Isolation, differentiation and characterization of vascular cells derived from human embryonic stem cells.
Herein, we describe a protocol for the isolation of human embryonic stem cells (hESCs)-derived vascular cells at various stages of development. The cells are isolated from 10 to 15-d-old human embryoid bodies (EBs) cultured in suspension. After dissociation, cells are labeled with anti-CD34 or anti-CD31 (PECAM1) antibody and separated from the cell mixture by magnetic-activated cell separation (MACS) or fluorescent-activated cell sorting (FACS). Isolated vascular cells are then cultured in media conditions that support specific differentiation and expansion pathways. The resulting vascular cell populations contain >80% endothelial-like or smooth muscle-like cells. Assuming typical initial cell adhesion and proliferation rates, the entire procedure can be completed within 1.5 months. Vascular cells isolated and differentiated under the described conditions may constitute a potential cell source for therapeutic application toward repair of ischemic tissues, preparation of tissue-engineered vascular grafts and design of cellular kits for drug screening applications.
Faculty of Biomedical Engineering, Technion Institute of Technology, Haifa, Israel.
Shulamit
S
Levenberg
Lino S
LS
Ferreira
Limor
L
Chen-Konak
Thomas P
TP
Kraehenbuehl
Robert
R
Langer
D000818:Animals; D001483:Base Sequence; D002454:Cell Differentiation; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D017931:DNA Primers / Q000235:genetics; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D051379:Mice; D032389:Myocytes, Smooth Muscle / Q000166:cytology* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D023822:Tissue Engineering
D017931:DNA Primers