title: Differentiation of induced pluripotent stem cells of swine into rod photoreceptors and their integration into the retina.
abstract: Absence of a regenerative pathway for damaged retina following injury or disease has led to experiments using stem cell transplantation for retinal repair, and encouraging results have been obtained in rodents. The swine eye is a closer anatomical and physiological match to the human eye, but embryonic stem cells have not been isolated from pig, and photoreceptor differentiation has not been demonstrated with induced pluripotent stem cells (iPSCs) of swine. Here, we subjected iPSCs of swine to a rod photoreceptor differentiation protocol consisting of floating culture as embryoid bodies followed by differentiation in adherent culture. Real-time PCR and immunostaining of differentiated cells demonstrated loss of expression of the pluripotent genes POU5F1, NANOG, and SOX2 and induction of rod photoreceptor genes RCVRN, NRL, RHO, and ROM1. While these differentiated cells displayed neuronal morphology, culturing on a Matrigel substratum triggered a further morphological change resulting in concentration of rhodopsin (RHO) and rod outer segment-specific membrane protein 1 in outer segment-like projections resembling those on primary cultures of rod photoreceptors. The differentiated cells were transplanted into the subretinal space of pigs treated with iodoacetic acid to eliminate rod photoreceptors. Three weeks after transplantation, engrafted RHO+ cells were evident in the outer nuclear layer where photoreceptors normally reside. A portion of these transplanted cells had generated projections resembling outer segments. These results demonstrate that iPSCs of swine can differentiate into photoreceptors in culture, and these cells can integrate into the damaged swine neural retina, thus, laying a foundation for future studies using the pig as a model for retinal stem cell transplantation.
authors.affiliation: Department of Ophthalmology, The second Xiangya Hospital, Central South University, Changsha, Hunan, People's Republic of China.
authors.forename: Liang
authors.initials: L
authors.lastname: Zhou
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.forename: Yongqing
authors.initials: Y
authors.lastname: Liu
authors.forename: Juan
authors.initials: J
authors.lastname: Fernandez de Castro
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Ezashi
authors.forename: Bhanu Prakash V L
authors.initials: BP
authors.lastname: Telugu
authors.forename: R Michael
authors.initials: RM
authors.lastname: Roberts
authors.forename: Henry J
authors.initials: HJ
authors.lastname: Kaplan
authors.forename: Douglas C
authors.initials: DC
authors.lastname: Dean
mesh_terms: D000818:Animals; D000943:Antigens, Differentiation / Q000378:metabolism; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002478:Cells, Cultured; D018920:Coculture Techniques; D003094:Collagen / Q000378:metabolism; D004338:Drug Combinations; D058732:Embryoid Bodies / Q000166:cytology / Q000378:metabolism; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism / Q000637:transplantation; D007797:Laminin / Q000378:metabolism; D011509:Proteoglycans / Q000378:metabolism; D051603:Recoverin / Q000378:metabolism; D012160:Retina / Q000187:drug effects / Q000473:pathology*; D017948:Retinal Rod Photoreceptor Cells / Q000166:cytology* / Q000378:metabolism; D012177:Retinol-Binding Proteins / Q000378:metabolism; D012243:Rhodopsin / Q000378:metabolism; D013552:Swine; D014404:Tubulin / Q000378:metabolism
chemical_list: D000943:Antigens, Differentiation; D004338:Drug Combinations; D007797:Laminin; D011509:Proteoglycans; D012177:Retinol-Binding Proteins; D014404:Tubulin; C059485:matrigel; D051603:Recoverin; D003094:Collagen; D012243:Rhodopsin