Abstract: BACKGROUND: Red light irradiation and silver ion dressing are mostly used to treat chronic difficult healing wounds clinically, but the optimal irradiation time of red light irradiation and silver ion dressing for chronic non-healing wounds, and the combination of different silver ion dressings have not been determined. 
OBJECTIVE: To investigate the optimal irradiation time and dressing combination of red light and silver ion dressing in the therapy of chronic non-healing wounds.
METHODS: The chronic non-healing wound model was made by applying Staphylococcus aureus on the whole skin defect and subcutaneous hydrocortisone injection in SD rats. 72 rat models were randomly divided into 4 groups with 18 rats in each group by random number table method. The rats were treated on the basis of standard dressing change and the following therapy: A1B1 group (red irradiation 20 minutes + lipid hydrocolloidal silver sulfate dressing), A1B2 group (red light irradiation 20 minutes + calcium alginate fiber dressing), A2B1 group (red light irradiation 30 minutes + lipid hydrocolloidal silver sulfate dressing), and A2B2 group (red light irradiation 30 minutes + calcium alginate fiber dressing); change dressing, irradiate once, and change dressing every 24 hours. After 14 days of continuous treatment, wound healing rate, bacterial colony number, inflammatory response, histomorphology and angiogenesis were detected in each group. 
RESULTS AND CONCLUSION: (1) With the extension of treatment time, the wound healing rate of rats in the four groups was increased, and the wound healing rate of rats in the A2B2 group at 3, 7, and 14 days after treatment was higher than that in the other three groups (P < 0.05). (2) The wound bacterial culture results on day 7 after treatment demonstrated that the number of bacterial colonies in the A2B2 group was lower than that in the other three groups (P < 0.05). Western blot assay exhibited that with the extension of treatment time, the protein expressions of tumor necrosis factor α and interleukin-6 in wound tissue of rats in the four groups were decreased, while the protein expressions of interleukin-10 were increased. The protein expressions of tumor necrosis factor α and interleukin-6 in the A2B2 group were lower than those in the other three groups (P < 0.05). The protein expression of interleukin-10 in the A2B2 group was higher than that of the other three groups (P < 0.05). (3) The wound hematoxylin-eosin staining on day 14 after treatment demonstrated that a large number of collagen fibers in the A2B2 group were parallel distributed and the most closely connected, which was significantly better than the other three groups. (4) The results of immunofluorescence staining indicated that the fluorescence intensity expression of CD31 in the A2B2 group was higher than that in the A1B1, A1B2 and A2B1 groups (P < 0.05). q-PCR detection at 3, 7, and 14 days after treatment exhibited that the mRNA expressions of vascular endothelial growth factor a and vascular endothelial growth factor receptor 2 in the A2B2 group were higher than those in the other three groups (P < 0.05). Western blot assay at 3, 7 and 14 days after treatment revealed that the protein expressions of vascular endothelial growth factor a and vascular endothelial growth factor receptor 2 in the A2B2 group were higher than those in the other three groups (P < 0.05). (5) These findings confirm that 30 minutes of red light irradiation combined with silver alginate fiber dressing has better results in treatment of chronic non-healing wounds. 

Key words: red light, silver ion dressing, factorial design, chronic non-healing wound, K value, angiogenesis, interaction, vascular endothelial growth factor