活化型c-Src对整合素αvβ3相关肿瘤行为的影响  

Effect of active c-Src on integrin αvβ3-related tumor behavior

作　　者：刘萍[1] 周玉兰[1] 黄建松[1] 崔雄鹰[1] 施小凤[1] 阮铮[1] 奚晓东[1] 

机构地区：[1]上海交通大学医学院附属瑞金医院上海血液学研究所 医学基因组学国家重点实验室,上海200025

出　　处：《诊断学理论与实践》2013年第4期419-425,共7页Journal of Diagnostics Concepts & Practice

基　　金：国家自然科学基金(81070414;81270594)

摘　　要：目的:探讨活化型c-Src对整合素αvβ3相关肿瘤行为的影响及其分子机制,为寻找治疗肿瘤的分子靶点提供思路。方法:通过定点突变的方法构建活化型c-Src真核表达载体,应用脂质体法转染至表达有整合素αvβ3的中国仓鼠卵巢(CHO)细胞,即CHO-αvβ3细胞中建立稳定细胞株。通过流式细胞术、蛋白印迹法检测细胞中目的蛋白的表达,检测稳定细胞株在固相化αvβ3配体玻连蛋白(Vn)上的黏附、伸展功能,利用人工重组基底膜侵袭小室,观察细胞株的迁移能力,并通过蛋白免疫印迹法检测细胞中整合素β3酪氨酸磷酸化的水平。结果:流式细胞术和蛋白印迹法检测结果均证实,CHO-αvβ3中成功表达活化型c-Src。活化型c-Src对CHO-αvβ3细胞在固相化Vn上的黏附、伸展能力无明显影响,但增强了细胞迁移能力及整合素β3的磷酸化水平。结论:活化型c-Src参与αvβ3相关肿瘤行为的调节,可能与增强整合素β3磷酸化水平相关,为进一步深入了解相关机制奠定基础。Objective： To investigate the effect of active c-Src on integrin ctv133-related tumor behavior and its molecular mechanism for providing insights to the searching of molecular targets for tumor therapy. Methods： Active mutant c-Sre （e-SrcY530F, in which the C-terminal regulatory tyrosine was replaced by a phenylalanine） was produced by site-directed mutagenesis and was transfected into CHO-αvβ3 cells via Lipofectamine 2000. Stable CHO cell line co- expressing human integrin ctv133 and active c-Src was established. The expression of integrin 133 and c-Src was detected by flow cytometry and Western blotting. Spreading and adhesion of the CHO-αvβ3 and CHO-αvβ3/SreYF cells on immobilized vitronectin were examined, and transwell assay was performed to evaluate migration ability. The phosphorylation of integrin 133 was detected by Western blotting. Results： Eukaryotie expression vector of active c-Src was constructed and transfected successfully into CHO-αvβ3 ceils. Active c-Src enhanced the migration ability on immobilized vitronectin and the level of integrin 133 phosphorylation, while had no effect on adhesion and spreading. Conclusions： Active c-Src is involved in regulating αvβ3-related tumor behavior, which might have a relationship with the enhancing of integrin 133 phosphorylation, and lay a foundation for further research on related mechanism.

关 键 词：活化型c-Src 整合素ΑVΒ3 迁移 黏附 磷酸化 

分 类 号：R730.1[医药卫生—肿瘤]