(772c) A Microfluidic Device to Measure Traction Forces During Confined Chemotactic Migration

Authors: 
Paul, C. D., University of Arkansas
Stroka, K. M., Johns Hopkins University
Konstantopoulos, K., Johns Hopkins University
Raman, P. S., Johns Hopkins University


Introduction: There is a wide heterogeneity of environments in which tumor cells migrate in vivo. In many cases, alignment of fibers in the ECM or cell-mediated matrix remodeling creates microchannel-like tunnels in the tumor stroma1,2. Previous work has suggested a reduced importance of actomyosin contractility for tumor cell migration in highly confining microchannels3. Traction forces are essential for two-dimensional cell migration and can be measured using deflectable PDMS microposts4 or deformable gels5. However, force measurement in three-dimensional environments, particularly those showing the architectural heterogeneity seen in the body, is much more difficult. Therefore, we engineered a device to measure traction force exertion during confined migration.

Materials and Methods:To quantify force exertion during chemotactic migration, we designed and built a microfluidic device containing a ladder-like array of microchannels with deflectable PDMS microposts at their bases. NIH-3T3 fibroblasts and HOS cells were induced to migrate on top of the posts, and the post deflections were quantified and converted to time-dependent forces using the beam bending theory. To explore the effects of modulating actomyosin contractility on force exertion in confining and non-confining environments, we treated cells with 50 µM blebbistatin, a myosin II inhibitor, or 0.1 nM calyculin A, which promotes myosin light chain phosphorylation. Migration speeds in microchannels containing flat bases were measured under these same conditions to explore migration efficiency. Statistical significance was assessed with non-paired Student’s t-test.

Results and Discussion: Initial experiments were performed with 3T3 fibroblasts, a common model used for cell migration studies, to demonstrate the utility of our device. In wide channels, 3T3 force exertion was consistent with measurements made on beds of microposts4. Remarkably, measured traction forces were significantly reduced in narrow channels vs. wide channels for both 3T3 and HOS cells, as shown in Figure 1. To explore the extent of actomyosin participation in the confined migration of cancer cells, drug treatments were used to increase or decrease cell contractility. Blebbistatin treatment significantly reduced migration speed and exerted force in wide channels but did not affect either in narrow channels. Treatment with calyculin A increased forces and migration speed in wide channels but not narrow channels. Together, these data strongly suggest that actomyosin contraction plays a reduced role in migration through confined spaces.

Conclusions: Understanding the diverse mechanisms of cell migration is vital for abrogating cancer cell migration. Here, we present the fabrication of a novel device to measure force exertion in well-defined, heterogeneous environments and quantitatively demonstrate that confined cells use a migration mechanism that is less dependent on actomyosin contractility than was previously appreciated. Future work will focus on determining how modulation of cell contractility affects migration through confined spaces for a variety of cell types and how this modulation could provide clues for inhibiting tumor cell migration.

References: (1) Wolf, K. et al., Sem Cell Devel Biol, 2009, 20, 931-941; (2) Fisher, K.E. et al., J Cell Sci, 2009, 122, 4558-4569; (3) Balzer, E.M. et al., FASEB J, 2012, 26, 4045-4056; (4) Han, S.J. et al., Biophys J, 2012, 103, 640-648; (5) Dembo, M. et al., Biophys J, 1999, 76, 2307-2316.