Engineering a GelMA–dECM-based 3D bioprinted liver fibrosis model: methotrexate-induced functional and molecular validation

Abstract

Three-dimensional (3D) bioprinting represents a cutting-edge advancement in additive manufacturing, offering unprecedented precision in fabricating in vitro models that recapitulate native tissue architecture and function. Here, we present the fabrication of a bioprinted hepatic construct using a composite bioink composed of in-house synthesized gelatin methacryloyl (GelMA), rat liver-derived decellularized extracellular matrix (dECM), and HepG2 cells. GelMA imparted mechanical integrity and biocompatibility, while the liver-specific dECM provided bioactive cues critical for recapitulating the hepatic microenvironment. Constructs were crosslinked using microbial transglutaminase and a photoinitiator to ensure structural stability and shape fidelity. Functional characterization included cytocompatibility assays (MTT, live/dead), metabolic activity assays (albumin and urea secretion), and liver-specific enzyme analysis (LDH, ALT, and ALP), alongside gene expression profiling, all of which confirmed hepatic function within the constructs. This synergy enhances cellular functionality and supports accurate fibrosis modeling for translational research. To establish a fibrosis model, methotrexate (MTX) was introduced, resulting in functional decline and upregulation of fibrosis-associated genes, thereby validating the fibrotic phenotype. This study demonstrates the development of a robust and physiologically relevant 3D bioprinted in vitro platform for modeling MTX-induced liver fibrosis, providing a promising tool for preclinical drug screening and translational research in hepatic disease. This study demonstrates a robust and physiologically relevant 3D bioprinted in vitro model of methotrexate-induced liver fibrosis, offering a valuable platform for translational applications in drug screening and hepatic disease modelling.