Controlled degradation of HepG2 Laden microspheres induced spheroid formation in alginate hydrogel.

Abstract

Liver tissue engineering has been an emerging field that shows promising advances to regenerate the liver and treat prevalent liver diseases in order to cope with the shortage of liver donors. Conventional approaches focus on cell-based therapies such as the fabrication of a three-dimensional (3D) liver construct to complement cell transplantation and this 3D environment is needed to promote liver cell viability and maintenance of their phenotype. Previous studies have demonstrated a cell delivery composite model with the use of genipin crosslinked gelatin microspheres (GC/GM) as carriers to mediate hepatocellular aggregation, however, with uncontrolled size. In this study, alginate is employed to encapsulate HepG2 laden microspheres in a 3D hydrogel bulk where a collagenase, MMP-9 is introduced to degrade the GC/GM and create well-defined spaces within the alginate to induce spheroid formation. This microcarrier/alginate composite model has allowed for spheroid formation in a size and shape controllable manner where the spheroids take up the size range of the GC/GM (75-160 μm) with a maximum size of 180 μm achieved after two weeks of culture. Spheroids obtained produced higher cell viability and superior functionalities in terms of albumin and urea secretion as well as gene expression of albumin and cytochrome P450. In contrast, HepG2 in control without GC/GM degradation limits growth and has no presence of spheroid. This model not only can be used for generating functional liver spheroids in a large-scale but also can immobilize cell laden microspheres with alginate at the site of injection.