Transforming growth factor β1 (TGF-β1) is a known pleiotropic growth factor in cell proliferation, migration and phenotypic transition. Fibroblasts are considered the most reactive in the vascular wall. We aimed to explore the effect and mechanism of TGF-β1 on aortal adventitial fibroblasts (AFs) induced by TGF-β1. AFs were cultured in vitro by tissue explants. After stimulation by TGF-β1 and treatment with small interfering RNA (siRNA)-Smad2 and siRNA-Smad3, MTT and the transwell chamber techniques were used for assessing AF proliferation and migration. The expression of phospho (pho)-Smad2, pho-Smad3, Smad7, α-smooth muscle actin (SMA) and collagen I and III were evaluated by Western blot analysis and real-time RT-PCR. After stimulation by TGF-β1 for 24 h, the proliferation and migration of treated AFs were higher compared to those of untreated AFs, as was the expression of Smad2, Smad3, pho-Smad2, pho-Smad3, SMA and collagen I and III (P< 0.05), but not Smad7 (P> 0.05). Knockdown of Smad2 and Smad3 inhibited proliferation and migration of AFs and down-regulated the expression of SMA and collagen I and III (P< 0.05). TGF-β1 plays a key role in remodeling processes by contributing to the proliferation, migration and phenotypic modulation of AFs and collagen composition. The mechanism may be related to both the Smad2 and Smad3 signaling pathways.