Cells on arrays of microsprings: An approach to achieve triaxial control of substrate stiffness

Microenvironmental biophysical stimuli influence diverse cellular functions, such as directional motility and stem cell differentiation. Previously, researchers have tuned the linear stiffness of microposts to investigate cell mechanobiological processes and direct cellular behavior; however, microposts suffer from an inherent, yet critical drawback - regulation of micropost stiffness is fundamentally limited to “biaxial” control. To overcome this issue, here we utilize three-dimensional (3D) direct-write laser lithography processes to fabricate arrays of microscale springs (μSprings). By adjusting the geometric characteristics of individual μSprings, the x-, y-, and z-axis stiffness of the cellular substrate can be customized at the microscale. COMSOL simulations were performed to characterize the theoretical “triaxial” stiffness associated with a variety of μSpring designs. Endothelial cells seeded on μSpring arrays were found to successfully deform the μSprings via cell-generated forces. By enabling user-control over the triaxial stiffness of discrete, microscale substrate features, the presented μSpring methodology could offer a powerful platform for cellular studies and applications in fields including tissue engineering, biomaterials, and regenerative medicine.