Noninvasive Reporter Gene Imaging of Human Oct4 (Pluripotency) Dynamics During the Differentiation of Embryonic Stem Cells in Living Subjects
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Human pluripotency gene networks (PGNs), controlled in part by Oct4, are central to understanding pluripotent stem cells, but current fluorescent reporter genes (RGs) preclude noninvasive assessment of Oct4 dynamics in living subjects.
To assess Oc4 activity noninvasively, we engineered a mouse embryonic stem cell line which encoded both a pOct4-hrluc (humanized renilla luciferase) reporter and a pUbi-hfluc2-gfp (humanized firefly luciferase 2 fused to green fluorescent protein) reporter.
In cell culture, pOct4-hRLUC activity demonstrated a peak at 48 h (day 2) and significant downregulation by 72 h (day 3) (p=0.0001). Studies in living subjects demonstrated significant downregulation in pOct4-hRLUC activity between 12 and 144 h (p = 0.001) and between 12 and 168 h (p = 0.0003). pOct4-hRLUC signal dynamics after implantation was complex, characterized by transient upregulation after initial downregulation in all experiments (n = 10, p = 0.01). As expected, cell culture differentiation of the engineered mouse embryonic stem cell line demonstrated activation of mesendodermal, mesodermal, endodermal, and ectodermal master regulators of differentiation, indicating potency to form all three germ layers.
We conclude that the Oct4-hrluc RG system enables noninvasive Oct4 imaging in cell culture and in living subjects.
- Noninvasive Reporter Gene Imaging of Human Oct4 (Pluripotency) Dynamics During the Differentiation of Embryonic Stem Cells in Living Subjects
Molecular Imaging and Biology
Volume 16, Issue 6 , pp 865-876
- Cover Date
- Print ISSN
- Online ISSN
- Springer US
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- Embryonic stem cells
- Pluripotent stem cells
- Human Oct4
- Reporter gene imaging
- Gene networks
- Molecular imaging
- In vivo imaging
- In vivo differentiation
- Industry Sectors
- Author Affiliations
- 1. Molecular Imaging Program @Stanford (MIPS), Department of Radiology, Division of Nuclear Medicine, James H. Clark Center, Stanford School of Medicine, Stanford University, 318 Campus Drive, E153, Stanford, CA, 94305-5427, USA
- 2. Department of Nuclear Medicine, Kyungpook National University Medical School, Kyungpook National University Hospital, 50 Samduck 2Ga, Chung Gu, Daegu, 700-712, South Korea
- 7. Institute of Regenerative Medicine and Stem Cells, Department of Obstetrics, Gynecology, and Reproductive Sciences, University of California San Francisco (UCSF) School of Medicine, RMB 902 E, 35 Medical Center Way, San Francisco, CA, 94143, USA
- 3. GE Global Research Center, General Electric, Niskayuna, NY, USA
- 4. Canary Center for Early Detection of Cancer, Palo Alto, CA, USA
- 5. Department of Bioengineering, Stanford School of Medicine, Stanford, CA, USA
- 6. Department of Materials Science and Engineering, Stanford University, Stanford, CA, USA