Characterization of oral keratinocyte stem cells and prospects of its differentiation to oral epithelial equivalents

Vol. 51 No. 4, 2010

ROMANIAN JOURNAL of MORPHOLOGY and EMBRYOLOGY

B. Calenic, N. Ishkitiev, K. Yaegaki, T. Imai, Mariana Costache, Mihaela Tovaru, S. Tovaru, Ioanina Parlatescu

Objective: Although oral keratinocyte stem cells play a key role in tissue homeostasis, wound healing, and neoplasia, they remain difficult to identify and characterize. The specific aim of the present study is to characterize an oral keratinocyte stem-cell population separated using a magnetic technique. Material and Methods: Oral human keratinocytes obtained from keratinized oral mucosa were magnetically separated using a proliferation-related marker, CD71 and alpha6beta4 integrin. The expression of different stem cell markers: CD44H, Nestin, Nanog, Oct 3/4, CD117 was checked by immunofluorescence. The ability of alpha6beta4pos CD71neg fraction to form oral epithelial equivalents was also assayed. Results: Three different oral keratinocyte subpopulations were obtained following magnetic separation: alpha6beta4pos CD71neg, alpha6beta4pos CD71pos and alpha6beta4neg. Our alpha6beta4pos CD71neg stem cell fraction was positive for Oct 3/4, CD44H and cytokeratin 19 while Nanog, Nestin and CD117 expression was absent. At the same time, the other two cell fractions alpha6beta4pos CD71pos and alpha6beta4neg were negative for all stem cell markers. Also, alpha6beta4pos CD71neg fraction was able to regenerate a well stratified and organized oral epithelial equivalent. The distribution of cytokeratin 19 and involucrin in the oral epithelial equivalent reflected the in vivo situation in oral gingival epithelium. Conclusions: The human gingival alpha6beta4pos CD71neg fraction was strongly positive for a panel of stem cell markers and could form oral epithelial equivalent. It is also suggested that a magnetic system may be an important tool in acquiring oral keratinocyte stem cells for research.

Corresponding author: Ken Yaegaki, DDS, PhD, e-mail: yaegaki-k@tky.ndu.ac.jp

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