title: Lentiviral transduction and clonal selection of hESCs with endothelial-specific transgenic reporters.
abstract: Generation of vascular endothelial cells (EC) from human embryonic stem cells (hESC) is a vital component of cell-based strategies for treatment of cardiovascular disease. Before hESC-derived ECs can be administered in therapeutic modalities, however, chemically defined culture conditions must be developed that reproducibly and robustly induce vascular differentiation. One approach to screening for culture conditions that support differentiation of hESCs to any cell type is their genetic modification with exogenous DNA sequence comprising a tissue-specific gene promoter driving reporters such as fluorescent protein or antibiotic drug resistance. The protocols herein provide instructions for the generation of clonal hESC lines containing a reporter transgene that is specifically expressed in vascular endothelial derivatives. Additionally, they demonstrate the methodology employed to assess vascular differentiation from clonal lines. Together, these protocols provide a solid foundation for study of vascular differentiation, and may also be applied, in principle, to studies of other specialized cell types derived from hESCs.
authors.affiliation: Department of Genetic Medicine, Weill Cornell Medical College, New York, New York, USA.
authors.forename: Daylon
authors.initials: D
authors.lastname: James
authors.forename: Qiansheng
authors.initials: Q
authors.lastname: Zhan
authors.forename: Christopher
authors.initials: C
authors.lastname: Kloss
authors.forename: Nikica
authors.initials: N
authors.lastname: Zaninovic
authors.forename: Zev
authors.initials: Z
authors.lastname: Rosenwaks
authors.forename: Shahin
authors.initials: S
authors.lastname: Rafii
mesh_terms: D002460:Cell Line; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism*; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D017930:Genes, Reporter; D006801:Humans; D016086:Lentivirus; D009928:Organ Specificity; D014161:Transduction, Genetic / Q000379:methods*; D019076:Transgenes