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目的 通过观察表皮细胞体外增殖与老化规律 ,为选择合适的组织工程化皮肤种子细胞提供依据。方法 取正常年轻人表皮细胞进行传代培养 ,以不同代龄细胞为实验对象 ,采用形态学观察、群体倍增时间(PDT)、免疫细胞化学及β 半乳糖苷酶染色等一系列方法 ,检测表皮细胞老化规律。结果 体外单层培养 9代 ,P2的PDT最短 ,前 5代增殖能力较强 ,P5以后PDT明显延长 ,P8细胞不再增殖 ;随着细胞的连续传代培养 ,SA β Gal表达呈现从弱 (在年轻细胞中占 9% )到强 (在老化细胞中占 6 5 % )的趋势。结论 建立了体外培养正常人表皮细胞的衰老模型 ,第 1~ 5代表皮细胞 (供者年龄 16~ 18岁 )可作为构建组织工程化皮肤的种子细胞
Objective To observe the proliferation and aging of epidermal cells in vitro and provide basis for selecting suitable tissue-engineered skin seed cells. Methods Normal young human epidermal cells were subcultured and different generations of the cells were used as experimental subjects. A series of methods including morphological observation, population doubling time (PDT), immunocytochemistry and β-galactosidase staining were used to detect the number of epidermal cells Aging laws. Results After cultured for 9 passages in vitro, the PDT of P2 was the shortest, and the proliferation ability of the first 5 passages was strong. After PD5, the PDT prolonged significantly and P8 cells no longer proliferated. With the continuous subculture of cells, the expression of SAβ- 9% of young cells) to strong (65% of aging cells). Conclusion The established model of human epidermal cell aging in vitro is established. The first to fifth representative epidermal cells (donor age 16-18 years old) can be used as seed cells for the construction of tissue-engineered skin