A novel NIH/3T3 duplex feeder system to engineer corneal epithelial sheets with enhanced cytokeratin 15-positive progenitor populations

Hideyuki Miyashita, Shigeto Shimmura, Kazunari Higa, Satoru Yoshida, Tetsuya Kawakita, Jun Shimazaki, Kazuo Tsubota

研究成果: Article査読

30 被引用数 (Scopus)

抄録

Corneal epithelial cell sheets co-cultivated with feeder cells are used to reconstruct the ocular surface in stem cell-depleted eyes. The present study was conducted to investigate the optimal method of using feeder cells in the interest of preserving progenitor cells in cultivated sheets. We compared the phenotype and secondary colony forming efficiency (CFE) of cell sheets that were engineered using 3T3 feeder cells as a separate layer or as a contact layer. We also devised a novel "duplex feeder" system that consists of two separate layers of feeder cells. After cells reached confluence, cells were cultured at the air-liquid interface to allow full stratification. Stratified sheets were then analyzed using immunohistochemistry and secondary colony formation. Contact feeder cultures and duplex feeder cultures yielded epithelial sheets with small, cuboid basal cells with strong expression of keratin (K)3, K12, and K15. Furthermore, only duplex feeder layers reproduced the basal K15, suprabasal K12 limbal phenotype where epithelial stem cells reside. A similar effect was observed when cornea stroma-derived progenitor cells were used as feeder cells. Duplex feeder sheets also produced significantly more secondary colonies than cells dissociated from single layer sheets, suggesting that the duplex feeder system produces transplantable sheets with a higher yield of progenitor cells.

本文言語English
ページ(範囲)1275-1282
ページ数8
ジャーナルTissue Engineering - Part A.
14
7
DOI
出版ステータスPublished - 2008 7月 1

ASJC Scopus subject areas

  • バイオエンジニアリング
  • 生化学
  • 生体材料
  • 生体医工学

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