Objective: The purpose of this study was to investigate the effects of different fluences of Q-switched 1064 nm Nd:YAG laser on skin repair and barrier, and clarify its mechanisms. Background data: The Q-switched 1064 nm Nd:YAG laser is widely used for rejuvenation, which needs appropriate fluence data to optimize efficacy and minimize side effects, and for elucidation of action mechanism. Materials and methods: The dorsal skin of BABL/c mice was administered 0, 1, 1.5, and 2 J/cm(2) energy level laser, twice a week for 4 weeks. Immediately, 7, 14, 21, and 28 days after last treatment, the skin elasticity, moisture content, and transepidermal water loss (TEWL) were measured; 7, 14, 21, and 28 days after last treatment, the hydroxyproline content, mRNA level of procollagen types I and III, protein level of keratin-10 (K-10), filaggrin, transforming growth factor beta receptor II (TGFbRII), Smad2, and p65 were detected. Results: Compared with the unirradiated control, the laser treatments decreased skin elasticity immediately, but increased skin moisture content in the 2 J/cm(2) group, and then from day 21 to day 28, the skin elasticity, moisture content, hydroxyproline content, and gene expression of types I and III procollagen increased significantly. The TEWL value of the irradiated group significantly increased after irradiation immediately and 7 days after, K-10 and filaggrin were also decreased at 7 days after. The phosphorylation of TGFbRII (p-TGFbRII) increased at days 7 and 21, and phosphorylation of Smad2 (p-Smad2) was induced at 21 days. Conclusions: Irradiation of 1064 nm Q-switched Nd:YAG laser was able to markedly promote repair of mouse skin within 28 days through stimulation of collagen synthesis, with less skin barrier dysfunction, especially at the 1.5 J/cm(2) fluence, and the activation of TGFb1-signaling pathways seemed to play an important role in repair.