The two main components are ОІCD-AAm and Ad-AAm. Figure 1A shows a schematic representation of the composition and of the photopolymerization procedure. In the first step, we developed a photoresist that can be used for 3D laser lithography. RESULTS Stimuli-responsive microstructures based on host-guest complexes We further demonstrate this aspect by modified structures, which asymmetrically stretch cells on one side only, and we study the response to this localized stimulation. In the last part, we emphasize the flexibility of our approach to changing and adapting the geometrical design of the scaffolds according to the specific application. We further show that this reorganization crucially depends on the activity of myosin II molecular motors. We highlight this by visualizing a strong reorganization of the actin cytoskeleton as a response to the external displacement. Another major advantage of this system is the ability to chemically fix the cells and the scaffolds at any point during the experiment, thus allowing us to stain the cells and their subcellular compartments while keeping them in the stretched state. Furthermore, this fabrication technique allows us to integrate nonresponsive scaffolds as controls in all experiments. Our approach is inherently scalable and enables to simultaneously study a large number of individual cells in well-defined microenvironments. Using this approach, we stretch single cells in a spatially and temporally well-defined manner and precisely track their response as a function of time via digital image correlation. Supramolecular polymers ( 22, 23) could provide an advantage over the abovementioned materials, if appropriate host or guest molecules are selected for the stimulation ( 24). Despite a number of studies demonstrating the dynamic control of cells using hydrogels responsive to temperature ( 18), pH ( 19), enzymes ( 20), or illumination with ultraviolet light ( 21), these applications are limited because the formation and cleavage of bonds are often performed under harsh conditions.

One crucial constraint for the application in cell biology is a specific physiological stimulus of the material that does not influence or alter the behavior of the cells. In recent years, a large number of these material systems with numerous applications in the macroscopic ( 13) and microscopic ( 14, 15) regime have been investigated and extensively reviewed ( 16, 17). However, the transition from passive to active systems requires responsive materials that can be stimulated on demand ( 12). In the past, 3D laser lithography has successfully been used to manufacture cell scaffolds with tailored geometry and spatially functionalized surfaces ( 11). Pharmacological inhibition or knockout of nonmuscle myosin 2A prevents these adjustments, suggesting that cellular tensional homeostasis strongly depends on functional myosin motors. When the stretch is released, traction forces gradually decrease until the initial set point is retrieved. After application of an equibiaxial stretch of up to 25%, cells remodel their actin cytoskeleton, double their traction forces, and equilibrate at a new dynamic set point within 30 min. Cells adhering in these scaffolds build up initial traction forces of ~80 nN.

This allows reversible actuation under physiological conditions by application of soluble competitive guests. The key material is a stimuli-responsive photoresist containing cross-links formed by noncovalent, directional interactions between ОІ-cyclodextrin (host) and adamantane (guest). Here, we introduce stimuli-responsive composite scaffolds fabricated by three-dimensional (3D) laser lithography to simultaneously stretch large numbers of single cells in tailored 3D microenvironments. Many essential cellular processes are regulated by mechanical properties of their microenvironment.

Mechanical equilibrium 3d