Abstract：
BACKGROUND: Restenosis frequently occurs after percutaneous angioplasty in patients with vascular occlusion and seriously threatens their health. Substantial evidence has revealed that preventing vascular smooth muscle cell proliferation using a drug-eluting stent is an effective approach to improve restenosis. Cucurbitacins have been demonstrated to exert an anti-proliferation effect in various tumors and a hypotensive effect. This study aims to investigate the role of cucurbitacins extracted from Cucumis melo L. (CuECs) and cucurbitacin B (CuB) on restenosis.
METHODS: C57BL/6 mice were subjected to left carotid artery ligation and subcutaneously injected with CuECs or CuB for 4 weeks. Hematoxylin-Eosin, immunofluorescence and immunohistochemistry staining were used to evaluate the effect of CuECs and CuB on neointimal hyperplasia. Western blot, real-time PCR, flow cytometry analysis, EdU staining and cellular immunofluorescence assay were employed to measure the effects of CuECs and CuB on cell proliferation and the cell cycle in vitro. The potential interactions of CuECs with cyclin A2 were performed by molecular docking.
RESULTS: The results demonstrated that both CuECs and CuB exhibited significant inhibitory effects on neointimal hyperplasia and proliferation of vascular smooth muscle cells. Furthermore, CuECs and CuB mediated cell cycle arrest at the S phase. Autodocking analysis demonstrated that CuB, CuD, CuE and CuI had high binding energy for cyclin A2. Our study also showed that CuECs and CuB dramatically inhibited FBS-induced cyclin A2 expression. Moreover, the expression of cyclin A2 in CuEC- and CuB-treated neointima was downregulated.
CONCLUSIONS: CuECs, especially CuB, exert an anti-proliferation effect in VSMCs and may be potential drugs to prevent restenosis.
METHODS: C57BL/6 mice were subjected to left carotid artery ligation and subcutaneously injected with CuECs or CuB for 4 weeks. Hematoxylin-Eosin, immunofluorescence and immunohistochemistry staining were used to evaluate the effect of CuECs and CuB on neointimal hyperplasia. Western blot, real-time PCR, flow cytometry analysis, EdU staining and cellular immunofluorescence assay were employed to measure the effects of CuECs and CuB on cell proliferation and the cell cycle in vitro. The potential interactions of CuECs with cyclin A2 were performed by molecular docking.
RESULTS: The results demonstrated that both CuECs and CuB exhibited significant inhibitory effects on neointimal hyperplasia and proliferation of vascular smooth muscle cells. Furthermore, CuECs and CuB mediated cell cycle arrest at the S phase. Autodocking analysis demonstrated that CuB, CuD, CuE and CuI had high binding energy for cyclin A2. Our study also showed that CuECs and CuB dramatically inhibited FBS-induced cyclin A2 expression. Moreover, the expression of cyclin A2 in CuEC- and CuB-treated neointima was downregulated.
CONCLUSIONS: CuECs, especially CuB, exert an anti-proliferation effect in VSMCs and may be potential drugs to prevent restenosis.
摘要：
背景：血管闭塞患者经皮血管成形术后经常出现再狭窄，严重威胁其健康。大量证据表明，使用药物洗脱支架预防血管平滑肌细胞增殖是改善再狭窄的有效方法。已经证明葫芦素在各种肿瘤中发挥抗增殖作用和降压作用。本研究旨在探讨从甜瓜中提取的葫芦素（CuECs）和葫芦素B（CuB）对再狭窄的作用。
方法:C57BL/6小鼠进行左颈动脉结扎并皮下注射CuECs或CuB4周。苏木精-伊红,免疫荧光和免疫组织化学染色用于评估CuECs和CuB对新生内膜增生的影响。蛋白质印迹,实时PCR，流式细胞术分析,采用EdU染色和细胞免疫荧光法检测CuECs和CuB对细胞增殖和细胞周期的影响。CuEC与细胞周期蛋白A2的潜在相互作用通过分子对接进行。
结果：结果表明，CuECs和CuB均对血管平滑肌细胞的新生内膜增生和增殖具有明显的抑制作用。此外，CuECs和CuB介导的细胞周期阻滞在S期。自动对接分析表明，CuB，CuD,CuE和CuI对细胞周期蛋白A2具有高结合能。我们的研究还显示CuECs和CuB显著抑制FBS诱导的细胞周期蛋白A2表达。此外，cyclinA2在CuEC和CuB处理的新内膜中的表达下调。
结论：CuEC，尤其是CuB,在VSMC中发挥抗增殖作用，可能是预防再狭窄的潜在药物。
方法:C57BL/6小鼠进行左颈动脉结扎并皮下注射CuECs或CuB4周。苏木精-伊红,免疫荧光和免疫组织化学染色用于评估CuECs和CuB对新生内膜增生的影响。蛋白质印迹,实时PCR，流式细胞术分析,采用EdU染色和细胞免疫荧光法检测CuECs和CuB对细胞增殖和细胞周期的影响。CuEC与细胞周期蛋白A2的潜在相互作用通过分子对接进行。
结果：结果表明，CuECs和CuB均对血管平滑肌细胞的新生内膜增生和增殖具有明显的抑制作用。此外，CuECs和CuB介导的细胞周期阻滞在S期。自动对接分析表明，CuB，CuD,CuE和CuI对细胞周期蛋白A2具有高结合能。我们的研究还显示CuECs和CuB显著抑制FBS诱导的细胞周期蛋白A2表达。此外，cyclinA2在CuEC和CuB处理的新内膜中的表达下调。
结论：CuEC，尤其是CuB,在VSMC中发挥抗增殖作用，可能是预防再狭窄的潜在药物。
