Supporting data for "Developing Advanced Scaffolds and Constructs for Liver Tissue Engineering"

The liver is a vital organ in human bodies, maintaining body’s homeostasis. It has an extraordinary regeneration ability, allowing it to re-grow even after substantial resection. However, when this regenerative capability is overwhelmed by excessive alcohol use, viral infection, cancer, etc., chronic or acute liver failure occurs. Liver diseases/failures are a growing global problem. Currently, liver transplantation is the only viable treatment for severely compromised liver. But the scarcity of donor organs heavily limits liver transplantation. Consequently, liver tissue engineering (LTE) has gained momentum and develops artificial structures to support liver healing and regeneration. 3D printing and bioprinting, the powerful manufacturing/biomanufacturing platforms, can create complex tissue engineering (TE) scaffolds and cell-laden living structures and thus significantly advance functional liver tissue regeneration. This project aims to develop advanced 3D printed scaffolds and bioprinted cell-scaffold constructs for LTE by mimicking liver’s native environment, integrating therapeutic functions such as localized drug delivery, antioxidation activity, and antibacterial protection, and promoting hepatocyte viability and functions.First, as having suitable inks for extrusion-based 3D printing in LTE is still a challenge, the research focused on developing new printing inks by chemically modifying natural polymers such as hyaluronic acid (HA) and collagen. A series of double-bond modified and oxidatively modified natural polymers were synthesized and their properties were studied. The new hydrogel inks were screened through rheological analyses and 3D printing into complex-structure scaffolds, e.g., hexagonal honeycomb hepatic lobule (the smallest unit of liver). Printability, mechanical strength and porosity were optimized.Second, as excessive oxidative stress and bacteria impede healing of liver wounds, antioxidative and antibacterial hydrogels capable of eliminating excessive reactive oxygen species (ROS) and hence oxidative stress (which is generated during liver injury or regeneration) and bacteria were 3D printed into LTE scaffolds. These new hydrogel scaffolds used a natural antioxidant, tannic acid, and antimicrobial quaternized chitosan. In vitro assessments showed excellent antioxidation effects and antimicrobial properties. These functionalized scaffolds also exhibited excellent biocompatibility for LTE.Third, anticancer drug-containing scaffolds were 3D printed for postoperative hepatocellular carcinoma patients. Printing inks were blends of double-bond modified carboxymethyl, oxidized dextran and xanthan gum dispersed with sorafenib-containing poly(lactic-co-glycolic acid) microspheres, as well as doxorubicin-containing blends of oxidized dextran and gelatin methacryloyl (GelMA). These inks could form pH-response hydrogel scaffolds via Schiff bond. Drug encapsulation and release were optimized to achieve localized and sustained delivery. In vitro results showed released drugs could effectively kill liver cancer cells. Also, hepatocytes proliferated well on scaffolds. These scaffolds provide dual functions of preventing tumour recurrence and assisting tissue regeneration.Finally, 3D bioprinting was used to develop cell-scaffold constructs for LTE, eliminating cell seeding and penetration problems in conventional TE strategies. Printing inks were composed of glycidyl methacrylate modified HA (HA-GMA), gelatin and GelMA. Bioinks were formed by mixing hepatocytes with printing inks. 3D printed hydrogel scaffolds were more conducive to cell attachment and growth than control. As observed through live/dead assay, albumin secretion, urea secretion, etc., 3D bioprinted liver tissue-mimicking constructs showed good hepatocyte viability in constructs and also liver function characteristics.