Pirfenidone (AMR69) inhibits the production of CCL2 and CCL12 in fibroblasts and also decreases TGF-β2 protein levels. Pirfenidone is an antifibrotic agent that is commonly used in studies related to pulmonary fibrosis and also has anti-inflammatory activity.

方法: 小鼠巨噬细胞样细胞系 RAW264.7 用 lipopolysaccharide (1 µg/mL) 和 Pirfenidone (30-300 µg/mL) 处理 8 h，使用 ELISA assay 检测 TNF-α。
结果: 将 RAW264.7 细胞暴露于 Pirfenidone 显著抑制细胞相关和分泌的 TNF-α 水平。[1]
方法: 对照肺的三种基质细胞系和来源于特发性肺纤维化 (IPF) 的三种细胞系用 Pirfenidone (0.1-1 mM) 处理 6 天，使用 MTT assay 检测细胞活力。
结果: Pirfenidone 以剂量依赖的方式抑制细胞增殖。1 mM Pirfenidone 处理第 6 天，对照细胞的增殖减少到 47%，IPF 细胞为 42%。[2]

方法: 为检测体对肺纤维化的作用，将 Pirfenidone (30-100 mg/kg，0.5% carboxymethylcellulose) 口服给药给 bleomycin 诱导肺纤维化的 ICR 小鼠，每天三次，持续 42 天。
结果: Pirfenidone 可抑制肺部炎症性水肿，显著抑制肺纤维化。在 bleomycin 诱导的小鼠肺纤维化发展过程中，Pirfenidone 通过调节肺 IFN-γ、bFGF 和 TGF-β1 水平发挥抗纤维化作用。[3]

Pirfenidone (PFD) is dissolved in DMSO and stored, and then diluted with appropriate media before use[3]. HLECs are seeded in 96-well plates (1×104 cells/well) for 24 hours in α-MEM/10% FBS/1%NEAA, and are cultured in stationary tubes in serum-free medium for 24 hours. And then the culture medium is removed and cells are bathed in α-MEM with 10% FBS and 1% NEAA supplemented with 0, 0.01, 0.1, 0.2, 0.3, 0.5, or 1 mg/mL Pirfenidone for 0, 4, 12, 24, 48, or 72 hours. After incubation with 180 μL α-MEM and 20 μL of 5 mg/mL MTT for 4 hours at 37°C, the MTT solution is discarded. The Formosan precipitates are dissolved in 180 μL DMSO by agitating the dishes for 10 minutes at 200 rpm on an orbital shaker. The absorbance at 490 nm in each well is read with a micro plate reader. We further examined the effects of PFD by refining the concentrations at 0.2, 0.25, 0.3, 0.4, 0.5 and 0.6 mg/mL using the MTT assay[3].