Human embryonic stem cells which express hrGFP in the undifferentiated state and during dopaminergic differentiation.
OBJECTIVE
Human embryonic stem cells (hESCs) which express a reporter gene consistently during all phases of differentiation would be valuable for basic research on cell transplantation. In this study, we describe karyotypically-abnormal variant hESCs, BGO1V2-EFG, which express hrGFP driven by the EF1 promoter.


METHODS
BGO1V2-EFG cells were analyzed by using immunocytochemistry, single cell-based confocal image, and in vitro differentiation, including dopaminergic differentiation.


RESULTS
Undifferentiated BGO1V2-EFG cells expressed pluripotent ESC markers and retained the ability to differentiate into cell types of all three germ layers. BGO1V2-EFG cells maintained stable and robust hrGFP expression in vitro in the undifferentiated state and during differentiation. The EF1 promoter retained activity during dopaminergic differentiation, as 76% of tyrosine hydroxlase (TH)-positive cells co-expressed hrGFP by confocal analysis. Treated with sodium butyrate (0.02 mM to 2.0 mM), an inhibitor of histone deacetylase (HDAC), during differentiation did not affect hrGFP expression, although TH expression was reduced by higher concentrations of sodium butyrate.


CONCLUSIONS
BGO1V2-EFG cells maintain stable and robust hrGFP expression in the undifferentiated state and during neural differentiation. Especially, the EF1 promoter was effective in driving hrGFP expression during dopaminergic differentiation. BGO1V2-EFG cells may be useful for transplantation studies in Parkinson disease animal models.
Chen
Jia
J
Cellular Neurobiology Research Branch, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, Department of Health and Human Services, Baltimore, MD 21224, USA. jichen@mail.nih.gov
Sai
Shang-Yi T
SY
Vazin
Tandis
T
Coggiano
Mark
M
Freed
William J
WJ
D002087:Butyrates / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D004298:Dopamine / Q000235:genetics / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D053595:Embryonic Stem Cells / Q000187:drug effects / Q000502:physiology*; D004791:Enzyme Inhibitors; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics*; D006801:Humans; D018613:Microscopy, Confocal / Q000379:methods; D020648:Peptide Elongation Factor 1 / Q000235:genetics; D011401:Promoter Regions, Genetic; D014162:Transfection / Q000379:methods; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism
D002087:Butyrates; D004791:Enzyme Inhibitors; D020648:Peptide Elongation Factor 1; D049452:Green Fluorescent Proteins; D014446:Tyrosine 3-Monooxygenase; D004298:Dopamine