Department of Nuclear Medicine, School of Medicine, Kyungpook National University, Daegu, Republic of Korea; BK21 Plus KNU Biomedical Convergence Program, Department of Biomedical Science, School of Medicine, Kyungpook National University, Daegu, Republic of Korea.
Department of Nuclear Medicine, School of Medicine, Kyungpook National University, Daegu, Republic of Korea.
Exp Cell Res. 2020 Sep 15;394(2):112146. doi: 10.1016/j.yexcr.2020.112146. Epub 2020 Jun 17.
Ischemia is the partial or complete blockage of blood supply to tissues. Extracellular vesicles (EVs) are emerging as a therapeutic tool for ischemic diseases. Most EV-based ischemia therapies are based on various stem cells. Here, we propose an alternative cell source for the isolation of pro-angiogenic EVs.
EVs were isolated from a mouse macrophage cell line (Raw 264.7). The characteristic features of the macrophage-derived EVs (MAC-EVs) were assessed using transmission electron microscopy, nanoparticle tracking analysis, and Western blotting (WB) analysis. WB and qRT-PCR were performed to identify the pro-angiogenic VEGF and Wnt3a proteins and microRNAs (miR-210, miR-126, and miR-130a) in the MAC-EVs. In vitro and in vivo Matrigel plug assays were performed to investigate the capacity of the MAC-EVs for tube (blood vessel-like) formation and new blood vessel formation and assessed by histology.
The MAC-EVs was positive for ALIX and negative for calnexin, with a round shape and an average size of 189 ± 65.1 nm. WB and qRT-PCR results revealed that VEGF, Wnt3a and miR-130a were more abundant in the MAC-EVs than cells. MAC-EVs treatment resulted in increased endothelial cellular proliferation, migration, and tube formation in vitro. In vivo assay results revealed that MAC-EVs increased the formation of new and larger blood vessels in the Matrigel plug of mice compared to the formation in the control group.
Our results suggest that MAC-EVs have the potential to induce angiogenesis in vitro and in vivo, could serve as a pro-angiogenic alternative for ischemic diseases.
缺血是组织部分或完全的血液供应阻塞。细胞外囊泡（EVs）正成为缺血性疾病的治疗工具。大多数基于 EV 的缺血治疗方法都是基于各种干细胞。在这里，我们提出了一种替代细胞来源，用于分离促血管生成的 EVs。
从鼠巨噬细胞系（Raw 264.7）中分离 EVs。使用透射电子显微镜、纳米颗粒跟踪分析和 Western blot（WB）分析评估巨噬细胞衍生 EVs（MAC-EVs）的特征。进行 WB 和 qRT-PCR 以鉴定 MAC-EVs 中的促血管生成 VEGF 和 Wnt3a 蛋白和 microRNAs（miR-210、miR-126 和 miR-130a）。进行体外和体内 Matrigel 塞植入物实验以研究 MAC-EVs 形成管状（血管样）结构和新血管形成的能力，并通过组织学进行评估。
MAC-EVs 阳性表达 ALIX，阴性表达 calnexin，呈圆形，平均大小为 189±65.1nm。WB 和 qRT-PCR 结果显示，VEGF、Wnt3a 和 miR-130a 在 MAC-EVs 中的含量高于细胞。MAC-EVs 处理可促进内皮细胞在体外增殖、迁移和管状形成。体内实验结果显示，与对照组相比，MAC-EVs 可增加小鼠 Matrigel 塞中新的和更大血管的形成。
我们的结果表明，MAC-EVs 具有在体外和体内诱导血管生成的潜力，可作为缺血性疾病的促血管生成替代物。
