Faculty of Chinese Medicine, State Key Laboratory of Quality Research in Chinese Medicine, Macau University of Science and Technology, Avenida Wai Long, Taipa, Macao.
Department of Endocrinology and Metabolism, Metabolic Vascular Disease Key Laboratory of Sichuan Province, Sichuan Clinical Research Center for Nephropathy, Luzhou, Sichuan, People's Republic of China.
Cell Biol Toxicol. 2023 Aug;39(4):1577-1591. doi: 10.1007/s10565-022-09748-8. Epub 2022 Aug 19.
Diabetic foot ulcer (DFU) is a devastating component of diabetes progression, leading to decreased quality of life and increased mortality in diabetic patients. The underlying mechanism of DFU is not completely understood. Hence, this study aims to elucidate the mechanism involved in wound healing in mouse models of DFU. Gain- and loss-of-function studies were performed to study the roles that WDR74 and TGF-β play in mouse models of DFU and primary bone marrow-derived mouse macrophages. M1 and M2 macrophage phenotypic markers, extracellular matrix (ECM) components, and angiogenic makers were determined by RT-qPCR and/or Western blot analysis. Localization of these proteins was determined by immunofluorescence staining and/or immunohistochemistry. Interaction between WDR74 with Smad2/3 in macrophages was detected by co-immunoprecipitation. We found that WDR74 and M2 macrophages were decreased in wound tissues from DFU mice. TGF-β/Smad pathway activation increased the expression of M2 macrophage markers (arginase-1 and YM1), IL-4, while decreased expression of M1 macrophage marker (iNOS). TGF-β/Smad pathway activation also increased the production of ECM and promoted the wound closure in diabetic mice. We also noticed that WDR74 overexpression increased Smad2/3 phosphorylation, elevated the population of M2 macrophage and ECM production, and alleviated DFU. LY2109761 treatment normalized effects of TGF-β or WDR74 overexpression. In conclusion, WDR74 promoted M2 macrophage polarization, leading to improved DFU in mice, through activation of the TGF-β/Smad pathway. Graphical Headlights 1. WDR74 promotes M2 macrophage polarization and ECM production. 2. WDR74 activates the TGF-β/Smad signaling pathway. 3. TGF-β/Smad activation promotes M2 macrophage polarization in murine DFU. 4. WDR74 enhances wound healing in murine DFU.
糖尿病足溃疡（DFU）是糖尿病进展的一个严重组成部分，导致糖尿病患者的生活质量下降和死亡率增加。DFU 的潜在机制尚未完全阐明。因此，本研究旨在阐明在 DFU 小鼠模型中伤口愈合涉及的机制。进行了增益和损耗功能研究，以研究 WDR74 和 TGF-β在 DFU 小鼠模型和原代骨髓来源的小鼠巨噬细胞中的作用。通过 RT-qPCR 和/或 Western blot 分析测定 M1 和 M2 巨噬细胞表型标志物、细胞外基质（ECM）成分和血管生成标志物。通过免疫荧光染色和/或免疫组织化学测定这些蛋白质的定位。通过共免疫沉淀检测 WDR74 与巨噬细胞中 Smad2/3 的相互作用。我们发现，DFU 小鼠伤口组织中 WDR74 和 M2 巨噬细胞减少。TGF-β/Smad 通路激活增加了 M2 巨噬细胞标志物（精氨酸酶-1 和 YM1）、IL-4 的表达，同时降低了 M1 巨噬细胞标志物（iNOS）的表达。TGF-β/Smad 通路激活还增加了 ECM 的产生，并促进了糖尿病小鼠的伤口闭合。我们还注意到，WDR74 过表达增加了 Smad2/3 磷酸化，增加了 M2 巨噬细胞和 ECM 产生的群体，并缓解了 DFU。LY2109761 治疗使 TGF-β 或 WDR74 过表达的作用正常化。总之，WDR74 通过激活 TGF-β/Smad 信号通路，促进 M2 巨噬细胞极化，改善小鼠的 DFU。
