Fujita, Jun

AU - Fujita, Chihana

AU - Yamaguchi, Miho

AU - Kanaami, Sayaka

AU - Ohno, Rei

AU - Sakamoto, Kazuho

AU - Kodama, Masami

AU - Kurokawa, Junko

AU - Kanazawa, Hideaki

AU - Seki, Tomohisa

AU - Kishino, Yoshikazu

AU - Okada, Marina

AU - Nakajima, Kazuaki

AU - Tanosaki, Sho

AU - Someya, Shota

AU - Hirano, Akinori

AU - Kawaguchi, Shinji

AU - Kobayashi, Eiji

AU - Fukuda, Keiichi

PY - 2017

Y1 - 2017

N2 - Cardiac regenerative therapies utilizing human induced pluripotent stem cells (hiPSCs) are hampered by ineffective large-scale culture. hiPSCs were cultured in multilayer culture plates (CPs) with active gas ventilation (AGV), resulting in stable proliferation and pluripotency. Seeding of 1 × 106 hiPSCs per layer yielded 7.2 × 108 hiPSCs in 4-layer CPs and 1.7 × 109 hiPSCs in 10-layer CPs with pluripotency. hiPSCs were sequentially differentiated into cardiomyocytes (CMs) in a two-dimensional (2D) differentiation protocol. The efficiency of cardiac differentiation using 10-layer CPs with AGV was 66%-87%. Approximately 6.2-7.0 × 108 cells (4-layer) and 1.5-2.8 × 109 cells (10-layer) were obtained with AGV. After metabolic purification with glucose- and glutamine-depleted and lactate-supplemented media, a massive amount of purified CMs was prepared. Here, we present a scalable 2D culture system using multilayer CPs with AGV for hiPSC-derived CMs, which will facilitate clinical applications for severe heart failure in the near future. In this article, Fujita and colleagues show that human induced pluripotent stem cells (hiPSCs) and hiPSC-derived cardiomyocytes (hiPSC-CMs) are stably and efficiently produced using multilayer culture plates with an active gas ventilation system because of stable CO2 and O2 concentration in culture media. Large numbers of obtained hiPSC-CMs are metabolically purified under glucose- and glutamine-depleted and lactate-supplemented conditions.

AB - Cardiac regenerative therapies utilizing human induced pluripotent stem cells (hiPSCs) are hampered by ineffective large-scale culture. hiPSCs were cultured in multilayer culture plates (CPs) with active gas ventilation (AGV), resulting in stable proliferation and pluripotency. Seeding of 1 × 106 hiPSCs per layer yielded 7.2 × 108 hiPSCs in 4-layer CPs and 1.7 × 109 hiPSCs in 10-layer CPs with pluripotency. hiPSCs were sequentially differentiated into cardiomyocytes (CMs) in a two-dimensional (2D) differentiation protocol. The efficiency of cardiac differentiation using 10-layer CPs with AGV was 66%-87%. Approximately 6.2-7.0 × 108 cells (4-layer) and 1.5-2.8 × 109 cells (10-layer) were obtained with AGV. After metabolic purification with glucose- and glutamine-depleted and lactate-supplemented media, a massive amount of purified CMs was prepared. Here, we present a scalable 2D culture system using multilayer CPs with AGV for hiPSC-derived CMs, which will facilitate clinical applications for severe heart failure in the near future. In this article, Fujita and colleagues show that human induced pluripotent stem cells (hiPSCs) and hiPSC-derived cardiomyocytes (hiPSC-CMs) are stably and efficiently produced using multilayer culture plates with an active gas ventilation system because of stable CO2 and O2 concentration in culture media. Large numbers of obtained hiPSC-CMs are metabolically purified under glucose- and glutamine-depleted and lactate-supplemented conditions.

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KW - Purification

KW - Regenerative medicine

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