1. Self-organizing optic-cup
morphogenesis in three-
dimensional culture
Mototsugu Eiraku, Nozomu Takata, Hiroki Ishibashi, Masako Kawada,
Eriko Sakakura, Satoru Okuda, Kiyotoshi Sekiguchi, Taiji Adachi &
Yoshiki Sasai
Nature 472, 51–56, 2011

2. Intro
• In vivo eye development in mouse
• Self-organizing optic cup morphogenesis
• Self-patterning of specific neural retinal
domains
• Self-directed stratification of neural retina
tissue
• Conclusions
• Discussion

3. In Vivo Eye Development (Mouse)
E8.5 E9.5 E10.5 E11.5
• Neuroectoderm – optic vesicle (optic cup,
neural retina, retinal pigment epithelium),
optic nerve
• Surface ectoderm - Lens

4. Molecular mechanisms of optic development
(Adler, Canto-Soler 2007)

5. Molecular mechanisms of optic development
(Adler, Canto-Soler 2007)

6. Optic-cup self-formation in 3D ES cell
culture
• What does self-organizing morphogenesis of
the optic cup look like?
• What factors may or may not play a role? (i.e.
lens/surface ectoderm, neuroepithelium)
• What are the characteristics of the
differentiated cells?
• In vitro vs. in vivo (self-assembly, patterning,
morphogenesis)

7. What does self-organizing morphogenesis
of the optic cup look like?
• ES cell aggregate Rx+
GFP+
neuroepithelium
– Activin, laminin, entactin, Nodal
• See what happens…

8. Evagination of optic vesicles

9. Evagination of optic vesicles
Rx-GFP+
vesicles are Pax+
Sox1-
, consistent with retinal
marker expression in mouse embryos

10. Flattening and invagination of optic vesicle

11. Flattening and invagination of optic vesicle
• Differentiation of the RPE and NR
• RPE –Pax6, Mitf expression,
observable pigment, ↓ Rx
expression
• NR – Rx, Chx10 expression
• Apical (aPKC) – Basal (Laminin)
polarity, same as in vivo

12. Overview of self-organizing morphogenesis

13. RPE, NR, and hinge cells
• Invagination occurs as cells
proliferate, differentiate
• Distinct morphologies, as in vivo
• Distinct actomyosin activation, RPE
vs. NR
• Distinct gene-expression

14. RPE, NR, and hinge cells

15. Self-Patterning into neural retina and RPE domains
• Vesicles with neighboring
neuroectodermal
epithelium invaginated and
formed optic cup
• (-)NE vesicles – no RPE
differentiation, NR develops
• Wnt3 expressing cells
significantly rescue RPE
differentiation
• RPE patterning is dependent
on tissue interactions
• NR develops autonomously

16. Self-Patterning into neural retina and RPE domains

17. Self-directed stratification of neural retina tissue
• Excise  prolonged culture

18. Self-directed stratification of neural retina tissue

19. Self-directed stratification of neural retina tissue
ONL – outer nuclear
layer
INL – inner nuclear layer
GCL – ganglion cell layer
PR – photoreceptors
BP – bipolar cells

20. Self-directed stratification of neural retina tissue
ES neural retina tissue

21. Conclusions
• Development of the optic cup does not depend on forces (chemical
or physical) of external structures (lens, surface ectoderm)
• Self-formed retinal epithelium domains have distinct
morphological, mechanical, and gene-expressing properties
• RPE differentiation is dependent upon induction factors from
neuroepithelium, NR differentiation is autonomous
• ES neural retina tissue self-forms in a manner consistent with the
spatiotemporal order seen in vivo
• Assembly, patterning, and morphogenesis of the retinal optic cup is
self-directed and self-organized.

22. Discussion
• What does this mean? How can it be useful?
• In vitro vs. in vivo, can organoids fully
resemble functional organs?
• Future research? Experimental approach?