Carter W Bradford, Niu Guilian, Ward Michael D, Small Gregory, Hahn Julianne E, Muffly Barbara J
Don & Erika Wallace Comprehensive Breast Program at H. Lee Moffitt Cancer Center and Research Institute and Department of Interdisciplinary Oncology, University of South Florida, Tampa, FL, USA.
Ann Surg Oncol. 2007 Oct;14(10):2971-8. doi: 10.1245/s10434-007-9442-4. Epub 2007 Jun 26.
HER2 overexpression imparts a metastatic advantage in breast cancer. We have shown that HER2 signaling in breast cancer cells induces adjacent endothelial cell (EC) retraction, disrupting endothelial integrity. Because endothelial integrity is dependent on the adherens junctions, we postulated that the mechanism of tumor cell-induced EC retraction involves dissociation of catenin proteins from vascular endothelial (VE) cadherin. In this study, we report a loss of VE-cadherin in tumor-associated EC. We also tested for a change of catenin dissociation from VE-cadherin by manipulating HER2 signaling in tumor cells.
We tested confluent monolayers of human EC for downregulation of VE cadherin and dissociation of catenins from VE cadherin after exposure to breast cancer cells or conditioned media. Using immunoprecipitation, we quantitated the remaining complexed catenins to VE-cadherin in tumor-associated EC after different treatments to manipulate HER2 signaling.
Treatment of EC with conditioned media from MCF-7 cells expressing HER2 induced a loss of VE-cadherin expression, and time-dependent dissociation of catenins from VE cadherin. Catenin dissociation from VE-cadherin was enhanced by Heregulin beta1 (P < .05) stimulation and decreased by trastuzumab (P < .05) blockade of HER2 signaling in cancer cells. An increase in EC phosphoSrc (Tyr 416) was seen by 8 hours.
Our data suggest that HER2 induction of EC retraction involves both down-regulation of VE-cadherin and dissociation of catenins. HER2 signaling appears to regulate this potential metastatic mechanism. Further, Src phosphorylation suggests that this pathway may be involved in this mechanism.
HER2过表达赋予乳腺癌转移优势。我们已经表明，乳腺癌细胞中的HER2信号传导会诱导相邻内皮细胞（EC）收缩，破坏内皮完整性。由于内皮完整性依赖于黏附连接，我们推测肿瘤细胞诱导的EC收缩机制涉及连环蛋白从血管内皮（VE）钙黏蛋白的解离。在本研究中，我们报告了肿瘤相关EC中VE钙黏蛋白的缺失。我们还通过操纵肿瘤细胞中的HER2信号传导来测试连环蛋白与VE钙黏蛋白解离的变化。
我们测试了人EC汇合单层在暴露于乳腺癌细胞或条件培养基后VE钙黏蛋白的下调以及连环蛋白与VE钙黏蛋白的解离。使用免疫沉淀法，我们在不同处理以操纵HER2信号传导后，定量肿瘤相关EC中与VE钙黏蛋白剩余的复合连环蛋白。
用来自表达HER2的MCF-7细胞的条件培养基处理EC会导致VE钙黏蛋白表达缺失，以及连环蛋白与VE钙黏蛋白的时间依赖性解离。在癌细胞中，Heregulin beta1（P <.05）刺激增强了连环蛋白与VE钙黏蛋白的解离，而曲妥珠单抗（P <.05）阻断HER2信号传导则使其减少。8小时时可见EC磷酸化Src（Tyr 416）增加。
我们的数据表明，HER2诱导的EC收缩涉及VE钙黏蛋白的下调和连环蛋白的解离。HER2信号传导似乎调节这种潜在的转移机制。此外，Src磷酸化表明该途径可能参与这一机制。
