Nicotinamide mononucleotide-loaded thermosensitive hydrogel promotes diabetic wound healing via antibacterial and antioxidant synergy in a mouse model

Figure 1 PF/polyhexamethylene biguanide-nicotinamide mononucleotide hydrogel exhibits favorable biocompatibility and antibacterial activity. A: The viability of HaCaT cells and EA.hy926 cells was detected by the CCK-8 assay; B and C: Representative images and quantitative analysis of the hemolysis assay for each group of hydrogel extracts under different dilution conditions; D and E: Representative images and corresponding quantification of Staphylococcus aureus colony formation treated with hydrogel extracts from each group; F and G: Representative images and corresponding quantification of Escherichia coli colony formation treated with hydrogel extracts from each group. S. aureus: Staphylococcus aureus; E. coil: Escherichia coli; NMN: Nicotinamide mononucleotide; NS: Not significant; PBS: Phosphate buffered saline; PHMB: Polyhexamethylene biguanide.

Figure 2 PF/polyhexamethylene biguanide-nicotinamide mononucleotide hydrogel enhances cell migration and vasculogenic tubule formation capacity. A and B: Wound healing assay was used to detect the migration ability and corresponding quantification of HaCaT and EA.hy926 cells treated with hydrogel extracts from each group; C: Angiogenesis assay of EA.hy926 cells treated with hydrogel extracts from each group and images analyzed by the angiogenesis plugin in Image J; D: Quantification of meshes and branches in the angiogenesis assay. NMN: Nicotinamide mononucleotide; PHMB: Polyhexamethylene biguanide.

Figure 3 PF/polyhexamethylene biguanide-nicotinamide mononucleotide hydrogel promotes wound healing in diabetic mice. Representative images of wounds in diabetic mice treated with different dressings on days 1, 3, 7, 10, 14, and 21. NMN: Nicotinamide mononucleotide; PHMB: Polyhexamethylene biguanide.

Figure 4 PF/polyhexamethylene biguanide-nicotinamide mononucleotide hydrogel enhances collagen deposition and vascular proliferation in mouse skin wounds. A and B: Hematoxylin and eosin and masson-stained wound images of diabetic mice with different dressings; C: Immunofluorescence staining of diabetic mouse wound tissues [Ki67 (pink) and CD31 (green)]. Scale bar: 100 μm. HE: Hematoxylin and eosin; NMN: Nicotinamide mononucleotide; PHMB: Polyhexamethylene biguanide.

Figure 5 PF/polyhexamethylene biguanide-nicotinamide mononucleotide hydrogel upregulates the expression of vascular endothelial growth factor and transforming growth factor-β1. A and B: Detection of VEGF and TGF-β1 mRNA expression in diabetic mouse wound tissues treated with different dressings by quantitative real-time-polymerase chain reaction; C: Western-blot analysis of vascular endothelial growth factor and transforming growth factor-β1 protein levels in diabetic mouse wound tissues treated with different dressings, with β-actin as the internal reference; D and E: Represent the quantification of protein expression levels. NMN: Nicotinamide mononucleotide; PHMB: Polyhexamethylene biguanide; TGF-β1: Transforming growth factor-β1; VEGF: Vascular endothelial growth factor.