3D bioprinted thick hepatic constructs with vascular network as a physiologically relevant in vitro organ model

Establishing adequate vascularization to engineered organs remains a significant challenge that must be addressed. This study presents a novel approach to fabricating viable thick metabolic tissue (>1 cm³) for applications in human physiology, fundamental biology, and medicine. We designed a tissue construct with a gyroid-shaped architecture to enable uniform flow and surface shear stress that adequately covers the inner surfaces of cell-laden constructs. The constructs (1 × 1 × 1 cm³) were fabricated using a digital light projection (DLP) printer with a cell-laden poly(ethylene glycol) (PEG)/gelatin methacryloyl (GelMA) bioink combined with human hepatocytes (HepG2), followed by coating the interconnected vascular channels with human endothelial cells (ECs). These constructs were then placed in flow chambers connected to a medium reservoir for continuous perfusion for up to 30 days. The constructs retained their original dimensions, and the cells maintained a greater than 85 % viability at all time points. Immunofluorescent staining confirmed hepatocytes and ECs using cell-specific markers (HNF4-α/albumin for hepatocytes and vWF for ECs). The EC layer effectively lined the vascular lumens, while viable hepatocyte aggregates populated the interior of the constructs. Functional assays demonstrated that the hepatocytes produced albumin and bilirubin at levels comparable to those observed in humans, validating the metabolic functionality of the hepatic tissue constructs. This study successfully developed thick, vascularized human hepatic tissue in an <i>in vitro</i> environment, maintaining functionality comparable to native liver cells over 30 days. The innovative gyroid design applied in these organ constructs represents a significant advancement in developing physiologically relevant <i>in vitro</i> vascularized organ models.