Isolation,cultivation and identification of dedifferentiation derived epithelial stem cells
- VernacularTitle:去分化来源表皮干细胞的分离与鉴定
- Author:
Xiaoyan SUN
;
Bo WEI
;
Xiaobing FU
;
Huiling LIU
- Publication Type:Journal Article
- Keywords:
epidermis;
stem cells;
cell dedifferentiation;
immunohistochemistry;
flow cytometry
- From:
Medical Journal of Chinese People's Liberation Army
1982;0(03):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To introduce an efficient method to isolate and identify the dedifferentiation derived epidermal stem cells. Methods HEKa cells obtained from Casacade (USA) were induced to reverse their differentiated process and produce immature, stem-like cells-the dedifferentiation derived epidermal stem cells (dHEK cells)-by basic fibroblasts growth factors (bFGF) in vitro. Improved collagen Ⅳ-coated adhesion method was used to isolate and cultivate the dHEK cells. Subsequently, morphological features and phenotypic changes of these immature dHEK cells were immunochemically stained and studied with confocal microscopy and flow FACS array. Results Immunocytochemical analysis proved that dHEK cells and their subunits were positive for ?1 integrin, keratin 19 and 14, yet negative for the expression of keratin 10, which was considered as an established marker of differentiated cells. Double staining immunofluorescence demonstrated that markers such as ?6 integrin and CD71 were co-expressed in dHEK cells, and it was shown that there was important regional differences in the distribution of ?6 integrin and CD71 in dHEK cells and their subpopulations. More importantly, two-color flow cytometric analysis of ?6 integrin and CD71 consistently revealed two phenotypically discrete populations of cells in the isolated dHEK cells, i.e. a major subpopulation exhibiting high levels of both ?6 and CD71 expression (?6+CD71+ representing 66.97%?5.04% of the total cells) and a minor population characterized by high level of ?6 expression and low level of CD71 expression (?6+CD71- representing 24.52%?7.88% of the total cells) (n=3). Conclusions The dHEK cells isolated by collagen Ⅳ-coated adhesion method have some characteristics of native epidermal stem cells, and perhaps it may act as a new vehicle for non-genetic manipulation and therapy for both skin disorders and systemic deficiencies. Additionally, although viewed as an ideal substitute of ESCs, the safety of dHEK cells and their functional difference with native epidermal stem cells are still unclear, which need further research.
