Homo sapiens
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA233965 | ENA |
REPOSITORIES: ENA
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Project description:Limbal epithelial stem cell (LESC) deficiency represents a significant clinical problem especially in bilateral cases. Induced pluripotent stem cells (iPSC) may be a promising source of LESC, allowing standardized and continual propagation and banking. The objective of this study was to generate iPSC from human limbal epithelial cultures and differentiate them back into limbal epithelial cells using substrata mimicking the natural LESC niche. Using Yamanaka’s episomal vectors limbal-derived iPSC were reprogrammed from LESC cultured from donor corneoscleral rims and from human skin fibroblasts. A clone from limbal-derived iPSC expressed stemness markers, had a diploid karyotype, and produced teratomas in nude mice representing three germ layers. Compared to parental LESC, this clone had fewer specific gene methylation changes revealed using the Illumina Infinium Methylation 450k Beadchips than compared to skin fibroblasts. The expression of putative LESC markers was examined by quantitative RT-PCR and immunostaining in limbal-derived and fibroblast-derived iPSC cultured on denuded human amniotic membrane or denuded cornea. Limbal-derived iPSC had markedly stronger expression of PAX6, ABCG2, Np63, keratins 14, 15, 17, and N-cadherin than fibroblast-derived iPSC. On denuded corneas, limbal-derived iPSC showed the expression of differentiated corneal keratins 3 and 12. The data suggest that iPSC differentiation to a desired lineage may be facilitated by their generation from the same tissue. This may be related to preservation of parental tissue epigenetic methylation signatures in iPSC and use of biological substrata similar to the natural niche of parental cells. The data pave the way for generating transplantable LESC from limbal-derived iPSC. Bisulphite converted DNA from the 12 samples were hybridised to the Illumina Infinium 450k Human Methylation Beadchip
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Project description:Transplantation of ex vivo expanded limbal stem cells (LSC) is the main treatment for limbal stem cell deficiency though the clinical problem of donor tissues shortage. Recently, as the development of tissue engineering, embryonic stem cells (ESC) derived corneal epithelial-like cells (ESC-CEC) has become a new direction to this issue.Our group successfully induced ESC into corneal epithelial-like cells, and in the present study we explored various aspects of physiological properties of ESC-CEC. The experiment included three samples: hES, the human embryonic stem cell line H1, RA_SB, the corneal epithelial-like cells derived from hES by differentiation with RA and SB, epithelial_cell, the primary human limbal stem cells from cadaver eyes. hES, the human embryonic stem cell line H1, RA_SB, the corneal epithelial-like cells derived from hES by differentiation with RA and SB, epithelial_cell, the primary human limbal stem cells from cadaver eyes.
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