Department of Pharmacognosy, University of Vienna, Althanstraße 14, Vienna 1090, Austria; vasopharm GmbH, Friedrich-Bergius-Ring 15, 97076 Würzburg, Germany.
Department of Pharmacognosy, University of Vienna, Althanstraße 14, Vienna 1090, Austria.
Phytomedicine. 2019 Jul;60:152938. doi: 10.1016/j.phymed.2019.152938. Epub 2019 Apr 23.
Tylophorine (TYL) is an alkaloid with antiproliferative action in cancer cells. Vascular smooth muscle cell (VSMC) proliferation and neointima formation contribute to restenosis after percutaneous coronary interventions.
HYPOTHESIS/PURPOSE: Our goal was to examine the potential of TYL to inhibit VSMC proliferation and migration, and to dissect underlying signaling pathways.
TYL was administered to platelet-derived growth factor (PDGF-BB)-stimulated, serum-stimulated, quiescent and unsynchronized VSMC of rat and human origin. BrdU incorporation and resazurin conversion were used to assess cell proliferation. Cell cycle progression was analyzed by flow cytometry of propidium iodide-stained nuclei. Expression profiles of proteins and mRNAs were determined using western blot analysis and RT-qPCR. The Click-iT OPP Alexa Fluor 488 assay was used to monitor protein biosynthesis.
TYL inhibited PDGF-BB-induced proliferation of rat aortic VSMCs by arresting cells in G1 phase of the cell cycle with an IC of 0.13 µmol/l. The lack of retinoblastoma protein phosphorylation and cyclin D1 downregulation corroborated a G1 arrest. Inhibition of proliferation and cyclin D1 downregulation were species- and stimulus-independent. TYL also decreased levels of p21 and p27 proteins, although at later time points than observed for cyclin D1. Co-treatment of VSMC with TYL and MG132 or cycloheximide (CHX) excluded proteasome activation by TYL as the mechanism of action. Comparable time-dependent downregulation of cyclin D1, p21 and p27 in TYL- or CHX-treated cells, together with decreased protein synthesis observed in the Click-iT assay, suggests that TYL is a protein synthesis inhibitor. Besides proliferation, TYL also suppressed migration of PDGF-activated VSMC. In a human saphenous vein organ culture model for graft disease, TYL potently inhibited intimal hyperplasia.
This unique activity profile renders TYL an interesting lead for the treatment of vasculo-proliferative disorders, such as restenosis.
千里光菲碱（TYL）是一种具有抗癌细胞增殖作用的生物碱。血管平滑肌细胞（VSMC）增殖和新生内膜形成是经皮冠状动脉介入治疗后再狭窄的原因。
假设/目的：我们的目标是研究 TYL 抑制 VSMC 增殖和迁移的潜力，并剖析潜在的信号通路。
将 TYL 给予血小板衍生生长因子（PDGF-BB）刺激、血清刺激、静止和未同步的大鼠和人源性 VSMC。通过 BrdU 掺入和 Resazurin 转化来评估细胞增殖。通过碘化丙啶染色核的流式细胞术分析细胞周期进程。使用 Western blot 分析和 RT-qPCR 确定蛋白质和 mRNA 的表达谱。使用 Click-iT OPP Alexa Fluor 488 测定法监测蛋白质生物合成。
TYL 通过将细胞周期阻滞在 G1 期，以 0.13µmol/L 的 IC 抑制 PDGF-BB 诱导的大鼠主动脉 VSMC 增殖。视网膜母细胞瘤蛋白磷酸化和 cyclin D1 下调的缺失证实了 G1 期阻滞。增殖抑制和 cyclin D1 下调与物种和刺激无关。TYL 还降低了 p21 和 p27 蛋白的水平，尽管比 cyclin D1 观察到的时间晚。TYL 与 MG132 或环己酰亚胺（CHX）共同处理 VSMC 可排除 TYL 的作用机制是通过蛋白酶体激活。在 TYL 或 CHX 处理的细胞中观察到 cyclin D1、p21 和 p27 的类似时依赖性下调，以及在 Click-iT 测定中观察到的蛋白质合成减少，表明 TYL 是一种蛋白质合成抑制剂。除了增殖，TYL 还抑制了 PDGF 激活的 VSMC 的迁移。在用于移植疾病的人隐静脉器官培养模型中，TYL 强力抑制内膜增生。
这种独特的活性谱使 TYL 成为治疗血管增殖性疾病（如再狭窄）的有趣先导化合物。
