A Benchmark Side-by-Side Comparison of Two Well-Established Protocols for in vitro Hematopoietic Differentiation From Human Pluripotent Stem Cells
Gutiérrez-Agüera, Francisco (Institut Germans Trias i Pujol. Institut de Recerca contra la Leucèmia Josep Carreras)
Rodriguez-Cortez, Virginia Carolina (Institut Germans Trias i Pujol. Institut de Recerca contra la Leucèmia Josep Carreras)
Petazzi, Paolo (Institut Germans Trias i Pujol. Institut de Recerca contra la Leucèmia Josep Carreras)
Bueno, Clara (Institut Germans Trias i Pujol. Institut de Recerca contra la Leucèmia Josep Carreras)
Menéndez Bujan, Pablo (Institut Germans Trias i Pujol. Institut de Recerca contra la Leucèmia Josep Carreras)
Universitat Autònoma de Barcelona

Date:	2021
Abstract:	The generation of transplantable hematopoietic stem cells (HSCs) from human pluripotent stem cells (hPSCs) remains challenging. Current differentiation protocols from hPSCs generate mostly hematopoietic progenitors of the primitive HSC-independent program, and it remains unclear what is the best combination of cytokines and hematopoietic growth factors (HGFs) for obtaining functional hematopoietic cells in vitro. Here, we have used the AND1 and H9 hESC lines and the H9:dual-reporter RUNX1C-GFP-SOX17-Cherry to compare the hematopoietic differentiation in vitro based on the treatment of embryoid bodies (EBs) with the ventral mesoderm inducer BMP4 plus HGFs in the absence (protocol 1) or presence (protocol 2) of stage-specific activation of Wnt/β-catenin and inhibition of Activin/Nodal. Despite a slight trend in favor of protocol 1, no statistically significant differences were observed between protocols at any time point analyzed throughout EB development regarding the frequency of hemogenic endothelial (HE) precursors; CD43+ CD45-, CD45+, and CD45 + CD34 + hematopoietic derivatives; or the output of clonogenic progenitors. Similarly, the kinetics of emergence throughout EB development of both SOX17 + HE and RUNX1C + definitive hematopoiesis was very similar for both protocols. The expression of the early master mesendodermal transcription factors Brachyury, MIXL1, and KDR revealed similar gene expression kinetics prior to the emergence of RUNX1C + definitive hematopoiesis for both protocols. Collectively, the simpler protocol 1 is, at least, as efficient as protocol 2, suggesting that supplementation with additional morphogens/HGFs and modulation of Activin/Nodal and Wnt/β-catenin pathways seem dispensable for in vitro hematopoietic differentiation of hPSCs.
Grants:	Ministerio de Economía y Competitividad RTC2018-4603-1 Instituto de Salud Carlos III PI20/00822
Rights:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.
Language:	Anglès
Document:	Article ; recerca ; Versió publicada
Published in:	Frontiers in Cell and Developmental Biology, Vol. 9 (21 2021) , p. 636704, ISSN 2296-634X

DOI: 10.3389/fcell.2021.636704
PMID: 34095110

7 p, 1.4 MB

The record appears in these collections:
Research literature > UAB research groups literature > Research Centres and Groups (research output) > Health sciences and biosciences > Institut d'Investigació en Ciencies de la Salut Germans Trias i Pujol (IGTP) > Josep Carreras Leukaemia Research Institute
Articles > Research articles
Articles > Published articles