Cholangiocyte and hepatocyte plasticity in human chronic liver disease

Liver disease is on the rise, which has created the urgent need for new treatments. An attractive therapeutic approach is the understanding and manipulation of the regenerative pathways of the liver. The regenerative capability of the liver is dependent on the nature of the injury. It is known that mild injury and hepatectomy induce hepatocyte proliferation driven regeneration. However, hepatocyte proliferation is impaired during chronic injury and secondary mechanisms of regeneration might exist. Indeed, studies in animal models have revealed several regenerative processes, which might take place in chronic disease: 1) liver stem cells activation, 2) dedifferentiation/redifferentiation of cholangiocytes/hepatocytes, 3) transdifferentiation between cholangiocytes and hepatocytes. There is little knowledge of the above processes in human and the current knowledge is mainly derived from histopathology analyses. Here we aim to define the mechanisms behind epithelial plasticity in the diseased liver. To understand the liver response to chronic disease, we have collected liver biopsies from just under 50 patients across the spectrum of non-alcoholic fatty liver disease and we performed state of the art 3D imaging and single nuclei RNA sequencing (snRNA). In depth, computational analysis has allowed us to dissect the cellular composition of the liver during the course of chronic liver disease. We further used liver organoids as an in vitro model to validate our snRNAseq findings. The analysis has revealed that disease progression is accompanied by tissue remodelling, loss of zonation and extensive ductular reaction. Further analyses, captured both at the transcriptomic and protein level the presence of cells sharing characteristics from cholangiocytes and hepatocytes, termed biphenotypic cells. Gene enrichment analysis revealed signalling pathways likely involved in their generation. In vitro modelling using NAFLD human derived liver organoids validated these molecular pathways and their importance in the generation of bi-phenotypic cells. In conclusion, we use snRNAseq analysis, to demonstrate the presence of a molecular pathway involved in liver epithelial cell plasticity during chronic liver disease. This study paves the way for the development of new therapies promoting tissue repair to improve organ function.