(248d) Control of Cell Migration Using a Mechanistically Tunable Fibrous Environment

Cells attach to and interact with their immediate fibrous microenvironment known as the extracellular matrix (ECM), which provides region-specific biochemical and biophysical cues that drive cell behavior.  Despite recent studies indicating the importance of controlling these factors which cause dramatic alterations in cell attachment, motility, and differentiation, most cell studies are still performed on substrates of flat tissue cultured glass, plastic, or gel.  Here we present cell behavior the STEP (Spinneret-based Tunable Engineered Parameters) platform, which deposits high aspect ratio nano/micro fibers with tight control on diameter, spacing, porosity, and orientation.  The resulting fibrous scaffolds, which mimic the mechanistic environment of ECM by providing biomechanical cues, can be tuned to elicit a desired cellular response.  In this study, C2C12 mouse myoblast cells are seeded onto highly aligned fibrous STEP scaffolds and their migration speeds are shown to directly result from varying substrate stiffness in forms of fiber diameter and fiber length.  Polystyrene fibers of three different diameters (200, 500, and 800 nm) were spun onto two substrate types (suspended single-layer (SS) and suspended double-layer (SD)) in addition to flat glass control.  Using standard cell culture techniques, C2C12 mouse myoblasts were seeded onto the scaffolds and attached within 4-6 hours.  After attaching, 2 mL DMEM media were added to keep cells viable over a 24 hour period.  During this time, cells on fiber networks were time-lapse imaged by an incubating microscope.  Since cells do not travel at a constant velocity, cell migration speeds were calculated by tracking the position of the cell’s nucleus every hour and noting the maximum distance traveled per hour in a 16-hour window.  Cells were observed to attach to and elongate along the fiber axis.  On flat glass, cells were observed to travel at an average velocity of 30 μm/hr.  Cells constricted to SS fibers traveled at the highest speeds (55 – 80 μm/hr) and onwe SD fibers were observed to probe each possible migration direction, thus, decreasing their overall migration velocity compared to SS substrates (40 – 50 μm/hr).  Similary, both fiber diameter and fiber length were observed to play a role in cell migration speed.  In each case, the conditions which resulted in the lower fiber stiffness (smaller diameter, longer length) allowed cells to travel at the top end of the speed ranges previously listed.  Greater fiber stiffness corresponds to an increased ability for the cell to form stable focal adhesions and a lower propensity to migrate at high speeds.  STEP platform provides a unique tool to fine-tune the mechanical environment to which native cells attach and interact with so that scaffolds and drug delivery therapies may be improved upon in the future.