Whole Liver Derived Acellular Extracellular Matrix for Bioengineering of Liver Constructs: An Updated Review

Abstract

Biomaterial templates play a critical role in establishing and bioinstructing three-dimensional cellular growth, proliferation and spatial morphogenetic processes that culminate in the development of physiologically relevant in vitro liver models. Various natural and synthetic polymeric biomaterials are currently available to construct biomimetic cell culture environments to investigate hepatic cell-matrix interactions, drug response assessment, toxicity, and disease mechanisms. One specific class of natural biomaterials consists of the decellularized liver extracellular matrix (dECM) derived from xenogeneic or allogeneic sources, which is rich in bioconstituents essential for the ultrastructural stability, function, repair, and regeneration of tissues/organs. Considering the significance of the key design blueprints of organ-specific acellular substrates for physiologically active graft reconstruction, herein we showcased the latest updates in the field of liver decellularization-recellularization technologies. Overall, this review highlights the potential of acellular matrix as a promising biomaterial in light of recent advances in the preparation of liver-specific whole organ scaffolds. The review concludes with a discussion of the challenges and future prospects of liver-specific decellularized materials in the direction of translational research.

Keywords: decellularization; liver; recellularization; scaffolds; tissue and organoids.

Figure 1

Schematic diagram of extracellular matrix components showing biochemical signaling molecules. Figure 1 is reproduced with copyright permission from [66], Wiley.

Figure 2

Macroscopic images for the preparation and characterization of native and decellularized rat liver (a). (a1) Fesh liver, (a2) frozen liver-after 24 h, (a3) liver perfused with distilled water over 30 min through both portal vein and bile duct systems, (a4) liver perfused with 4% TritonX-100 solution over 3 h and (a5) liver perfused with 0.5% SDS solution over 3 h. (b) Cross sections of histological and immunofluorescence images of native and decellularized livers stained with hematoxylin-eosin, fibronectin (red), collagen I (green), laminin (red), and collagen IV (red) showing the overall structure, sulfated GAG, and collagen, respectively. (c) Scanning electron microscopy images showing ultrastructure of normal and decellularized livers treated with 0.4% Triton and 0.5% SDS based protocols. SEM images of (c1–c3) normal liver and (c4–c6) decellularized liver. Intact and smooth vessel wall (c5) and extracellular matrix parenchyma (c6) with hepatocyte-sized free space in the decellularized liver matrix can be clearly observed. Yellow triangles in indicate the vessel walls native and decellularized liver. (d) Appearance of over perfused decellularized liver left lobes, with vasculature preserved. (e) Confirmation of DNA removal native and decellularized rat liver. Data are expressed as means ± SD (n = 3). *** p < 0.001. Figure 2 is adopted with copyright permission from [136], Elsevier.

Figure 3

Decellularized rat liver scaffold after reendothelialization. (A) Adhesion of endothelial cells to the vascular walls and infiltration towards the extravascular. (B) Anastomosis of the PV and IVC of the reendothelialized liver scaffold to the abdominal aorta and IVC of the recipient rat. (C) Ultrasound imaging showed the blood flow into the transplanted scaffold 8 days post-transplantation. Figure 3 is reproduced with copyright permission from [149], Wiley.

Figure 4

Transplantation (auxiliary) of the bioengineered human liver graft recellularized with induced pluripotent stem cells (iPSCs). (A) Schematic description of the surgical techniques for the transplantation of auxiliary liver grafts or human bioengineered liver grafts: (1) after right nephrectomy, (2) PV and IVC were exposed. (3) IVC anastomosis (end to side). (4) PV anastomosis (end to side). (5) After reperfusion. (6) Before closing abdomen. (B) Microscopic images of the engineered liver graft three–four days post-implantation. (C) immunofluorescence staining of recellularized auxiliary graft post-transplantation (left), compared to human adult liver tissue (middle), and rat recipient liver (right). H&E, hematoxylin and eosin; h-ALB, human-specific albumin; h-CD31, human-specific CD31; h-CK7, human-specific cytokeratin 7. Sections were counterstained with Hoechst (blue stain). (D) The serum concentration of human specific A1AT and human-specific ALB was measured by ELISA at day 4 post-transplantation. Abbreviation: N.D: Not determined. Figure 4 is adopted with copyright permission from [150], Elsevier.

Figure 5

Engraftment (in vivo) of the bioengineered liver graft at different time points (postoperative days 14 and 28). (A,B) CT images of the transplanted graft on postoperative 14 (A) and postoperative 28 (B). Angiography through the intraportal infusion catheter on postoperative 14 (C) and postoperative 28 (D). Reconstructed three-dimensional CT image of the remnant native liver and the transplanted graft on postoperative 14 (E) and postoperative 28 (F). Time courses of the CT images for the transplanted graft on postoperative 14 (G) and postoperative 28 (H), Blue tube depicts IVC and HV, and pink tube depicts PV. Time course of the mean CT numbers for the graft and remnant native liver on postoperative 14 (I) and postoperative 28 (J). Intraoperative images of the graft on postoperative 28 (K,L). Macroscopic images of the procured graft on postoperative 28 (M). The arrowheads indicate PV in the graft. Macroscopic image of the stenosed anastomosis site of the procured graft on POD 28 (N). Patency rate was estimated to be approximately 11.9%. The dashed red and white lines show the anastomosis site and patent area, respectively. Weight of the procured graft on postoperative 28 (O). Figure 5 is adopted with copyright permission from [156], Elsevier.