Liver ductal organoids reconstruct intrahepatic biliary trees in decellularized liver grafts.

Three-dimensional scaffolds decellularized from native organs are a promising technique to establish engineered liver grafts and overcome the current shortage of donor organs. However, limited sources of bile duct cells and inappropriate cell distribution in bioengineered liver grafts have hindered their practical application. Organoid technology is anticipated to be an excellent tool for the advancement of regenerative medicine. In the present study, we reconstructed intrahepatic bile ducts in a rat decellularized liver graft by recellularization with liver ductal organoids. Using an ex vivo perfusion culture system, we demonstrated the biliary characteristics of repopulated mouse liver organoids, which maintained bile duct markers and reconstructed biliary tree-like networks with luminal structures. We also established a method for the co-recellularization with engineered bile ducts and primary hepatocytes, revealing the appropriate cell distribution to mimic the native liver. We then utilized this model in human organoids to demonstrate the reconstructed bile ducts. Our results show that liver ductal organoids are a potential cell source for bile ducts from bioengineered liver grafts using three-dimensional scaffolds.

Decellularized liver-regenerative 3D printing biomaterials for cell-based liver therapies via a designed procedure combined with papain-containing reagent treatments and supercritical fluids

BACKGROUND: The biologic scaffolds derived from decellularized tissues and organs have been successfully developed in a variety of preclinical and/or clinical studies. OBJECTIVE: The new decellularized liver-regenerative 3D printing biomaterials were designed and prepared for cell-based liver therapies. METHODS: An extraction process was employed to remove the tissue and cellular molecules from porcine liver via pretreatment of supercritical fluid of carbon dioxide (ScCO2). Varying porosities of the decellularized liver tissues were created using papain-containing reagent treatments after ScCO2. RESULTS: The resulting liver-regenerative 3D printing biomaterials of decellularized liver collagen scaffolds were characterized by Fourier transform infrared spectroscopy, thermo-gravimetric analysis, differential scanning calorimetry and scanning electron microscopy. CONCLUSIONS: The decellularized liver collagen scaffolds with good thermal stability (>150 °C) were obtained and employed as liver-regenerative 3D printing biomaterials for cell-based liver therapies.

Implanted subcutaneous versus intraperitoneal bioscaffold seeded with hepatocyte-like cells: functional evaluation

Background and objectives The X-linked bleeding disorder, hemophilia A, is caused by defective production of factor VIII (FVIII). Hemophilic patients require regular FVIII infusions. Recombinant factor replacement poses the safest line of therapy. However, its main drawbacks are high expenses and the higher liability for formation of inhibitors. Recent studies confirmed the ability of bone marrow-derived stem cells to secrete FVIII. This study aims to generate bioscaffold from decellularized liver and subsequently seed it with trans-differentiated human stem cells into hepatic-like cells. This scaffold can then be implanted intraperitoneally or subcutaneously to provide FVIII. Methods After generation of the bioscaffold, seeding of discoid scaffolds with trans-differentiated human hepatocyte-like cells was performed. Then, the generated organoid was implanted into peritoneal cavity or subcutaneous tissue of experimental rats. Results Serum human FVIII was significantly increased in rats subjected to subcutaneous implantation compared intraperitoneal implantation. Immunostaining for detecting Cytokeratin 19 and human anti-globulin confirmed the presence of mature human hepatocytes that were significantly increased in subcutaneous implanted scaffold compared to the intraperitoneal one. Conclusion Implantation of decellularized bioscaffold seeded with trans-differentiated stem cells in rats was successful to establish production of FVIII. Subcutaneous implantation showed higher FVIII levels than intraperitoneal implantation.

Biocompatibility of a decellularized liver scaffold in studies in vitro

The development of multicomponent threedimensional structures based on decellularized tissue is a perspective alternative for organ transplantation in end-stage liver disease. The technology of rat liver decellularization is presented which consist in sequential perfusion of organ through the portal vein and use of 0.1 % sodium dodecyl sulfate as an active solution. The absence of the cytotoxic effect of decellularized scaffolds on allogeneic splenocytes and multipotent mesenchymal stromal cells was established. The obtained liver scaffolds are biocompatible in cells cultures and correspond criteria for cell carriers.