表皮角质形成细胞分化无血清培养方法的建立

Establishment of differentiation of epidermal keratinocyte in serum free medium

目的建立一种研究角质形成细胞分化的细胞培养方法。方法细胞培养采用无血清无钙离子的角质形成细胞生长培养基(keratinocytegrowthmedium,KGM),通过改变培养基中的钙离子浓度来调节角质形成细胞的分化状态,并对分化标记物K10以及在银屑病患者受损表皮中表达异常增高的组织型纤溶酶原激活剂(tissue-typeplasminogenactiva-tor,tPA)通过免疫细胞化学技术(immunocytochemistry,ICC)分别进行检测。结果低钙(0·09mmol/L)培养条件下,细胞处于未分化状态,K10染色为阴性,tPA呈弱阳性表达;高钙(1·5mmol/L)条件下,细胞出现分层分化,K10染色为阳性,tPA的表达明显增强。结论通过改变KGM的钙离子浓度来调节角质形成细胞的分化,可用于角质形成细胞分化的研究。

Objective To establish a cell culture method for differentiation of keratinocyte in serum free medium mediated by the Ca 2+ concentration. Methods Keratinocytes were cultured in serum and Ca 2+ free keratinocyte growth medium (KGM). Differentiation of keratinocytes was regulated by the change of Ca 2+ concentration in medium, and the expression of K10 and tPA that is elevated in psoriasis were detected by immunocytochemistry (ICC). Results When Ca 2+ concentration in medium was 0.09 mmol/L, keratinocytes were in the state of undifferentiation, K10 expression was negative, and tPA expressed weakly. When cells were exposed to 1.5 mmol/L Ca 2+ medium, terminal differentiation of keratinocyte was induced, K10 expression was positive, and tPA expression was significantly increased. Conclusion The changes of Ca 2+ concentration in mediating the keratinocyte differentiation can be used in the study on keratinocyte differentiation in KGM.