Department of Vascular Surgery, Wuxi Second People's Hospital, Wuxi, China.
Department of General Surgery, Wuxi Second People's Hospital, Wuxi, China.
Pharmacology. 2023;108(4):331-343. doi: 10.1159/000527812. Epub 2023 May 9.
Endothelial progenitor cells (EPCs) have been proven to exhibit a therapeutic effect in deep vein thrombosis, but are susceptible to microenvironment. Besides, Matrine has promotive effects on EPCs, but its effects on microRNA (miR)-126 remain obscure, which are therefore discussed in the study.
The cultured EPCs were extracted from Sprague-Dawley rats and identified by immunofluorescence assay. After being treated with Matrine or transfected with miR-126b inhibitor and small interfering RNA targeting forkhead box (FOXO) 4, the viability and apoptosis of EPCs were determined by cell counting kit-8 assay and flow cytometry. The migration, invasion, and tube formation abilities were detected by scratch, Transwell, and tube formation assays. The target genes of miR-126b were predicted by TargetScan, and verified by dual-luciferase reporter assay. The expressions of miR-126b, FOXO4, matrix metalloproteinase (MMP) 2, MMP9, and vascular endothelial growth factor (VEGF) A were determined by quantitative real-time polymerase chain reaction and Western blot.
The EPCs were successfully extracted and cultured, as evidenced by positive reaction cluster of differentiation (CD) 34 and CD133. Matrine promoted the viability, migration, invasion, and tube formation while inhibiting the apoptosis of EPCs, and upregulated the expression of miR-126b. Besides, miR-126b inhibitor reversed the effects of Matrine on EPCs and downregulated the expression levels of MMP2, MMP9, and VEGFA. MiR-126b targeted the FOXO4, and siFOXO4 reversed the abovementioned effects of miR-126b inhibitor on EPCs.
Matrine protects EPCs from apoptosis and promotes their migration, invasion, and tube formation abilities via regulating miR-126b/FOXO4 axis.
内皮祖细胞 (EPCs) 已被证明在深静脉血栓形成中具有治疗作用，但易受微环境影响。苦参碱对 EPCs 有促进作用，但对 microRNA (miR)-126 的作用尚不清楚，因此在本研究中进行了讨论。
从 Sprague-Dawley 大鼠中提取培养的 EPCs，并用免疫荧光法鉴定。苦参碱处理或转染 miR-126b 抑制剂和靶向叉头框 (FOXO) 4 的小干扰 RNA 后，通过细胞计数试剂盒-8 检测和流式细胞术检测 EPCs 的活力和凋亡。划痕、Transwell 和管形成试验检测迁移、侵袭和管形成能力。通过双荧光素酶报告基因检测预测 miR-126b 的靶基因，并通过验证。定量实时聚合酶链反应和 Western blot 检测 miR-126b、FOXO4、基质金属蛋白酶 (MMP) 2、MMP9 和血管内皮生长因子 (VEGF) A 的表达。
EPCs 成功提取和培养，分化（CD）34 和 CD133 阳性反应簇证实了这一点。苦参碱促进 EPCs 的活力、迁移、侵袭和管形成，同时抑制 EPCs 的凋亡，并上调 miR-126b 的表达。此外，miR-126b 抑制剂逆转了苦参碱对 EPCs 的作用，并下调了 MMP2、MMP9 和 VEGFA 的表达水平。miR-126b 靶向 FOXO4，siFOXO4 逆转了 miR-126b 抑制剂对 EPCs 的上述作用。
苦参碱通过调节 miR-126b/FOXO4 轴保护 EPCs 免于凋亡，并促进其迁移、侵袭和管形成能力。
