组织块法分离山羊皮肤干细胞研究

Study on the isolation of goat skin stem cells by explant

采用组织块法从表皮和真皮中分离山羊皮肤干细胞,通过形态学观察、免疫组化染色以及克隆形成率等方法检测两种来源的皮肤干细胞体外生长特性。结果表明,从表皮和真皮中均能分离得到皮肤干细胞;表皮来源的皮肤干细胞体外可传11代,真皮来源的皮肤干细胞体外可传17代;表皮来源的皮肤干细胞第1代K 19和in tegrin-β1阳性细胞率及克隆形成率分别为(34.0±1.62)%,(37.5±2.12)%,(19.4±1.77)%,而真皮来源的皮肤干细胞分别为(29.2±3.12)%,(33.0±1.12)%,(16.6±2.60)%;表皮来源的皮肤干细胞第3代K 19和in te-grin-β1阳性细胞率及克隆形成率分别为(46.4±1.82)%,(55.3±1.98)%,(25.3±1.08)%,真皮来源的皮肤干细胞分别为(53.7±1.17)%,(63.0±1.12)%,(30.9±2.16)%;随着传代次数增加,真皮来源的干细胞的体外活力以及免疫组化染色阳性率、克隆形成率均极显著高于表皮来源的皮肤干细胞(P<0.01)。

The goat skin stem cells were isolated and cultured by two different methods of epidermal explant and dermal explant for the purpose of comparing the effeciency of the two methods on the cultivation and proliferation of skin stem cells in vitro. The resultant cells were observed and identified by cell growth curve, immnohistochemical staining and colony forming efficiency. The results showed the skin stem cells can be separated by both methods;the skin stem cells from epidermal explant could be subcultured 11 times but the stem cells from dermal explant could be subcuhured 17 times in vitro in the same culture condition;The positive expressions of K19,integrin-β1 and CFE of skin stem cells in the first passage by epidermal explant were （34.0±1.62）%,（37.5±2.12）%,（19.4±1.77）% respectively,and those in the first passage by dermal explant were （29.2±3.12）%,（33.0±1.12）%,（16.6±2.60）% respectively;The positive expressions of K19,integrin-β1 and CFE of skin stem cells in the third passage by epidermal explant were （46.4±1.82）%, （55.3± 1.98） %, （25.3± 1.08） % respectively, and those in the third passage by dermal explant were （53.7±1.17）% （63.0± 1.12）%, （30.9±2.16）% respectively;and the stem cells from dermal explant had higher efficiency in immnohistochemical staining,CFE and subculture ability were higher than those of epidermal explant when they were subcultured in vitrv （P〈0. 01 ）.