Matrix stiffening promotes a tumor vasculature phenotype

Significance Dysregulation of both vascular architecture and function is a hallmark of numerous diseases, including cancer. This dysregulation is currently largely attributed to up-regulated proangiogenic growth factors. Here, we show that the stiffness of the underlying extracellular matrix also plays a central role in promoting angiogenesis and a characteristic tumor-like vasculature both in vitro and in vivo. The matrix stiffness-mediated angiogenesis is dependent on increased matrix metalloprotease activity. In addition, increased matrix cross-linking disrupts endothelial cell–cell junctional integrity and results in leakier vasculature. These results suggest that altered tissue mechanics, which are characteristic of solid tumors, directly influence vascular phenotype and, subsequently, may impair therapeutic delivery and efficacy. Tumor microvasculature tends to be malformed, more permeable, and more tortuous than vessels in healthy tissue, effects that have been largely attributed to up-regulated VEGF expression. However, tumor tissue tends to stiffen during solid tumor progression, and tissue stiffness is known to alter cell behaviors including proliferation, migration, and cell–cell adhesion, which are all requisite for angiogenesis. Using in vitro, in vivo, and ex ovo models, we investigated the effects of matrix stiffness on vessel growth and integrity during angiogenesis. Our data indicate that angiogenic outgrowth, invasion, and neovessel branching increase with matrix cross-linking. These effects are caused by increased matrix stiffness independent of matrix density, because increased matrix density results in decreased angiogenesis. Notably, matrix stiffness up-regulates matrix metalloproteinase (MMP) activity, and inhibiting MMPs significantly reduces angiogenic outgrowth in stiffer cross-linked gels. To investigate the functional significance of altered endothelial cell behavior in response to matrix stiffness, we measured endothelial cell barrier function on substrates mimicking the stiffness of healthy and tumor tissue. Our data indicate that barrier function is impaired and the localization of vascular endothelial cadherin is altered as function of matrix stiffness. These results demonstrate that matrix stiffness, separately from matrix density, can alter vascular growth and integrity, mimicking the changes that exist in tumor vasculature. These data suggest that therapeutically targeting tumor stiffness or the endothelial cell response to tumor stiffening may help restore vessel structure, minimize metastasis, and aid in drug delivery.