The hypertrophic cartilage induction influences the building-block capacity of human adipose stem/stromal cell spheroids for biofabrication

Abstract

As an alternative to the classical tissue engineering approach, bottom-up tissue engineering emerges using building blocks in bioassembly technologies. Spheroids can be used as building blocks to reach a highly complex ordered tissue by their fusion (bioassembly), representing the foundation of biofabrication. In this study, we analyzed the biomechanical properties and the fusion capacity of human adipose stem/stromal cell (ASC) we spheroids during an in vitro model of hypertrophic cartilage established by our research group. Hypertrophic induced-ASC spheroids showed a statistically significant higher Young's modulus at weeks 2 (P < .001) and 3 (P < .0005) compared with non-induced. After fusion, non-induced and induced-ASC spheroids increased the contact area and decreased their pairs' total length. At weeks 3 and 5, induced-ASC spheroids did not fuse completely, and the cells migrate preferentially in the fusion contact region. Alizarin red O staining showed the highest intensity of staining in the fused induced-ASC spheroids at week 5, together with intense staining for collagen type I and osteocalcin. Transmission electron microscopy and element content analysis (X-ray Energy Dispersive Spectroscopy) revealed in the fused quartet at week 3 a crystal-like structure. Hypertrophic induction interferes with the intrinsic capacity of spheroids to fuse. The measurements of contact between spheroids during the fusion process, together with the change in viscoelastic profile to the plastic, will impact the establishment of bioassembly protocols using hypertrophic induced-ASC spheroids as building blocks in biofabrication.

Keywords: adipose stromal/stem cells; bioassembly; biofabrication; biomechanical; building blocks; fusion; hypertrophic cartilage; osteogenesis; spheroids.