姜黄素对环孢素A诱导激活的大鼠牙龈成纤维细胞TGF-β1/Smad3通路的抑制作用

目的：体外实验研究姜黄素（curcumin,Cur）对环孢素A（cyclosporine A,CsA）作用大鼠牙龈成纤维细胞TGF-β1/Smad3通路的影响,为进一步探讨Cur抑制CsA所致药物性牙龈增生的发生机制提供理论依据。方法：使用CCK-8法观察0、5、10、20、30 μmol/L Cur对大鼠牙龈成纤维细胞增殖的影响, 20 μmol/L Cur+200 ng/mL CsA共同作用对大鼠牙龈成纤维细胞增殖的影响。实时荧光定量PCR检测20 μmol/L Cur+200 ng/mL CsA共同作用下,牙龈成纤维细胞中转化生长因子TGF-β1、Smad3、平滑肌肌动蛋白α-SMA和I型胶原蛋白COL-I的mRNA表达变化;蛋白免疫印迹实验检测TGF-β1、Smad3、p-Smad3、α-SMA和COL-I的蛋白表达变化。细胞划痕实验观察20 μmol/L Cur+200 ng/mL CsA 对牙龈成纤维细胞迁移能力的影响。采用SPSS 23.0 软件包对数据进行统计学分析。结果：大鼠牙龈成纤维细胞在20 μmol/L Cur+200 ng/mL CsA共同作用下,细胞增殖和迁移能力明显降低;20 μmol/L Cur显著下调了牙龈成纤维细胞中TGF-β1、α-SMA 和COL-I的mRNA 表达,蛋白免疫印迹实验提示,TGF-β1、p-Smad3、α-SMA 和COL-I的表达同样显著下调。结论：Cur可能通过抑制CsA激活的牙龈成纤维细胞TGF-β1/Smad3 信号通路,从而降低牙龈成纤维细胞增殖、迁移、平滑肌肌动蛋白和胶原分泌,改善牙龈增生。

Effect of curcumin on TGF-β1 /Smad3 pathway in rat gingival fibroblast treated with cyclosporine A

PURPOSE: The aim of the present study was to investigate the effect of curcumin (Cur) on TGF-β1/Smad3 pathway of rat gingival fibroblast treated with cyclosporine A (CsA) in vitro, and to provide theoretical basis for the mechanism of curcumin inhibiting drug-induced gingival hyperplasia induced by CsA. METHODS: Healthy Sprague-Dawley rat gingival fibroblasts were cultured with different concentrations of Cur (0, 5, 10, 20, 30 μmol/L) and Cur (20 μmol/L)+CsA(200 ng/mL), cell proliferation was assessed with CCK-8 assay. The mRNA levels of TGF-β1, Smad3, α-SMA and collagen type Ⅰ in gingival fibroblasts were detected by real-time PCR under Cur(20 μmol/L)+CsA(200 ng/mL); the protein level of TGF-β1, Smad3, p-Smad3, α-SMA and collagen type Ⅰ were determined through Western blot. The effect of Cur(20 μmol/L)+CsA(200 ng/mL) on migration ability of gingival fibroblasts was observed through Scratch wound-healing assay. The data were analyzed with SPSS 23.0 software package. RESULTS: Cell proliferation and migration ability of rats gingival fibroblasts were significantly reduced under Cur(20 μmol/L)+CsA(200 ng/mL). 20 μmol/L Cur significantly decreased mRNA expression of TGF-β1, α-SMA and collagen type Ⅰ in gingival fibroblasts, and Western blot suggested significantly down-regulated expression of TGF-β1, p-Smad3, α-SMA, and collagen typeⅠ. CONCLUSIONS: Cur may inhibit TGF-β1/Smad3 signaling pathway of gingival fibroblasts activated by CsA, thereby weakening proliferation and migration, reducing secretion of smooth muscle actin and collagen of gingival fibroblasts, and ameliorating gingival hyperplasia.