Endothelial Induced EMT in Breast Epithelial Cells with Stem Cell Properties

Epithelial to mesenchymal transition (EMT) is a critical event in cancer progression and is closely linked to the breast epithelial cancer stem cell phenotype. Given the close interaction between the vascular endothelium and cancer cells, especially at the invasive front, we asked whether endothelial cells might play a role in EMT. Using a 3D culture model we demonstrate that endothelial cells are potent inducers of EMT in D492 an immortalized breast epithelial cell line with stem cell properties. Endothelial induced mesenchymal-like cells (D492M) derived from D492, show reduced expression of keratins, a switch from E-Cadherin (E-Cad) to N-Cadherin (N-Cad) and enhanced migration. Acquisition of cancer stem cell associated characteristics like increased CD44high/CD24low ratio, resistance to apoptosis and anchorage independent growth was also seen in D492M cells. Endothelial induced EMT in D492 was partially blocked by inhibition of HGF signaling. Basal-like breast cancer, a vascular rich cancer with stem cell properties and adverse prognosis has been linked with EMT. We immunostained several basal-like breast cancer samples for endothelial and EMT markers. Cancer cells close to the vascular rich areas show no or decreased expression of E-Cad and increased N-Cad expression suggesting EMT. Collectively, we have shown in a 3D culture model that endothelial cells are potent inducers of EMT in breast epithelial cells with stem cell properties. Furthermore, we demonstrate that basal-like breast cancer contains cells with an EMT phenotype, most prominently close to vascular rich areas of these tumors. We conclude that endothelial cells are potent inducers of EMT and may play a role in progression of basal-like breast cancer.上皮间质转化（Epithelial to mesenchymal transition, EMT）是癌症进展中的关键事件，且与乳腺上皮癌症干细胞表型密切相关。鉴于血管内皮细胞与癌细胞，尤其是侵袭前沿的癌细胞之间存在紧密互作，我们探究了内皮细胞是否可能在上皮间质转化中发挥作用。通过三维培养模型，我们证实内皮细胞是具有干细胞特性的永生化乳腺上皮细胞系D492发生上皮间质转化的强效诱导剂。由D492衍生的内皮诱导型间质样细胞（D492M）表现出角蛋白表达降低、从E-钙粘蛋白（E-Cadherin, E-Cad）向N-钙粘蛋白（N-Cadherin, N-Cad）转换以及迁移能力增强的特征。D492M细胞还获得了癌症干细胞相关特性，包括CD44高表达/CD24低表达比例升高、抗凋亡能力增强以及非锚定依赖性生长能力提升。抑制肝细胞生长因子（Hepatocyte Growth Factor, HGF）信号通路可部分阻断内皮细胞诱导的D492细胞发生上皮间质转化。基底样乳腺癌是一类富含血管、具备干细胞特性且预后不良的癌症，其发生与上皮间质转化密切相关。我们对多份基底样乳腺癌样本开展了内皮细胞与上皮间质转化标志物的免疫染色实验，结果显示，紧邻富含血管区域的癌细胞的E-钙粘蛋白表达缺失或降低，而N-钙粘蛋白表达升高，提示此类癌细胞发生了上皮间质转化。综上，我们在三维培养模型中证实，内皮细胞是具有干细胞特性的乳腺上皮细胞发生上皮间质转化的强效诱导剂。此外，我们发现基底样乳腺癌中存在具有上皮间质转化表型的癌细胞，且这些细胞主要富集于肿瘤内富含血管的区域附近。我们由此得出结论：内皮细胞是上皮间质转化的强效诱导剂，可能在基底样乳腺癌的进展过程中发挥重要作用。