(a) Representative images of 3T3 fibroblasts on soft and stiff static substrates, as well as a dynamically stiffened soft-to-stiff substrate (stiffening performed on day 1). 3T3 fibroblast spreading quantification on (b) day 1 and (c) day 2. Stiff substrates were used as a positive control as photocrosslinking was applied before cell seeding and static soft substrates did not receive light exposure. The moduli for the Soft and Stiff substrates were ~3 and ~37 kPa (measured by AFM indentation), respectively. The modulus of the Soft-to-Stiff substrate was changed from ~3 to 37 kPa after stiffening. Thus, the final moduli of the Soft, Soft-to-Stiff and Stiff substrates were ~3, ~37 and ~37 kPa, respectively. n > 40 cells per group. ***p < 0.001, n.s.: not significant.