Epithelial cell plasticity in chronic liver disease (MASLD/NAFLD)

Abstract

It has been proposed that liver regeneration in chronic liver disease, such as metabolic dysfunction-associated steatotic liver disease, previously known as non-alcoholic fatty liver disease, is driven by hepatocyte renewal in the early stages, but when hepatocyte proliferation is impaired due to disease burden, alternative regenerative mechanisms emerge to repair the liver parenchyma (Gadd et al., 2020). A potential mechanism of interest is the transdifferentiation of cholangiocytes into hepatocytes via either direct differentiation or via a progenitor cell population (Michalopoulos & Khan, 2015). However, the reality of this process in a disease context especially in human remains to be fully demonstrated while the molecular mechanisms involved have not been investigated in detail. Here, I address these questions by firstly using human liver tissue from chronic liver disease to describe the morphological and transcriptomic changes observed in cholangiocytes and secondly, I take advantage of cholangiocyte organoids as an in vitro model to assess their plasticity in end stage cirrhotic liver from patients undergoing liver transplantation. For the first part of the dissertation, I perform immunostaining analyses on tissue section from explanted liver to confirm the presence of cells displaying both cholangiocytes and hepatocytes markers thereby reinforcing the transdifferentiation hypothesis. I then use single nuclei RNA sequencing analysis from liver biopsies to capture and characterise the transcriptomic signature of those biphenotypic cells. Next, for the second part of the dissertation, I confirm that cholangiocyte organoids can be derived from explanted liver and express hepatocytes markers when grown in defined culture conditions, making them an ideal in vitro model for biphenotypic cells. Finally, I employ single nuclei RNA and ATAC multiomics analysis to characterise in detail the in vitro process of cholangiocytes differentiation towards hepatocyte-like cells. Overall, the results propose that the diseased liver is exhibiting significant epithelial plasticity and uncover molecular mechanisms involved in these processes. Also, they validate the interest of cholangiocytes organoids to study the regenerative process occurring in end stage liver disease.