Guiding cell migration in 3D: A collagen matrix with graded directional stiffness

While matrix stiffness has been implicated in cell adhesion and migration, most studies have focused on the effects of substrate stiffness in 2D. The present work describes a novel continuous stiffness gradient model for studying such processes in 3D. Wedge-shaped collagen scaffolds were compressed to produce sheets of a desired (0.1 mm) uniform thickness, but with increasing collagen density along the length of the sheet. Dynamic mechanical analysis, carried out on 1 mm wide strips obtained from the two ends and the middle of each sheet, showed that the elastic modulus increased from 1057 +/- 487 kPa to 2305 +/- 693 kPa at the soft and stiff end respectively and was 1835 +/- 31 kPa in the middle. In constructs seeded with agarose marker beads prior to compression, mean agarose bead density rose from 10 +/- 1 to 71 +/- 12 at the soft and stiff end respectively and was 19 +/- 5 in the middle, indicating successful engineering of a density gradient corresponding to the measured stiffness gradient. Growth-arrested human dermal fibroblasts, initially seeded evenly within such constructs, accumulated preferentially towards the stiff part of the gradient after 3 and 6 days in culture. Durotactic migration was significant after 6 days. This model provides a new means for studying cellular mechanotaxis and patterning cells which is controllable, biomimetic and in 3D.