Proinflammatory signals are insufficient to drive definitive hematopoietic specification of human HSCs in vitro.

Abstract

Recent studies in zebrafish and mice have revealed that proinflammatory signaling is a positive regulator of definitive hematopoietic development. Whether proinflammatory signaling also regulates human hematopoietic specification remains unknown. Here, we explored the impact of the proinflammatory cytokines tumor necrosis factor-α (TNFα), interferon-γ (IFNγ), and interleukin-1β (IL1β) on in vitro hematopoietic differentiation using human pluripotent stem cells. Gene expression analysis and enzyme-linked immunosorbent assay revealed the absence of a proinflammatory signature during hematopoietic development of human pluripotent stem cells. Functionally, the emergence of hemogenic endothelial progenitors (CD31+CD34+CD45- or CD34+CD43-CD73-) and hematopoietic cells (CD43+CD45+) was not affected by treatment with increasing doses of TNFα, IFNγ, and IL1β irrespective of the developmental window or the differentiation protocol used (embryoid body or OP9 co-culture based). Similarly, knockdown of endogenous NF-kB signaling had no impact on hematopoietic differentiation of human pluripotent stem cells. This study serves as a demonstration that TNFα, IFNγ, and IL1β signals do not improve hematopoietic differentiation of human pluripotent stem cells using current protocols and suggests that proinflammatory signaling is insufficient to drive definitive hematopoietic specification of human hematopoietic stem cells in vitro.

This work was supported by the European Research Council (646903 to PM) and the Spanish Ministry of Economy and Competitiveness (SAF2013-43072R to AG, SAF2013-43065R to PM, and ISCIII/FEDER-PI14/01119 to CB). CB is supported by a Miguel Servet contract (CPII13/00011). AG is supported by the Ramon y Cajal Program (RyC-2013-13221). PM also acknowledges support from Obra Social La Caixa-Fundacio Josep Carreras and Generalitat de Catalunya (SGR330).