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Method Article
Raphael Lis
Division of Regenerative Medicine, Ansary Stem Cell Institute, Weill Cornell Medicine
Corresponding Author
Shahin Rafii
Division of Regenerative Medicine, Ansary Stem Cell Institute, Weill Cornell Medicine
Corresponding Author
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This work is licensed under a CC BY 4.0 License. Read Full License
In this method article, we are describing a multi-phasic approach for converting adult murine endothelial cells \(ECs) to bona fide haematopoietic stem cells \(HSCs) \(rEC-HSCs) via transient expression of four transcription factors FosB, Gfi1, Runx1, and Spi1 \(FGRS) and stimulation with angiocrine factors supplied by the vascular niche. Adult mouse ECs were isolated from Runx1-IRES-GFP; Rosa26-rtTa and maintain in culture with EC growth factor and tgfb inhibition. Induction phase \(day 0-8) of conversion is initiated by expressing FGRS in mature ECs resulting in endogenous Runx1 expression. During specification phase \(day 8-20), endogenous Runx1+ FGRS-transduced ECs commit to a haematopoietic fate and no longer require FGRS expression. The vascular-niche drives robust self-renewal and expansion of rEC-HSCs \(day 20-28). Upon conversion, rEC-HSCs are endowed with a transcriptomic signature and long-term self-renewal capacity similar to adult HSCs, and are competent for clonal engraftment and multi-lineage reconstituting potential, including antigen-dependent adaptive immune function. Our work provides a tractable strategy to interrogate the generation of engraftable hematopoietic cell, and advances the mechanistic understanding of hematopoietic development and HSC self-renewal.
Keywords
Direct reprogramming, Haematopoietic stem cells, Vascular niche, Endothelial cells, Haemogenic endothelium
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S.R. is the founder and a non-paid consultant to Angiocrine Bioscience (AB), San Diego, CA.
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Associated Publications
Conversion of adult endothelium to immunocompetent haematopoietic stem cells
by Raphael Lis, Charles C. Karrasch, Michael G. Poulos, +12
Nature (22 May, 2017)
Method Article
Raphael Lis
Division of Regenerative Medicine, Ansary Stem Cell Institute, Weill Cornell Medicine
Corresponding Author
Shahin Rafii
Division of Regenerative Medicine, Ansary Stem Cell Institute, Weill Cornell Medicine
Corresponding Author
Version History
CURRENT STATUS: Posted
Version 1
Posted 22 May, 2017
No community comments so far
Metrics
Comments: 0
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
In this method article, we are describing a multi-phasic approach for converting adult murine endothelial cells \(ECs) to bona fide haematopoietic stem cells \(HSCs) \(rEC-HSCs) via transient expression of four transcription factors FosB, Gfi1, Runx1, and Spi1 \(FGRS) and stimulation with angiocrine factors supplied by the vascular niche. Adult mouse ECs were isolated from Runx1-IRES-GFP; Rosa26-rtTa and maintain in culture with EC growth factor and tgfb inhibition. Induction phase \(day 0-8) of conversion is initiated by expressing FGRS in mature ECs resulting in endogenous Runx1 expression. During specification phase \(day 8-20), endogenous Runx1+ FGRS-transduced ECs commit to a haematopoietic fate and no longer require FGRS expression. The vascular-niche drives robust self-renewal and expansion of rEC-HSCs \(day 20-28). Upon conversion, rEC-HSCs are endowed with a transcriptomic signature and long-term self-renewal capacity similar to adult HSCs, and are competent for clonal engraftment and multi-lineage reconstituting potential, including antigen-dependent adaptive immune function. Our work provides a tractable strategy to interrogate the generation of engraftable hematopoietic cell, and advances the mechanistic understanding of hematopoietic development and HSC self-renewal.
Figure 1
Figure 2
Figure 3
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Associated Publications
Conversion of adult endothelium to immunocompetent haematopoietic stem cells
by Raphael Lis, Charles C. Karrasch, Michael G. Poulos, +12
Nature (22 May, 2017)