胚胎成纤维细胞和Ⅳ型胶原黏附分选表皮干细胞的比较研究

目的比较以Ⅳ型胶原和胚胎成纤维细胞黏附分选表皮干细胞的效率，探索从皮肤中快速表皮干细胞分选的有效方法。方法使用DispaseⅡ和CollagenaseⅠ混合酶、胰酶分步消化皮肤得到的细胞悬液，实验组用胚胎成纤维细胞，对照组使用Ⅳ型胶原，分别作为黏附基质，在细胞培养板中对表皮干细胞进行黏附筛选，培养5d后统计各组表皮干细胞克隆形成数量，评价筛选效率。结果经抗β-整合素单抗和抗角蛋白K19单抗免疫细胞染色确证形成克隆的细胞是表皮干细胞，在两个细胞密度水平，实验组和对照组培养出的表皮干细胞克隆数量差异无统计学意义（P〉0．05）。结论使用成纤维细胞分选表皮干细胞与使用Ⅳ型胶原方法分选效率相当，成纤维细胞条件培养液对黏附效率没有影响，但能促进表皮干细胞生长。

Isolation of epidermal stem cells adhered by embryo fibroblasts and type Ⅳ collagen

Objective To explore a rapid isolating epidermal stem cells method based on adhered type Ⅳ collagen and embryo fibroblasts, respectively. Methods Epidermal cells digested by dispase Ⅱ , Collagenase Ⅰ and trypsin from skin were dropped to cell culture plate in two different cell density. Fibroblasts digested by trypsin from abortion embryo tissue was laid on the plate in experiment group, while type Ⅳ collagen was coated in the plate in control group. Condition medium was applied in control group Ⅰ , but free in control group Ⅱ. The amount of cell clones in each group was counted after being cultured for 5 days. Results The cell clones were proved to be epidermal stem cells by immunity stained with anti-β integrin and anti-keratin 19 antibody. There was no distinctive variation between all groups in each culture density levels（ P 〉 0.05 ）. Conclusion That effect on isolating epidermal stem cells is similar by applying fibroblasts or type Ⅳ collagen. Fibroblasts condition medium could not improve the isolating effect of epidermal stem cells, but could make them grow more rapidly.