Self-Formation of Optic Cups and Storable Stratified Neural Retina from Human ESCs

Abstract

In this report, we demonstrate that an optic cup structure can form by self-organization in human ESC culture. The human ESC-derived optic cup is much larger than the mouse ESC-derived one, presumably reflecting the species differences. The neural retina in human ESC culture is thick and spontaneously curves in an apically convex manner, which is not seen in mouse ESC culture. In addition, human ESC-derived neural retina grows into multilayered tissue containing both rods and cones, whereas cone differentiation is rare in mouse ESC culture. The accumulation of photoreceptors in human ESC culture can be greatly accelerated by Notch inhibition. In addition, we show that an optimized vitrification method enables en bloc cryopreservation of stratified neural retina of human origin. This storage method at an intermediate step during the time-consuming differentiation process provides a versatile solution for quality control in large-scale preparation of clinical-grade retinal tissues.

Human hepatic organoids for the analysis of human genetic diseases

“…In brief, iPSCs were first dissociated into single cells and induced to form endoderm spheres in response to growth factors and chemicals added to the culture according to a modified protocol (30). The endodermal spheres then formed posterior foregut-like structures when cultured in the presence of a low concentration (1%-2%) of a Matrigel scaffold that supports the formation of 3D structures from stem cells (31,32), and by addition of various concentrations of FGF10, which is known to promote the differentiation of foregut endoderm into hepatic and gallbladder cells during organogenesis (33). The developing foregut structures were transferred on day 9 into a medium with other growth factors to promote their maturation into HOs.…”

Analysis of opo cis-regulatory landscape uncovers Vsx2 requirement in early eye morphogenesis

“…Recent data in mice have shown that the invagination of the lens ectoderm is achieved through an apical constriction mechanism driven by the RhoA-dependent activation of the actin regulator Shroom3 (refs 30,31). In contrast, the inward folding of the neural ectoderm has been described, both in mammals and teleosts, as a tissue-autonomous mechanism depending on integrin function 28,32 . Mutant analysis in medaka (Oryzias latipes) has identified the vertebrate-specific transmembrane protein Opo, encoded by the gene ojoplano (opo), as an essential component of the molecular machinery involved in neural retina morphogenesis 28,33 .…”

Light‐Patterned RNA Interference of 3D‐Cultured Human Embryonic Stem Cells

“…[2,4,6,7] Advances in this area include the controlled differentiation of pluripotent or multipotent stem cells cultured in 3D matrices followed by self-organization into tissues in vitro. [1,[8][9][10][11] However, current methods for organoid formation have insufficient control over the course of the morphogenetic processes and the resulting structures inadequately replicate native tissues. [2,4] Microfluidic and nanotopographic patterning approaches establish some organ-level functions ex vivo, but lack the dimensionality to approximate developing tissues.…”

“…We are currently applying this technology to generate an in vitro optic cup for retinal tissue growth by spatial control of Wnt signaling to pattern the differentiation of hESC to neural retina and retinal pigment epithelium. [11,40] Moreover, the two photon excitation approach can also be combined with in vivo imaging [41] to become a promising means for investigation of developmental processes in vivo.…”