Induction of Functional 3D Ciliary Epithelium-Like Structure From Mouse Induced Pluripotent Stem Cells

Invest Ophthalmol Vis Sci. 2016 Jan 1;57(1):153-61. doi: 10.1167/iovs.15-17610.


Purpose: To generate ciliary epithelium (CE) from mouse induced pluripotent stem (iPS) cells.

Methods: Recently, a protocol for self-organizing optic cup morphogenesis in three-dimensional culture was reported, and it was suggested that ocular tissue derived from neural ectoderm could be differentiated. We demonstrated that a CE-like double-layered structure could be induced in simple culture by using a modified Eiraku differentiation protocol.

Results: Differentiation of a CE-like double-layered structure could be promoted by glycogen synthase kinase 3β (GSK-3β) inhibitor. Connexin43 and aquaporin1 were expressed in both thin layers, and induced CE-like cells expressed ciliary marker genes, such as cyclinD2, zic1, tgfb2, aldh1a3, wfdc1, otx1, BMP4, and BMP7. Increases in cytoplasmic and nuclear β-catenin in aggregates of the CE-like double-layered structure were confirmed by Western blot analysis. In addition, tankyrase inhibitor prevented the induction of the CE-like double-layered structure by GSK-3β inhibitor. Dye movement from pigmented cells to nonpigmented cells in the mouse iPS cell-derived CE-like structure was observed in a fluid movement experiment, consistent with the physiological function of CE in vivo.

Conclusions: We could differentiate CE from mouse iPS cells in the present study. In the future, we hope that this CE-like complex will become useful as a graft for transplantation therapy in pathologic ocular hypotension due to CE dysfunction, and as a screening tool for the development of drugs for diseases associated with CE function.

MeSH terms

  • Animals
  • Blotting, Western
  • Cell Differentiation
  • Cell Proliferation
  • Cells, Cultured
  • Ciliary Body / cytology*
  • Disease Models, Animal
  • Epithelial Cells / cytology*
  • Imaging, Three-Dimensional*
  • Immunohistochemistry
  • Induced Pluripotent Stem Cells / cytology*
  • Mice
  • Mice, Inbred C57BL