6-O-羧甲基壳聚糖对人增生性瘢痕成纤维细胞TGF-β_1表达的影响

Effects of 6-O-carboxymethyl-chitosan on the expression of TGF-β_1 in human hypertrophic scar fibroblasts

目的:观察不同浓度6-O-羧甲基壳聚糖(6-O-CMC)对体外培养的人增生性瘢痕成纤维细胞(hypertrophic scar fibrob lasts,HSFBs)转化生长因子-β1(transforming growth factor-beta1,TGF-β1)蛋白及mRNA表达的影响,在分子水平上探讨其抑制增生性瘢痕生长的作用机制。方法:体外培养人HSFBs,选取第3～5代细胞进行下一步实验。实验分组采用含不同浓度6-O-CMC(0,100,200,400μg/ml)的DMEM培养液分别进行培养。培养24h后,显微镜下观察各组HSFBs形态及生长情况。收集细胞培养上清,运用双抗体夹心ELISA法检测各组TGF-β1蛋白表达水平。运用荧光定量RT-PCR技术检测各组TGF-β1 mRNA表达水平。结果:与空白组相比,三个药物剂量组HSFBs TGF-β1蛋白及mRNA的表达均减少(P<0.05),药物浓度越高表达量越低(P<0.05),且TGF-β1蛋白及mRNA表达的变化具有一致性。结论:6-O-CMC有抑制体外培养的人HSFBs TGF-β1表达的作用,初步探讨了6-O-CMC发挥抑制增生性瘢痕生长的作用机制,为羧甲基壳聚糖类药物的开发利用提供理论依据。

Objective To observe the effects of 6-O-carboxymethyl-chitosan（6-O-CMC） with different concentrations on the expression of transforming growth factor-beta1（TGF-β1） in human hypertrophic scar fibroblasts（HSFBs） in vitro.Methods HSFBs were cultured in vitro,and the 3～5 generation cells were chose to the next experiment.The DMEM medium with different concentrations of 6-O-CMC（100,200,400ug/ml） were used in the experimental groups,and the control group without 6-O-CMC.After 24 hours,cell morphology and growth state of HSFBs were observed under the microscope.The expression of TGF-β1 protein was detected by ELISA.The expression of TGF-β1 mRNA was was detected by Real-time RT-PCR.Results Compared with the control group,the expressions of HSFBs TGF-β1 protein and mRNA in the other groups all reduced obviously（P0.05）.Concentration and expression were in inverse proportion（P0.05）.The expression of TGF-β1 protein and mRNA are on the same direction.Conclusions 6-O-CMC could suppress the expression of HSFBs TGF-β1.We exploring the mechanism of 6-O-CMC in treating hypertrophic scar,which provides theoretical basis for the development and utilization of new drugs.