体外培养人角质形成细胞周期与表皮生长因子受体表达检测

[摘 要] 目的 比较人角质形成细胞（KC）在角质形成细胞无血清培养基（K-SFM）和限制性角质形成细胞无血清培养基（DK-SFM）中的生长、增殖、传代和细胞性状变化的情况。方法 取健康男性包皮，将KC用K-SFM和DK-SFM培养，倒置显微镜下观察两组的细胞形态、数量、传代情况;用流式细胞仪检测细胞周期和细胞表皮生长因子受体（EGFR）的含量。结果 原代培养天12天，K-SFM培养基KC的细胞数量(304.70±83.08)明显多于DK-SFM(75.30±26.08),有统计学差异（P＜0.01），K-SFM培养基KC细胞集落数(2.20±1.23)明显多于DK-SFM(0.00±0.00),有统计学差异（P＜0.01）。K-SFM组细胞可以传4代，DK-SFM组无法传代。K-SFM组G0-G1期细胞比例65.40%, DK-SFM组G0-G1期细胞比例84.10%；K-SFM组G2-M期细胞比例17.02%，DK-SFM组G2-M期细胞比例0.07%； K-SFM组S期细胞比例17.57%,D K-SFM组S期细胞比例组15.82%；K-SFM组细胞凋亡率1.04%,D-SFM组细胞凋亡率18.69%。K-SFM组细胞EGFR表达率为33.73%；DK-SFM组细胞EGFR表达率为3.19%。结论K-SFM培养基中KC细胞生长性状、增殖活性、细胞集落数、传代次数、EGFR表达量等指标均优于DK-SFM培养基。

Measurement of cell cycle of and expression of epidermal growth factor receptor in cultured human keratinocytes in vitro

Investigation of the culture methods of human keratinocytes in vitroBu Xiao-lin, Lu hongguang（Department of Dermatology, Affiliated hospital,Guiyang Medical College, Guiyang 550004, China）Abstract: Objective To compare the biological characteristics of human keratinocytes (KC) in keratinocyte serum-free medium(K-SFM) and defined keratinocyte serum-free medium(DK-SFM).Methods The keratinocytes were harvested from foreskin. A single cell suspension were dissociated by Dispase and trypsin and cultured respectively in the medium of K-SFM and DK-SFM. The morphology of KC was observed under light microscope. The cell cycle and expression of epidermal growth factor receptor (EGFR) were determined with flow cytometry.Result KC grew fast and the numbers of cells and colony appeared much more signifcant high in K-SFM than in DK-SFM. The cells in K-SFM could be maintained in culture up to 4 passages but couldn’t pass into the second generation in DK-SFM. There were also higher expression of EGFR were recorded in the cells of K-SFM than in DK-SFM. Conclusion In contrast to DK-SFM, K-SFM seem to take advantage of the cultivation of human KC in vitro.