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J Cell Physiol. 2016 Dec;231(12):2555-62. doi: 10.1002/jcp.25374. Epub 2016 Jul 11.

Generation of Human Lens Epithelial-Like Cells From Patient-Specific Induced Pluripotent Stem Cells.

Li D1,2,3, Qiu X2,4, Yang J2,4, Liu T5, Luo Y2,4, Lu Y2,4.

Author information

1
Research Center, Eye & ENT Hospital of Fudan University, Shanghai, China.
2
Key Laboratory of Myopia, Ministry of Health, Shanghai, China.
3
State Key Laboratory of Molecular Engineering of Polymers, Fudan University, Shanghai, China.
4
Department of Ophthalmology, Eye & ENT Hospital of Fudan University, Shanghai, China.
5
Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy for Sciences, Shanghai, China.

Abstract

Cataractogenesis begins from the dynamic lens epithelial cells (LECs) and adjacent fiber cells. LECs derived from cell lines cannot maintain the crystalline expression as the primary LECs. The current study aimed to efficiently generate large numbers of human LECs from patient-specific induced pluripotent stem cells (iPSCs). Anterior lens capsules were collected from cataract surgery and were used to culture primary hLECs. iPSCs were induced from these primary hLECs by lentiviral transduction of Oct4, Sox2, Klf4, and c-Myc. Then, the generated iPSCs were re-differentiated into hLECs by the 3-step addition of defined factor combinations (Noggin, BMP4/7, bFGF, and EGF) modified from an established method. During the re-differentiation process, colonies of interest were isolated using a glass picking tool and cloning cylinders based on the colony morphology. After two steps of isolation, populations of LEC-like cells (LLCs) were generated and identified by the expression of lens marker genes by qPCR, western blot and immunofluorescence staining. The study introduced a modified protocol to isolate LLCs from iPSCs by defined factors in a short time frame. This technique could be useful for mechanistic studies of lens-related diseases. J. Cell. Physiol. 231: 2555-2562, 2016.

PMID:
26991066
DOI:
10.1002/jcp.25374
[Indexed for MEDLINE]

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