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A Quick and Efficient Protocol for RBC Production from hPSC

Review of  “High-Efficiency Serum-Free Feeder-Free Erythroid Differentiation of Human Pluripotent Stem Cells Using Small Molecules” from Stem Cells Translational Medicine by Stuart P. Atkinson

The production of large amounts of red blood cells (RBCs or erythrocytes) from human pluripotent stem cells (hPSCs) represents an exciting strategy with both basic and translational research applications. While previous studies have reported varying levels of success [1-5], none have ticked all the boxes required for the clinical application of hPSC-derived RBCs.

In a new Stem Cells Translational Medicine study, intrepid researchers from the laboratory of Joanne C. Mountford (University of Glasgow, UK) have now described a quick and efficient feeder- and serum-free protocol for the large scale production of RBCs from hPSCs [6]. Will this strategy bring hPSC-derived RBCs closer to the clinic?

This new paper describes (in great detail) the combination of various cytokines and small molecule drugs into a multistep month-long protocol (See Figure for some of the detail). Following this methodology, Olivier et al hoped to produce large amounts of RBCs from hPSCs by mimicking known developmental processes and doing away with any requirement for laborious and expensive post-differentiation cell selection/sorting steps.

This newly described strategy helped the authors to generate a cell population with an extraordinary level of cell purity; nearly all cells expressed an erythroid marker by the end of the 4-week protocol. Even though only around 10% of cells underwent spontaneous enucleation, the massive expansion of cell numbers (50,000–200,000 erythroid cells for each hPSC employed) led to the production of a significant number of morphologically mature RBCs.

At the molecular level, a high percentage of the erythroid cells still expressed fetal hemoglobins (90-95%). However, the addition of various small molecules, such as inhibitor VIII (GSK3b inhibitor), IBMX (inhibitor of cAMP and cGMP phosphodiesterases), SR1 (antagonist of the aryl hydrocarbon receptor), and Pluripotin, increased the proportion of adult globins and decreased the proportion of embryonic globins. The authors noted that this globin expression pattern resembled that observed in umbilical cord blood and, therefore, should make RBCs produced via this new protocol suitable for use in transfusions.

This quick and efficient feeder- and serum-free defined protocol represents a grand step towards the clinical application of hPSC-derived erythrocytes. However, this improved methodology also provides researchers a tractable system to pull apart the regulatory mechanisms that control the various stages of hematopoiesis and erythropoiesis. Furthermore, the application of disease- and patient-specific induced pluripotent stem cell (iPSC) with this protocol may also prove useful in the modeling of diseases and for drug screening. 


  1. Lu SJ, Feng Q, Park JS, et al. Biologic properties and enucleation of red blood cells from human embryonic stem cells. Blood 2008;112:4475-4484.
  2. Salvagiotto G, Burton S, Daigh CA, et al. A defined, feeder-free, serum-free system to generate in vitro hematopoietic progenitors and differentiated blood cells from hESCs and hiPSCs. PLoS One 2011;6:e17829.
  3. Feng Q, Lu SJ, Klimanskaya I, et al. Hemangioblastic derivatives from human induced pluripotent stem cells exhibit limited expansion and early senescence. Stem Cells 2010;28:704-712.
  4. Dias J, Gumenyuk M, Kang H, et al. Generation of red blood cells from human induced pluripotent stem cells. Stem Cells Dev 2011;20:1639-1647.
  5. Kobari L, Yates F, Oudrhiri N, et al. Human induced pluripotent stem cells can reach complete terminal maturation: in vivo and in vitro evidence in the erythropoietic differentiation model. Haematologica 2012;97:1795-1803.
  6. Olivier EN, Marenah L, McCahill A, et al. High-Efficiency Serum-Free Feeder-Free Erythroid Differentiation of Human Pluripotent Stem Cells Using Small Molecules. Stem Cells Transl Med 2016;5:1394-1405.