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Chondrogenic Differentiation of Human-Induced Pluripotent Stem Cells

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Cartilage Tissue Engineering

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2598))

Abstract

The generation of large quantities of genetically defined human chondrocytes remains a critical step for the development of tissue engineering strategies for cartilage regeneration and high-throughput drug screening. This protocol describes chondrogenic differentiation of human-induced pluripotent stem cells (hiPSCs), which can undergo genetic modification and the capacity for extensive cell expansion. The hiPSCs are differentiated in a stepwise manner in monolayer through the mesodermal lineage for 12 days using defined growth factors and small molecules. This is followed by 28 days of chondrogenic differentiation in a 3D pellet culture system using transforming growth factor beta 3 and specific compounds to inhibit off-target differentiation. The 6-week protocol results in hiPSC-derived cartilaginous tissue that can be characterized by histology, immunohistochemistry, and gene expression or enzymatically digested to isolate chondrocyte-like cells. Investigators can use this protocol for experiments including genetic engineering, in vitro disease modeling, or tissue engineering.

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Acknowledgements

This work was supported by the Shriners Hospitals for Children, Nancy Taylor Foundation, Arthritis Foundation, NIH (AG46927, AG15768, AR075899, AR072999, AR074992, AR073752, AR080902, T32 DK108742, T32 EB018266), and Taiwan GSSA Scholarship.

Conflicts of Interest

FG is an employee of Cytex Therapeutics, Inc. The other authors declare that they have no competing interests.

Author Contribution Statement

ARD, NS, CLW, and FG were involved in the development, testing, and troubleshooting of these protocols and the writing of this paper.

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Author notes

  1. Farshid Guilak and Chia-Lung Wu contributed equally with all other contributors.

Authors and Affiliations

  1. Department of Orthopaedic Surgery, Washington University, St. Louis, MO, USA

    Amanda R. Dicks, Nancy Steward & Farshid Guilak

  2. Shriners Hospitals for Children – St. Louis, St. Louis, MO, USA

    Amanda R. Dicks, Nancy Steward & Farshid Guilak

  3. Department of Biomedical Engineering, Washington University, St. Louis, MO, USA

    Amanda R. Dicks & Farshid Guilak

  4. Center of Regenerative Medicine, Washington University, St. Louis, MO, USA

    Amanda R. Dicks, Nancy Steward & Farshid Guilak

  5. Department of Orthopaedic Surgery and Rehabilitation, Center for Musculoskeletal Research, University of Rochester, Rochester, NY, USA

    Chia-Lung Wu

Authors
  1. Amanda R. Dicks
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  2. Nancy Steward
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  3. Farshid Guilak
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  4. Chia-Lung Wu
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Corresponding authors

Correspondence to Farshid Guilak or Chia-Lung Wu .

Editor information

Editors and Affiliations

  1. AO Research Institute Davos, Davos Platz, Switzerland

    Martin J. Stoddart

  2. AO Research Institute Davos, Davos Platz, Switzerland

    Elena Della Bella

  3. UCB Pharma, Slough, UK

    Angela R. Armiento

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Dicks, A.R., Steward, N., Guilak, F., Wu, CL. (2023). Chondrogenic Differentiation of Human-Induced Pluripotent Stem Cells. In: Stoddart, M.J., Della Bella, E., Armiento, A.R. (eds) Cartilage Tissue Engineering. Methods in Molecular Biology, vol 2598. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2839-3_8

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  • DOI: https://doi.org/10.1007/978-1-0716-2839-3_8

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2838-6

  • Online ISBN: 978-1-0716-2839-3

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