Fmoc-3F-Phe-Arg-NH2 and Fmoc-3F-Phe-Asp-OH dipeptides undergo coassembly to form two-component nanofibril hydrogels. These hydrogels support the viability and growth of NIH 3T3 fibroblast cells. The supramolecular display of Arg and Asp at the nanofibril surface effectively mimics the integrin-binding RGD peptide of fibronectin, without covalent connection between the Arg and Asp functionality.

Thermoresponsive poly(N-isopropyl acrylamide) (PNIPAM) microgels were patterned on polystyrene substrates via dip coating, creating cytocompatible substrates that provided spatial control over cell adhesion. This simple dip coating method, which exploits variable substrate withdrawal speeds form particle suspension formed stripes of densely-packed PNIPAM microgels, while spacings between the stripes contained sparsely-distributed PNIPAM microgels. The assembly of three different PNIPAM microgel patterns, namely patterns composed of 50 μm stripes/50 μm spacings, 50 μm stripes/100 μm spacings, and 100 μm stripes/100 μm spacings was verified using high-resolution optical micrographs and ImageJ analysis. PNIPAM microgels existed as monolayers within stripes and spacings, as revealed by atomic force microscopy (AFM). Upon cell seeding on PNIPAM micropatterned substrates, NIH3T3 fibroblast cells preferentially adhered within spacings to form cell patterns. Three days after cell seeding, cells proliferated to form confluent cell layers. The thermoresponsiveness of the underlying PNIPAM microgels was then utilized to recover fibroblast cell sheets from substrates simply by lowering the temperature, without disrupting the underlying PNIPAM microgel patterns. Harvested cell sheets similar to these have been used for multiple tissue engineering applications. Also, this simple, low cost, template-free dip coating technique can be utilized to micropattern multifunctional PNIPAM microgels, generating complex stimuli-responsive substrates to study cell-material interactions and allow drug delivery to cells in a spatially and temporally-controlled manners.

For many tissue engineering applications and studies to understand how materials fundamentally affect cellular functions, it is important to have the ability to synthesize biomaterials that can mimic elements of native cellextracellular matrix interactions. Hydrogels possess many properties that are desirable for studying cell behavior. For example, hydrogels are biocompatible and can be biochemically and mechanically altered by exploiting the presentation of cell adhesive epitopes or by changing hydrogel crosslinking density. To establish physical and biochemical tunability, hydrogels can be engineered to alter their properties upon interaction with external driving forces such as pH, temperature, electric current, as well as exposure to cytocompatible irradiation. Additionally, hydrogels can be engineered to respond to enzymes secreted by cells, such as matrix metalloproteinases and hyaluronidases. This review details different strategies and mechanisms by which biomaterials, specifically hydrogels, can be manipulated dynamically to affect cell behavior. By employing the appropriate combination of stimuli and hydrogel composition and architecture, cell behavior such as adhesion, migration, proliferation, and differentiation can be controlled in real time. This three-dimensional control in cell behavior can help create programmable cell niches that can be useful for fundamental cell studies and in a variety of tissue engineering applications.

Protein organization on biomembranes and their dynamics are essential for cellular function. It is not clear, however, how protein binding may influence the assembly of underlying lipids or how the membrane structure leads to functional protein organization. Toward this goal, we investigated the effects of annexin a5 binding to biomimetic membranes using fluorescence imaging and correlation spectroscopy. Annexin a5 (anx a5), a peripheral intracellular protein that plays a membrane remodeling role in addition to other functions, binds specifically and tightly to anionic (e.g., phosphatidylserine)-containing membranes in the presence of calcium ion. Our fluorescence microscopy reveals that annexin likely forms assemblies, along with a more dispersed population, upon binding to anionic biomembranes in the presence of calcium ion, which is reflected in its two-component Brownian motion. To investigate the effects of annexin binding on the underlying lipids, we used specific acyl chain-labeled phospholipid analogs, NBD-phosphatidylcholine (NBD-PC) and NBD-phosphatidylserine (NBD-PS). We find that both NBD-labeled lipids cluster under anx a5 assemblies, as compared with when they are found under the dispersed annexin population, and NBD-PS exhibits two-component lateral diffusion under the annexin assemblies. In contrast, NBD-PC diffusion is slower by an order of magnitude under the annexin assemblies in contrast to its diffusion when not localized under anx a5 assemblies. Our results indicate that upon binding to membranes, the peripheral protein annexin organizes the underlying lipids into domains, which may have functional implications in vivo.