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目的 观察体外培养不同代次人成纤维细胞Ⅰ、Ⅲ型胶原基因的表达及对EGF刺激的反应性。方法 将正常人二倍体成纤维细胞进行连续传代培养 ,以不同代次的细胞为实验对象 ,应用RT PCR、3 H TdR掺入量测定等方法对不同代次细胞进行检测。结果 随细胞代次的增加 ,Ⅰ、Ⅲ型胶原mRNA含量逐渐降低 ,3 H TdR掺入量降低 ,EGF促进P10 培养的成纤维细胞增殖 ,对衰老成纤维细胞 (P60 )无生长增殖的调控能力。结论 为建立人成纤维细胞衰老模型提供部分实验依据。
OBJECTIVE: To observe the expression of collagen type Ⅰ and type Ⅲ collagen gene in different generations of human fibroblasts cultured in vitro and its reactivity to EGF stimulation. Methods The normal human diploid fibroblasts were subcultured in succession. The cells of different generations were used as experimental subjects. RT-PCR and 3H-TdR incorporation assay were used to detect different generations of cells. Results With the increase of cell number, the mRNA contents of type Ⅰ and type Ⅲ collagen gradually decreased and the incorporation of 3 H TdR decreased. EGF promoted the proliferation of fibroblasts cultured in P10 and had no effect on the growth and proliferation of aged fibroblasts (P60) . Conclusion Some experimental evidences for establishing human fibroblast cell senescence model are provided.