Spatial heterogeneity analysis of seeding of human induced pluripotent stem cells for neuroectodermal differentiation
Introduction: Preparing a uniform cell population in high–density seeding of adherent human induced pluripotent stem cells (hiPSC) requires stable culture conditions and consistent culture operation. In this study, we evaluated cell distribution patterns by changing cell seeding operations and their...
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Elsevier
2024-06-01
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| Series: | Regenerative Therapy |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2352320424001846 |
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| author | Ali Ahmed Issa Qatan Shinji Tanbara Masakazu Inamori Kazuhiro Fukumori Masahiro Kino–oka |
| author_facet | Ali Ahmed Issa Qatan Shinji Tanbara Masakazu Inamori Kazuhiro Fukumori Masahiro Kino–oka |
| author_sort | Ali Ahmed Issa Qatan |
| collection | DOAJ |
| description | Introduction: Preparing a uniform cell population in high–density seeding of adherent human induced pluripotent stem cells (hiPSC) requires stable culture conditions and consistent culture operation. In this study, we evaluated cell distribution patterns by changing cell seeding operations and their impact on differentiation toward the neuroectodermal lineage. Methods: The hiPSC line 201B7 was seeded at 1.23 × 105 cells/cm2 following a conventional operation, prolongated time of cell seeding suspension or vessel tilting during cell seeding operation. Fluorescent imaging of cell nuclei was performed 24 h following cell seeding and used for spatial heterogeneity analysis. Flow cytometric analysis was also performed seven days after cell differentiation induction toward neuroectodermal lineage. Results: Indices for spatial heterogeneity following high–density cell seeding were proposed to assess cell distribution patterns. Global heterogeneity (HG) was shown to be mostly affected by vessel tilting during cell seeding operation, while local heterogeneity (HL) was affected by prolongated time of cell seeding suspension. Changes in both spatial heterogeneities in the hiPSC population resulted in a lower yield of target neuroectodermal cells compared with the control operation. Conclusion: High–density hiPSC seeding is critical for achieving a higher yield of target cells of neuroectodermal lineage. Understanding the spatial heterogeneity in early stages detects errors in cell culture motion and predicts cell fate in later stages of cell culture. |
| format | Article |
| id | doaj-art-0370f517a1a8460b98f072575d4a770f |
| institution | OA Journals |
| issn | 2352-3204 |
| language | English |
| publishDate | 2024-06-01 |
| publisher | Elsevier |
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| series | Regenerative Therapy |
| spelling | doaj-art-0370f517a1a8460b98f072575d4a770f2025-08-20T02:07:31ZengElsevierRegenerative Therapy2352-32042024-06-012692293110.1016/j.reth.2024.10.006Spatial heterogeneity analysis of seeding of human induced pluripotent stem cells for neuroectodermal differentiationAli Ahmed Issa Qatan0Shinji Tanbara1Masakazu Inamori2Kazuhiro Fukumori3Masahiro Kino–oka4Department of Biotechnology, Graduate School of Engineering, Osaka University, 2–1 Yamadaoka, Suita, Osaka 565–0871, JapanDepartment of Biotechnology, Graduate School of Engineering, Osaka University, 2–1 Yamadaoka, Suita, Osaka 565–0871, JapanCell Manufacturing Systems Engineering (Healios) Joint Research Chair, Graduate School of Engineering, Osaka University, 2–1 Yamadaoka, Suita, Osaka 565–0871, JapanDepartment of Biotechnology, Graduate School of Engineering, Osaka University, 2–1 Yamadaoka, Suita, Osaka 565–0871, JapanDepartment of Biotechnology, Graduate School of Engineering, Osaka University, 2–1 Yamadaoka, Suita, Osaka 565–0871, Japan; Cell Manufacturing Systems Engineering (Healios) Joint Research Chair, Graduate School of Engineering, Osaka University, 2–1 Yamadaoka, Suita, Osaka 565–0871, Japan; Research Base for Cell Manufacturability, TechnoArena, Graduate School of Engineering, Osaka University, 2–1 Yamadaoka, Suita, Osaka 565–0871, Japan; Corresponding author. Cell Manufacturing Systems Engineering (Healios) Joint Research Chair, Graduate School of Engineering, Osaka University, 2–1 Yamadaoka, Suita, Osaka 565–0871, Japan.Introduction: Preparing a uniform cell population in high–density seeding of adherent human induced pluripotent stem cells (hiPSC) requires stable culture conditions and consistent culture operation. In this study, we evaluated cell distribution patterns by changing cell seeding operations and their impact on differentiation toward the neuroectodermal lineage. Methods: The hiPSC line 201B7 was seeded at 1.23 × 105 cells/cm2 following a conventional operation, prolongated time of cell seeding suspension or vessel tilting during cell seeding operation. Fluorescent imaging of cell nuclei was performed 24 h following cell seeding and used for spatial heterogeneity analysis. Flow cytometric analysis was also performed seven days after cell differentiation induction toward neuroectodermal lineage. Results: Indices for spatial heterogeneity following high–density cell seeding were proposed to assess cell distribution patterns. Global heterogeneity (HG) was shown to be mostly affected by vessel tilting during cell seeding operation, while local heterogeneity (HL) was affected by prolongated time of cell seeding suspension. Changes in both spatial heterogeneities in the hiPSC population resulted in a lower yield of target neuroectodermal cells compared with the control operation. Conclusion: High–density hiPSC seeding is critical for achieving a higher yield of target cells of neuroectodermal lineage. Understanding the spatial heterogeneity in early stages detects errors in cell culture motion and predicts cell fate in later stages of cell culture.http://www.sciencedirect.com/science/article/pii/S2352320424001846Spatial heterogeneity analysisHigh–density cell seedingCell seeding operationHuman induced pluripotent stem cellsNeuroectodermal differentiation |
| spellingShingle | Ali Ahmed Issa Qatan Shinji Tanbara Masakazu Inamori Kazuhiro Fukumori Masahiro Kino–oka Spatial heterogeneity analysis of seeding of human induced pluripotent stem cells for neuroectodermal differentiation Regenerative Therapy Spatial heterogeneity analysis High–density cell seeding Cell seeding operation Human induced pluripotent stem cells Neuroectodermal differentiation |
| title | Spatial heterogeneity analysis of seeding of human induced pluripotent stem cells for neuroectodermal differentiation |
| title_full | Spatial heterogeneity analysis of seeding of human induced pluripotent stem cells for neuroectodermal differentiation |
| title_fullStr | Spatial heterogeneity analysis of seeding of human induced pluripotent stem cells for neuroectodermal differentiation |
| title_full_unstemmed | Spatial heterogeneity analysis of seeding of human induced pluripotent stem cells for neuroectodermal differentiation |
| title_short | Spatial heterogeneity analysis of seeding of human induced pluripotent stem cells for neuroectodermal differentiation |
| title_sort | spatial heterogeneity analysis of seeding of human induced pluripotent stem cells for neuroectodermal differentiation |
| topic | Spatial heterogeneity analysis High–density cell seeding Cell seeding operation Human induced pluripotent stem cells Neuroectodermal differentiation |
| url | http://www.sciencedirect.com/science/article/pii/S2352320424001846 |
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