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目的 观察重组谷胱甘肽S -转移酶与饰胶蛋白的融合蛋白 (GST -Decorin)对转化生长因子 β1 刺激增生性瘢痕成纤维细胞的作用 ,探索瘢痕治疗的新方法。 方法 采用RT -PCR技术克隆饰胶蛋白 (Decorin)cDNA并连接到pGEX - 4T - 1载体上 ,大肠杆菌内表达GST -Decorin ;体外分离培养增生性瘢痕成纤维细胞 ,加入 1∶6 .2 5 ,1∶12 .5 0 ,1∶2 5 .0 0 ,1∶5 0 .0 0 ,1∶10 0 .0 0稀释度的GST -Decorin ,经四唑盐 (MTT)比色法测定细胞增殖速度 ;进一步将 2mg L的TGF - β1 或同时与 1∶12 .5 0的GST -Decorin加入培养基内 ,比较细胞增殖速度的变化 ,并应用原位杂交及图像分析观察细胞Ⅰ、Ⅲ型前胶原mRNA的表达。 结果 稀释度为 1∶6 .2 5～ 1∶2 5的GST -Decorin可有效地抑制瘢痕成纤维细胞的增殖速度 (P <0 .0 1或P <0 .0 5 ) ;1∶12 .5 0的GST -Decorin能完全拮抗 2mg L的TGF - β1 刺激细胞增殖及Ⅰ、Ⅲ型前胶原的mRNA表达。 结论 GST -Decorin可抑制体外培养的增生性瘢痕成纤维细胞增殖并拮抗TGF - β1 ,提示其具有潜在的治疗瘢痕增生的作用
Objective To investigate the effect of recombinant GST-Decorin (GST-Decorin) on hypertrophic scar fibroblasts stimulated by transforming growth factor β1 (TGFβ1) and to explore a new method for the treatment of scar. Methods Decorin cDNA was cloned by RT - PCR and ligated into pGEX - 4T - 1 vector. GST - Decorin was expressed in E. coli. Hypertrophic scar fibroblasts were isolated and cultured in vitro. , 1:12 .5 0, 1:2 5 .0 0, 1: 50.0 .0, 1:10 0. 0 0 dilution of GST-Decorin, as determined by MTT colorimetric assay Proliferation rate; further 2mg L of TGF - β1 or at the same time with 1:12 .5 0 GST -Decorin added to the medium, comparing cell proliferation rate changes, and application of in situ hybridization and image analysis of cells Ⅰ, Ⅲ Procollagen mRNA expression. Results GST-decorin diluted 1:6. 25 ~ 1: 25 could effectively inhibit the proliferation of scar fibroblasts (P <0.01 or P <0.05); 1:12. 50 GST-Decorin completely antagonized the 2 mg L TGF - β1 stimulated cell proliferation and type Ⅰ, Ⅲ procollagen mRNA expression. Conclusion GST-Decorin can inhibit the proliferation of hypertrophic scar fibroblasts in vitro and antagonize the expression of TGF - β1, which suggests that GST-Decorin has a potential role in the treatment of scar hyperplasia