Materials called hydrogels have potential applications as scaffolds for tissue engineering, but methods are needed to assemble them into complex structures that mimic those found in nature. Just such a method has now been reported.
Hydrogels are jelly-like materials that are made up of water-absorbing, 3D polymer networks. Their excellent properties, such as deformability, biocompatibility and diverse chemical functionalities, have led to them being widely studied as materials for use in foods and medical implants, as scaffolds for cell culture and as drug-delivery vehicles. However, it is still challenging to construct complex, hierarchical architectures from them. Writing in Science Advances, Chiang et al.1 report a sophisticated method for the programmable assembly of high-order hydrogel architectures.
One reason for the interest in hydrogel assembly is that the resulting structures could be used to mimic the morphology of living tissues. Such tissues often contain many cell types that form hierarchical structures from various microscopic modular units, such as kidney nephrons, liver lobules and pancreatic islets. In the past few years, 'bottom-up' strategies — in which microscopic cellular building blocks are assembled into dense hierarchical 3D constructs — have attracted attention for tissue construction2,3. The building blocks can be categorized into three standard shapes: points, lines and planes4. Chiang and colleagues use their method to make a cell-laden hydrogel architecture that belongs to the plane category, which can stack or roll up into higher-order structures.
The authors developed an electromicrofluidic device that effectively allowed them to assemble hydrogels on a chip. More specifically, they used a fluid-handling technique called electrowetting to dispense, mix, transport and position single droplets (0.1 microlitres in volume) of a polymer solution into designated spots. They then used a technique called dielectrophoresis to manipulate dielectric (insulator) particles suspended within the droplets into a controlled pattern. They used positive dielectrophoresis to gather the particles in the high region of an electric field; and negative dielectrophoresis to gather those in the low electric-field region. In the final step of the process, the authors converted the droplets into hydrogels by crosslinking the dissolved polymer molecules using one of three methods (light- or heat-induced crosslinking, or chemical crosslinking).
In one example, they manipulated nine droplets, each containing a different fluorescent dye, into a 3 × 3 tiled pattern. They then crosslinked the polymers in the droplets using ultraviolet light to obtain a seamless heterogeneous hydrogel architecture (Fig. 1a) that could be handled with tweezers. Chiang et al. also used their platform to arrange hydrogels into controlled patterns after crosslinking.
The researchers used their system to arrange and culture cells (fibroblasts and cardiomyocytes) on biocompatible hydrogels such as poly(ethylene glycol) diacrylate (PEGDA) and gelatin methacryloyl (GelMA). They used electrowetting to move droplets containing both pre-crosslinked GelMA and cells into a hexagonal pattern, used dielectrophoresis to arrange the cells, and then immobilized the cells by crosslinking the polymers to form hydrogels (Fig. 1b). The cells survived the process well, suggesting that the procedure is able to manipulate and assemble cells without severely damaging them.
One advantage of Chiang and colleagues' method is that the technology provides a highly programmable 3D microenvironment for studying cell behaviour. For example, the authors constructed assemblies that contain hydrogels of different stiffnesses to observe the effect on cultured cells. They found that, when cardiomyocytes from newborn mice were seeded onto an assembly containing both GelMA and PEGDA hydrogels, the cells adhered mainly to the stiffer GelMA, where they exhibited their characteristic beating behaviour (contracting and lengthening). And when fibroblasts were cultured in GelMA hydrogels of different stiffnesses, the cells on the stiffer gels tended to exhibit increased polarization (that is, an asymmetrical shape or asymmetrical organization of cellular components) and aspect ratios (the ratio of length to width).
Many other systems have been reported for handling droplets or particles using electrowetting5 and dielectrophoresis6 — such technology is known as digital microfluidics7. But Chiang et al. have combined the two methods into a single platform. Their work also offers an advance for tissue engineering. In general, previously reported methods for tissue engineering precisely controlled the size of the cellular building blocks to be assembled, but the internal morphology of the blocks depended on self-assembly processes. By contrast, Chiang and colleagues' approach actively controls internal morphology, and might be an effective tool for rapidly building physiologically relevant tissue constructs.
Future work could make this technology even more useful by improving the resolution of the particle positioning, extending the process from two to three dimensions, scaling it up from the micrometre to the centimetre scale and finding ways to organize different types of cell in a single droplet. Because the assembly process is highly controllable, it could also become a powerful technology for 3D bioprinting and additive manufacturing — the industrial version of 3D printing. Footnote 1
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Advanced Materials Interfaces (2018)