Fig. 1 Cell index (CI) values of respective treatment groups on A549 cells at different time points (5th, 10th, 15th, and 20th hours)
Scan for full text
Department of Biomedical Science, Faculty of Medicine, University of Malaya, Kuala Lumpur 50603, Malaysia
1.Department of Biomedical Science, Faculty of Medicine, University of Malaya, Kuala Lumpur 50603, Malaysia
2.School of Biosciences, Taylor’s University, Lakeside Campus, Subang Jaya, Selangor Darul Ehsan 47500, Malaysia
3.Forest Research Institute Malaysia, Kepong, Selangor Darul Ehsan 52109, Malaysia
Published: 2020-09 ,
Received: 27 May 2020 ,
Accepted: 16 July 2020
Cite this article
Mee Lee LOOI, Alwyn Khai Howe Wong, Shelly Anne Gnapragasan, et al. Anti-migratory effects of
Mee Lee LOOI, Alwyn Khai Howe Wong, Shelly Anne Gnapragasan, et al. Anti-migratory effects of
Department of Biomedical Science, Faculty of Medicine, University of Malaya, Kuala Lumpur 50603, Malaysia
Piper betle (PB), also known as “betel” in Malay language, is a tropical Asian vine. PB leaves are commonly chewed by Asians along with betel quid. It contains phenols such as eugenol and hydroxychavicol along with chlorophyll, β-carotene, and vitamin C (Salehi et al.,
本文研究了槟榔(Piper betle,PB)水提取物对癌细胞转移能力的调节作用及其对微管蛋白结构和网络的影响.采用亚毒剂量对PB和5-氟尿嘧啶(5-FU)处理的癌细胞进行抗迁移研究,观察细胞迁移20小时.在所有处理组中,100 µg/mL PB处理的癌细胞均显示出最大的抗迁移作用(P=0.016).总体上,PB对癌细胞的抗迁移作用高于5-FU.通过细胞形态观察发现,PB处理的细胞表现出与标准微管抑制剂(紫杉醇)相似的细胞特征,并在PB和紫杉醇处理的癌细胞中发现M期细胞群.通过对微管结构和网络进一步研究,我们发现PB和紫杉醇处理的癌细胞表现出长期破坏的纺锤体.因此,我们认为PB具有对癌细胞的抗迁移作用,并可能改变蛋白结构和网络.
Piper betle;
Cell migration;
Microtubules;
Tubulin network
Piper betle (PB), also known as “betel” in Malay language, is a tropical Asian vine. PB leaves are commonly chewed by Asians along with betel quid. It contains phenols such as eugenol and hydroxychavicol along with chlorophyll, β-carotene, and vitamin C (Salehi et al.,
The ability of cancer cells to disseminate to distant regions is partly the result of microtubule dynamics. Microtubules are part of the cell cytoskeleton, contribute to shape and dynamics, and are essential for cell movement and directionality (Ganguly et al.,
The sub-toxic levels of PB and 5-FU were used to ensure no cell cytotoxicity in the cell migration analysis. After 24 h of respective treatment, values of IC20 (20% inhibitory concentration) and IC30 (30% inhibitory concentration) recorded for PB on human lung adenocarcinoma (A549) cells were 20 and 100 μg/mL, respectively, whereas IC20 and IC30 for 5-FU on A549 cells were determined at 4.0 and 12.5 μmol/L, respectively. The anti-migration effects of PB and 5-FU were observed real-time on A549 cells for 20 h using xCELLigence (ACEA Biosciences, USA). A high cell index (CI) value indicates a positive migration effect as more cells adhere to the gold microelectrodes, causing high impedance. The negative control group (cells with serum-free media) showed CI values less than 0.5 (Fig.
Fig. 1 Cell index (CI) values of respective treatment groups on A549 cells at different time points (5th, 10th, 15th, and 20th hours)
5-FU: 5-fluorouracil; PB: Piper betle. *P<0.05, compared with the positive control group; #P<0.05, compared with the negative control group
After 5 h of respective treatments, no significant differences in CI values were seen between groups (F=3.107, P=0.060). However, significant differences were observed between treatment groups after 10 h (F=6.114, P=0.008), 15 h (F=10.984, P=0.001), and 20 h (F=10.555, P=0.001). Indeed, after 10 h, cells in the positive control wells migrated towards the chemo-attractant, leading to high CI values (Fig.
We further observed cell morphology, structure, and the tubulin network in PB-, paclitaxel-, and 5-FU-treated human colorectal adenocarcinoma cells (HT29). We observed apoptotic cells in HT29 treated with high-dose 5-FU (Fig.
Fig. 2 Cell morphologies and microtubule networks of untreated, 10 μg/mL paclitaxel-, 130 μmol/L 5-FU-, and 200 μg/mL PB-treated HT29 cells after 24 h
(a, b) Untreated; (c, d) 10 μg/mL paclitaxel; (e, f) 130 μmol/L 5-FU; (g, h) 200 μg/mL PB. Cell morphologies were observed under 400×magnification. Red arrows indicate apoptotic cells, while green arrows indicate M phase cells. Microtubule networks were observed under 1000×magnification. Tubulin was stained with anti-tubulin antibody at 1:1000 dilution followed by goat anti-mouse IgG conjugated with DyLight 488 at 1:1000 dilution (green signal). Nuclei were counterstained with DAPI (blue signal). Paclitaxel-and PB-treated HT29 cells have long spindles (yellow arrows). 5-FU: 5-fluorouracil; PB: Piper betle; DAPI: 4',6-diamidino-2-phenylindole, dihydrochloride
It has previously been reported that microtubule-targeting drugs inhibit cell motility, triggering both anti-migratory and anti-angiogenic effects (Belotti et al.,
Detailed methods are provided in the electronic supplementary materials of this paper.
We thank Taylor’s University and Forest Research Institute Malaysia (FRIM), Malaysia, for the laboratory facilities.
Compliance with ethics guidelines
Mee Lee LOOI, Alwyn Khai Howe WONG, Shelly Anne GNAPRAGASAN, Anis Zafirah JAPRI, Aiysvariyah RAJEDADRAM, and Kar Yong PIN declare that they have no conflict of interest.
This article does not contain any studies with human or animal subjects performed by any of the authors.
D Belotti, V Vergani, T Drudis, 等. The microtubule-affecting drug paclitaxel has antiangiogenic activity. Clin Cancer Res, 1996. 2(11):1843-1849. 9816139. [Baidu Scholar]
AL Blajeski, VA Phan, TJ Kottke, 等. G1 and G2 cell-cycle arrest following microtubule depolymerization in human breast cancer cells. J Clin Invest, 2002. 110(1):91-99. 12093892DOI:10.1172/JCI13275. [Baidu Scholar]
C Cadart, E Zlotek-Zlotkiewicz, M le Berre, 等. Exploring the function of cell shape and size during mitosis. Dev Cell, 2014. 29(2):159-169. 24780736DOI:10.1016/j.devcel.2014.04.009. [Baidu Scholar]
A Gandalovičová, D Rosel, M Fernandes, 等. Migrastatics–anti-metastatic and anti-invasion drugs: promises and challenges. Trends Cancer, 2017. 3(6):391-406. 28670628DOI:10.1016/j.trecan.2017.04.008. [Baidu Scholar]
A Ganguly, HL Yang, R Sharma, 等. The role of microtubules and their dynamics in cell migration. J Biol Chem, 2012. 287(52):43359-43369. 23135278DOI:10.1074/jbc.M112.423905. [Baidu Scholar]
A Ganguly, F Cabral, HL Yang, 等. Peloruside A is a microtubule-stabilizing agent with exceptional anti-migratory properties in human endothelial cells. Oncoscience, 2015. 2(6):585-595. 26244166. [Baidu Scholar]
A Guha Majumdar, M Subramanian. Hydroxychavicol from Piper betle induces apoptosis, cell cycle arrest, and inhibits epithelial-mesenchymal transition in pancreatic cancer cells. Biochem Pharmacol, 2019. 166274-291. 31154000DOI:10.1016/j.bcp.2019.05.025. [Baidu Scholar]
C Hayot, O Debeir, P van Ham, 等. Characterization of the activities of actin-affecting drugs on tumor cell migration. Toxicol Appl Pharmacol, 2006. 211(1):30-40. 16005926DOI:10.1016/j.taap.2005.06.006. [Baidu Scholar]
E Mukhtar, VM Adhami, H Mukhtar. Targeting microtubules by natural agents for cancer therapy. Mol Cancer Ther, 2014. 13(2):275-284. 24435445DOI:10.1158/1535-7163.MCT-13-0791. [Baidu Scholar]
PL Ng, NF Rajab, SM Then, 等. Piper betle leaf extract enhances the cytotoxicity effect of 5-fluorouracil in inhibiting the growth of HT29 and HCT116 colon cancer cells. J Zhejiang Univ-Sci B (Biomed & Biotechnol), 2014. 15(8):692-700. DOI:10.1631/jzus.B1300303. [Baidu Scholar]
EA Perez. Microtubule inhibitors: differentiating tubulin-inhibiting agents based on mechanisms of action, clinical activity, and resistance. Mol Cancer Ther, 2009. 8(8):2086-2095. 19671735DOI:10.1158/1535-7163.Mct-09-0366. [Baidu Scholar]
B Salehi, ZA Zakaria, R Gyawali, 等. Piper species: a comprehensive review on their phytochemistry, biological activities and applications. Molecules, 2019. 24(7):1364DOI:10.3390/molecules24071364. [Baidu Scholar]
M Shakibaei, P Kraehe, B Popper, 等. Curcumin potentiates antitumor activity of 5-fluorouracil in a 3D alginate tumor microenvironment of colorectal cancer. BMC Cancer, 15:250, 2015. DOI:10.1186/s12885-015-1291-0. [Baidu Scholar]
M Théry, M Bornens. Get round and stiff for mitosis. HFSP J, 2008. 2(2):65-71. 19404473DOI:10.2976/1.2895661. [Baidu Scholar]
PF Wu, HC Tseng, CC Chyau, 等. Piper betle leaf extracts induced human hepatocellular carcinoma HEP3B cell death via MAPKS regulating the p73 pathway in vitro and in vivo. Food Funct, 2014. 5(12):3320-3328. 25371988DOI:10.1039/C4FO00810C. [Baidu Scholar]
244
Views
2
Downloads
0
CSCD
Publicity Resources
Related Articles
Related Author
Related Institution