Abstract
Balanced organogenesis requires the orchestration of multiple cellular interactions to create the collective cell behaviours that progressively shape developing tissues. It is currently unclear how individual, localized parts are able to coordinate with each other to develop a whole organ shape. Here we report the dynamic, autonomous formation of the optic cup (retinal primordium) structure from a three-dimensional culture of mouse embryonic stem cell aggregates. Embryonic-stem-cell-derived retinal epithelium spontaneously formed hemispherical epithelial vesicles that became patterned along their proximal–distal axis. Whereas the proximal portion differentiated into mechanically rigid pigment epithelium, the flexible distal portion progressively folded inward to form a shape reminiscent of the embryonic optic cup, exhibited interkinetic nuclear migration and generated stratified neural retinal tissue, as seen in vivo. We demonstrate that optic-cup morphogenesis in this simple cell culture depends on an intrinsic self-organizing program involving stepwise and domain-specific regulation of local epithelial properties.
This is a preview of subscription content, access via your institution
Access options
Subscribe to this journal
Receive 51 print issues and online access
$199.00 per year
only $3.90 per issue
Buy this article
- Purchase on Springer Link
- Instant access to full article PDF
Prices may be subject to local taxes which are calculated during checkout
Similar content being viewed by others
References
Spemann, H. Ueber korrelationen in der entwicklung des auges. Verh. Anat. Ges. 15, 61–79 (1901)
Lewis, W. H. Experimental studies on the development of the eye in Amphibia I. On the origin of the lens in Rana palustrius . Am. J. Anat. 3, 505–536 (1904)
Nakagawa, S., Takada, S., Takada, R. & Takeichi, M. Identification of the laminar-inducing factor: Wnt-signal from the anterior rim induces correct laminar formation of the neural retina in vitro . Dev. Biol. 260, 414–425 (2003)
Martinez-Morales, J. R., Rodrigo, I. & Bovolenta, P. Eye development: a view from the retina pigmented epithelium. Bioessays 26, 766–777 (2004)
Mu, X. & Klein, W. H. A gene regulatory hierarchy for retinal ganglion cell specification and differentiation. Semin. Cell Dev. Biol. 15, 115–123 (2004)
Wilson, S. W. & Houart, C. Early steps in the development of the forebrain. Dev. Cell 6, 167–181 (2004)
Rembold, M., Loosli, F., Adams, R. J. & Wittbrodt, J. Individual cell migration serves as the driving force for optic vesicle evagination. Science 313, 1130–1134 (2006)
Cayouette, M., Poggi, L. & Harris, W. A. Lineage in the vertebrate retina. Trends Neurosci. 29, 563–570 (2006)
Adler, R. & Canto-Soler, M. V. Molecular mechanisms of optic vesicle development: complexities, ambiguities and controversies. Dev. Biol. 305, 1–13 (2007)
Martinez-Morales, J. R. & Wittbrodt, J. Shaping the vertebrate eye. Curr. Opin. Genet. Dev. 19, 511–517 (2009)
Picker, A. et al. Dynamic coupling of pattern formation and morphogenesis in the developing vertebrate retina. PLoS Biol. 7, e1000214 (2009)
Fuhrmann, S. Eye morphogenesis and patterning of the optic vesicle. Curr. Top. Dev. Biol. 93, 61–84 (2010)
Livesey, F. J. & Cepko, C. L. Vertebrate neural cell-fate determination: lessons from the retina. Nature Rev. Neurosci. 2, 109–118 (2001)
Bailey, T. J. et al. Regulation of vertebrate eye development by Rx genes. Int. J. Dev. Biol. 48, 761–770 (2004)
Lopashov, G. Developmental Mechanism of Vertebrate Eye Rudiments (Pergammon, 1963)
Hyer, J., Mima, T. & Mikawa, T. FGF1 patterns the optic vesicle by directing the placement of the neural retina domain. Development 125, 869–877 (1998)
Hyer, J., Kuhlman, J., Afif, E. & Mikawa, T. Optic cup morphogenesis requires pre-lens ectoderm but not lens differentiation. Dev. Biol. 259, 351–363 (2003)
Smith, A. N., Miller, L. A., Radice, G., Ashery-Padan, R. & Lang, R. A. Stage-dependent modes of Pax6-Sox2 epistasis regulate lens development and eye morphogenesis. Development 136, 2977–2985 (2009)
Wataya, T. et al. Minimization of exogenous signals in ES cell culture induces rostral hypothalamic differentiation. Proc. Natl Acad. Sci. USA 105, 11796–11801 (2008)
Eiraku, M. et al. Self-organized formation of polarized cortical tissues from ESCs and its active manipulation by extrinsic signals. Cell Stem Cell 3, 519–532 (2008)
Au, E. & Fishell, G. Cortex shatters the glass ceiling. Cell Stem Cell 3, 472–474 (2008)
Ikeda, H. et al. Generation of Rx+/Pax6+ neural retinal precursors from embryonic stem cells. Proc. Natl Acad. Sci. USA 102, 11331–11336 (2005)
Fujiwara, H. et al. Regulation of mesodermal differentiation of mouse embryonic stem cells by basement membranes. J. Biol. Chem. 282, 29701–29711 (2007)
Lagutin, O. et al. Six3 promotes the formation of ectopic optic vesicle-like structures in mouse embryos. Dev. Dyn. 221, 342–349 (2001)
Tang, K. et al. COUP-TFs regulate eye development by controlling factors essential for optic vesicle morphogenesis. Development 137, 725–734 (2010)
Hilfer, S. R. & Yang, J.-J. W. Accumulation of CPC-precipitable material at apical cell surfaces during formation of the optic cup. Anat. Rec. 197, 423–433 (1980)
Sawyer, J. M. et al. Apical constriction: a cell shape change that can drive morphogenesis. Dev. Biol. 341, 5–19 (2010)
Kinoshita, N., Sasai, N., Misaki, K. & Yonemura, S. Apical accumulation of Rho in the neural plate is important for neural plate cell shape change and neural tube formation. Mol. Biol. Cell 19, 2289–2299 (2008)
Agius, E. et al. Converse control of oligodendrocyte and astrocyte lineage development by Sonic hedgehog in the chick spinal cord. Dev. Biol. 270, 308–321 (2004)
Amano, M. et al. Phosphorylation and activation of myosin by Rho-associated kinase (Rho-kinase). J. Biol. Chem. 271, 20246–20249 (1996)
Schoenwolf, G. C. & Smith, J. L. Mechanisms of neurulation: traditional viewpoint and recent advances. Development 109, 243–270 (1990)
Haigo, S. L., Hildebrand, J. D., Harland, R. M. & Wallingford, J. B. Shroom induces apical constriction and is required for hingepoint formation during neural tube closure. Curr. Biol. 13, 2125–2137 (2003)
Hildebrand, J. D. Shroom regulates epithelial cell shape via the apical positioning of an actomyosin network. J. Cell Sci. 118, 5191–5203 (2005)
Nishimura, T. & Takeichi, M. Shroom3-mediated recruitment of Rho kinases to the apical cell junctions regulates epithelial and neuroepithelial planar remodeling. Development 135, 1493–1502 (2008)
Rico, F. et al. Probing mechanical properties of living cells by atomic force microscopy with blunted pyramidal cantilever tips. Phys. Rev. E 72, 021914 (2005)
Krieg, M. et al. Tensile forces govern germ-layer organization in zebrafish. Nature Cell Biol. 10, 429–436 (2008)
Mascaro, A. L., Sacconi, L. & Pavone, F. S. Multi-photon nanosurgery in live brain. Front. Neuroenergetics 2, (2010)
Fuhrmann, S. Wnt signaling in eye organogenesis. Organogenesis 4, 60–67 (2008)
Westenskow, P., Piccolo, S. & Fuhrmann, S. β-catenin controls differentiation of the retinal pigment epithelium in the mouse optic cup by regulating Mitf and Otx2 expression. Development 136, 2505–2510 (2009)
Liu, W., Lagutin, O., Swindell, E., Jamrich, M. & Oliver, G. Neuroretina specification in mouse embryos requires Six3-mediated suppression of Wnt8b in the anterior neural plate. J. Clin. Invest. 120, 3568–3577 (2010)
Fujimura, N., Taketo, M. M., Mori, M., Korinek, V. & Kozmik, Z. Spatial and temporal regulation of Wnt/β-catenin signaling is essential for development of the retinal pigment epithelium. Dev. Biol. 334, 31–45 (2009)
Yang, X. J. Roles of cell-extrinsic growth factors in vertebrate eye pattern formation and retinogenesis. Semin. Cell Dev. Biol. 15, 91–103 (2004)
Diep, D. B., Hoen, N., Backman, M., Machon, O. & Krauss, S. Characterisation of the Wnt antagonists and their response to conditionally activated Wnt signalling in the developing mouse forebrain. Brain Res. Dev. Brain Res. 153, 261–270 (2004)
Norden, C., Young, S., Link, B. A. & Harris, W. A. Actomyosin is the main driver of interkinetic nuclear migration in the retina. Cell 138, 1195–1208 (2009)
Pinzón-Duarte, G., Kohler, K., Arango-Gonzalez, B. & Guenther, E. Cell differentiation, synaptogenesis, and influence of the retinal pigment epithelium in a rat neonatal organotypic retina culture. Vision Res. 40, 3455–3465 (2000)
Matsuda, T. & Cepko, C. L. Controlled expression of transgenes introduced by in vivo electroporation. Proc. Natl Acad. Sci. USA 104, 1027–1032 (2007)
Stella, S. L., Jr, Li, S., Sabatini, A., Vila, A. & Brecha, N. C. Comparison of the ontogeny of the vesicular glutamate transporter 3 (VGLUT3) with VGLUT1 and VGLUT2 in the rat retina. Brain Res. 1215, 20–29 (2008)
Fuhrmann, S., Levine, E. M. & Reh, T. A. Extraocular mesenchyme patterns the optic vesicle during early eye development in the embryonic chick. Development 127, 4599–4609 (2000)
Cavodeassi, F. et al. Early stages of zebrafish eye formation require the coordinated activity of Wnt11, Fz5, and the Wnt/β-catenin pathway. Neuron 47, 43–56 (2005)
Seiler, M. J. et al. Visual restoration and transplant connectivity in degenerate rats implanted with retinal progenitor sheets. Eur. J. Neurosci. 31, 508–520 (2010)
Honda, H., Tanemura, M. & Nagai, T. A three-dimensional vertex dynamics cell model of space-filling polyhedra simulating cell behaviour in a cell aggregate. J. Theor. Biol. 226, 439–453 (2004)
Nagai, T. & Honda, H. Computer simulation of wound closure in epithelial tissues: cell-basal-lamina adhesion. Phys. Rev. E 80, 061903 (2009)
Inoue, Y. & Adachi, T. Coarse-grained Brownian ratchet model of membrane protrusion on cellular scale. Biomech. Model. Mechanobiol. 10.1007/s10237-010-0250-6 (19 August 2010)
Acknowledgements
We are grateful to S. Nakanishi, S. Yonemura, R. Ladher, K. Muguruma, H. Inomata and M. Ohgushi for comments and to members of the Sasai laboratory for discussion. We also thank Olympus, particularly Y. Saito, M. Suzuki and Y. Imai, for their help and discussion regarding the design, assembly and optimized utility of the incubator-combined multi-photon and confocal optic systems, and T. Sugitate and N. Saito at JPK Instruments for technical advice on the AFM assay. This work was supported by grants-in-aid from MEXT (Y.S., M.E., T.A.), the Knowledge Cluster Initiative at Kobe and the S-Innovation Project (Y.S., K.S.), and the Leading Project for Realization of Regenerative Medicine (Y.S).
Author information
Authors and Affiliations
Contributions
M.E. and Y.S. designed the project and wrote the manuscript. M.E., N.T., M.K. and E.S performed experiments. K.S. provided critical technical information on matrix experiments. H.I., S.O. and T.A. performed computer simulation by discussing details with M.E. and Y.S.
Corresponding authors
Ethics declarations
Competing interests
The authors declare no competing financial interests.
Supplementary information
Supplementary Information
This file contains Supplementary Figures 1-6 with legends and Legends for Supplementary Movies 1-8. (PDF 1055 kb)
Supplementary Movie 1
This movie shows evagination of Rx+ vesicles from an SFEBq-cultured ESC aggregate. (MOV 2195 kb)
Supplementary Movie 2
This movie shows eye-cup morphogenesis of ESC-derived retinal tissues in 3D live imaging (MOV 7884 kb)
Supplementary Movie 3
This movie shows inhibition of invagination by aphidicolin treatment. (MOV 774 kb)
Supplementary Movie 4
This movie shows tissue dynamics responses to 3D-pinpointed cell ablation by multi photon laser. (MOV 14187 kb)
Supplementary Movie 5
This movie shows invagination in the ESC-derived retinal epithelium isolated and cocultured with Wnt-expressing cells. (MOV 2382 kb)
Supplementary Movie 6
This movie shows interkinetic nuclear migration in ESC-derived NR tissues. (MOV 6564 kb)
Supplementary Movie 7
This movie shows eversion of the RPE-hinge portion of the Phase-4 cup occurring after excision at the proximal hinge. (MOV 509 kb)
Supplementary Movie 8
This movie shows computer-simulated animation of the invagination process. (MOV 2129 kb)
Rights and permissions
About this article
Cite this article
Eiraku, M., Takata, N., Ishibashi, H. et al. Self-organizing optic-cup morphogenesis in three-dimensional culture. Nature 472, 51–56 (2011). https://doi.org/10.1038/nature09941
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1038/nature09941
This article is cited by
-
Generation of sarconoids from angiosarcoma patients as a systematic-based rational approach to treatment
Journal of Hematology & Oncology (2024)
-
Transplanted human photoreceptors transfer cytoplasmic material but not to the recipient mouse retina
Stem Cell Research & Therapy (2024)
-
The Case Against Organoid Consciousness
Neuroethics (2024)
-
Retinal Organoids: A Next-Generation Platform for High-Throughput Drug Discovery
Stem Cell Reviews and Reports (2024)
-
Innovative explorations: unveiling the potential of organoids for investigating environmental pollutant exposure
Environmental Science and Pollution Research (2024)
Comments
By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.