Evaluation of topical methylene blue nanoemulsion for wound healing in diabetic mice

Abstract

Context: Diabetic wounds (DW) are a complication of diabetes and slow wound healing is the main manifestation. Methylene blue (MB) has been shown to exhibit therapeutic effects on diabetes-related diseases. Objective: To investigate the mechanisms of action of MB-nanoemulsion (NE) in the treatment of DW. Materials and methods: The concentration of MB-NE used in the in vivo and in vitro experiments was 0.1 mg/mL. Streptozocin-induced diabetic mice were used as models. The mice were separated into nondiabetic, diabetic, MB-NE treated, and NE-treated groups. Intervention of high glucose-induced human umbilical vein endothelial cells using MB-NE. The mechanism by which MB-NE promotes DW healing is investigated by combining histological analysis, immunofluorescence analysis, TUNEL and ROS assays and western blotting. Results: In diabetic mice, the MB-NE accelerated DW healing (p < 0.05), promoted the expression of endothelial cell markers (α-SMA, CD31 and VEGF) (p < 0.05), and reduced TUNEL levels. In vitro, MB accelerated the migration rate of cells (p < 0.05); promoted the expression of CD31, VEGF, anti-apoptotic protein Bcl2 (p < 0.05) and decreased the expression of the pro-apoptotic proteins cleaved caspase-3 and Bax (p < 0.05). MB upregulated the expression of Nrf2, catalase, HO-1 and SOD2 (p < 0.05). In addition, MB reduced the immunofluorescence intensity of TUNEL and ROS in cells and reduced apoptosis. The therapeutic effect of MB was attenuated after treatment with an Nrf2 inhibitor (ML385). Discussion and conclusion: This study provides a foundation for the application of MB-NE in the treatment of DW.