The regulation of cellular morphogenesis and differentiation via the physical properties of the provisional extracellular matrix (ECM) is poorly understood and our group has been working towards elucidating the dominant physical factors of the ECM that influence cell spreading, migration and differentiation in 3-D culture. We apply a biosynthetic PEG-protein hydrogel as an ECM-analog for cell culture, with highly defined and precisely controllable density, microarchitecture, proteolytic susceptibility, compliance and biofunctionality. The matrix is used to encapsulate mesenchymal cells while pseudo-independently altering biochemical and physical properties of the microenvironment using simple compositional modifications to the bio-synthetic constituents. We have shown that the proteolytic resistance and compliance of the matrix have a profound influence on the regulation of cell morphogenesis and phenotype determination. Beyond the control over the intrinsic physical attributes of the hydrogel, our laboratory has recently developed an optical 3-D micro-patterning approach to non-invasively create any prescribed geometrical feature having submicron spatial resolution in situ, anywhere within the PEG-protein hydrogel biomaterial. The micropatterns are made using a simple but highly effective application of computer-guided laser micro-ablation that creates localized imperfections in the hydrogel architecture. These imperfections are used to guide anisotropic cellular development within the amorphous material, including preferentially guiding neural cellular development in the hydrogels based on contact guidance and differential mechanical resistance of the scaffolding. Precisely controlled bulk material properties and custom 3-D landscaping with micropatterning are collectively used to elucidate the dominant and influential physical factors affecting morphogenesis patterns, phenotypic states, and differentiation of various cell types.
