Surface wrinkling of materials offers a simple yet elegant approach to fabricating cell culture substrates with highly ordered topographies for investigating cell mechanobiology. In this study we present a tunable shape memory polymer (SMP) bilayer system that is programmed to form, under cell compatible conditions, wrinkles with feature sizes on the micron and sub-micron length scale. We found that with increasing deformation fixed into the SMP substrate, wrinkled topographies with increasing amplitudes, decreasing wavelengths, and increasing degree of wrinkle orientation were achieved.Analysis of the cellular response to previously wrinkled (static) substrates revealed that cell nuclear alignment increased as SMP deformation increased. Analysis of the cellular response to an actively wrinkling substrate demonstrated that cell alignment can be controlled by triggering wrinkle formation. These findings demonstrate that the amount of deformation fixed (and later recovered) in an SMP bilayer system can be used to control the resulting wrinkle characteristics and cell mechanobiological response. The tailored and dynamic substrate functionality provided by this approach is expected to enable new investigation and understanding of cell mechanobiology.

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ResearchCite this article: Baker RM, Brasch ME, Manning ML, Henderson JH. 2014 Automated, contour-based tracking and analysis of cell behaviour over long time scales in environments of varying complexity and cell density. J. R. Soc. Understanding single and collective cell motility in model environments is foundational to many current research efforts in biology and bioengineering. To elucidate subtle differences in cell behaviour despite cell-to-cell variability, we introduce an algorithm for tracking large numbers of cells for long time periods and present a set of physics-based metrics that quantify differences in cell trajectories. Our algorithm, termed automated contour-based tracking for in vitro environments (ACTIVE), was designed for adherent cell populations subject to nuclear staining or transfection. ACTIVE is distinct from existing tracking software because it accommodates both variability in image intensity and multi-cell interactions, such as divisions and occlusions. When applied to low-contrast images from live-cell experiments, ACTIVE reduced error in analysing cell occlusion events by as much as 43% compared with a benchmark-tracking program while simultaneously tracking cell divisions and resulting daughter-daughter cell relationships. The large dataset generated by ACTIVE allowed us to develop metrics that capture subtle differences between cell trajectories on different substrates. We present cell motility data for thousands of cells studied at varying densities on shape-memory-polymer--based nanotopographies and identify several quantitative differences, including an unanticipated difference between two 'control' substrates. We expect that ACTIVE will be immediately useful to researchers who require accurate, long-time-scale motility data for many cells.

Soft, anisotropic materials, such as myocardium in the heart and the extracellular matrix surrounding cells, are commonly found in nature. This anisotropy leads to specialized responses and is imperative to material functionality, yet few soft materials exhibiting similar anisotropy have been developed. Our group introduced an anisotropic shape memory elastomeric composite (A-SMEC) composed of non-woven, aligned polymer fibers embedded in an elastomeric matrix. The composite exhibited shape memory (SM) behavior with significant anisotropy in room-temperature shape fixing. Here, we exploit this anisotropy by bonding together laminates with oblique anisotropy such that tensile deformation at room temperature - mechanical programming - results in coiling. This response is a breakthrough in mechanical programming, since non-affine shape change is achieved by simply stretching the layered A-SMECs at room temperature. We will show that pitch and curvature of curled geometries depend on fiber orientations and the degree of strain programmed into the material. To validate experimental results, a model was developed that captures the viscoplastic response of A-SMECs. Theoretical results correlated well with experimental data, supporting our conclusions and ensuring attainability of predictable curling geometries. We envision these smart, soft, shape changing materials will have aerospace and medical applications.

Recent advances in shape memory polymers have enabled the study of programmable, shape-changing, cytocompatible tissue engineering scaffolds. For treatment of bone defects, scaffolds with shape memory functionality have been studied for their potential for minimally invasive delivery, conformal fitting to defect margins, and defect stabilization. However, the extent to which the osteogenic differentiation capacity of stem cells resident in shape memory scaffolds is preserved following programmed shape change has not yet been determined. As a result, the feasibility of shape memory polymer scaffolds being employed in stem cell-based treatment strategies remains unclear. To test the hypothesis that stem cell osteogenic differentiation can be preserved during and following triggering of programmed architectural changes in shape memory polymer scaffolds, human adipose-derived stem cells were seeded in shape memory polymer foam scaffolds or in shape memory polymer fibrous scaffolds programmed to expand or contract, respectively, when warmed to body temperature. Osteogenic differentiation in shape-changing and control scaffolds was compared using mineral deposition, protein production, and gene expression assays. For both shape-changing and control scaffolds, qualitatively and quantitatively comparable amounts of mineral deposition were observed; comparable levels of alkaline phosphatase activity were measured; and no significant differences in the expression of genetic markers of osteogenesis were detected. These findings support the feasibility of employing shape memory in scaffolds for stem cell-based therapies for bone repair.

Foams prepared from shape memory polymers (SMPs) offer the potential for low density materials that can be triggered to deploy with a large volume change, unlike their solid counterparts that do so at near-constant volume. While examples of shape memory foams have been reported in the past, they have been limited to dual SMPs: those polymers featuring one switching transition between an arbitrarily programmed shape and a single permanent shape established by constituent crosslinks. Meanwhile, advances by SMP researchers have led to several approaches toward triple- or multi-shape polymers that feature more than one switching phase and thus a multitude of temporary shapes allowing for a complex sequence of shape deployments. Here, we report the design, preparation, and characterization of a triple shape memory polymeric foam that is open cell in nature and features a two phase, crosslinked SMP with a glass transition temperature of one phase at a temperature lower than a melting transition of the second phase. The soft materials were observed to feature high fidelity, repeatable triple shape behavior, characterized in compression and demonstrated for complex deployment by fixing a combination of foam compression and bending. We further explored the wettability of the foams, revealing composition-dependent behavior favorable for future work in biomedical investigations.

The ability of a three-dimensional scaffold to support cell seeding prior to implantation is a critical criterion for many scaffold-based tissue engineering and regenerative medicine strategies. Shape memory polymer functionality may present important new opportunities and challenges in cell seeding, but the extent to which shape memory activation can positively or negatively affect cell seeding has yet to be reported. The goal of this study was to determine whether shape memory activation can affect cell seeding. The hypothesis was that shape memory activation of porous scaffolds during cell seeding can affect both the number of cells seeded in a scaffold and the distribution (in terms of average infiltration distance) of cells following seeding. Here, we used a porous shape memory foam scaffold programmed to expand when triggered to study cell number and average cell infiltration distance following shape memory activation. We found that shape memory activation can affect both the number of cells and the average cell infiltration distance. The effect was found to be a function of rate of shape change and scaffold pore interconnectivity. Magnitude of shape change had no effect. Only reductions in cell number and infiltration distance (relative to control and benchmark) were observed. The findings suggest that strategies for tissue engineering and regenerative medicine that involve shape memory activation in the presence of a cell-containing medium in vitro or in vivo should consider how recovery rate and scaffold pore interconnectivity may ultimately impact cell seeding.