Department of Emergency, Tongji Hospital, Tongji University School of Medicine, Shanghai, China.
Biosci Rep. 2020 Oct 30;40(10). doi: 10.1042/BSR20200557.
Transplantion of bone marrow-derived endothelial progenitor cells (EPCs) may be a novel treatment for deep venous thrombosis (DVT). The present study probed into the role of microRNA (miR)-361-5p in EPCs and DVT recanalization. EPCs were isolated from male Sprague-Dawley (SD) rats and identified using confocal microscopy and flow cytometry. The viability, migration and tube formation of EPCs were examined using MTT assay, wound-healing assay and tube formation assay, respectively. Target gene and potential binding sites between miR-361-5p and fibroblast growth factor 1 (FGF1) were predicted by StarBase and confirmed by dual-luciferase reporter assay. Relative expressions of miR-361-5p and FGF1 were detected using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot as needed. A DVT model in SD rats was established to investigate the role of EPC with miR-361-5p antagomir in DVT by Hematoxylin-Eosin (H&E) staining. EPC was identified as 87.1% positive for cluster of difference (CD)31, 2.17% positive for CD133, 85.6% positive for von Willebrand factor (vWF) and 94.8% positive for vascular endothelial growth factor receptor-2 (VEGFR2). MiR-361-5p antagomir promoted proliferation, migration and tube formation of EPCs and up-regulated FGF1 expression, thereby dissolving thrombus in the vein of DVT rats. FGF1 was the target of miR-361-5p, and overexpressed FGF1 reversed the effects of up-regulating miR-361-5p on suppressing EPCs. Down-regulation of miR-361-5p enhanced thrombus resolution in vivo and promoted EPC viability, migration and angiogenesis in vitro through targeting FGF1. Therefore, miR-361-5p may be a potential therapeutic target for DVT recanalization.
骨髓源性内皮祖细胞（EPC）移植可能是治疗深静脉血栓形成（DVT）的一种新方法。本研究探讨了微小 RNA（miR）-361-5p 在 EPC 和 DVT 再通中的作用。从雄性 Sprague-Dawley（SD）大鼠中分离 EPC，并通过共聚焦显微镜和流式细胞术进行鉴定。通过 MTT 测定、划痕愈合测定和管形成测定分别检测 EPC 的活力、迁移和管形成。通过 StarBase 预测 miR-361-5p 和成纤维细胞生长因子 1（FGF1）之间的靶基因和潜在结合位点，并通过双荧光素酶报告基因测定进行验证。根据需要，通过实时定量聚合酶链反应（qRT-PCR）和 Western blot 检测 miR-361-5p 和 FGF1 的相对表达。通过苏木精-伊红（H&E）染色建立 SD 大鼠 DVT 模型，研究 miR-361-5p 反义寡核苷酸对 DVT 中 EPC 的作用。EPC 鉴定为 CD31 阳性 87.1%，CD133 阳性 2.17%，血管性血友病因子（vWF）阳性 85.6%，血管内皮生长因子受体 2（VEGFR2）阳性 94.8%。miR-361-5p 反义寡核苷酸促进 EPC 的增殖、迁移和管形成，并上调 FGF1 表达，从而溶解 DVT 大鼠静脉中的血栓。FGF1 是 miR-361-5p 的靶基因，过表达 FGF1 逆转了上调 miR-361-5p 对抑制 EPC 的作用。miR-361-5p 的下调通过靶向 FGF1 增强了体内血栓溶解，并促进了体外 EPC 的活力、迁移和血管生成。因此，miR-361-5p 可能是 DVT 再通的潜在治疗靶点。
