Contactless 3D acoustic assembly of liquid capsules for bottom-up tissue engineering

In recent years, considerable efforts have been directed towards developing systems that replicate native tissue microarchitecture, enhancing cell viability and achieving close-to-native cellular organization. Despite advancements in various assembly methods, scalability and cell viability remain challenging due to the time consuming nature of certain approaches. Acoustic assembly has emerged as a powerful technology for modular units' assembly, leveraging sound waves to achieve rapid, contactless spatial arrangement by fine-tuning parameters such as frequency, amplitude, and chamber geometry. Here we present a system that employs acoustic waves to generate spatial patterns of liquid-core microcapsules, encapsulating poly-caprolactone surface-functionalized microparticles and umbilical cord-derived mesenchymal stem cells. The microcapsules were produced using electrohydrodynamic atomization in conjugation with an aqueous two-phase system and subsequently embedded in gelatin methacrylate. Acoustic waves were then applied to assemble the liquid-core microcapsules in well-defined patterns within the hydrogel precursor followed by crosslinking for structural stability. This approach allows us to define spatial patterns with precision, aligning with simulation predictions. The liquid nature of the microcapsules' core permits the organization of cells within the space towards the formation of microtissues decoupled from the external environment. The patterned constructs maintained cell viability for 14 days, facilitating the formation of microaggregates within liquid-core microcapsules and maintained organized microstructures. To explore the versatility of this system, we successfully patterned and stacked multiple layers of microcapsules, increasing structural complexity. Furthermore, we demonstrated its ability to support co-culture by seeding human umbilical vein endothelial cells onto the constructs as a proof of concept for promoting enhanced cellular interactions. This platform offers a scalable, versatile solution for developing tissue-mimetic multiscale constructs with tunable complexity, enabling rapid and non-contact assembly, making it a valuable tool for advancing in vitro models and studying complex cellular interactions.