吡非尼酮对转化生长因子-β_1诱导的大鼠心肌成纤维细胞增殖和胶原蛋白合成的影响

Effect of Pirfenidone on TGF-β_1 induced proliferation and collagen synthesis of rat cardiac fibroblasts

目的探讨吡非尼酮(PFD)对转化生长因子-β1(TGF-β1)诱导的乳鼠心肌成纤维细胞(CFB)增殖和Ⅰ型胶原蛋白(ColⅠ)合成的影响,探讨其抗纤维化作用机制。方法 1体外培养CFB,不同浓度(0、100、200、400、800和1 600μg/ml)PFD加入CFB培养液中24、48和72 h,四甲基偶氮唑盐(MTT)比色法检测CFB增殖,筛选PFD最佳作用浓度范围和作用时间;2根据以上实验结果,选用400和1 600μg/ml PFD加入CFB培养液中48 h作为观察浓度和干预时间,实时定量逆转录-聚合酶链反应(RT-q PCR)和Western blot检测PFD对TGF-β1诱导CFB增殖、ColⅠm RNA和ColⅠ表达的影响。结果与对照组比较,400和1 600μg/ml PFD能显著抑制TGF-β1诱导CFB增殖、ColⅠm RNA和ColⅠ表达,400μg/ml即能起显著抑制作用。结论 PFD对CFB增殖和胶原蛋白合成有抑制作用,PFD的抗纤维化作用与其有关。

【Objective】 To investigate the effect of Pirfenidone（PFD） on transforming growth factor-β1（TGF-β1） induced proliferation and type Ⅰcollagen（Col Ⅰ） synthesis in rat cardiac fibroblasts（CFB） and explore its anti-fibrotic mechanism. 【Methods】 Cardiac fibroblasts were treated with PFD of different concentrations（0, 100, 200, 400, 800 and 1,600 μg/ml） for 24, 48 and 72 hours. Cell proliferation activity was detected by MTT assay and then the appropriate concentration and time of Pirfenidone were found out.The cultured cardiac fibroblasts were stimulated with TGF-β1and then treated with PFD of different concentrations（400 and 1,600 μg/ml） for 48 hours. Cell proliferation activity was detected by MTT assay, the expression of type Ⅰ collagen m RNA was determined by real-time quantitative reverse transcription-polymerase chain reaction（RT-q PCR） and the amount of type Ⅰ collagen in cultured cells was measured by Western bolt. 【Results】 Pirfenidone inhibited cardiac fibroblast proliferation stimulated with TGF-β1, the expression of type Ⅰ collagen m RNA and the amount of type Ⅰ collagen. The effective concentration of Pirfenidone was 400 ~ 1,600 μg/ml for 48 h after Pirfenidone was added into culture medium, the concentration of 400 μg/ml worked significantly. 【Conclusion】 Pirfenidone can inhibit cardiac fibroblast proliferation stimulated with TGF-β1and collagen production which may contribute to its anti-fibrotic mechanism.