Costunolide (Costus lactone) has anti-inflammatory and anti-oxidant properties and mediates apoptosis.

Telomerase (MCF-7 cells):90 μM, Telomerase (MDA-MB-231 cells):65 μM, FPTase:20 μM

Costunolide可降低小鼠角膜微囊模型中由乙烯基酯树脂鳞片胶泥诱导的新生血管形成.Costunolide通过阻断血管生长因子信号转导通路来抑制血管生成.

Costunolide连续处理人肿瘤细胞(A549,SK-OV-3,SK-MEL-2,XF498和HCT-1)48小时,可以剂量依赖性抑制细胞增殖。Costunolide通过ROS介导的线粒体通透性转换和细胞色素C在HL-60人类白血病细胞中释放细胞溶质来诱导细胞凋亡。Costunolide具有明显的抗真菌活性,包括毛癣菌 、猴毛癣菌,红色毛癣菌等。Costunolide抑制MCF-7和MDA-MB-231端粒酶活性和细胞增殖,该抑制作用呈浓度和时间依赖性。Costunolide也剂量依赖性抑制由法尼基蛋白转移酶诱导的人核纤层蛋白B法尼基化过程。

Telomerase activity assay: The telomerase activity is measured by the TRAP assay using the TRAPez Telomerase Detection Kit, which includes primers of a 36-bp internal control (IC) for quantifying the amplification of telomerase activity within a linear range close to 2.5 logs. For RNase treatment, 10μL of extract are incubated with 1μg of RNase at 37 °C for 20 minutes. The products of the TRAP assay are resolved by electrophoresis in a nondenaturing12% PAGE in a buffer containing 0.5 × Tris–borate EDTA and detected by autoradiograph. For quantification of TRAP products, the dried gels are exposed to Fuji Imaging Plate at room temperature. Results are corrected for background, and a standard value of 100 is given to the untreated control cell signal. Signal intensities of Costunolide-treated cells are compared to the standard and are expressed as a fraction of the maximum value of 100. [1]

1) Plate 500-10,000 cells in 200 μL media per well in a 96 well plate. Leave 8 wells empty for blank controls. 2) Incubate (37 °C, 5% CO2) overnight to allow the cells to attach to the wells. 3) Add 2 μL of Costunolide dissolved in DMSO to each well. Place on a shaking table, 150 rpm for 5 minutes, to thoroughly mix the samples into the media. 5) Incubate (37 °C, 5% CO2) for 48 hours to allow Costunolide to take effect. 6) Make 2 mL or more of MTT solution per 96 well plate at 5 mg/mL in PBS. Do not make a stock as MTT in solution is not stable long-term. 7) Add 20 μL MTT solution to each well. Place on a shaking table, 150 rpm for 5 minutes, to thoroughly mix the MTT into the media. 8) Incubate (37 °C, 5% CO2) for 1-5 hours to allow the MTT to be metabolized. 9) Dump off the media. (Dry plate on paper towels to remove residue if necessary. 10) Resuspend formazan (MTT metabolic product) in 200 μL DMSO. Place on a shaking table, 150 rpm for 5 minutes, to thoroughly mix the formazan into the solvent. 11) Read optical density at 560 nm and subtract background at 670 nm. Optical density should be directly correlated with cell quantity. (Only for Reference)