Cellular Crosstalk Promotes Hepatic Progenitor Cell Proliferation and Stellate Cell Activation in 3D Co-culture

Maya W Haaker; Jung-Chin Chang; Brian K Chung; Tobias S Pieper; Falko Noé; Tongtong Wang; Niels Geijsen; Martin Houweling; Christian Wolfrum; Arie B Vaandrager; Espen Melum; Bart Spee; J Bernd Helms

doi:10.1016/j.jcmgh.2025.101472

Cellular Crosstalk Promotes Hepatic Progenitor Cell Proliferation and Stellate Cell Activation in 3D Co-culture

Cell Mol Gastroenterol Hepatol. 2025 Jan 30;19(5):101472. doi: 10.1016/j.jcmgh.2025.101472. Online ahead of print.

Authors

Affiliations

¹ Department of Biomolecular Health Sciences, Faculty of Veterinary Medicine, Utrecht University, The Netherlands.
² Norwegian PSC Research Center, Department of Transplantation Medicine, Division of Surgery and Specialized Medicine, eDivision of Surgery and Specialized Medicine, Oslo University Hospital Rikshospitalet, Oslo, Norway; Institute of Clinical Medicine, Faculty of Medicine, University of Oslo, Oslo, Norway; Research Institute of Internal Medicine, Oslo University Hospital, Rikshospitalet, Oslo, Norway.
³ Institute of Food, Nutrition and Health, ETH Zurich, Schwerzenbach, Switzerland.
⁴ Department of Anatomy and Embryology, Leiden University Medical Center, Leiden, The Netherlands.
⁵ Norwegian PSC Research Center, Department of Transplantation Medicine, Division of Surgery and Specialized Medicine, eDivision of Surgery and Specialized Medicine, Oslo University Hospital Rikshospitalet, Oslo, Norway; Institute of Clinical Medicine, Faculty of Medicine, University of Oslo, Oslo, Norway; Research Institute of Internal Medicine, Oslo University Hospital, Rikshospitalet, Oslo, Norway; Section of Gastroenterology, Department of Transplantation Medicine, Division of Surgery and Specialized Medicine, Oslo University Hospital Rikshospitalet, Oslo, Norway; Hybrid Technology Hub-Centre of Excellence, Institute of Basic Medical Sciences, Faculty of Medicine, University of Oslo, Oslo, Norway.
⁶ Department of Clinical Sciences, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands.
⁷ Department of Biomolecular Health Sciences, Faculty of Veterinary Medicine, Utrecht University, The Netherlands. Electronic address: j.b.helms@uu.nl.

PMID: 39892785
DOI: 10.1016/j.jcmgh.2025.101472

Abstract

Background & aims: Following liver damage, ductular reaction often coincides with liver fibrosis. Proliferation of hepatic progenitor cells is observed in ductular reaction, whereas activated hepatic stellate cells (HSCs) are the main drivers of liver fibrosis. These observations may suggest a functional interaction between these 2 cell types. Here, we report on an in vitro co-culture system to examine these interactions and validate their co-expression in human liver explants.

Methods: In a 3D organoid co-culture system, we combined freshly isolated quiescent mouse HSCs and fluorescently labeled progenitor cells (undifferentiated intrahepatic cholangiocyte organoids), permitting real-time observation of cell morphology and behavior. After 7 days, cells were sorted based on the fluorescent label and analyzed for changes in gene expression.

Results: In the 3D co-culture system, the proliferation of progenitor cells is enhanced, and HSCs are activated, recapitulating the cellular events observed in the patient liver. Both effects in 3D co-culture require close contact between the 2 different cell types. HSC activation during 3D co-culture differs from quiescent (3D mono-cultured) HSCs and activated HSCs on plastic (2D mono-culture). Upregulation of a cluster of genes containing Aldh1a2, Cthrc1, and several genes related to frizzled binding/Wnt signaling were exclusively observed in 3D co-cultured HSCs. The localized co-expression of specific genes was confirmed by spatial transcriptomics in human liver explants.

Conclusion: An in vitro 3D co-culture system provides evidence for direct interactions between HSCs and progenitor cells, which are sufficient to drive responses that are similar to those seen during ductular reaction and fibrosis. This model paves the way for further research into the cellular basis of liver pathology.

Keywords: Ductular Reaction; Intrahepatic Cholangiocyte Organoid; Liver Fibrosis; Retinoic Acid.