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First published online May 10, 2007
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2006-0677v1
25/8/1995    most recent
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Submitted on October 25, 2006
Accepted on May 4, 2007

TISSUE-SPECIFIC STEM CELLS

Human Amniotic Epithelial Cells as Novel Feeder Layers for Promoting Ex Vivo Expansion of Limbal Epithelial Progenitor Cells

Ying Ting Chen 1, Wei Li 2, Yasutaka Hayashida 2, Hua He 2, Szu Yu Chen 2, David Y. Tseng 2, Ahmad Kheirkhah 2, Scheffer C. G. Tseng 2*

1 From TissueTech, Inc. and Ocular Surface Center, Miami, Florida, USA; Department of Ophthalmology and Institute of Clinical Medicine, College of Medicine, National Cheng Kung University, Tainan, Taiwan
2 From TissueTech, Inc. and Ocular Surface Center, Miami, Florida, USA

* To whom correspondence should be addressed. E-mail: stseng{at}ocularsurface.com.


  Abstract

Human amniotic epithelial cells (HAECs) are a unique embryonic cell source that potentially can be used as feeder layers for expanding different types of stem cells. In vivo, HAECs uniformly expressed pan-cytokeratins (pan-CK) and heterogeneously expressed vimentin (Vim). The two phenotypes expressing either pan-CK (+)/Vim(+) or pan-CK(+)/Vim(-) were maintained in serum-free media with high calcium. In contrast, all HAECs became pan-CK (+)/Vim(+) in serum-containing media, which also promoted HAEC proliferation for at least 8 passages especially supplemented with EGF and insulin. Mitomycin C-arrested HAEC feeder layers were more effective in promoting clonal growth of human limbal epithelial progenitors than conventional 3T3 murine feeder layers. Cells in HAEC-supported clones were uniformly smaller, sustained more proliferation, and expressed less CK12 and connexin 43, but higher levels of stem cell-associated markers such as p63, Musashi-1 and ABCG2 than those of 3T3-supported clones. Subculturing of clonally expanded limbal progenitors from HAEC feeder layers, but not from 3T3 feeder layers, gave rise to uniformly p63-positive epithelial progenitor cells as well as nestin-positive neuronal-like progenitors. Collectively, these results indicated that HAECs can be used as a human feeder layer equivalent for more effective ex vivo expansion of adult epithelial stem cells from the human limbus.

Key Words. ABCG2, amniotic membrane, amniotic epithelial cells, ex vivo expansion, feeder layers, connexin 43 cytokeratins, K12, limbus, Musashi-1, neuronal progenitors, plasticity, progenitor cells, p63, stem cells




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[Abstract] [Full Text] [PDF]


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