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Self-organizing optic-cup morphogenesis in three-
dimensional cultureMototsugu Eiraku, Nozomu Takata, Hiroki Ishibashi,
Masako Kawada, Eriko Sakakura, Satoru Okuda, Kiyotoshi Sekiguchi, Taiji Adachi & Yoshiki Sasai
Nature 472, 51–56, 2011
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
In Vivo Eye Development (Mouse)
E8.5 E9.5 E10.5E11.5
• Neuroectoderm – optic vesicle (optic cup, neural retina, retinal pigment epithelium), optic nerve
• Surface ectoderm - Lens
Molecular mechanisms of optic development (Adler, Canto-Soler 2007)
Molecular mechanisms of optic development (Adler, Canto-Soler 2007)
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)
What does self-organizing morphogenesis of the optic cup look like?
• ES cell aggregate Rx+ GFP+ neuroepithelium – Activin, laminin, entactin, Nodal
• See what happens…
Evagination of optic vesicles
Evagination of optic vesicles
Rx-GFP+ vesicles are Pax+ Sox1-, consistent with retinal marker expression in mouse embryos
Flattening and invagination of optic vesicle
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
Overview of self-organizing morphogenesis
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
RPE, NR, and hinge cells
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
Self-Patterning into neural retina and RPE domains
Self-directed stratification of neural retina tissue
• Excise prolonged culture
Self-directed stratification of neural retina tissue
Self-directed stratification of neural retina tissue
ONL – outer nuclear layer
INL – inner nuclear layer
GCL – ganglion cell layer
PR – photoreceptors
BP – bipolar cells
Self-directed stratification of neural retina tissue
ES neural retina tissue
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.
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?