Aims: Cold atmospheric plasma (CAP) interacted with tissue and had potential effects on wound healing and tissue regeneration. The aim of this study is to investigate the efficacy and safety of CAP as a novel therapy for diabetic wounds in vitro and in vivo. Methods: The plasma consists of ionized helium gas that is produced by a high-voltage (8.5kV) and high-frequency (17kHz) power supply. Eight-week-old male db/db mice were treated with conventional wound dressing only (control group), additional 90’s CAP (low-dose group) or 180’s CAP (high-dose group) for 2 weeks, respectively. Skin samples around wound in 3, 7, 14 days and blood samples were collected and analyzed in three groups. We carried out in vitro analysis included scratch wound healing assays in immortalized human epidermal HaCaT cells. Results: After 14 days of treatment, CAP could obviously promote diabetic wound healing because of inflammation inhibition and angiogenesis increase. Wound- closure rates of two CAP groups were significantly faster than that of control group. The protein expression of IL-6, TNF-α, iNOS and SOD significantly decreased while the protein level of VEGF, TGF-β in two CAP groups significantly increased compared to those in control group (all p < 0.05). And such changes showed a good consistency with the change in mRNA level (all p < 0.05). In vitro, scratch wound healing assays showed that plasma treatment could effectively accelerate the wound healing within 3 minutes exposure (all p < 0.05). Additionally, there were no significant differences in histological observation and the level of serum ALT, AST, BUN, CREA and WBC among the three groups. Conclusions: CAP treatment for 3 min daily improves diabetic wound healing by inhibiting inflammation, reducing oxidative stress and enhancing angiogenesis without toxicity to liver and kidney. Disclosure R. He: None. Q. Li: None. M. Yu: None. T. Wang: None. H. Lu: None. J. Lu: None. W. Zhu: None. M. Luo: None. J. Zhang: None. H. Gao: None. W. Xing: None. D. Wang: None. F. Liu: None. Funding National Natural Science Fund of China; Shanghai Science & Technology; Shanghai Municipal Education Commission