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Generation of pure lymphatic endothelial cells from human pluripotent stem cells and their therapeutic effects on wound repair.

ABSTRACT: Human pluripotent stem cells (hPSCs) have emerged as an important source for cell therapy. However, to date, no studies demonstrated generation of purified hPSC-derived lymphatic endothelial cells (LECs) and tested their therapeutic potential in disease models. Here we sought to differentiate hPSCs into the LEC lineage, purify them with LEC markers, and evaluate their therapeutic effects. We found that an OP9-assisted culture system reinforced by addition of VEGF-A, VEGF-C, and EGF most efficiently generated LECs, which were then isolated via FACS-sorting with LYVE-1 and PODOPLANIN. These hPSC-derived LYVE-1(+)PODOPLANIN(+)cells showed a pure committed LEC phenotype, formed new lymphatic vessels, and expressed lymphangiogenic factors at high levels. These hPSC-derived LECs enhanced wound healing through lymphangiogenesis and lymphvasculogenesis. Here we report, for the first time, that LECs can be selectively isolated from differentiating hPSCs, and that these cells are potent for lymphatic vessel formation in vivo and wound healing. This system and the purified hPSC-derived LECs can serve as a new platform for studying LEC development as well as for cell therapy.

SUBMITTER: Lee SJ

PROVIDER: S-EPMC4464258 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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Generation of pure lymphatic endothelial cells from human pluripotent stem cells and their therapeutic effects on wound repair.

Lee Shin-Jeong SJ Park Changwon C Lee Ji Yoon JY Kim Sangsung S Kwon Pil Jae PJ Kim Woansang W Jeon Yong Heui YH Lee Eugine E Yoon Young-sup YS

Scientific reports 20150612

Human pluripotent stem cells (hPSCs) have emerged as an important source for cell therapy. However, to date, no studies demonstrated generation of purified hPSC-derived lymphatic endothelial cells (LECs) and tested their therapeutic potential in disease models. Here we sought to differentiate hPSCs into the LEC lineage, purify them with LEC markers, and evaluate their therapeutic effects. We found that an OP9-assisted culture system reinforced by addition of VEGF-A, VEGF-C, and EGF most efficien ...[more]

PMID: 26066093

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Project description:Corneal endothelial cells (CECs) do not proliferate or recover after illness or injury, resulting in decreased cell density and loss of pump/barrier function. Considering the shortage of donor cornea, it is vital to establish robust methods to generate CECs from induced pluripotent stem cells (iPSCs). We investigated the efficacy and safety of transplantation of iPSC-derived CECs into a corneal endothelial dysfunction (CED) rabbit model. iPSCs were generated from human fibroblasts. We characterized iPSCs by demonstrating the gene expression of the PSC markers OCT4, SOX2, TRA-1-60, and NANOG, teratoma formation, and differentiation into three germ layers. Differentiation of iPSCs into CECs was induced via neural crest cell (NCC) induction. CEC markers were detected using immunofluorescence and gene expression was analyzed using quantitative real-time PCR (qRT-PCR). After culturing iPSC-derived NCCs, we found the expression of zona occludens-1 (ZO-1) and Na+/K+ ATPase and a hexagonal morphology. ATP1A1, COL8A1, and AQP1 mRNA expression was higher in iPSC-derived CECs than in iPSCs and NCCs. We performed an injection of iPSC-derived CECs into the anterior chamber of a CED rabbit model and found improved levels of corneal transparency. We also found increased numbers of ZO-1- and ATP1A1-positive cells in rabbit corneas in the iPSC-derived CEC transplantation group. Usage of the coating material vitronectin (VTN) and fasudil resulted in good levels of CEC marker expression, demonstrated with Western blotting and immunocytochemistry. Combination of the VTN coating material and fasudil, instead of FNC mixture and Y27632, afforded the best results in terms of CEC differentiation's in vitro and in vivo efficacy. Successful transplantation of CEC-like cells into a CED animal model confirms the therapeutic efficacy of these cells, demonstrated by the restoration of corneal clarity. Our results suggest that iPSC-derived CECs can be a promising cellular resource for the treatment of CED.

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Generation of pure lymphatic endothelial cells from human pluripotent stem cells and their therapeutic effects on wound repair.

Publications

Generation of pure lymphatic endothelial cells from human pluripotent stem cells and their therapeutic effects on wound repair.

Similar Datasets

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