西亚试剂：Expansion and maintenance of human embryonic stem cell–deri

Expansion and maintenance of human embryonic stem cell–derived endothelial cells by TGFβ inhibition is Id1 dependent

Daylon James1, Hyung-song Nam2,7,8, Marco Seandel1,3,8, Daniel Nolan1, Tyler Janovitz1, Mark Tomishima4, Lorenz Studer4, Gabsang Lee4, David Lyden1, Robert Benezra2, Nikica Zaninovic5, Zev Rosenwaks5, Sina Y Rabbany1,6 & Shahin Rafii1

Previous efforts to differentiate human embryonic stem cells (hESCs) into endothelial cells have not achieved sustained expansion and stability of vascular cells. To define vasculogenic developmental pathways and enhance differentiation, we used an endothelial cell–specific VE-cadherin promoter driving green fluorescent protein (GFP) (hVPr-GFP) to screen for factors that promote vascular commitment. In phase 1 of our method, inhibition of transforming growth factor (TGF)β at day 7 of differentiation increases hVPr-GFP+ cells by tenfold. In phase 2, TGFβ inhibition maintains the proliferation and vascular identity of purified endothelial cells, resulting in a net 36-fold expansion of endothelial cells in homogenous monolayers, which exhibited a transcriptional profile of Id1highVEGFR2highVE-cadherin+ ephrinB2+. Using an Id1-YFP hESC reporter line, we showed that TGFβ inhibition sustains Id1 expression in hESC-derived endothelial cells and that Id1 is required for increased proliferation and preservation of endothelial cell commitment. Our approach provides a serum-free method for differentiation and long-term maintenance of hESC-derived endothelial cells at a scale relevant to clinical application.

1 Howard Hughes Medical Institute, Ansary Stem Cell Institute, Department of Genetic Medicine, Weill Cornell Medical College, New York, New York, USA.
2 Program in Cancer Biology and Genetics, Memorial Sloan-Kettering Cancer Center, New York, New York, USA.
3 Division of Medical Oncology, Department of Medicine, Memorial Sloan-Kettering Cancer Center, New York, New York, USA.
4 Developmental Biology Program, Memorial Sloan-Kettering Cancer Center, New York, New York, USA.
5 Ronald O. Perelman and Claudia Cohen Center for Reproductive Medicine, New York, New York, USA.
6 Bioengineering Program, Hofstra University, Hempstead, New York, USA.
7 Present address: Weill Cornell Medical College, New York, New York, USA.
8 These authors contributed equally to this work.

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