(789c) Engineering of Surface Gradients On Biopolymeric Films for the Spatial Presentation of Growth Factor and Modulation of Physical Properties

The native microenvironment of cells contains physical and biochemical cues that control cellular processes, which ultimately dictate cell fate. Biochemical cues, such as growth factors, are often presented in vivo in a “matrix-bound” fashion. Moreover, these cues vary significantly through the dimensions of the microenvironment presenting themselves as gradients. Thus biomaterials that exhibit a gradient in physical (i.e. stiffness) or biochemical (cytokines) cues would be beneficial to further the understanding of cell-material interactions. In this work, surface gradients of stiffness and of the growth factor bone morphonogenic protein 2 (BMP-2) were generated on polyelectrolyte multilayers (PEM) films composed of hyaluronan (HA) and poly(l-lysine) (PLL) by means of microfluidics. A gradient of water-soluble carbodiimide as cross-linking agent was used to generate the stiffness gradient. FTIR spectroscopy confirmed that cross-linking was anisotropic and followed the gradient of cross-linker. It was characterized by a decrease of the COO⁻ bands. Fluorescently labeled BMP-2 was used to validate gradient formation and to calculate surface concentration. BMP-2 surface gradients exhibited a profile with three distinct zones: a high-constant concentration region, an area with a linearly decreasing gradient, and a low-constant concentration region. C2C12 skeletal muscle cells were used to investigate early cellular processes such as adhesion, migration and differentiation. C2C12 cells adhered better and spread more in regions of high cross-linking; while in low cross-linking regions they behave poorly with regards to adhesion and spreading. Time-lapse microscopy was used to investigate cell migration along the BMP-2 gradient. Effective bioactivity of the BMP-2 gradients was assessed by measuring the SMAD signaling pathway of C2C12 cells in response to BMP-2. This technology may be a useful tool to better understand cell-material interactions in vitro.