补骨脂素抗增生性瘢痕作用机制研究

The mechanism of anti-proliferative scar of psoralen

目的探讨补骨脂素抗增生性瘢痕的作用机制。方法体外培养成纤维细胞,按随机数字表法分为正常组(培养正常成纤维细胞)、瘢痕组(培养增生性瘢痕成纤维细胞)、TGF-β1组(10 ng/ml TGF-β1处理增生性瘢痕成纤维细胞5 min^12 h)、Smurf2 RNA干扰组[Smad泛素化调节因子2(Smad ubiquitin regulatory factor2,Smurf2)siRNA转染增生性瘢痕成纤维细胞72 h]、补骨脂素组(10μmol/L补骨脂素处理增生性瘢痕成纤维细胞继续培养72 h)、补骨脂素+TGF-β1组(增生性瘢痕成纤维细胞加入补骨脂素培养72 h后加入TGF-β1培养6 h)。采用Western blot法检测Smurf2、α-平滑肌肌动蛋白(α-actin SMA,α-SMA)蛋白表达;RT-PCR法检测Ⅰ型胶原蛋白mRNA表达;ELISA法检测TGF-β1蛋白分泌。结果与正常组比较,瘢痕组Smurf2蛋白[(0.83±0.08)比(0.38±0.07)]表达增加(P<0.05);与瘢痕组比较,Smurf2 RNA干扰组TGF-β1[(2.2±0.18)比(4.2±0.47)]表达降低(P<0.05);TGF-β1组Smurf2[(0.71±0.06)比(0.42±0.04)]、α-SMA[(1.42±0.12)比(0.91±0.09)]蛋白表达增加(P<0.05),Ⅰ型胶原蛋白mRNA[(0.72±0.09)比(0.41±0.07)]表达增加(P<0.05);补骨脂素组Smurf2[(0.05±0.01)比(0.42±0.04)]、α-SMA[(0.71±0.07)比(0.91±0.09)]蛋白表达降低(P<0.05),Ⅰ型胶原蛋白mRNA表达[(0.12±0.04)比(0.41±0.07)]降低(P<0.05)。结论补骨脂素可能通过TGF-β1/Smurf2信号通路抑制α-SMA蛋白表达,从而降低Ⅰ型胶原蛋白表达,起到抑制瘢痕形成的作用。

Objective To investigate the mechanism of anti-proliferative scar of psoralen.Methods Fibroblasts were cultured in vitro.The fibroblasts were divided into normal group(normal fibroblasts),scar group(hypertrophic scar fibroblasts),and TGF-β1 group(10 ng/ml TGF-β1 treated hypertrophic scar fibrils cells 5 min-12 hours),Smurf2 RNA interference group(Smurf2 siRNA transfected hypertrophic scar fibroblasts for 72 hours),psoralen group(10μmol/L psoralen treatment of hypertrophic scar fibroblasts for 72 hours).The psoralen+TGF-β1 group(proliferative scar fibroblasts were added to the psoralen for 72 hours and then added to TGF-β1 for 6 hours).The expressions of Smurf2 andα-SMA protein were detected by Western blot method.The expression of type I collagen mRNA was detected by RT-PCR.The secretion of TGF-β1 protein was detected by ELISA.Results Compared with the normal group,the expression of Smurf2 protein(0.83±0.08 vs.0.38±0.07)in the scar group significantly increased(P<0.05).Compared with the scar group,the expression of TGF-β1(2.2±0.18 vs.4.2±0.47)significantly decreased in Smurf2 RNA interference group(P<0.05).The expression of Smurf2(0.71±0.06 vs.0.42±0.04)andα-SMA(1.42±0.12 vs.0.91±0.09)proteinin the TGF-β1 group significantly increased(P<0.05),and the expression of type I collagen mRNA(0.72±0.09 vs.0.41±0.07)significantly increased(P<0.05).The expression of Smurf2(0.05±0.01 vs.psoralen group significantly decreased(P<0.05).The expression of collagen mRNA(0.12±0.04 vs.0.41±0.07)significantly decreased(P<0.05).Conclusions Psoralen may inhibit the expression ofα-SMA protein and decrease the expression of type I collagen by TGF-β1/Smurf2 signaling to inhibit scar formation.