Raptor通过Wnt3a/β-catenin信号通路对乳腺癌侵袭与转移的作用

目的探讨Raptor是否可以通过Wnt3a/β-catenin信号通路促进乳腺癌细胞的侵袭与转移。方法采用免疫印迹法(Western blotting)检测乳腺癌细胞MCF-7和MDA-MB-231(MDA231)中Raptor蛋白的表达情况;采用小RNA(RNAi)干扰技术将质粒转染MDA231,将过表达质粒转染MCF-7;趋化运动实验检测乳腺癌细胞的运动能力;Transwell侵袭实验检测乳腺癌细胞的侵袭能力;Western blotting检测上皮细胞-间充质转化(EMT)标志物的表达情况;核质分离实验检测β-联蛋白(β-catenin)的转核情况。结果 Western blotting法检测结果显示,Raptor在低转移细胞系MCF-7中低表达,而在高转移细胞系MDA231中高表达。转染后,SiRaptor/MDA231细胞组中Raptor蛋白的表达明显下降,MCF-7/Raptor细胞组中Raptor蛋白的表达明显升高,表明转染成功。用Wnt3a刺激处理后,趋化运动和Transwell侵袭实验结果显示,SiRaptor/MDA231细胞组运动和侵袭能力明显降低,MCF-7/Raptor细胞组运动和侵袭能力明显增强。Western blotting法检测结果显示,用Wnt3a和抑制剂Dkk1不同处理后,SiRaptor/MDA231细胞组中上皮-钙黏蛋白(E-cadherin)上调,波形蛋白(vimentin)下调,细胞核中β-catenin的表达降低,而MCF-7/Raptor细胞组中E-cadherin蛋白下调,vimentin蛋白上调,β-catenin蛋白的表达升高。结论 Raptor能通过Wnt3a/β-catenin信号通路促进EMT的发生,进而促进乳腺癌细胞的侵袭与转移。

Raptor promoting invasion and metastasis of breast cancer through the signaling pathway Wnt3a/β-catenin

Objective To explore whether the Raptor can promote invasion and metastasis of breast cancer cells through Wnt3a/-catenin signaling pathway. Methods Using Western blotting to detect the protein expression of Raptor in breast cancer cells MDA-MB-231(MDA231) and MCF-7; Using RNAi technology to transfect small RNA interference plasmid into MDA231 breast cancer cells transiently. Overexpression of plasmid was transfected into MCF-7 breast cancer cells. Transwell and chemotaxis assay were used to detect the invasion and chemotaxis ability of breast cancer cells. Western blotting detected the expression of epithelial-mesenchymal transition(EMT) markers; Karyoplasm separation experiment tested the expression of β-catenin in nuclei. Results Western blotting result showed that the expression of Raptor was low in MCF-7, and high in MDA231. The protein expression of Raptor in SiRaptor/MDA231 was decreased obviously, and the protein expression of Raptor in MCF-7/Raptor was obviously higher. Transwell invasion and chemotaxis assay result showed that the invasion and chemotaxis ability of SiRaptor/MDA231 was decreased with the stimulation of Wnt3a, however, its capability of invasion and chemotaxis of MCF-7/Raptor was significantly enhanced. According to the result of Western blotting method with unused Wnt3a or at the same time stimulated with Wnt3a and Dkk1, the expression of E-cadherin was higher, and the expressions of vimentin and β-catenin were reduced in SiRaptor/MDA231, while in the MCF-7/Raptor, the expression of E-cadherin was reduced, and the expressions of vimentin and β-catenin were increased. Conclusion Through Wnt3a/β-catenin signaling pathway Raptor promotes the occurrence of EMT, thus to promote the invasion and metastasis of breast cancer cells.