Differentiation and Transplantation of Mouse ES and iPS Derived Retina-like Sheets in Retinal Degeneration Mouse (rd1)

Abstract

Purpose: : To characterize the 3-dimentional differentiation of the mES and miPS derived retinal cell sheets derived from Rx-GFP mES and Nrl-GFP miPS, and to evaluate the viability and maturation of retinal sheets transplanted into the wild type and degenerative mouse (rd1) retina.

Methods: : Rx-GFP mES and Nrl-GFP miPS were differentiated in three-dimensional culture into retina tissue by a modified method by Eiraku et al (Nature, April 2011). The retinal progenitors were evaluated immunohistochemically along differentiation. The embryonic retina (E17) and postnatal retina (p0, p4) and mES or miPS derived retina were also transplanted into the subretinal space of wild type (Bl6) and RD1 retina. Their survival and maturation was immunohistologically evaluated.

Results: : The three-dimentional culture of mES or miPS cells produced optic vesicle-like structures strongly expressing Rx-GFP (ES line) co-labeled with Pax6 on d.d.9. These vesicles formed well organized retinal neuroepithelial like layers positive for Crx and Recoverin on d.d.16, followed by rhodopsin expression around d.d. 25. These retina-like structure partially presented outer limiting membrane (OLM) like structure labeled with ZO-1 and GS.E17 and p0 retina tended to form rosettes strongly expressing rhodopsin whereas p4 retina often lost structure but ingetrated into the ONL of wild type host retina. Retina-like sheets of <d.d.20 derived from mES and miPS also tended to form rosette, and seemed to mature in subretinal space.

Conclusions: : Retina-like sheets derived from mES and Nrl-GFP miPS could serve as graft in retina sheets transplantation.

Keywords: transplantation • differentiation • retina