Guest, Rachel

Boulter, Luke

McCaffrey, Caitlin

2026-05-29T14:34:25Z

2026-05-29

Liver regeneration following injury occurs through several well-defined mechanisms, including the hypertrophy or hyperplasia (proliferation) of epithelial cells and the recruitment and differentiation of hepatic progenitor cells. In specific contexts, experimental evidence suggests that mature hepatocytes have the capacity for transdifferentiation in order to contribute to biliary repair; however, it remains unknown whether all hepatocytes possess this propensity or whether lineage plasticity is zonated within the lobule. This study aimed to investigate whether the position of hepatocytes along the axis of the lobule is a determinant of hepatocyte plasticity. This was achieved through two independent approaches, the first being the design and breeding of two mouse models, which facilitated in vivo lineage tracing of hepatocyte populations from opposite poles of the liver lobule. These mouse models enabled fluorescent labelling of Glutaminase (Gls2)-expressing periportal (zone 1/2) or Glutamine Synthetase (GS)-expressing pericentral (zone 3) hepatocytes, utilising a dual-fluorescent mT/mG CreER-loxP system. Based on established methods described in published studies, intrahepatic cholangiocarcinoma (iCC) was induced via hydrodynamic tail vein delivery of Notch intracellular domain 1 (NICD1) and Akt overexpression, thereby stimulating cell fate change in the two hepatocyte populations of interest. The second approach involved the development of a spheroid culture model from primary murine hepatocytes (PMH) isolated from control mice and lineage-traced Gls2CreER-mT/mG and GSCreER-mT/mG mouse models. This complementary approach enabled the in vitro modelling of zonated hepatocyte gene expression, providing a platform to evaluate hepatocyte plasticity via NICD1/Akt upregulation ex vivo. The data presented in this study reveal that periportal hepatocytes, originating from zone 1/2, specifically those expressing Gls2, undergo transdifferentiation and phenotypic changes that contribute to biliary repair more readily than GS-expressing hepatocytes from the Wnt-high pericentral zone. Additionally, primary murine hepatocyte (PMH) spheroid models not only preserve zonated gene expression profiles of constituent hepatocytes ex vivo but also demonstrate that the zonated phenotypic markers can be influenced through changes in culture media composition. The PMH spheroid platform appeared to recapitulate the transdifferentiation seen in vivo upon NICD1/Akt upregulation, with gene expression analyses indicating a shift from a hepatocyte to a biliary lineage, closely mirroring changes in targeted hepatocyte populations in the mouse models. Importantly, these findings suggest that hepatocyte plasticity in biliary repair is zonally restricted and can be modelled and experimentally manipulated ex vivo, providing new insight into the determinants and modulation of hepatocyte fate in regeneration and disease.

https://era.ed.ac.uk/handle/1842/44774

https://doi.org/10.7488/era/7288

en

Liver

Hepatocyte

Plasticity

Transdifferentiation

Notch signalling

Akt1

Defining metabolic zonation as a determinant of hepatocyte plasticity

Thesis

Doctoral

PhD Doctor of Philosophy