Cell adhesion, spreading and migration require the dynamic formation and dispersal of contacts with the extracellular matrix (ECM). In vivo, the number, availability and distribution of ECM binding sites dictate the shape of a cell and determine its mobility. To analyse the geometrical limits of ECM binding sites required for cell attachment and spreading, we used microcontact printing to produce regular patterns of ECM protein dots of defined size separated by nonadhesive regions. Cells cultured on these substrata adhere to and spread on ECM regions as small as 0.1 microm2, when spacing between dots is less than 5 microm. Spacing of 5-25 microm induces a cell to adapt its shape to the ECM pattern. The ability to spread and migrate on dots > or =1 microm2 ceases when the dot separation is > or =30 microm. The extent of cell spreading is directly correlated to the total substratum coverage with ECM-proteins, but irrespective of the geometrical pattern. An optimal spreading extent is reached at a surface coating above 15%. Knowledge of these geometrical limits is essential for an understanding of cell adhesion and migration, and for the design of artificial surfaces that optimally interact with cells in a living tissue

For both cells and tissues, shape is closely correlated with function presumably via geometry-dependent distribution of tension. In this study, we identify common shape determinants spanning cell and tissue scales. For cells whose sites of adhesion are restricted to small adhesive islands on a micropatterned substrate, shape resembles a sequence of inward-curved circular arcs. The same shape is observed for fibroblast-populated collagen gels that are pinned to a flat substrate. Quantitative image analysis reveals that, in both cases, arc radii increase with the spanning distance between the pinning points. Although the Laplace law for interfaces under tension predicts circular arcs, it cannot explain the observed dependence on the spanning distance. Computer simulations and theoretical modeling demonstrate that filamentous network mechanics and contractility give rise to a modified Laplace law that quantitatively explains our experimental findings on both cell and tissue scales. Our model in conjunction with actomyosin inhibition experiments further suggests that cell shape is regulated by two different control modes related to motor contractility and structural changes in the actin cytoskeleton.

The cellular environment impacts a myriad of cellular functions by providing signals that can modulate cell phenotype and function. Physical cues such as topography, roughness, gradients, and elasticity are of particular importance. Thus, synthetic substrates can be potentially useful tools for exploring the influence of the aforementioned physical properties on cellular function. Many micro- and nanofabrication processes have been employed to control substrate characteristics in both 2D and 3D environments. This review highlights strategies for modulating the physical properties of surfaces, the influence of these changes on cell responses, and the promise and limitations of these surfaces in in-vitro settings. While both hard and soft materials are discussed, emphasis is placed on soft substrates. Moreover, methods for creating synthetic substrates for cell studies, substrate properties, and impact of substrate properties on cell behavior are the main focus of this review.

Cell behavior is governed by interactions with the cellular environment. [1][2][3] These interactions include cell-cell as well as cell-extracellular matrix (ECM) contacts and act, in addition to soluble growth factors, as key regulators of cell survival, proliferation, and differentiation. However, not only the molecular composition of the contact sites, but also their spatial distributions impact cell behavior. [ 4 , 5 ] Realizing cell-culture scaffolds that mirror the complex in vivo arrangement of ECM components is an active area of biomaterials engineering. Corresponding 2D lithographically defi ned micropatterned structures are widely used and have even become commercially available. [6][7][8][9] In addition to patterned ligand distributions, mechanical interactions between cells and their environment also play an important role in regulating cellular functions. [ 10 ] Consequently, corresponding fl exible substrates were developed that allow measurements of cellular forces in 2D on planar substrates or pillar arrays. [ 11 , 12 ] However, discrepancies between cell behavior in vivo and in artifi cial 2D environments have become evident. [ 5 , 13 , 14 ] Therefore, devices that capture more of the structural complexity present in 3D tissues are highly desirable.Fibrous collagen or matrigel matrices are commonly used to study 3D cell behavior, [ 15 , 16 ] but these matrices have a random pore size and are structurally and chemically ill-defi ned. We [ 17 ] and others [18][19][20] have recently shown that direct laser writing (DLW) is a versatile technique to fabricate tailored 3D cell-culture scaffolds in the micrometer to nanometer range. By using an adequate photoresist, elastic 3D scaffolds for cell force measurements have also been realized. [ 17 ] These DLW scaffolds have been homogeneously coated with ECM molecules. Ideally, they should have an adjustable distribution of cell-substrate contact sites to manipulate cell adhesion and cell shape in all three dimensions. In this communication, we report on tailored 3D scaffolds with a controlled ECM distribution. By sequential DLW of two different photoresists, composite-polymer scaffolds with distinct protein-binding properties are fabricated and selectively biofunctionalized thereafter. We further demonstrate that cells cultured in these scaffolds selectively form cell adhesion sites with the functionalized parts, allowing for control of cell adhesion and cell shape in three dimensions, forming the basis for future designer tissue-culture scaffolds.To fabricate micrometer-scale composite-polymer scaffolds with distinct protein-binding properties as depicted in Figure 1 , we fi rst selected and characterized appropriate materials. For the small protein-binding cubes in the scaffolds we selected Ormocomp, a member of the inorganic (Si-O-Si)-organic hybrid polymer Ormocer family (Fraunhofer Institute for Silica Research, Würzburg, Germany). Ormocomp is a biocompatible photoresist that has previously been used in our lab to manufacture elastic 3D scaffolds...

Stimuli-responsive microstructures are critical to create adaptable systems in soft robotics and biosciences. For such applications, the materials must be compatible with aqueous environments and enable the manufacturing of three-dimensional structures. Poly(N-isopropylacrylamide) (pNIPAM) is a well-established polymer, exhibiting a substantial response to changes in temperature close to its lower critical solution temperature. To create complex actuation patterns, materials that react differently with respect to a stimulus are required. Here, we introduce functional three-dimensional hetero-microstructures based on pNIPAM. By variation of the local exposure dose in three-dimensional laser lithography, we demonstrate that the material parameters can be altered on demand in a single resist formulation. We explore this concept for sophisticated three-dimensional architectures with large-amplitude and complex responses. The experimental results are consistent with numerical calculations, able to predict the actuation response. Furthermore, a spatially controlled response is achieved by inducing a local temperature increase by two-photon absorption of focused light.