Decellularized Liver Matrix-Modified Cryogel Scaffolds as Potential Hepatocyte Carriers in Bioartificial Liver Support Systems and Implantable Liver Constructs

Abstract

Recent progress in the use of decellularized organ scaffolds as regenerative matrices for tissue engineering holds great promise in addressing the issue of donor organ shortage. Decellularization preserves the mechanical integrity, composition, and microvasculature critical for zonation of hepatocytes in the liver. Earlier studies have reported the possibility of repopulating decellularized matrices with hepatic cell lines or stem cells to improve liver regeneration. In this work, we study the versatility of the decellularized liver matrix as a substrate coating of three-dimensional cryogel scaffolds. The coated cryogels were analyzed for their ability to maintain hepatic cell growth and functionality in vitro, which was found to be significantly better than the uncoated cryogel scaffolds. The decellularized liver matrix-coated cryogel scaffolds were evaluated for their potential application as a cell-loaded bioreactor for bioartificial liver support and as an implantable liver construct. Extracorporeal connection of the coated cryogel bioreactor to a liver failure model showed improvement in liver function parameters. Additionally, offline clinical evaluation of the bioreactor using patient-derived liver failure plasma showed its efficacy in improving liver failure conditions by approximately 30-60%. Furthermore, implantation of the decellularized matrix-coated cryogel showed complete integration with the native tissue as confirmed by hematoxylin and eosin staining of tissue sections. HepG2 cells and primary human hepatocytes seeded in the coated cryogel scaffolds implanted in the liver failure model maintained functionality in terms of albumin synthesis and cytochrome P450 activity post 2 weeks of implantation. In addition, a 20-60% improvement in liver function parameters was observed post implantation. These results, put together, suggest a possibility of using the decellularized matrix-coated cryogel scaffolds for liver tissue engineering applications.

Decellularized liver matrix as substrates for rescue of acute hepatocytes toxicity

“…Despite the DLM‐based biomaterial providing the hepatocytes with a better 3D architecture, the hepatocytes are unable to maintain a steady growth rate for extended periods of time, and even lose their viability and functions after 5 days of culture. Similar results were also found in a previous study (Damania et al, ). Hepatocytes spheroid, which can create an efficient 3D tissue construct for maintaining hepatic functions for a long‐term culture may be carried out as a suitable cell source for further optimization of this promising research.…”

“…Glt‐HPA‐DLM as a 3D culture environment can effectively enhance the viability and functions of hepatocytes (Figure ). Previous study has been reported that decellularized liver ECM‐based scaffold can alleviate the injury conditions after partial hepatectomy treatment to some extent; besides, AST and ALT were decreased whereas bilirubin and albumin were increased after transplantation in vivo (Damania et al, ). In this study, the efficiency of DLM‐based biomaterial for ameliorating hepatotoxicants‐induced damaged hepatocytes is summarized below:…”

“…On the other hand, the decellularization procedure without using harsh detergents may result in structural integrity of ECM (Maghsoudlou et al, ); moreover, Triton X‐100, a mild detergent and typically used for liver decellularization (Badylak, Taylor, & Uygun, ), can only reduce cell component/DNA content without depleting any glycosaminoglycans (GAGs) during the washing process (Bruyneel & Carr, ). The effect could be attributed that GAGs and other components (collagen, proteoglycans) constitute the liver extracellular microenvironment, and also play an important role in hepatic functions and viability (Damania et al, ; Lee et al, ); furthermore, the wound ECM of GaIN‐damaged cells can be subsequently replaced by mature ECM and return to the preinjury state (Jonker, Dijkhuis, Boes, Hardonk, & Grond, ). Therefore, Glt‐HPA‐DLM can more easily ameliorate GaIN‐injured hepatocytes, rather than CCl 4 or CHCl 3 ‐injured hepatocytes.…”

A minimally‐invasive cryogel based approach for the development of human ectopic liver in a mouse model

“…Thus, hepatic carcinoma cells are competent alternative which are easy and inexpensive to culture. Further, in our previous study, we found no significant differences between primary hepatocytes and HepG2 cell lines in terms of relevant cell functionalities, when grown on a cryogel 3D matrix and implanted in vivo (Damania et al, 2018). Therefore, to demonstrate proof of concept, we used HepG2 cells to establish an ectopic liver using injectable approach due to the limited supply of freshly isolated primary human hepatocytes and minimizing animal use.…”

Tissue Engineering in Liver Regenerative Medicine: Insights into Novel Translational Technologies

“…By using the ECM of an acellularized liver, which can be integrated into the scaffold matrix, the highest similarity with the in vivo conditions can be achieved [50]. During decellularization, cells and other immunogenic factors are removed and only the natural scaffold of tissue remains.…”