Scaffolds obtained from decellularized human extrahepatic bile ducts support organoids to establish functional biliary tissue in a dish

Abstract

Biliary disorders can lead to life‐threatening disease and are also a challenging complication of liver transplantation. As there are limited treatment options, tissue engineered bile ducts could be employed to replace or repair damaged bile ducts. We explored how these constructs can be created by seeding hepatobiliary LGR5+ organoids onto tissue‐specific scaffold. For this, we decellularized discarded human extrahepatic bile ducts (EBD) that we recellularized with organoids of different origin, that is, liver biopsies, extrahepatic bile duct biopsies, and bile samples. Here, we demonstrate efficient decellularization of EBD tissue. Recellularization of the EBD extracellular matrix (ECM) with the organoids of extrahepatic origin (EBD tissue and bile derived organoids) showed more profound repopulation of the ductal ECM when compared with liver tissue (intrahepatic bile duct) derived organoids. The bile duct constructs that were repopulated with extrahepatic organoids expressed mature cholangiocyte‐markers and had increased electrical resistance, indicating restoration of the barrier function. Therefore, the organoids of extrahepatic sources are identified to be the optimal candidate for the development of personalized tissue engineered EBD constructs.

Cholangiocyte organoids from human bile retain a local phenotype and can repopulate bile ducts in vitro

“…These empty scaffolds were created as a model for bile duct damage using a method called decellularization. 26 We demonstrate that recellularization by BCOs results in the formation of a confluent monolayer with cholangiocyte‐like cells. Importantly, BCOs are less proliferative as well as lose their LGR5 expression when cultured upon EHBD scaffolds.…”

“…Since we recently showed that our repopulated scaffolds with BCOs are properly polarized by cilia staining and that they become functional constructs by demonstrating transepithelial electrical resistance (TEER) and ion‐channel functionality, 26 the next goal would be to create a 3D construct. Cholangiocyte‐organoids cultured in non‐canonical WNT‐stimulated conditions have previously shown feasibility to repopulate 3D constructs and function as EHBD in vivo.…”

“…EHBD scaffolds were created according to the previously published protocol by Willemse et al. 26 using a Triton‐X‐100‐based decellularization method. EHBD scaffolds were reseeded with single‐cells derived from ERCP‐BCOs (passage 5–9) and cultured for 21 days.…”

Research Progress, Challenges, and Breakthroughs of Organoids as Disease Models

“…Additives enhance the contact between the matrix and organoids and enhance some properties of the hydrogel, which are more conducive to the formation and development of organoids. 2) The hydrogel has a structure similar to that of the environment in vivo , such as the decellularized matrix of homologous tissue ( Giobbe et al, 2019 ; Mollica et al, 2019 ; Bi et al, 2020a ; Bi et al, 2020b ; Meran et al, 2020 ; Willemse et al, 2021 ), a culture hydrogel of intestinal organs composed of a collagen V-coated porous substrate, and a collagen I gel overlay ( Tong et al, 2018 ); 3D collagen type I scaffolds mimicking the double cortical layer ultrastructure of the thymus ( Bortolomai et al, 2019 ). Different tissues have different structures.…”

Bile Duct Repair in Human Liver Grafts: Effective Cholangiocyte Organoid Engraftment and Plasticity

“…SOX17) and gained expression of intrahepatic cholangiocyte-related genes (SOX4). Whether this plastisity is also observed when intrahepatic cholangiocyte organoids engraft downstream was not shown, however a recent in vitro study demonstrated that repopulation of the extrahepatic bile duct matrix by intrahepatic cholangiocyte organoids was less profound than their extrahepatic counterparts (7). Interestingly, when freshly isolated human cholangioctes were infused limited engraftment and survival benefits were observed.…”