Please wait a minute...
浙江大学学报（医学版）
J Zhejiang Univ (Med Sci)  2019, Vol. 48 Issue (6): 631-637    DOI: 10.3785/j.issn.1008-9292.2019.12.07
    
Chloroxoquinoline inhibits invasion in breast cancer via down-regulating Rho/Rho kinase signaling pathway
LIU Jingwen1(),YANG Xinglian1,SHEN Kaili1,ZENG Linghui1,*(),SUN Yan2,*()
1. School of Medicine, Zhejiang University City College, Hangzhou 310015, China
2. Department of Oncology, Zhejiang Cancer Hospital, Hangzhou 310022, China
Download: HTML( 6 )   PDF(8655KB)
Export: BibTeX | EndNote (RIS)      

Abstract  

Objective: To investigate the effect of chloroxoquinoline on cytoskeleton of breast cancer cells and its relation with Rho/Rho kinase signaling pathway. Methods: Breast cancer Bcap37 and MDA-MB-453 cells were treated with different concentrations of chloroxoquinoline. Wound healing and Transwell assay were conducted to detect cell migration and invasion, respectively. Rhodamine-phalloidin staining and immunofluorescent staining were used to observe the polymerization state of F-actin and the expression of α-Tublin in breast cancer cells, respectively. Western blot was used to detect the phosphorylation level of key protein in Rho/Rho kinase signaling pathway. Results: Compared with the control group, chloroxoquinoline treatment induced dose-dependent decrease in cell migration and invasion, and Bcap37 and MDA-MB-453 cells treated with chloroxoquinoline showed dose-dependent changes in cell morphology and decrease in cell body. The staining of F-actin and α-Tublin was irregular and clustered. Furthermore, treatment of chloroxoquinoline down-regulated the phosphorylation of the Rho/Rho kinase signaling proteins Cofilin, Limk and Rock2 (all P < 0.01). Conclusions: Chloroxoquinoline inhibits the cytoskeleton in breast cancer Bcap37 and MDA-MB-453 cells and inhibits cell migration. This effect may be associated with down-regulation of Rho/Rho kinase signaling pathway.

Key words： Breast neoplasms/pathology      Cytoskeleton      rho-Associated kinases/antagonists & inhibitors      Signal transduction/drug effects      Actins      Tubulin      Antineoplastic agents/pharmacology     
Received: 15 July 2019      Published: 19 January 2020
CLC:  R737.9  
Corresponding Authors: ZENG Linghui,SUN Yan     E-mail: laby186640@outlook.com;zenglh@zucc.edu.cn;sunyan_19750205@163.com
Service
E-mail this article
Add to my bookshelf
Add to citation manager
E-mail Alert
RSS
Articles by authors
LIU Jingwen
YANG Xinglian
SHEN Kaili
ZENG Linghui
SUN Yan
Cite this article:

LIU Jingwen,YANG Xinglian,SHEN Kaili,ZENG Linghui,SUN Yan. Chloroxoquinoline inhibits invasion in breast cancer via down-regulating Rho/Rho kinase signaling pathway. J Zhejiang Univ (Med Sci), 2019, 48(6): 631-637.

URL:

http://www.zjujournals.com/med/10.3785/j.issn.1008-9292.2019.12.07     OR     http://www.zjujournals.com/med/Y2019/V48/I6/631


氯氧喹通过下调Rho/Rho激酶信号通路抑制乳腺癌细胞转移

目的: 通过观察氯氧喹对乳腺癌细胞系细胞骨架的聚合状态及其对Rho/Rho激酶信号通路相关靶点蛋白磷酸化的作用，明确氯氧喹治疗乳腺癌的作用机制。方法: 以不同浓度氯氧喹处理乳腺癌Bcap37和MDA-MB-453细胞系，划痕试验检测细胞迁移，Transwell试验检测细胞侵袭，罗丹明标记鬼笔环肽染色法观察F肌动蛋白的聚解状态，免疫荧光法观察α微管蛋白的表达，蛋白质印迹法检测Rho/Rho激酶信号通路关键蛋白磷酸化水平。结果: 氯氧喹可剂量依赖性地抑制细胞迁移和侵袭；Bcap37和MDA-MB-453细胞经氯氧喹处理后其细胞形态发生改变，细胞体积减少；F肌动蛋白和α微管蛋白染色形态不规则，荧光染色聚集成团，且呈现剂量依赖性；氯氧喹可下调Rho/Rho激酶信号通路中肌动蛋白解聚因子Cofilin、磷酸化蛋白激酶（Limk）、Rho关联卷曲蛋白激酶2（Rock2）磷酸化表达（均P < 0.01）。结论: 氯氧喹可抑制乳腺癌细胞骨架聚合从而抑制细胞迁移，该作用可能与抑制Rho/Rho激酶信号通路相关。


关键词： 乳腺肿瘤/病理学,  细胞骨架,  rho相关激酶类/拮抗剂和抑制剂,  信号传导/药物作用,  肌动蛋白类,  微管蛋白,  抗肿瘤药/药理学 
Fig 1 The results of cell scratch test in MDA-MB- 453 cells
Fig 2 The results of Transwell test in MDA-MB- 453 cells
Fig 3 Distribution of cytoskeletal actin in Bcap37 and MDA-MB- 453 cells (Phalloidin-iFluor 488)
Fig 4 Distribution of α-microtubulin in Bcap37 and MDA-MB- 453 cells (immunofluorescence staining)
Fig 5 Representative blots of protein expression of Rho/Rho kinase signaling in MDA-MB- 453 cells
Fig 6 Statistic analysis of phosphorylated protein expression of Rho/Rho kinase signaling protein in MDA-MB- 453 cells (n=6)
[1]   YOULDEN D R , CRAMB S M , YIP C H et al. Incidence and mortality of female breast cancer in the Asia-Pacific region[J]. Cancer Biol Med, 2014, 11 (2): 101- 115
[2]   SUNG H , ROSENBERG P S , CHEN W Q et al. Female breast cancer incidence among Asian and Western populations:more similar than expected[J]. J Natl Cancer Inst, 2015, 107 (7): pii:djv107
[3]   耿维凤 . 抗癌新药-氯氧喹胶囊[J]. 中国医院药学杂志, 2005, 25 (10): 994
GENG Weifeng . Anticancer drug-chloroxoquinoline capsule[J]. Chinese Journal of Hospital Pharmacy, 2005, 25 (10): 994
doi: 10.3321/j.issn:1001-5213.2005.10.055
[4]   孙燕, 冯琳迤, 雷蕾 et al. 氯氧喹对不同类型乳腺癌细胞系的抑制作用及机制[J]. 中国药理学通报, 2019, 35 (3): 347- 353
SUN Yan , FENG Linyi , LEI Lei et al. Inhibitory effect of chloroxoquinoline in different breast cancer cells[J]. Chinese Pharmacological Bulletin, 2019, 35 (3): 347- 353
doi: 10.3969/j.issn.1001-1978.2019.03.011
[5]   OLSON M F . Applications for ROCK kinase inhibition[J]. Current Opinion in Cell Biology, 2008, 20 (2): 242- 248
doi: 10.1016/j.ceb.2008.01.002
[6]   肖英, 程爱兰 . 细胞骨架在肿瘤侵袭转移中的研究进展[J]. 中国肿瘤临床, 2016, 43 (22): 1007- 1011
XIAO Ying , CHENG Ailan . Cell cytoskeleton-the movers in cancer metastasis[J]. Chinese Journal of Clinical Oncology, 2016, 43 (22): 1007- 1011
doi: 10.3969/j.issn.1000-8179.2016.22.828
[7]   ROTTNER K , STRADAL T E . Actin dynamics and turnover in cell motility[J]. Curr Opin Cell Biol, 2011, 23 (5): 569- 578
doi: 10.1016/j.ceb.2011.07.003
[8]   AYLETT C H , L?WE J , AMOS L A . New insights into the mechanisms of cytomotive actin and tubulin filaments[J]. Int Rev Cell Mol Biol, 2011, 292 1- 71
doi: 10.1016/B978-0-12-386033-0.00001-3
[9]   FRANCIS R, XU X, PARK H, et al. Connexin43 modulates cell polarity and directional cell migration by regulating microtubule dynamics[J/OL]. PLoS One, 2011, 6(10): e26379.
[10]   KATSETOS C D , DRáBEROVá E , LEGIDO A et al. Tubulin targets in the pathobiology and therapy of glioblastoma multiforme. Ⅱ. γ-Tubulin[J]. J Cell Physiol, 2009, 221 (3): 514- 520
doi: 10.1002/jcp.21884
[11]   PRICE L S , COLLARD J G . Regulation of the cytoskeleton by Rho-familyGTPases:implications for tumour cell invasion[J]. Semin Cancer Biol, 2001, 11 (2): 167- 173
doi: 10.1006/scbi.2000.0367
[12]   TAKAHASHI N , NOBUSUE H , SHIMIZU T et al. ROCK inhibition induces terminal adipocyte differentiation and suppresses tumorigenesis in chemoresistant osteosarcoma cells[J]. Cancer Res, 2019, 79 (12): 3088- 3099
doi: 10.1158/0008-5472.CAN-18-2693
[13]   CASCIONE M , DE MATTEIS V , TOMA C C et al. Morphomechanical and structural changes induced by ROCK inhibitor in breast cancer cells[J]. Exp Cell Res, 2017, 360 (2): 303- 309
doi: 10.1016/j.yexcr.2017.09.020
[14]   HUANG G X , WANG Y , SU J et al. Up-regulation of Rho-associated kinase 1/2 by glucocorticoids promotes migration, invasion and metastasis of melanoma[J]. Cancer Lett, 2017, 410 1- 11
doi: 10.1016/j.canlet.2017.09.005
[1] XU A'qiao,HE Hongqin,SHI Qiujun,LI Zhiqing,ZHANG Shengjian. Digital breast tomosynthesis in diagnosis of dense breast lesions[J]. J Zhejiang Univ (Med Sci), 2019, 48(2): 186-192.
[2] PAN Jingying, HE Mengye, KE Wei, HU Menglin, WANG Meifang, SHEN Peng. Advances on correlation of PET-CT findings with breast cancer molecular subtypes, treatment response and prognosis[J]. J Zhejiang Univ (Med Sci), 2017, 46(5): 473-480.
[3] CHEN Zhiqiang, WANG Ying, MI Xianjun, DUAN Lifeng, CHEN Ang, HUANG Huayong. Volume of 4% neutral buffered formaldehyde affects the results of HER2 gene detected by fluorescence in situ hybridization in primary invasive breast cancer[J]. J Zhejiang Univ (Med Sci), 2017, 46(4): 439-444.
[4] TIAN Hua, CHEN Yang, ZHAO Jiangang, LIU Daren, LIANG Gang, GONG Weihua, CHEN Li, WU Yulian. Effects of siRNAs targeting CD97 immune epitopes on biological behavior in breast cancer cell line MDA-MB231[J]. J Zhejiang Univ (Med Sci), 2017, 46(4): 341-348.
[5] XIA Guangfa, ZHU Juanying, YUAN Jun, CAO Bo, TANG Jie, CHEN Yiding. Efficacy of adjuvant endocrine therapy in breast cancer patients with a positive-to-negative switch of hormone receptor status after neoadjuvant chemotherapy[J]. J Zhejiang Univ (Med Sci), 2016, 45(6): 614-619.
[6] WANG Jiaochen, WENG Shouxiang, JIN Xiaofen, YU Wenjie, ZHOU Tao, GAN Meifu. Neuroendocrine differentiation and Wilms' tumor protein-1 expression in breast mucinous carcinoma and their significance[J]. J Zhejiang Univ (Med Sci), 2016, 45(1): 45-50.
[7] CAO Pei-long，et al. Expressions of FOXC1 and MMP-7 in molecular subtypes of breast cancer and their association with clinicopathological characteristics [J]. J Zhejiang Univ (Med Sci), 2014, 43(4): 406-412.
[8] ZHOU Mei-qi，et al. Prospect and guideline update of sentinel lymph node biopsy for patients with early-stage breast carcinoma[J]. J Zhejiang Univ (Med Sci), 2014, 43(4): 381-387.
[9] ZHANG Ru-yi，DING Yue-min，ZHANG Li-huang. LIM kinases and their roles in the nervous system[J]. J Zhejiang Univ (Med Sci), 2014, 43(1): 119-125.
[10] LIU Jia，WANG Hongyan，DENG Yuan，CAO Peilong，LI Xiaofeng. Expression of EZH2 in breast cancer and its clinicopathological significance[J]. J Zhejiang Univ (Med Sci), 2013, 42(4): 443-.
[11] XU Shui-Ling, XU Yan, HUANG Jia, FAN Hong-Yan, JIN Meng-Mei. Role of phospholipase C in cytoskeleton rearrangements of dendritic cells invaded by Mycobacterium tuberculosis[J]. J Zhejiang Univ (Med Sci), 2013, 42(2): 184-191.
[12] . Effects of snakegourd root polysaccharide on apoptosis of MCF-7 cells[J]. J Zhejiang Univ (Med Sci), 2012, 41(5): 527-534.
[13] . Effect of Evn-50 on cell growth and apoptosis in tamoxifen-resistance human breast cancer cell line MCF-7/TAM-R[J]. J Zhejiang Univ (Med Sci), 2012, 41(5): 498-505.
[14] . Effects of 8-Methoxypsoralen on intracellular [Ca2+]i and cytoskeleton actin organization in human melanocytes in vitro[J]. J Zhejiang Univ (Med Sci), 2009, 38(4): 348-351.