Mechanotransduction of fluid stresses governs 3D cell migration

Abstract

Solid tumors are characterized by high interstitial fluid pressure, which drives fluid efflux from the tumor core. Tumor-associated interstitial flow (IF) at a rate of ∼3 μm/s has been shown to induce cell migration in the upstream direction (rheotaxis). However, the molecular biophysical mechanism that underlies upstream cell polarization and rheotaxis remains unclear. We developed a microfluidic platform to investigate the effects of IF fluid stresses imparted on cells embedded within a collagen type I hydrogel, and we demonstrate that IF stresses result in a transcellular gradient in β1-integrin activation with vinculin, focal adhesion kinase (FAK), FAK PY397 , F actin, and paxillindependent protrusion formation localizing to the upstream side of the cell, where matrix adhesions are under maximum tension. This previously unknown mechanism is the result of a force balance between fluid drag on the cell and matrix adhesion tension and is therefore a fundamental, but previously unknown, stimulus for directing cell movement within porous extracellular matrix.mechanobiology | breast cancer | metastasis I ntegrins and associated focal adhesion (FA) proteins form a tension-sensitive mechanical link between the extracellular matrix (ECM) and the cytoskeleton, and serve as key components in the signaling cascade by which cells transduce mechanical signals into biological responses (mechanotransduction) (1, 2). Contractile stresses generated by the cell are balanced by tractions at cell-substrate adhesions, and the FA protein vinculin accumulates at regions of high substrate stress (3, 4). The FA protein paxillin colocalizes with vinculin (4) and mediates β1-integrin FA turnover through interaction with FA kinase (FAK) (5). The FAK-paxillin signaling axis recruits vinculin to β1 integrins at regions of high matrix adhesion tension (6), and paxillin-a key mechanosensor (7)-mediates protrusion formation at regions of high stress on 2D substrates (8), and FAK-paxillin-vinculin signaling is required for mechanosensing and durotaxis (9).The tumor microenvironment imparts mechanical and chemical signals on tumor and stromal cells (10), and advanced breast carcinomas are characterized by high interstitial fluid pressure (11), an indicator of poor prognosis (12). This elevated fluid pressure drives interstitial flow (IF) and alters chemical transport within the tumor (13), and IF influences tumor cell migration through the generation of autocrine chemokine gradients (14). Equally important, although not as well understood, is the physical drag imparted on the ECM and constitutive cells (15) by IF, which is analogous to the FA-activating shear stresses generated on endothelial cells by hemodynamic forces (16). With endothelial cells, shear stress can be the dominant mechanical stimulus that induces FAK activation and cytoskeletal remodeling; however, for cells embedded within a porous matrix scaffold, the ratio of the force due to the pressure drop across the cell to the total shear force is inversely proportional to hydrogel p...

EMT Transition Alters Interstitial Fluid Flow–Induced Signaling in ERBB2-Positive Breast Cancer Cells

“…In the future, investigating the role of IFF on 3D DCIS structures in a more clinically relevant matrix will lead to better understanding of the role of IFF on breast cancer invasion. We must also consider the contribution of other proposed mechanisms of IFF mechanotransduction, including mechanosensors such as integrins and the tumor cell glycocalyx, which have both been implicated in IFF-induced cell invasion (17,19). Examining these other mechanisms may help identify the upstream mediator of PI3K phosphorylation in NeuN cells.…”

“…In metastatic cells, IFF altered extracellular chemokine gradients to accelerate tumor cell invasion via CXCR4-and CCR7-dependent chemotaxis (14,15), and increased MMP activity via shear stress sensing through the glycocalyx (17). More recently, IFF was shown to induce invasion against the direction of flow due to fluid drag forces and activation of integrins on the upstream side of the cell (19). Although we know some of its effects on invasive cells, the specific pathways triggered by IFF are still unclear, especially those that play a crucial role in the early stages of invasion.…”

Paxillin controls endothelial cell migration and tumor angiogenesis by altering neuropilin 2 expression

“…Mechanical forces also contribute to control of capillary cell growth, migration and fate determination Mammoto et al, 2012), and changes in mechanical forces elicited by altering ECM stiffness can regulate angiogenesis through p190RhoGAP . Given that paxillin mediates force-dependent control of cell behaviors (Sawada and Sheetz, 2002;Zaidel-Bar et al, 2007), including directional cell motility (Plotnikov et al, 2012;Polacheck et al, 2014;Sero et al, 2011), cell-derived mechanical forces might also contribute to the control of angiogenesis through paxillin.…”

Implications of Increase in Vascular Permeability in Tumors by VEGF: A Commentary on the Pioneering Work of Harold Dvorak

“…In human tumors, the collagen component can be even greater (13). The increase in collagen content within the extracellular matrix contributes to tumor cell movement (14) and metastasis.…”

“…Studies in human tumor xenografts have verified that effects of IFP on radiation response are independent of extent of hypoxia (28). Recent studies have revealed that IFP increases tumor cell aggression by activating signal transduction pathways that promote epithelial-mesenchymal transition (29) and cell motility (14).…”