Browse > Article
http://dx.doi.org/10.14348/molcells.2018.2210

Generation of Isthmic Organizer-Like Cells from Human Embryonic Stem Cells  

Lee, Junwon (Department of Physiology, Yonsei University College of Medicine)
Choi, Sang-Hwi (Department of Physiology, Yonsei University College of Medicine)
Lee, Dongjin R (Department of Physiology, Yonsei University College of Medicine)
Kim, Dae-Sung (Department of Biotechnology, Brain Korea 21 PLUS Project for Biotechnology, College of Life Sciences and Biotechnology, Korea University)
Kim, Dong-Wook (Department of Physiology, Yonsei University College of Medicine)
Publication Information
Molecules and Cells / v.41, no.2, 2018 , pp. 110-118 More about this Journal
Abstract
The objective of this study was to induce the production of isthmic organizer (IsO)-like cells capable of secreting fibroblast growth factor (FGF) 8 and WNT1 from human embryonic stem cells (ESCs). The precise modulation of canonical Wnt signaling was achieved in the presence of the small molecule CHIR99021 ($0.6{\mu}M$) during the neural induction of human ESCs, resulting in the differentiation of these cells into IsO-like cells having a midbrain-hindbrain border (MHB) fate in a manner that recapitulated their developmental course in vivo. Resultant cells showed upregulated expression levels of FGF8 and WNT1. The addition of exogenous FGF8 further increased WNT1 expression by 2.6 fold. Gene ontology following microarray analysis confirmed that IsO-like cells enriched the expression of MHB-related genes by 40 fold compared to control cells. Lysates and conditioned media of IsO-like cells contained functional FGF8 and WNT1 proteins that could induce MHB-related genes in differentiating ESCs. The method for generating functional IsO-like cells described in this study could be used to study human central nervous system development and congenital malformations of the midbrain and hindbrain.
Keywords
FGF; human pluripotent stem cells; isthmic organizer; neural differentiation; Wnt;
Citations & Related Records
연도 인용수 순위
  • Reference
1 Martinez, S., Crossley, P.H., Cobos, I., Rubenstein, J.L., and Martin, G.R. (1999). FGF8 induces formation of an ectopic isthmic organizer and isthmocerebellar development via a repressive effect on Otx2 expression. Development 126, 1189-1200.
2 Irving, C., and Mason, I. (2000). Signalling by FGF8 from the isthmus patterns anterior hindbrain and establishes the anterior limit of Hox gene expression. Development 127, 177-186.
3 Kiecker, C., and Lumsden, A. (2005). Compartments and their boundaries in vertebrate brain development. Nat. Rev. Neurosci. 6, 553-564.   DOI
4 Kiecker, C., and Lumsden, A. (2012). The role of organizers in patterning the nervous system. Annu. Rev. Neurosci. 35, 347-367.   DOI
5 Kim, D.S., Lee, J.S., Leem, J.W., Huh, Y.J., Kim, J.Y., Kim, H.S., Park, I.H., Daley, G.Q., Hwang, D.Y., and Kim, D.W. (2010). Robust enhancement of neural differentiation from human ES and iPS cells regardless of their innate difference in differentiation propensity. Stem Cell Rev. 6, 270-281.   DOI
6 Kirkeby, A., Grealish, S., Wolf, D.A., Nelander, J., Wood, J., Lundblad, M., Lindvall, O., and Parmar, M. (2012). Generation of regionally specified neural progenitors and functional neurons from human embryonic stem cells under defined conditions. Cell Rep. 1, 703-714.   DOI
7 Kiyasova, V., and Gaspar, P. (2011). Development of raphe serotonin neurons from specification to guidance. Eur. J. Neurosci. 34, 1553-1562.   DOI
8 Lancaster, M.A., Renner, M., Martin, C.A., Wenzel, D., Bicknell, L.S., Hurles, M.E., Homfray, T., Penninger, J.M., Jackson, A.P., and Knoblich, J.A. (2013). Cerebral organoids model human brain development and microcephaly. Nature 501, 373-379.   DOI
9 Barkovich, A.J., Millen, K.J., and Dobyns, W.B. (2009). A developmental and genetic classification for midbrain-hindbrain malformations. Brain 132, 3199-3230.   DOI
10 Basson, M.A., and Wingate, R.J. (2013). Congenital hypoplasia of the cerebellum: developmental causes and behavioral consequences. Front. Neuroanat. 7, 29.
11 Canning, C.A., Lee, L., Irving, C., Mason, I., and Jones, C.M. (2007). Sustained interactive Wnt and FGF signaling is required to maintain isthmic identity. Dev. Biol. 305, 276-286.   DOI
12 Benjamini, Y., and Yekutieli, D. (2001). The control of the false discovery rate in multiple testing under dependency. Ann. Stat. 29, 1165-1188.   DOI
13 Blaess, S., and Ang, S. (2015). Genetic control of midbrain dopaminergic neuron development. Wiley Interdisciplinary Reviews -Developmental Biology 4, 113-134.   DOI
14 Broccoli, V., Boncinelli, E., and Wurst, W. (1999). The caudal limit of Otx2 expression positions the isthmic organizer. Nature 401, 164-168.   DOI
15 Nordstrom, U., Jessell, T.M., and Edlund, T. (2002). Progressive induction of caudal neural character by graded Wnt signaling. Nat. Neurosci. 5, 525-532.   DOI
16 McMahon, A.P., Joyner, A.L., Bradley, A., and McMahon, J.A. (1992). The midbrain-hindbrain phenotype of Wnt-1-/Wnt-1- mice results from stepwise deletion of engrailed-expressing cells by 9.5 days postcoitum. Cell 69, 581-595.   DOI
17 Millet, S., Campbell, K., Epstein, D.J., Losos, K., Harris, E., and Joyner, A.L. (1999). A role for Gbx2 in repression of Otx2 and positioning the mid/hindbrain organizer. Nature 401, 161-164.   DOI
18 Naujok, O., Lentes, J., Diekmann, U., Davenport, C., and Lenzen, S. (2014). Cytotoxicity and activation of the Wnt/beta-catenin pathway in mouse embryonic stem cells treated with four GSK3 inhibitors. BMC Res. Notes 7, 273.   DOI
19 Olander, S., Nordstrom, U., Patthey, C., and Edlund, T. (2006). Convergent Wnt and FGF signaling at the gastrula stage induce the formation of the isthmic organizer. Mech. Dev. 123, 166-176.   DOI
20 Partanen, J. (2007). FGF signalling pathways in development of the midbrain and anterior hindbrain. J. Neurochem. 101, 1185-1193.   DOI
21 Pfaffl, M.W. (2001). A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res. 29, e45.   DOI
22 Wurst, W., and Bally-Cuif, L. (2001). Neural plate patterning: upstream and downstream of the isthmic organizer. Nat. Rev. Neurosci. 2, 99-108.   DOI
23 Ciani, L., and Salinas, P.C. (2005). WNTs in the vertebrate nervous system: from patterning to neuronal connectivity. Nat. Rev. Neurosci. 6, 351-362.
24 Rhinn, M., and Brand, M. (2001). The midbrain--hindbrain boundary organizer. Curr. Opin. Neurobiol. 11, 34-42.   DOI
25 Crossley, P.H., Martinez, S., and Martin, G.R. (1996). Midbrain development induced by FGF8 in the chick embryo. Nature 380, 66-68.   DOI
26 de Lau, W., Peng, W.C., Gros, P., and Clevers, H. (2014). The Rspondin/ Lgr5/Rnf43 module: regulator of Wnt signal strength. Genes Dev. 28, 305-316.   DOI
27 Shimamura, K., and Rubenstein, J.L. (1997). Inductive interactions direct early regionalization of the mouse forebrain. Development 124, 2709-2718.
28 Simeone, A. (2000). Positioning the isthmic organizer where Otx2 and Gbx2meet. Trends Genet. 16, 237-240.   DOI
29 Stern, C.D. (2001). Initial patterning of the central nervous system: how many organizers? Nat. Rev. Neurosci. 2, 92-98.   DOI
30 Lupo, G., Bertacchi, M., Carucci, N., Augusti-Tocco, G., Biagioni, S., and Cremisi, F. (2014). From pluripotency to forebrain patterning: an in vitro journey astride embryonic stem cells. Cell. Mol. Life Sci. 71, 2917-2930.   DOI