A 3Rs approach to tumour metastasis: investigating the role of cancer stem cells in the metastasis of oral squamous cell carcinoma using an in vitro m

肿瘤转移的3Rs方法：利用体外微流控模型研究肿瘤干细胞在口腔鳞癌转移中的作用每年诊断出30万例口腔鳞状细胞癌(OSCC)，约30%的患者表现出转移扩散。转移是一个复杂的过程，癌细胞通过这个过程移动到第二个部位形成新的肿瘤。以前的工作表明，癌症干细胞(CSCs)能够通过在上皮状态和间充质状态之间切换来驱动转移。然而，它们在与血管系统的相互作用中所起的作用仍不清楚，血管系统是肿瘤微环境(TME)的关键组成部分，被认为对转移有很大影响。在这里，我使用了微流控装置，一种很有前途的新的体外模型，来研究这些相互作用。我使用光刻和软光刻技术制造了这些设备，并使用它们在纤维蛋白凝胶中联合培养内皮细胞和口腔鳞状细胞癌细胞，以3D方式评估这两种细胞之间的相互作用。在开发和优化OSCC单芯片转移模型后，我发现了癌细胞和内皮细胞之间的双向通信，这极大地改变了这两种细胞的行为。最有趣的是，癌细胞的侵袭经历了显著的变化，并且在内皮细胞存在的情况下显著减少。对基质的成像显示，当内皮细胞在纤维蛋白凝胶中进行萌芽血管生成时，它会降解并重塑基质。这种重塑的环境限制了口腔鳞癌细胞在基质中的迁移，并抑制了完整的上皮向间充质转化(EMT)-相反，通过上皮性肿瘤细胞通过内皮细胞形成的预先存在的轨迹进行侵袭进展。侵袭流中细胞的免疫荧光染色显示了与转移性CSCs的部分EMT相关的特征。这些发现可能代表了血管生态位的产生，它改变了周围的TME，并通过这一点影响了侵袭性CSC的表型。

A 3Rs approach to tumour metastasis: investigating the role of cancer stem cells in the metastasis of oral squamous cell carcinoma using an in vitro microfluidic model 300,000 cases of oral squamous cell carcinoma (OSCC) are diagnosed every year, and around 30% of patients exhibit metastatic spread. Metastasis is a complex process by which cancer cells travel to a secondary site to form a new tumour. Previous work has suggested that cancer stem cells (CSCs) are capable of driving metastasis by switching between epithelial and mesenchymal states. However, the role they play in the interactions with the vascular system, a key component of the tumour microenvironment (TME) recognised to greatly influence metastasis, remains unclear. Here, I used microfluidic devices, a promising new in vitro model, to study these interactions. I fabricated these devices using photolithography and soft lithography techniques and used them to co-culture endothelial cells with OSCC cells in a fibrin gel, to assess the interactions between these two cell types in 3d. After developing and optimising this OSCC metastasis-on-a-chip model, I uncovered a two-way communication between cancer cells and endothelial cells that greatly changes the behaviour of both cell types. Most interestingly, cancer cell invasion undergoes marked changes and is significantly reduced in the presence of endothelial cells. Imaging of the matrix shows that endothelial cells degrade and remodel the matrix when undergoing sprouting angiogenesis in the fibrin gel. This remodelled environment restricts OSCC cell migration in the matrix, and full epithelial-to-mesenchymal transition (EMT) is suppressed - instead invasion progresses through streaming of epithelial tumour cells through pre-existing tracks made by endothelial cells. Immunofluorescent staining of the cells in the invasive streams demonstrates features that have been associated with partial EMT in metastatic CSCs. These findings may represent the generation of a vascular niche that acts to modify the surrounding TME and, through this, influences the invasive CSC phenotype.