Superior Red Blood Cell Generation from Human Pluripotent Stem Cells Through a Novel Microcarrier-Based Embryoid Body Platform

Tissue Eng Part C Methods. 2016 Aug;22(8):765-80. doi: 10.1089/ten.TEC.2015.0579. Epub 2016 Aug 1.

Abstract

In vitro generation of red blood cells (RBCs) from human embryonic stem cells and human induced pluripotent stem cells appears to be a promising alternate approach to circumvent shortages in donor-derived blood supplies for clinical applications. Conventional methods for hematopoietic differentiation of human pluripotent stem cells (hPSC) rely on embryoid body (EB) formation and/or coculture with xenogeneic cell lines. However, most current methods for hPSC expansion and EB formation are not amenable for scale-up to levels required for large-scale RBC generation. Moreover, differentiation methods that rely on xenogenic cell lines would face obstacles for future clinical translation. In this study, we report the development of a serum-free and chemically defined microcarrier-based suspension culture platform for scalable hPSC expansion and EB formation. Improved survival and better quality EBs generated with the microcarrier-based method resulted in significantly improved mesoderm induction and, when combined with hematopoietic differentiation, resulted in at least a 6-fold improvement in hematopoietic precursor expansion, potentially culminating in a 80-fold improvement in the yield of RBC generation compared to a conventional EB-based differentiation method. In addition, we report efficient terminal maturation and generation of mature enucleated RBCs using a coculture system that comprised primary human mesenchymal stromal cells. The microcarrier-based platform could prove to be an appealing strategy for future scale-up of hPSC culture, EB generation, and large-scale generation of RBCs under defined and xeno-free conditions.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Culture Techniques / instrumentation*
  • Cell Culture Techniques / methods*
  • Cell Differentiation*
  • Cells, Cultured
  • Coculture Techniques
  • Culture Media, Serum-Free
  • Embryoid Bodies / cytology*
  • Erythrocytes / cytology*
  • Humans
  • Pluripotent Stem Cells / cytology*

Substances

  • Culture Media, Serum-Free