Development of microfluidic-based 3D bioprinting systems to create hierarchical in vitro models of human tissues

Microfluidics is rapidly revolutionizing the scientific panorama, providing unmatched high-throughput platforms that find application in numerous areas of physics, chemistry, biology, and material science. In the context of tissue engineering, microfluidic chips have been proposed, in combination with bioactive materials, as promising tools for spinning cell-laden fibers with on-demand characteristics. However, cells encapsulated in filaments produced via microfluidic spinning technology (MST) are confined in a quasi-3D environment that fails to replicate the intricate 3D architecture of biological tissues. The recent synergistic combination of 3D bioprinting with microfluidic devices enables to create sophisticated microfibers and arrange them in 3D to realize three-dimensional functional replicas of human tissues. It is pivotal to mimic the structural heterogeneities of native tissues in terms of material composition, cell type, cell density and microstructural properties in order to realize functional 3D tissue models. In fact, these factors have a significant impact on both mechanical properties and the biological outcome. Recently, microfluidic-based systems have emerged as advanced tools to control in one-step the production of microfibers with tailored dimension, composition, and morphology. More specifically, microfluidic devices can be used to control fiber properties in real time through microchannels where various components can be mixed, split or emulsified. Ultimately, the macro-architectural control of 3D constructs is made possible by the coupling of microfluidic chips with a 3D printing system. In this work, we report the fabrication of a series of novel extrusion-based microfluidic printing heads that enable the spinning and deposition of microfibers with tailored characteristics in a three-dimensional environment. These tools allow for the 3D printing of fiber-based scaffolds with adjustable intra-fiber composition, cell density and microporosity. The study aims to realize functionally graded structures that simulate the change in properties (e.g., material composition, cell density, cell type population and microporosity) observed at the interface of two different tissues. Mechanical and morphological analysis have been carried out to evaluate the resulting internal microstructure, while the biological response of embedded cells was monitored with biological and biochemical assays that allow to investigate cell viability, expression of target proteins and production of specific enzymes, confirming the differentiation of bioprinted stem cells.