Fabrication Of Macroscopic Tissue Constructs With Different Shapes Via A Modular Approach

It remains challenges to fabricate macroscopic tissues of different shapes via traditional tissue engineering methodologies. In this work, we applied a "bottom-up" approach to engineer macroscopic tissues in a custom developed perfusion culture system, the effects of perfusion culture time, cell types and microcarrier types were investigated in details regarding tissue properties.It was shown that, when human fibroblasts (HF)(0.25×107cells/cm3) were dynamically seeded onto CytoporeTM microbeads in spinner flasks and cultivated for14days, cell density reached7.56×107cells/cm3at the end of culture, and cells occupied the whole surface area of the beads as well as the inner porous space while showing great viability based on scanning electron microscope (SEM) and live/dead staining assay. These microtissues were loaded onto the molds in custom-made perfusion culture system and cultivated for28days. It was found that the constructed macrotissues had high cell viability and homogeneous distribution of both cells and extracellular matrix (ECM) during the first14days; But at the time point of21d and28d, there existed dead cells and inhomogeneous distribution of cells and ECM. Moreover based on this finding, we successfully constructed macroscopic tissues with the shapes of alphabetical letters, round disk and tube with a14day perfusion culture. In addition, HF, mesenchymal stem cells (MSCs), and HepG2cells were also seeded onto either Cytopore or Cultispher microcarriers, and the formation of microtissues, as well as macroscopic tissues were compared, showing that HF and MSCs were more prone to tissue formation.Taken together, we demonstrated the feasibility of fabricating macroscopic tissues with different shapes using different cells and microcarriers though a modular approach. These studies prove the great potential of using cell-laden microbeads in tissue engineering applications.