重组核心蛋白多糖对人正常皮肤和增生性瘢痕成纤维细胞增殖及生物学活性的影响

Effects of recombinant human decorin on proliferation and biological activity of normal human skin and hypertrophic scar fibroblasts in vitro

目的 观察重组核心蛋白多糖 (Decorin)对人正常皮肤和增生性瘢痕成纤维细胞体外增殖和生物学活性的影响 ,以探讨Decorin在增生性瘢痕形成和成熟中的作用。 方法 分离培养人正常皮肤和增生性瘢痕成纤维细胞 ,分别加入不同质量浓度 (0 .0 1,0 .1,2 ,10 μg/ml)的重组Decorin ;培养 12 ,2 4 ,4 8h用二甲基噻唑二苯基四唑溴盐 (MTT)法测定细胞增殖速度 ;培养 4 8h用流式细胞仪检测细胞周期和凋亡 ;收集细胞培养上清 ,ELISA法检测TGF - β1蛋白水平 ,放射免疫方法检测Ⅰ、Ⅲ型前胶原蛋白 (PCⅠ、PCⅢ )含量。 结果 2 ,10 μg/ml的Decorin可有效抑制正常皮肤和增生性瘢痕成纤维细胞的增殖 (P <0 .0 5或P <0 .0 1) ,明显增加两种细胞处于G1期的百分比(P <0 .0 1) ;并抑制细胞分泌转化生长因子 (TGF) - β1和PCⅠ、PCⅢ ,下调PCⅠ /PCⅢ比值。实验组和对照组 (未加重组Decorin)均未检测到终末期凋亡细胞。 结论 Decorin可抑制正常皮肤和增生性瘢痕成纤维细胞的增殖及其生物学活性 ,可能在预防增生性瘢痕形成和促进瘢痕成熟中起一定的作用。

Objective To investigate effects of recombinant human decorin on proliferation and biological activity of normal human skin and hypertrophic scar fibroblasts in vitro so as to explore the role of decorin in formation and mature of hypertrophic scar. Methods Fibroblasts derived from normal human skin and hypertrophic scar were harvested and cultured in vitro with recombinant human decorin of serial diluted concentration (0.01 μg/ml,0.1 μg/ml,2 μg/ml and 10 μg/ml). After 12,24 and 48 hours of culture, the cell proliferation rates were measured by MTT colorimetric determination. After 48 hours of culture,cell cycle distributions and apoptosis were analyzed with flow cytometer,followed by collection of the cell supernatants. Contents of TGF-β1 and type of Ⅰ and Ⅲ pro-collagens (PCⅠ and PC Ⅲ) in the supernatants were detected by enzyme-linked immunoabsorbent assay and radioimmunoassay,respectively. Results The proliferations of normal human skin and hypertrophic scar fibroblasts were significantly inhibited and the percent of cells at the G1 phase significantly increased by 2 μg/ml decorin ( P <0.05 or P <0.01). Contents of TGF-β1 and PCⅠand PCⅢ and the ratio of PCⅠto PCⅢ in the supernatants were significantly decreased. Cells in late apoptosis were not found in control and groups treated with decorin. Conclusions Recombinant human decorin can inhibit the proliferation and biological activity of normal human skin and hypertrophic scar fibroblasts in vitro, suggesting that decorin may be effective in preventing hypertrophic scar and scar promotion from mature.