Hypoxia-Induced Extracellular Vesicles Derived from Human Umbilical Cord Mesenchymal Stem Cells Regulate Macrophage Polarization and Enhance Angiogenesis to Promote Diabetic Wound Healing

<b>Background</b>: Diabetic wound healing has always been a clinical challenge with minimal response or efficacy to standard treatment. This study aims to assess the therapeutic potential of hypoxia-induced extracellular vesicles (hy-EVs) produced by human umbilical cord mesenchymal stem cells (HUCMSCs) to treat diabetic wounds. <b>Methods</b>: HUCMSCs were isolated from umbilical cord tissue, cultured under hypoxic conditions to induce the release of extracellular vesicles (EVs) and compared with normoxia-induced extracellular vesicles (n-EVs). We assessed the functions of hy-EVs on human skin fibroblasts (HSFs) and human umbilical vein endothelial cells (HUVECs) in vitro. Simultaneously, we analyzed the pro-angiogenic effects of hy-EVs, their effects on macrophage polarization, and their ability to scavenge endogenous reactive oxygen species (ROS). In addition, a diabetic wound model was established to assess the curative effect of hy-EVs in diabetic wound healing. <b>Results</b>: We found by in vitro study that hy-EVs markedly improved the functional activities of HSFs, thus significantly promoting wound repair. Remarkably, it was determined that hy-EVs greatly enhanced the proliferation and migration ability as well as the angiogenic ability of HUVECs, while promoting the expression of hypoxia-inducible factor-1α (HIF-1α), vascular endothelial-generation-associated factor A (VEGFA), and platelet endothelial adhesion molecule (CD31), which suggested that hy-EVs can effectively activate the HIF-1α pathway to promote angiogenesis. Above all, we found that hy-EVs promoted the expression of CD206 while decreasing the expression of CD86, suggesting that hy-EVs could induce macrophages to shift from M1-type (pro-inflammatory) to M2-type (anti-inflammatory), thereby modulating the inflammatory response. Additionally, hy-EVs inhibited ROS production in both HSFs and HUVECs to reduce oxidative stress. In vivo results showed that hy-EVs enhanced collagen deposition and angiogenesis, modulated macrophage polarization, and inhibited immune response at the wound spot, which significantly enhanced diabetic wound healing. <b>Conclusions</b>: Our study shows that hy-EVs significantly promote angiogenesis through activation of the HIF-1α pathway, modulate macrophage polarization and attenuate cellular oxidative stress, possibly through delivery of specific miRNAs and proteins. Our discoveries offer a key theoretical basis and potential application to develop novel therapeutic strategies against diabetes-related tissue injury.