Highly functional 3D fibrin-embedded hepatocyte tissues with long-term functional stability constructed via the CAViTs method.

Construction of biomaterial-based in vitro liver model with native-like structural polarity and functions represents a critical challenge for conducting safety assessments. Conventional models often lack continuous bile canalicular networks or long-term stability, limiting their utility for such assessments. In this study, we developed fibrin-embedded hepatocyte tissue (FEHT) using a unique tissue engineering technique, the "Cell-Assembled Viscous Tissue by Sedimentation" (CAViTs) method. FEHT exhibited dense, interconnected three-dimensional bile canalicular networks that maintained structural and functional maturity for at least 30 days. Gene expression analyses demonstrated FEHT profiles significantly closer to native human liver tissue than conventional spheroid or sandwich cultures. Sensitivity to cyclosporine A-induced cholestatic toxicity was >20 times higher in FEHT than in the spheroid model. Further, FEHT maintained high viability and metabolic activity for over two weeks without medium replenishment. Thus, FEHT represents a robust in vitro model that mimics the physiological liver architecture and could provide a useful tool for long-term pharmacokinetic and safety assessments.