人角质形成细胞的体外改良培养及生物学特征

Improved methods for in vitro culturing human keratinocytes and biological features

目的:探讨体外人角质形成细胞的改良培养方法并观察其生物学特征。方法:取健康人包皮环切术包皮,采用EDTA预处理和冷消化法分离表皮,制成细胞悬液,通过倒置相差显微镜、电镜观察其形态学特征,绘制细胞生长曲线;免疫组化(ABC法)测定抗角蛋白单克隆抗体鉴定细胞。结果:体外培养的人皮肤角质形成细胞经过1～2天的潜伏期,即进入指数增生期,持续约5天,然后进入平台期。第7天左右细胞完全融合连成片状,形态呈扁平不规则多边形,免疫组化显示胞浆被染成棕黄色,为角蛋白阳性。透射电镜下见角质形成细胞胞浆内有大量束状张力纤维呈典型角质形成细胞特征。结论:采用改良体外培养方法获得的人角质形成细胞保持正常的生物学特征,从而建立一种简单易行的人角质形成细胞优化培养技术。

Objective：To explore methods to improve in vitro cultured human keratinocytes and observe biological characteristics. Methods： Keratinocytes isolated from healthy human foreskin were incubated in a humidified 5% carbon dioxide at 37℃. EDTA pretreatment and cool digestion were used to improve culture methods. Keratinocytes were then identified by their morphological features using light microscopy, transmission electron microscopy and immuno -histochemistry. Results：A large amount of human keratinocytes could be obtained ＇after 7 days. Keratinoeytes displayed typical morphology when observed by phase -contrast microscopy, electron microscopy, and immunohistochemical staining. Conelusion：Keratinocytes retain their normal biological characteristics while using improved culture methods, offering an easier technique for in vitro culturing human keratinoeytes.