Portal venous repopulation of decellularised rat liver scaffolds with syngeneic bone marrow stem cells

Abstract

Liver transplantation is the only life-saving treatment for end-stage liver failure but is limited by the organ shortage and consequences of immunosuppression. Repopulation of decellularised scaffolds with recipient cells provides a theoretical solution, allowing reliable and timely organ sourcing without the need for immunosuppression. Recellularisation of the vasculature of decellularised liver scaffolds was investigated as an essential prerequisite to the survival of other parenchymal components. Liver decellularisation was carried out by portal vein perfusion using a detergent-based solution. Decellularised scaffolds were placed in a sterile perfusion apparatus consisting of a sealed organ chamber, functioning at 37 C in normal atmospheric conditions. The scaffold was perfused via portal vein with culture medium. A total of 10 7 primary cultured bone marrow stem cells, selected by plastic adherence, were infused into the scaffold, after which repopulated scaffolds were perfused for up to 30 days. The cultured stem cells were assessed for key marker expression using fluorescence-activated cell sorting (FACS), and recellularised scaffolds were analysed by light, electron and immunofluorescence microscopy. Stem cells were engrafted in portal, sinusoidal and hepatic vein compartments, with cell alignment reminiscent of endothelium. Cell surface marker expression altered following engraftment, from haematopoietic to endothelial phenotype, and engrafted cells expressed sinusoidal endothelial endocytic receptors (mannose, Fc and stabilin receptors). These results represent one step towards complete recellularisation of the liver vasculature and progress towards the objective of generating transplantable neo-organs.

Vascular reconstruction of the decellularized biomatrix for whole-organ engineering—a critical perspective and future strategies

“…The recellularized organ was cultured for 30 days. The cells showed markers for hepatic sinusoidal endothelial cells (mannose, Fc, and stabilin receptors), along with cell alignment reminiscent of the endothelium (Harper et al, 2020). Alternatively, many mechanisms have been proven concerning the role of MSCs as a paracrine factory in promoting angiogenesis (Caplan and Dennis, 2006).…”

Liver regeneration biology: Implications for liver tumour therapies

“…Repopulation of decellularised livers : Repopulation of decellularised liver scaffolds with cells has offered a further refinement to the in vitro investigation of liver regeneration as well as potential therapeutic opportunities[ 118 ].…”

“…Others have demonstrated that implanting repopulated ECM liver scaffolds into rats which had undergone extended hepatectomy improved liver function and extended their mean lifespan from 16 to 72 h[ 128 ]. In the last 10 years, a variety of animal models, decellularisation techniques, repopulation routes and cell sources have been described, with promising outcomes in terms of vascular repopulation[ 118 , 129 , 130 ], hepatocyte survival[ 131 ] as well as formation of biliary duct-like structures and activation of liver detoxification enzymes[ 132 ]. One of the commonest sources of liver scaffolds is the rat[ 118 , 127 , 132 - 137 ] repopulated with rat hepatocytes (although cholangiocytes[ 136 ] and lineages from pluripotent stem cells, mesenchymal cells, and fibroblasts have also been described[ 137 ] usually via the portal vein.…”

“…In the last 10 years, a variety of animal models, decellularisation techniques, repopulation routes and cell sources have been described, with promising outcomes in terms of vascular repopulation[ 118 , 129 , 130 ], hepatocyte survival[ 131 ] as well as formation of biliary duct-like structures and activation of liver detoxification enzymes[ 132 ]. One of the commonest sources of liver scaffolds is the rat[ 118 , 127 , 132 - 137 ] repopulated with rat hepatocytes (although cholangiocytes[ 136 ] and lineages from pluripotent stem cells, mesenchymal cells, and fibroblasts have also been described[ 137 ] usually via the portal vein. With regards to human tissue, Verstegen et al [ 138 ] demonstrated that decellularised human livers can be repopulated with human umbilical vein endothelial cells, leading to re-endothelialisation of the vascular tree.…”

Cell Therapy and Bioengineering in Experimental Liver Regenerative Medicine: In Vivo Injury Models and Grafting Strategies

“…Different cell types could be used for repopulating decellularized livers, including primary hepatocytes, iPSCderived hepatocyte-like cells, stem cells, or mesenchymal stromal cells for recellularization of the parenchymal areas, and endothelial progenitor cells or human umbilical vein endothelial cells (HUVEC) for reendothelization [85,[128][129][130][131][132]. However, optimization of the recellularization process is still a challenge when more than one cell type per scaffold is used.…”