Three-Dimensional Co-culture of Human Hematopoietic Stem/Progenitor Cells and Mesenchymal Stem/Stromal Cells in a Biomimetic Hematopoietic Niche Microenvironment

  • Marta H. G. Costa
  • Tiago S. Monteiro
  • Susana Cardoso
  • Joaquim M. S. Cabral
  • Frederico Castelo Ferreira
  • Cláudia L. da SilvaEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 2002)


The development of cellular therapies to treat hematological malignancies has motivated researchers to investigate ex vivo culture systems capable of expanding the number of hematopoietic stem/progenitor cells (HSPC) before transplantation. The strategies exploited to achieve relevant cell numbers have relied on culture systems that lack biomimetic niche cues thought to be essential to promote HSPC maintenance and proliferation. Although stromal cells adhered to 2-D surfaces can be used to support the expansion of HSPC ex vivo, culture systems aiming to incorporate cell–cell interactions in a more intricate 3-D environment can better contribute to recapitulate the bone marrow (BM) hematopoietic niche in vitro.

Herein, we describe the development of a 3-D co-culture system of human umbilical cord blood (UCB)-derived CD34+ cells and BM mesenchymal stem/stromal cell (MSC) spheroids in a microwell-based platform that allows to attain large numbers of spheroids with uniform sizes. Further comparison with a traditional 2-D co-culture system exploiting the supportive features of feeder layers of MSC is provided, while functional in vitro assays to assess the features of HSPC expanded in the 2-D vs. 3-D MSC co-culture systems are suggested.


Co-culture Hematopoietic niche Hematopoietic stem/progenitor cells (HSPC) Mesenchymal stem/stromal cells (MSC) Microwells Spheroids 



MHC acknowledges Fundação para a Ciência e a Tecnologia (FCT), Portugal, for granting PhD scholarship SFRH/BD/52000/2012. Funding received by iBB—Institute for Bioengineering and Biosciences from FCT (UID/BIO/04565/2013) and from Programa Operacional Regional de Lisboa 2020 (Project N. 007317) is acknowledged. The authors also acknowledge the funding received from Programa Operacional Regional de Lisboa 2020 through the project PRECISE—Accelerating progress toward the new era of precision medicine (Project N. 16394) and from FCT through the project Design and operation of a prototype packed-bed reactor for the production of hematopoietic stem/progenitor cells (PTDC/QEQ-EPR/6623/2014).


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Copyright information

© Springer Science+Business Media New York 2018

Authors and Affiliations

  • Marta H. G. Costa
    • 1
  • Tiago S. Monteiro
    • 2
  • Susana Cardoso
    • 2
  • Joaquim M. S. Cabral
    • 1
    • 3
  • Frederico Castelo Ferreira
    • 1
    • 3
  • Cláudia L. da Silva
    • 1
    • 3
    Email author
  1. 1.Department of Bioengineering and IBB-Institute for Bioengineering and Biosciences, Instituto Superior TécnicoUniversidade de LisboaLisboaPortugal
  2. 2.Instituto de Engenharia de Sistemas de Computadores – Microsystems and Nanotechnology (INESC–MN)LisboaPortugal
  3. 3.The Discoveries Centre for Regenerative and Precision Medicine, Lisbon Campus, Instituto Superior TécnicoUniversidade de LisboaLisboaPortugal

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