Macrophage Phenotype Transition as the Biological Mechanism of Chronic Wound Healing Treated with Non-Thermal, Non-Cavitational Therapeutic Ultrasound

巨噬细胞表型转变作为非热、非空化超声治疗慢性伤口愈合的生物学机制

基本信息

批准号：
10373982
负责人：
Olivia Ngo
金额：
$ 2.71万
依托单位：
DREXEL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-04-01 至 2022-08-01
项目状态：
已结题

项目摘要

PROJECT SUMMARY/ ABSTRACT Chronic wounds affect approximately 6.5 million patients in the United States. Current standard protocols for wound management do not guarantee healing and focus on maintaining a wound environment that is conducive to passive self-healing. Hence, there is a need to develop alternative treatments that promote active healing and shorten healing time leading to reduced costs. We have previously reported that treatment with low-frequency (20-100 kHz), low-intensity (50-150 mW/cm2) ultrasound (LFLI US) significantly (p<0.03) reduces venous ulcer size in vivo as compared to wounds treated with a sham device. This proposal aims at determining the biological mechanisms by which LFLI US promotes chronic wound healing in vitro. There is evidence that the cause of impaired healing is the dysregulation of macrophage phenotype, especially the defective transition from pro- inflammatory (M1) to pro-healing (M2) macrophages. Our characterization of tissue debrided from chronic wounds has shown that healing chronic wounds contain higher proportions of M1-like than M2- like macrophages. Additionally, the signaling protein Rac2, downstream of integrin and focal adhesion kinase activation, is a key regulator of mechanotransduction in macrophages and facilitates the transition of macrophages from the M1 to M2 phenotype. The proposed study will systematically examine the effects of LFLI ultrasound on macrophage phenotype, using macrophages cultured in three-dimensional (3D) scaffolds. We hypothesize that LFLI US directly and indirectly stimulates the transition of pro-inflammatory M1 macrophages to pro-healing M2 macrophages via Rac2. This project will enhance our understanding of chronic wound healing and the potential of therapeutic ultrasound to accelerate healing. Aim 1 will elucidate the direct effects of LFLI US on macrophage function and phenotype by treating inflammatory macrophages directly with LFLI US and characterizing functional changes (proliferation, migration, and phagocytosis), protein/cytokine secretion, and gene expression. Concurrently, we will validate Rac2 as the potential mechanotransduction pathway which promotes M1 to M2 macrophages transition by analyzing integrins, focal adhesion kinases, and Rac2 via confocal microscopy and RNA characterization. [Aim 2 will validate the in vitro findings from Aim 1 using our previously developed diagnostic M1/M2 score on debrided tissue from chronic wound patients treated with LFLI US.] Aim 3 will elucidate the indirect effects of LFLI US on macrophage function and phenotype via a 3D macrophage fibroblast co-culture. The results of this study will inform the optimal design of LFLI ultrasound therapy protocols, lead to a personalized, active treatment for chronic wounds, accelerate chronic wound healing, and contribute to reduced annual wound care costs.

项目摘要/摘要慢性伤口影响美国约650万患者。当前标准伤口管理方案不能保证康复并专注于维持伤口有利于被动自我修复的环境。因此，有必要开发替代方案促进积极康复并缩短康复时间的治疗，导致成本降低。我们有先前报道了低频（20-100 kHz），低强度（50-150 mW/cm2）的治疗与伤口相比用假装置处理。该建议旨在确定生物学机制 LFLI US在体外促进慢性伤口愈合。有证据表明康复受损的原因是巨噬细胞表型的失调，尤其炎症（M1）促进治疗（M2）巨噬细胞。我们从慢性伤口表明，愈合的慢性伤口比M2-的比例更高。像巨噬细胞。此外，信号蛋白Rac2，整联蛋白和局灶性粘附的下游激酶激活是巨噬细胞机械转导的关键调节剂，并促进巨噬细胞从M1到M2表型的过渡。拟议的研究将系统地使用在三维（3D）脚手架。我们假设LFLI直接和间接地刺激了我们促炎性M1巨噬细胞通过RAC2过渡到促愈合M2巨噬细胞。这个项目将增强我们对慢性伤口愈合的理解以及治疗性超声的潜力加速治愈。 AIM 1将阐明Lfli US对巨噬细胞函数的直接影响和通过直接用LFLI US处理炎症性巨噬细胞并表征功能，表型通过变化（增殖，迁移和吞噬作用），蛋白/细胞因子分泌和基因表达。同时，我们将验证RAC2为促进M1的潜在机械转导途径通过分析整联蛋白，局灶性激酶和Rac2通过共聚焦分析M2巨噬细胞过渡显微镜和RNA表征。 [AIM 2将使用我们的AIM 1验证AIM 1的体外发现先前从经过治疗的慢性伤口患者的精明组织中开发了M1/M2的诊断M1/M2评分 [AIM 3与lfli US。通过3D巨噬细胞成纤维细胞共培养的表型。这项研究的结果将告知最佳 LFLI超声治疗方案的设计，导致慢性的个性化，主动治疗伤口，加速慢性伤口愈合，并导致年度伤口护理成本降低。