School of Pharmacy, Minzu University of China, Beijing 100081, PR China; Laboratory of Ethnomedicine of Ministry of Education, Minzu University of China, PR China.
Department of Biochemistry, Institute of Glycobiology, Dalian Medical University, Liaoning, Dalian 116044, PR China.
J Ethnopharmacol. 2018 Jun 12;219:161-172. doi: 10.1016/j.jep.2018.03.012. Epub 2018 Mar 12.
Gypenosides are major constituents in Gynostemma pentaphyllum (Thunb.) Makino. Previous studies have shown that gypenosides isolated from G. pentaphyllum possess inhibitory effect on the growth of cancer cells, especially A549 cells, with structure-activity relationship (SAR). However, the underlying mechanism of gypenoside-induced A549 cell death remains to be clarified.
To further investigate SAR and the underlying mechanism of gypenosides in A549 cells.
Gypenosides were isolated from G. pentaphyllum using chromatography methods and identified using MS and NMR data. The cytotoxicity was determined with CCK-8 assay. The effects of gypenosides on apoptosis, cell cycle and migration were investigated through cell morphology observation, flow cytometry analysis and key proteins detection.
Three gypenosides, 2α,3β,12β,20(S)-tetrahydroxydammar-24-ene-3-O-β-D-glucopyranoside-20-O-β-D-glucopyranoside, gypenoside L and gypenoside LI were isolated from G. pentaphyllum. Gypenoside stereoisomers, gypenoside L (S configuration at C20) and gypenoside LI (R configuration at C20) showed stronger activity against A549 cells. Furthermore, both induced A549 cell apoptosis through intrinsic and extrinsic pathways evidenced by reducing mitochondrial membrane potential (MMP), generating reactive oxygen species (ROS), releasing more cytochrome c and down-regulating procaspase 8. However, gypenoside L blocked A549 cells in G0/G1, while gypenoside LI induced G2/M arrest, which was further verified by different expression of CDK1, CDK2 and CDK4. In addition, both inhibited A549 cell migration, which was evidenced by down-regulation of MMP-2/9 as well as scratch wound assay and transwell assay.
C of gypenoside played an important role in A549 cell cytotoxicity and gypenoside stereoisomers could be used as potential multi-target chemopreventive agents for cancer.
绞股蓝皂苷是绞股蓝（Thunb.）Makino 的主要成分。先前的研究表明，从绞股蓝中分离出的绞股蓝皂苷对癌细胞的生长具有抑制作用，尤其是 A549 细胞，具有结构-活性关系（SAR）。然而，绞股蓝诱导 A549 细胞死亡的机制仍有待阐明。
进一步研究 SAR 以及绞股蓝皂苷在 A549 细胞中的作用机制。
采用色谱法从绞股蓝中分离出绞股蓝皂苷，并通过 MS 和 NMR 数据鉴定。用 CCK-8 法测定细胞毒性。通过细胞形态观察、流式细胞术分析和关键蛋白检测，研究绞股蓝皂苷对细胞凋亡、细胞周期和迁移的影响。
从绞股蓝中分离得到三种绞股蓝皂苷，2α,3β,12β,20(S)-四羟基达玛-24-烯-3-O-β-D-吡喃葡萄糖苷-20-O-β-D-吡喃葡萄糖苷、绞股蓝苷 L 和绞股蓝苷 LI。绞股蓝皂苷立体异构体，即 C20 构型为 S 的绞股蓝苷 L 和 C20 构型为 R 的绞股蓝苷 LI，对 A549 细胞的活性更强。此外，两者均通过降低线粒体膜电位（MMP）、产生活性氧物种（ROS）、释放更多细胞色素 c 和下调前半胱天冬酶 8，诱导 A549 细胞凋亡，通过内源性和外源性途径。然而，绞股蓝苷 L 阻止 A549 细胞进入 G0/G1 期，而绞股蓝苷 LI 诱导 G2/M 期阻滞，这进一步通过不同的 CDK1、CDK2 和 CDK4 表达得到证实。此外，两者均抑制 A549 细胞迁移，这表现在 MMP-2/9 的下调以及划痕实验和 Transwell 实验。
C 位的糖基在绞股蓝皂苷对 A549 细胞的细胞毒性中起着重要作用，绞股蓝苷立体异构体可作为癌症的潜在多靶化学预防剂。
