Scalable Production of Size-Controlled Cholangiocyte and Cholangiocarcinoma Organoids within Liver Extracellular Matrix-Containing Microcapsules

Gilles S. van Tienderen, Jorke Willemse, Bas van Loo, Eline V.A. van Hengel, Jeroen de Jonge, Luc J.W. van der Laan, Jeroen Leijten, Monique M.A. Verstegen*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

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Abstract

Advances in biomaterials, particularly in combination with encapsulation strategies, have provided excellent opportunities to increase reproducibility and standardization for cell culture applications. Herein, hybrid microcapsules are produced in a flow-focusing microfluidic droplet generator combined with enzymatic outside-in crosslinking of dextran-tyramine, enriched with human liver extracellular matrix (ECM). The microcapsules provide a physiologically relevant microenvironment for the culture of intrahepatic cholangiocyte organoids (ICO) and patient-derived cholangiocarcinoma organoids (CCAO). Micro-encapsulation allowed for the scalable and size-standardized production of organoids with sustained proliferation for at least 21 days in vitro. Healthy ICO (n = 5) expressed cholangiocyte markers, including KRT7 and KRT19, similar to standard basement membrane extract cultures. The CCAO microcapsules (n = 3) showed retention of stem cell phenotype and expressed LGR5 and PROM1. Furthermore, ITGB1 was upregulated, indicative of increased cell adhesion to ECM in microcapsules. Encapsulated CCAO were amendable to drug screening assays, showing a dose-response response to the clinically relevant anti-cancer drugs gemcitabine and cisplatin. High-throughput drug testing identified both pan-effective drugs as well as patient-specific resistance patterns. The results described herein show the feasibility of this one-step encapsulation approach to create size-standardized organoids for scalable production. The liver extracellular matrix-containing microcapsules can provide a powerful platform to build mini healthy and tumor tissues for potential future transplantation or personalized medicine applications.

Original languageEnglish
Article number3657
JournalCells
Volume11
Issue number22
DOIs
Publication statusPublished - 18 Nov 2022

Bibliographical note

Funding Information:
This project was partly funded by the Erasmus MC Human Disease Model Award 2018 (HDMA-380801, GT, LL, MV), the Medical Delta (Regenerative Medicine 4D, LL, MV), the TKI-LSH grant (EMC-LSH19002, JW, EH, MV, JJ, LL), the Dutch Cancer Society (KWF project number 14364, MV), the Dutch Research Council (ENW-XS project number OCENW.XS21.2.003, MV) and the Dutch Society for Gastroenterology and Hepatology (NVGE Gastrostart Vervolg 01-2022 MV, LL). JL acknowledges financial support from Dutch Research Council (Vidi, 17522) and European Research Council (Starting Grant, 759425).

Publisher Copyright:
© 2022 by the authors.

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