The Bidirectional Crosstalk between CD4⁺ T Cells and Cholangiocytes in a Patient-Derived Organoid Model

Abstract

This thesis explores how cholangiocytes, the epithelial cells lining the bile ducts, interact with CD4⁺ T cells in chronic liver disease. The study focuses mainly on primary sclerosing cholangitis (PSC) and compares it with alcohol-associated liver disease (ALD). Although cholangiocytes have often been considered passive targets of damage, increasing evidence suggests that they actively contribute to inflammation and immune regulation. To investigate this, patient-derived cholangiocyte organoids were used as a three-dimensional in vitro model. The first part of the work examined whether cholangiocyte organoids derived from PSC and ALD patients retain disease-related features. Transcriptomic analyses showed that the two types of organoids were overall similar, but PSC-derived organoids displayed subtle differences linked to a more immune-responsive and regenerative profile. In contrast, ALD-derived organoids were more associated with metabolic and homeostatic functions. After stimulation with IL-17A, both PSC and ALD organoids showed a strong and largely overlapping inflammatory response, suggesting that this cytokine induces a common epithelial activation program regardless of disease origin. The second part of the thesis investigated the interaction between cholangiocytes and CD4⁺ T cells. For this purpose, an optimized co-culture system was established to allow immune cells and organoids to interact under suitable experimental conditions. The results showed that soluble factors released by activated T cells were able to induce inflammatory changes in cholangiocytes, but direct cell–cell contact triggered a broader and more complex response. At the same time, cholangiocytes also influenced T-cell behaviour, indicating a bidirectional interaction. In particular, the PD-1/PD-L1 axis emerged as a relevant pathway in this crosstalk, supporting the idea that cholangiocytes may both promote and regulate immune responses. Overall, this thesis shows that cholangiocyte organoids are a useful model for studying biliary inflammation and epithelial–immune communication. The findings support the view that cholangiocytes are active participants in the immune processes that characterize PSC, rather than simple bystanders. More broadly, this work provides new insight into the role of epithelial–immune interactions in chronic cholangiopathies and offers a basis for future studies on disease mechanisms and therapeutic strategies.