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M.–H. Kao, C.–C. Cheng, D. Wang, J. Chen; Expression of Keratinocyte Differentiation Markers in Rabbit Cornea During Wound Healing . Invest. Ophthalmol. Vis. Sci. 2005;46(13):2082.
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Purpose: Limbal epithelial stem cells (LSC) are the progenitors of corneal epithelium. In normal cornea, the p63 positive cells are localized to the basal cells of the limbus but not in the central cornea. In wound–healing cornea, p63 positive cells are present in both corneal and limbal epithelium. The purpose of this study is to explore the possible roles of p63 in corneal wound repair. Methods: Immunofluorescent staining was used to examine the expression patterns of p63, PCNA and keratins 14 and 3 in freshly prepared wound–healing or normal corneal tissues. Corneal and limbal explants derived from normal and wound–healing cornea/limbus were cultured on amniotic membrane (AM) to examine their proliferative potential. Results: It has been well established that corneal epithelium is strongly positive with Keratin 3 and limbal epithelial basal layer is the site where Keratin 14–, PCNA– or p63–positive cells are resided. Here we show that in the epithelium of wound–healing cornea, the basal cells are positive with p63, keratin 14 and PCNA, and are negative with Keratin 3, characteristics of that of limbal basal cells. In contrast, Keratin 3 is expressed in cells located from suprabasal to superficcal layers. Interestingly, immunofluorescent staining of serial sections show that all p63 positive cells are also keratin 14 and PCNA positive. Similar results were obtained with ex vivo cultured cornea with or without wounding. In vivo wounded corneas were allowed to undergo repair to complete the reepithlization. On days 7th and 30th post–reepithelization, central corneas were excised and explanted on AM to evaluate their growth potential. Central corneal epithelium taken from day 7th post–reepithelization grew vigorously to form an epithelial outgrowth with a rate that was faster than unwounded limbal tissue. In contrast, central corneas taken from day 30th post–reepithelization were unable to grow and no epithelial outgrowth was formed. The growth potential of the wound repaired central cornea is correlated with the expression of p63, Keraint 14 and PCNA. Conclusions: Our results suggest that p63 is an important regulator for the proliferation of corneal epithelial cells.
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