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Embryonic Stem Cells/Induced Pluripotent Stem Cells
Article first published online: 24 JUL 2012
Copyright © 2012 AlphaMed Press
Volume 30, Issue 8, pages 1655–1663, August 2012
How to Cite
Xi, J., Liu, Y., Liu, H., Chen, H., Emborg, M. E. and Zhang, S.-C. (2012), Specification of Midbrain Dopamine Neurons from Primate Pluripotent Stem Cells. STEM CELLS, 30: 1655–1663. doi: 10.1002/stem.1152
Author contributions: J.X.: conception and design, collection and/or assembly of data, data analysis and interpretation, and manuscript writing; Y.L., H.L., and H.C.: collection and/or assembly of data and data analysis and interpretation; M.E.E.: collection of non-human primate data; S.-C.Z.: conception and design, financial support, collection and/or assembly of data, data analysis and interpretation, 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 June 13, 2012.
- Issue published online: 24 JUL 2012
- Article first published online: 24 JUL 2012
- Accepted manuscript online: 13 JUN 2012 02:55PM EST
- Manuscript Accepted: 8 MAY 2012
- Manuscript Received: 20 FEB 2012
- NIH-NINDS. Grant Number: NS045926
- Parkinson's Disease Foundation, Center for Stem Cells and Regenerative Medicine
- University of Wisconsin Madison
- NICHD. Grant Number: P30 HD03352
- National Natural Science Foundation of China. Grant Number: 30901615
- NIH-NCRR. Grant Number: P51 RR000167
- Wisconsin National Primate Research Center, University of Wisconsin-Madison
Additional Supporting Information may be found in the online version of this article.
|SC_12-0172_sm_supplFigure1.pdf||345K||Supplementary Fig. 1. Floor plate progenitors induced by SHH or CHIR. (A) The floor plate progenitors exhibited epithelial morphology in each individual colony on day 8. (B) Immunostaining for FoxA2, Otx2, and En1 at day 18 after treatment with 500ng/ml SHH. (C) Immunostaining for En1 (green), HoxA2 (red) and HoxB1 (blue) at day 18 after treatment with 2 or 3 uM of CHIR. (D) Immunostaining for En1 (green), OTX2 (red) and FOXA2 (blue) at day 18 after treatment with 1 uM of CHIR. (E) Immunostaining for TH (green) and 5HT (red) at day 36 after treatment with 0.4 or 1 uM of CHIR and subsequent treatment with FGF8. (F) Quantification of the data presented in E. (G) Immunostaining for TH (green) and GABA (red) at day 36. (H) Immunostaining for TH (green) and NESTIN (red) at day 36. Bar =50um|
|SC_12-0172_sm_supplFigure2.pdf||491K||Supplementary Fig. 2. Midbrain floor plate progenitors induced by SHH and CHIR. (A) Separate immunofluorescent channels correspond to merged images in Fig. 3d. (B) Immunostaining of midbrain floor plate progenitors for Nestin (green), Lmx1a (red) and FoxA2 (blue) at day 18. (C) Immunostaining for Ngn2 (green) and Mash1 (red) at day 22 after withdrawal of SHH and CHIR from day 13. Bar =50um|
|SC_12-0172_sm_supplFigure3.pdf||311K||Supplementary Fig. 3. Differentiation of midbrain floor plate progenitors and DA neurons from human iPSCs (line IMR90-4). (A) Immunostaining and quantification of markers for midbrain floor plate progenitors at day 18 (Corin, FoxA2, En1, Lmx1a and Otx2). (B) Co-staining of TH (green) with other midbrain DA neuronal markers FoxA2, En1, Lmx1a and Nurr1 (red) as well as quantification of the cell populations. Bar =50um|
|SC_12-0172_sm_supplFigure4.pdf||230K||Supplementary Fig. 4. Electrophysiological properties of DA neurons. (A) Electrophysiological characteristics of neurons that were differentiated for 10 weeks in vitro. (B) Inward Na+ and outward K+ currents were triggered upon - 50mV to + 50 mV voltage steps. The initial currents were enlarged in the lowest panel. (C) Action potentials were induced from − 60 pA to + 60 pA injected current steps. The lower panel shows the response upon + 60 pA current injection. (D) Both excitatory (inward current) and inhibitory (outward current) spontaneous postsynaptic currents are present which are inhibited by CNQX and bicuculine, respectively. The asterisks indicate individual excitatory (sEPSCs) and inhibitory (sIPSCs) events.|
|SC_12-0172_sm_supplFigure5.pdf||800K||Supplementary Fig. 5. Differentiation of midbrain floor plate progenitors and DA neurons from rhesus monkey iPSCs. (A) Experimental paradigm showing differentiation of midbrain DA neurons from rhesus monkey iPSCs. (B) Immunostaining of markers for midbrain floor plate progenitors at day 18 (Corin, FoxA2, En1, Lmx1a and Otx2). (C) Co-staining of TH (green) with other midbrain DA neuronal markers FoxA2, En1, Lmx1a and Nurr1 (red). (D) Quantification of midbrain floor plate progenitor populations among total cells. (E) Quantification of TH+ DA neuron populations and midbrain marker expression in TH+ cells. Bar =50um|
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