Ultrasound Combined with Microbubbles Promotes Diabetic Wound Healing by Regulating Macrophage Polarization

Abstract

One of the crucial reasons for impaired healing of diabetic wounds is the excessive and prolonged inflammatory reaction, resulting in the persistent accumulation of neutrophils and M1 macrophages in the wound bed. These cells secrete large amounts of pro-inflammatory cytokines, disrupting the equilibrium of macrophage polarization. As central regulators of inflammatory responses, the polarization status of macrophages is critical to the pathogenesis of wound healing impairment. This study aimed to investigate the therapeutic effect of low-intensity ultrasound combined with microbubbles (USMB) on diabetic wound healing and its underlying mechanism in regulating macrophage polarization. A diabetic rat wound model was established, and successfully modeled rats were randomly divided into three groups: the Model group, the Ultrasound (US) group, and the Ultrasound Combined with Microbubbles (USMB) group. Wound repair was assessed via wound healing rate calculation, histological examination, and immunohistochemical analysis. Macrophage polarization was analyzed using immunofluorescence staining (CD86, CD206) and qRT-PCR assays. Transcriptome sequencing of wound tissues from the USMB and Model groups was conducted on day 6, and the expression of key pathway molecules was validated by qRT-PCR and Western blot.The findings demonstrated that USMB treatment significantly accelerated wound healing in diabetic rats, facilitating granulation tissue formation, collagen deposition, cell proliferation, and angiogenesis. Meanwhile, in the USMB group, the expression of the M1 macrophage marker CD86 and pro-inflammatory cytokines was downregulated, whereas the expression of the M2 macrophage marker CD206 and anti-inflammatory cytokines was upregulated. Transcriptome sequencing identified 1725 differentially expressed genes (DEGs) between the two groups, with significant enrichment in the IL-17 signaling pathway. Further validation confirmed that the mRNA and protein levels of pathway-related molecules, including IL-17B, NF-κB, and TNF-α, were significantly lower in the USMB group than in the Model group. In summary, USMB treatment effectively promotes wound healing in diabetic rats. The underlying mechanism may involve targeted inhibition of the IL-17 signaling pathway, specifically targeting the IL-17B/NF-κB/TNF-α axis. This subsequently drives macrophage polarization from the M1 to M2 phenotype, mitigates excessive inflammatory responses, improves the wound inflammatory microenvironment, and ultimately accelerates tissue repair and regeneration.