title: Treatment with retinoic acid and lens epithelial cell-conditioned mediumin vitrodirected the differentiation of pluripotent stem cells towards corneal endothelial cell-like cells.
abstract: Embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) have extensive self-renewal capacity and the potential to differentiate into all tissue-specific cell lineages, including corneal endothelial cells (CECs). They are a promising prospect for the future of regenerative medicine. The method of derivation of CECs from ESCs and iPSCs, however, remains to be elucidated. In this study, mouse ESCs and iPSCs were induced to differentiate into CECs using CEC embryonic development events as a guide. All-transretinoic acid (RA) treatment during the embryoid body (EB) differentiation step was used to promote neural crest (NC) cell differentiation as first step and was followed by a second induction in CEC- or lens epithelial cell (LEC)-conditioned medium (CM) to ultimately generate CEC-like cells. During the corresponding differentiation stages, NC developmental markers and CEC differentiation markers were detected at the protein level using immunocytochemistry (ICC) and at the mRNA level by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). During the first stage, the data indicated that 4 days of treatment with 1 μM RA starting on day 4 of EB formation favored NC cell differentiation and that plating on gelatin-coated plates led to cell migration out of the EBs. The second-stage differentiation results showed that the CM, particularly the LEC-CM, enhanced the yield of polygonal cells with CEC-specific marker expression shown by ICC and RT-qPCR. This study demonstrates that mouse ESCs and iPSCs were induced and expressed CEC differentiation markers when subjected to a two-step inducement process, suggesting that they are a promising resource for corneal endothelium failure replacement therapy in the future.
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, P.R. China ; Department of Ophthalmology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, P.R. China.
authors.forename: Ping
authors.initials: P
authors.lastname: Chen
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, P.R. China.
authors.forename: Jun-Zhao
authors.initials: JZ
authors.lastname: Chen
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, P.R. China.
authors.forename: Chun-Yi
authors.initials: CY
authors.lastname: Shao
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, P.R. China.
authors.forename: Chuan-Yin
authors.initials: CY
authors.lastname: Li
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, P.R. China.
authors.forename: Yi-Dan
authors.initials: YD
authors.lastname: Zhang
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, P.R. China.
authors.forename: Wen-Juan
authors.initials: WJ
authors.lastname: Lu
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, P.R. China.
authors.forename: Yao
authors.initials: Y
authors.lastname: Fu
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, P.R. China.
authors.forename: Ping
authors.initials: P
authors.lastname: Gu
authors.affiliation: Department of Ophthalmology, Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, P.R. China.
authors.forename: Xianqun
authors.initials: X
authors.lastname: Fan
keywords: conditioned medium; corneal endothelium cell; differentiation; embryonic stem cell; induced pluripotent stem cell; retinoic acid