Interstitial flow influences direction of tumor cell migration through competing mechanisms

Abstract

Interstitial flow is the convective transport of fluid through tissue extracellular matrix. This creeping fluid flow has been shown to affect the morphology and migration of cells such as fibroblasts, cancer cells, endothelial cells, and mesenchymal stem cells. A microfluidic cell culture system was designed to apply stable pressure gradients and fluid flow and allow direct visualization of transient responses of cells seeded in a 3D collagen type I scaffold. We used this system to examine the effects of interstitial flow on cancer cell morphology and migration and to extend previous studies showing that interstitial flow increases the metastatic potential of MDA-MB-435S melanoma cells [Shields J, et al. (2007) Cancer Cell 11:526-538]. Using a breast carcinoma line (MDA-MB-231) we also observed cell migration along streamlines in the presence of flow; however, we further demonstrated that the strength of the flow as well as the cell density determined directional bias of migration along the streamline. In particular, we found that cells either at high seeding density or with the CCR-7 receptor inhibited migration against, rather than with the flow. We provide further evidence that CCR7-dependent autologous chemotaxis is the mechanism that leads to migration with the flow, but also demonstrate a competing CCR7-independent mechanism that causes migration against the flow. Data from experiments investigating the effects of cell concentration, interstitial flow rate, receptor activity, and focal adhesion kinase phosphorylation support our hypothesis that the competing stimulus is integrin mediated. This mechanism may play an important role in development of metastatic disease. mechanobiology | computational model | signaling | cell mechanics T issues are composed of cells residing in an extracellular matrix (ECM) containing interstitial fluid that transports nutrients and signaling molecules (1, 2). Osmotic and hydrostatic pressure gradients across tissues resulting from physiologic processes such as drainage toward lymphatics, inflammation, locally elevated pressures due to tumor growth or leaky microvessels, and muscle contraction each drive fluid flow through the ECM (2, 3). This fluid flow is termed interstitial flow and has long been recognized to be instrumental in tissue transport and physiology (1, 4, 5). Chary and Jain used fluorescence recovery after photobleaching to directly observe fluid flow in the tissue interstitium and determined typical flow velocities to be on the order of 0.1-2.0 μm/s, and more recent studies have demonstrated that flow can reach velocities as high as 4.0 μm/s (6, 7).Interstitial flow is particularly important in driving transport in the vicinity of tumors, as neoplastic tissue is often characterized by localized increases in interstitial pressure, leading to high interstitial pressure gradients at the tumor margin (8). Interstitial flow has hence emerged as a possible stimulus for guiding tumor cell migration in the formation of metastases (9-12).Shields et al. observed increase...

Mechanotransduction of fluid stresses governs 3D cell migration

“…In particular, adhesions containing paxillin elongate under applied traction (23). FAK also becomes autophosphorylated at Tyr 397 through binding with β1 integrins in endothelial cells in response to shear stress (24) and in breast adenocarcinoma cells when exposed to 3 μm/s IF as we have shown previously (17). Furthermore, phosphorylation of FAK at Tyr 397 is required for invadopodia formation for breast cancer cells (25), and FAK PY397 is differentially expressed in metastatic cervical carcinoma relative to noncancerous epithelium (26).…”

“…In addition to imparting fluid stresses, IF convects autocrine factors downstream, causing gradients of autologously secreted chemokines that have been shown to direct migration downstream in response to IF (14). Furthermore, we have shown previously that at high cell densities, autocrine chemokine fields overlap with fields from neighboring cells, reducing transcellular autocrine gradients and inducing preferential upstream migration (17). In this study, cells were seeded at a sufficiently high density (6 × 10 5 cells per mL) and flow rate to minimize contributions from autocrine gradients, leading to upstream migration.…”

“…mammary adenocarcinoma cells embedded in a collagen type I hydrogel (Fig. 1A) (17,20). By holding one medium channel (upstream) at a higher fluid pressure than the other channel (downstream), a stable and repeatable flow field can be generated through the collagen gel (Movie S1 and SI Appendix, Fig.…”

“…IF has been shown to induce rheotaxis when cells are seeded at sufficiently high density for paracrine chemokine fields to interfere with autocrine chemokine fields (17), and cells migrating upstream have been shown to migrate with increased persistence (35). We seeded cells at a density shown to induce preferential upstream migration of MDA-MB 231 (6 × 10 5 cells per mL) and applied 4.6 μm/s flow for 8 h, tracking the cells by taking images every 15 min.…”

Biomimetic Model of Tumor Microenvironment on Microfluidic Platform

“…Recently, several studies using microfluidic platform have been reported that recreate the 3D context of each aspect of the TME and metastasis in vitro. [10][11][12][13] However, to our knowledge, no versatile in vitro experimental model that reconstitutes direct interplay between constituents of the TME during tumorigenesis has yet been reported. Here we describe a biomimetic TME model to mimic the complex inter actions of the tumor, stromal, and endothe lial cells, all of which are essential components of the TME that hinder the effective treatment of cancers.…”