Xeno-Free 3D Bioprinted Liver Model for Hepatotoxicity Assessment

• Published in: International journal of molecular sciences Vol. 25; no. 3; p. 1811
• Main Authors: Ali, Ahmed S M, Berg, Johanna, Roehrs, Viola, Wu, Dongwei, Hackethal, Johannes, Braeuning, Albert, Woelken, Lisa, Rauh, Cornelia, Kurreck, Jens
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 01-02-2024
• Subjects: 3-D printers; 3D bioprinting; Adaptation; Analysis; Animal experimentation; Animal welfare; Animals; Bioprinting; Cell culture; Chemical and Drug Induced Liver Injury; chemically defined media; Collagen; Cryopreservation; Dengue fever; Drug-Related Side Effects and Adverse Reactions; Epidermal growth factor; Ethics; Ethylenediaminetetraacetic acid; FBS-free; Hepatitis; HuH-7 cell line; Laboratory animals; Liver; Liver diseases; liver model; Marine toxins; Polyvinyl alcohol; Printing, Three-Dimensional; Tissue Engineering; Tissue Scaffolds; Toxicity; Viruses; xeno-free; Germany; HuH-7 cell line; chemically defined media; 3D bioprinting; FBS-free; liver model; xeno-free
• ISSN: 1422-0067; 1661-6596; 1422-0067
• DOI: 10.3390/ijms25031811

Three-dimensional (3D) bioprinting is one of the most promising methodologies that are currently in development for the replacement of animal experiments. Bioprinting and most alternative technologies rely on animal-derived materials, which compromises the intent of animal welfare and results in the generation of chimeric systems of limited value. The current study therefore presents the first bioprinted liver model that is entirely void of animal-derived constituents. Initially, HuH-7 cells underwent adaptation to a chemically defined medium (CDM). The adapted cells exhibited high survival rates (85-92%) after cryopreservation in chemically defined freezing media, comparable to those preserved in standard medium (86-92%). Xeno-free bioink for 3D bioprinting yielded liver models with high relative cell viability (97-101%), akin to a Matrigel-based liver model (83-102%) after 15 days of culture. The established xeno-free model was used for toxicity testing of a marine biotoxin, okadaic acid (OA). In 2D culture, OA toxicity was virtually identical for cells cultured under standard conditions and in CDM. In the xeno-free bioprinted liver model, 3-fold higher concentrations of OA than in the respective monolayer culture were needed to induce cytotoxicity. In conclusion, this study describes for the first time the development of a xeno-free 3D bioprinted liver model and its applicability for research purposes.