# Ultrasound combined with microbubbles promotes diabetic wound healing by regulating macrophage polarization

## TL;DR

Using ultrasound with microbubbles helps heal diabetic wounds by reducing inflammation and shifting immune cells to a healing state.

## Contribution

This study reveals that USMB promotes diabetic wound healing by inhibiting the IL-17 signaling pathway and modulating macrophage polarization.

## Key findings

- USMB treatment accelerated wound healing and improved tissue regeneration in diabetic rats.
- USMB reduced M1 macrophage markers and pro-inflammatory cytokines while increasing M2 markers and anti-inflammatory cytokines.
- Transcriptomic analysis identified the IL-17 signaling pathway as a key target of USMB, with reduced IL-17B, NF-κB, and TNF-α expression.

## Abstract

One of the crucial reasons for the impaired healing of diabetic wounds is the excessive and prolonged inflammatory response at the wound site, which leads to the persistent accumulation of neutrophils and M1 macrophages, the release of abundant pro-inflammatory cytokines, and the disruption of the balance in macrophage polarization. Macrophages, as central regulators of inflammatory responses, play a pivotal role in wound healing impairment through their polarization states. This study aimed to investigate the therapeutic effect of low-intensity ultrasound combined with microbubbles (USMB) on diabetic wound healing and the molecular mechanisms by which USMB regulates macrophage polarization.

Diabetic rat wound models were established, and rats with successful modeling were divided into three groups: the Model group, the Ultrasound (US) group, and the Ultrasound Combined with Microbubbles (USMB) group. Wound repair efficacy was evaluated by calculating the wound healing rate, conducting histological examinations (HE and Masson staining), and performing immunohistochemical staining(CD31,Ki67).Macrophage polarization was detected using immunofluorescence staining (CD86, CD206) and qRT-PCR (pro-inflammatory/anti-inflammatory factors). Transcriptomic sequencing was conducted on wound tissues from the USMB and Model groups on day 6, followed by validation of key pathway molecules using qRT-PCR and Western blotting.

USMB treatment accelerated wound healing, enhanced granulation tissue formation, increased collagen deposition, stimulated cell proliferation, and promoted angiogenesis. Meanwhile, the USMB group exhibited decreased expression of M1-type macrophage marker CD86 and pro-inflammatory cytokines (IL-1β, IL-6), along with increased expression of M2-type marker CD206 and anti-inflammatory cytokines (IL-10, Arg1). Transcriptomic analysis identified 1725 differentially expressed genes between the two groups, with the IL-17 signaling pathway being significantly enriched. Further validation revealed that the mRNA and protein expressions of pathway-related molecules including IL-17B, NF-κB, and TNF-α in the USMB group were significantly lower than those in the Model group.

USMB treatment can promote wound healing in diabetic rats, and its potential mechanism may lie in the targeted inhibition of the IL-17 signaling pathway, particularly acting on the IL-17B/NF-κB/TNF-α axis. This thereby promotes the polarization of macrophages from the M1 to M2 phenotype, reduces excessive inflammatory responses, improves the inflammatory microenvironment, and further accelerates tissue repair and regeneration.