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浙江大学学报（医学版）
浙江大学学报（医学版）  2019, Vol. 48 Issue (6): 631-637    DOI: 10.3785/j.issn.1008-9292.2019.12.07
原著     
氯氧喹通过下调Rho/Rho激酶信号通路抑制乳腺癌细胞转移
刘婧雯1(),杨兴莲1,沈凯莉1,曾玲晖1,*(),孙燕2,*()
1. 浙江大学城市学院医学院, 浙江 杭州 310015
2. 浙江省肿瘤医院综合肿瘤内科, 浙江 杭州 310022
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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摘要：

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    
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
收稿日期: 2019-07-15 出版日期: 2020-01-19
:  R737.9  
基金资助: 浙江省医药卫生科技计划(2015RCB004);杭州市留学回国人员资助项目(2018)
通讯作者: 曾玲晖,孙燕     E-mail: laby186640@outlook.com;zenglh@zucc.edu.cn;sunyan_19750205@163.com
作者简介: 刘婧雯(1999—), 女, 大学生, 主要从事药理学研究, E-mail:laby186640@outlook.com; https://orcid.org/0000-0002-4710-1285
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引用本文:

刘婧雯,杨兴莲,沈凯莉,曾玲晖,孙燕. 氯氧喹通过下调Rho/Rho激酶信号通路抑制乳腺癌细胞转移[J]. 浙江大学学报（医学版）, 2019, 48(6): 631-637.

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.

链接本文:

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

图 1  各组MDA-MB- 453细胞划痕试验结果
图 2  各组MDA-MB- 453细胞Transwell小室试验结果
图 3  各组Bcap37细胞和MDA-MB- 453细胞骨架肌动蛋白分布情况(鬼笔环肽染色)
图 4  各组Bcap37细胞和MDA-MB- 453细胞α微管蛋白分布情况(免疫荧光染色)
图 5  各组MDA-MB- 453细胞中Rho/Rho激酶信号通路蛋白表达电泳图
图 6  各组MDA-MB- 453细胞中Rho/Rho激酶信号通路蛋白磷酸化水平比较(n=6)
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