Telephone: +61 3 9035 6740; Fax: +61 3 8349 4432
Embryonic Stem Cells/Induced Pluripotent Stem Cells
Version of Record online: 22 OCT 2012
Copyright © 2012 AlphaMed Press
Volume 30, Issue 11, pages 2400–2411, November 2012
How to Cite
Denham, M., Bye, C., Leung, J., Conley, B. J., Thompson, L. H. and Dottori, M. (2012), Glycogen Synthase Kinase 3β and Activin/Nodal Inhibition in Human Embryonic Stem Cells Induces a Pre-Neuroepithelial State That Is Required for Specification to a Floor Plate Cell Lineage. STEM CELLS, 30: 2400–2411. doi: 10.1002/stem.1204
Author contributions: M.D.: concept and design, financial support, collection and/or assembly of data, data analysis and interpretation, manuscript writing, and final approval of manuscript; C.B., J.L., B.J.C., and L.H.T.: collection and/or assembly of data and final approval of manuscript; M.D.: concept and design, data analysis and interpretation, financial support, manuscript writing, and final approval of manuscript.
Disclosure of potential conflicts of interest is found at the end of this article.
First published online in STEM CELLSEXPRESS August 21, 2012.
- Issue online: 22 OCT 2012
- Version of Record online: 22 OCT 2012
- Accepted manuscript online: 21 AUG 2012 08:18AM EST
- Manuscript Accepted: 11 JUL 2012
- Manuscript Received: 25 FEB 2012
- University of Melbourne
- National Health and Medical Research Council of Australia. Grant Number: NHMRC:520165
- Australian Research Council Initiative Stem Cells Australia
Additional Supporting Information may be found in the online version of this article.
|sc-12-0191_sm_SupplFigure1.pdf||744K||Supplementary Figure 1: NKX2.1 quantification of Condition A at day 11 by FACS. (A-B) The average proportion of NKX2.1 positive cells in Condition A was 75.18% ± 7.9 SEM (n=3). FACS analysis plots of NKX2.1 were set against the side scatter and gates set according to hESCs expression levels. (C-F) Low and high power image of NKX2.1 staining of Condition A at day 11. Scale bars: C-D = 500μm and E-F =100μm.|
|sc-12-0191_sm_SupplFigure2.tif||107K||Supplementary Figure 2: QPCR analysis of GLI1, GLI2 and GLI3 in Condition A at day 4. (A-C) Analysis of levels were compared to hESCs and the fold change levels for hESC set at 1. GLI1, GLI2 and GLI3 all showed significantly higher level in Condition A at day 4 (P < 0.05). Star indicates significance (P < 0.05).|
|sc-12-0191_sm_SupplFigure3.tif||970K||Supplementary Figure 3: FACS analysis of Condition CEarly (Day 4) for SOX2 and OCT4. (A-B) FACS analysis was performed and gates set according to hESCs for SOX2 and OCT4 expression. The average proportions of SOX2 and OCT4 positive cells for Condition CEarly were 89.17% ± 2.7 SEM and 0.91% ± 0.12 SEM (n=3), respectively.|
|sc-12-0191_sm_SupplFigure4.tif||994K||Supplementary Figure 4: FACS analysis of Conditions D-G for FOXA2. FACS analysis was performed and gates set according to hESCs. FACS analysis of Conditions D, F and G all yielded high percentages of FOXA2 positive cells, 86% (±2.8 SEM), 83.5% (±5.9 SEM) and 85.7% (±5.7 SEM) respectively. Condition E resulted in few (3.3%; ±1.2 SEM) FOXA2 positive cells, significantly lower than the other conditions (P < 0.0001). Sample size is n=3 for each condition.|
|sc-12-0191_sm_SupplFigure5.tif||83K||Supplementary Figure 5: QPCR analysis for mesoderm and endoderm markers in Condition D and E at day11. (A-B) BRACHYURY and SOX17 expression levels were analysed in Conditions D and E and compared to hESC with the fold change levels for hESC set at 1. No significant differences in expression levels were found.|
|sc-12-0191_sm_SupplFigure6.pdf||1859K||Supplementary Figure 6: Forebrain marker expression in Condition E at day 11. (AD) FOXG1 positive cells were detected throughout Condition E at day 11 and none were detected in Condition D. (E-H) OTX2+/PAX6+ cells were detected in Condition E at day 11. A-D Scale bar: 50μm|
Please note: Wiley Blackwell is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.