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Tetsuya Kawakita, Kazunari Higa, Shigeto Shimmura, Machiko Tomita, Kazuo Tsubota, Jun Shimazaki; Fate of Corneal Epithelial Cells Separated from Limbus In Vivo. Invest. Ophthalmol. Vis. Sci. 2011;52(11):8132-8137. doi: 10.1167/iovs.11-7984.
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To characterize corneal epithelial cells separated from limbus in vivo by transplantation of a stainless steel ring with or without creating a defect inside the ring.
A stainless steel ring (diameter, 8 mm; width, 300 μm; depth, 250 μm) was transplanted into rabbit corneal stroma using 10-0 nylon interrupted suture after cutting to a 250 μm depth by corneal vacuum trephine (diameter, 8.0 mm). Epithelial cells were removed inside the ring, and re-epithelization was evaluated after 1 week. Hematoxylin staining and immunostaining against p63, Ki67, and cytokeratin 3 were performed for phenotypic analysis of corneal epithelia. A corneal epithelial defect was centrally created inside the ring (4, 5, and 6 mm diameter) after transplantation. When re-epithelization was achieved, a central epithelial defect was continuously created until cells were exhausted within the ring. The number of created defects was also analyzed to assess the potential of re-epithelialization.
Ring-transplanted corneal stroma showed few signs of inflammation, and when epithelium was totally removed from inside the ring, complete epithelial defects were persistent for ≥ 1 month. Corneal sensation was significantly decreased in corneas with the ring (P < 0.05). Immunostaining demonstrated similar expression patterns for p63, Ki67, and cytokeratin3 as the controls. When rings were transplanted into the intact cornea, inside epithelia prevented epithelial defects in vivo for ≤ 6 months.
Transient-amplifying cells might maintain homeostasis for >1 month when separated from their limbus in vivo. This model will be useful for future stem cell research or wound healing models.
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