TISSUE ENGINEERED TUMOR MICROVESSELS TO STUDY THE ROLE OF FLOW SHEAR STRESS ON ENDOTHELIAL BARRIER FUNCTION

摘要

As solid tumors develop, a variety of physical stresses arise including growth induced compressive force, matrix stiffening due to desmoplasia, and increased interstitial fluid pressure and altered flow patterns due to leaky vasculature and poor lymphatic drainage [1]. These microenvironmental stresses likely contribute to the abnormal cell behavior that drives tumor progression, and have become an increasingly significant area of cancer research. Of particular importance, is the role of flow shear stress on tumor-endothelial signaling, vascular function, and angiogenesis. Compared to normal vasculature, blood vessels in tumors are poorly functional due to dysregulated expression of angiogenic growth factors, such as vascular endothelial growth factor (VEGF) or the angiopoietins. Also, because of the abnormal vessel structure, blood velocities can be an order of magnitude lower than that of normal microvessels. Recently published work utilizing intravital microscopy to measure blood velocities in mouse mammary fat pad tumors, demonstrated for the first time that shear rate gradients in tumors may help guide branching and growth of new vessels [2]. However, much still remains unknown about how shear stress regulates endothelial organization, permeability, or expression of growth factors within the context of the tumor microenvironment. In this study, we utilized a collagen based in vitro microfluidic tumor vascular model to investigate the biomechanical mechanisms that regulate endothelial barrier function. It is hypothesized that low shear stress may alter the tumor endothelial phenotype such that permeability is increased and angiogenic potential is enhanced. Target microvascular wall shear stresses (WSS) ranging from low (τ_W=1 dyn/cm~2), normal (τ_W=4 dyn/cm~2), and high (τ_W=10 dyn/cm~2) were estimated based on Poiseuille assumptions and experimentally measured using micro-particle image velocimetry (μ-PIV). Endothelial morphology, expression of tight junction proteins, and vascular permeability were measured as a function of WSS. Tumor expressed VEGFA, was also measured using quantitative RT-PCR. Results demonstrate that fluid flow increased endothelial alignment in the direction of flow and decreased vascular permeability as a function of WSS. This flow-mediated response was greatest during co-culture with tumor cells relative to endothelial mono-cultures. In addition, tumor-expressed VEGFA was significantly upregulated by both the presence of flow as well as co-culture with endothelial cells.