Clinically, increased breast tumor stiffness is associated with metastasis and poorer outcomes. Yet, in vitro studies of tumor cells in 3D scaffolds have found decreased invasion in stiffer environments. To resolve this apparent contradiction, MDA-MB-231 breast tumor spheroids were embedded in 'low' (2 kPa) and 'high' (12 kPa) stiffness 3D hydrogels comprised of methacrylated gelatin/collagen I, a material that allows for physiologically-relevant changes in stiffness while matrix density is held constant. Cells in high stiffness materials exhibited delayed invasion, but more abundant actin-enriched protrusions, compared to those in low stiffness. We find that cells in high stiffness had increased expression of Mena, an invadopodia protein associated with metastasis in breast cancer, as a result of EGFR and PLCγ1 activation. As invadopodia promote invasion through matrix remodeling, we examined matrix organization and determined that spheroids in high stiffness displayed a large fibronectin halo. Interestingly, this halo did not result from increased fibronectin production, but rather from Mena/α5 integrin dependent organization. In high stiffness environments, FN1 knockout inhibited invasion while addition of exogenous cellular fibronectin lessened the invasion delay. Analysis of fibronectin isoforms demonstrated that EDA-fibronectin promoted invasion and that clinical invasive breast cancer specimens displayed elevated EDA-fibronectin. Combined, our data support a mechanism by which breast cancer cells respond to stiffness and render the environment conducive to invasion. More broadly, these findings provide important insight on the roles of matrix stiffness, composition, and organization in promoting tumor invasion.

中文翻译：

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支架刚度通过 EGFR 相关的 Mena 上调和基质重塑影响乳腺癌细胞侵袭。


临床上，乳腺肿瘤硬度增加与转移和较差的结果相关。然而，对 3D 支架中肿瘤细胞的体外研究发现，在较僵硬的环境中，肿瘤细胞的侵袭会减少。为了解决这一明显的矛盾，MDA-MB-231乳腺肿瘤球体被嵌入“低”（2 kPa）和“高”（12 kPa）硬度的3D水凝胶中，该水凝胶由甲基丙烯酸明胶/胶原蛋白I组成，这种材料允许生理学上当基体密度保持不变时，刚度发生相关变化。与低硬度材料相比，高硬度材料中的细胞表现出延迟的侵袭，但富含肌动蛋白的突起更丰富。我们发现，由于 EGFR 和 PLCγ1 激活，高硬度细胞中 Mena 的表达增加，Mena 是一种与乳腺癌转移相关的侵袭伪足蛋白。由于侵袭伪足通过基质重塑促进侵袭，我们检查了基质组织并确定高硬度的球体显示出大的纤连蛋白晕。有趣的是，这个光环并不是由纤连蛋白产量增加造成的，而是由 Mena/α5 整合素依赖性组织造成的。在高硬度环境中，FN1 敲除抑制侵袭，而添加外源细胞纤连蛋白则减少侵袭延迟。纤连蛋白亚型分析表明，EDA-纤连蛋白促进侵袭，并且临床侵袭性乳腺癌标本显示 EDA-纤连蛋白升高。综合起来，我们的数据支持乳腺癌细胞对僵硬做出反应并使环境有利于侵袭的机制。更广泛地说，这些发现为基质硬度、成分和组织在促进肿瘤侵袭中的作用提供了重要的见解。