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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
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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 wordsBreast 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
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)
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