Incidence rates of obesity-related disorders like non-alcoholic steatohepatitis (NASH) have been rising globally during the last decades. Here, abnormal hepatic fat accumulation, inflammation, and fibrosis are seen, which, consequently, lead to disruption of hepatic tissue architecture and function, increasing the risk of chronic liver disease, cirrhosis, and hepatocellular carcinoma. The underlying mechanisms of NASH pathogenesis are still partially unknown and treatment options for patients worldwide are limited. Today, we know that NASH is driven by highly complex cellular interactions of multiple cell types, including hepatic stellate cells (HSCs) and infiltrating immune cells. How these cells participate in disease initiation and progression and which intercellular cross talk mechanisms that may drive these events are only partially described in literature.
The work presented in this thesis is divided into two parts. In part I (paper I), we aimed to determine HSC plasticity in NASH and map transcriptional networks controlling these dynamics. Here, we introduced NASH in mice through western diet and fructose feeding and isolated HSCs at different time points up until 24 weeks. RNA-seq experiments revealed time-resolved transcriptional changes in HSCs upon disease development and found a potential core transcriptional program controlling induction of the early activation state. Motif enrichment analysis at promoter regions of HSC-activated genes indicate ETS1 and RUNX1 as putative transcriptional regulators driving NASH-associated HSC plasticity. This was confirmed by loss-of-function experiments where decreased expression of fibrogenic genes like Col1a1 and Acta2 upon HSC activation in vitro was observed.
In part II (paper II), we wanted to investigate hepatic tissue heterogeneity and plasticity in advanced-NASH. To recapitulate human pathophysiology, we fed mice a western diet in combination with fructose for 52 weeks and compared them with aged-matched healthy controls. Single-cell RNA-seq experiments revealed 11 distinct cell populations with an overall alteration in liver cell composition in NASH compared to healthy livers. interestingly, we found an increase in the mononuclear phagocyte (MP) heterogeneity with expansion of infiltrating monocytes and monocyte-derived macrophages. In line, RNA velocities predicted an altered plasticity of MPs in NASH, showing kupffer cell (KC)-like cells as both monocyte- and KC-derived. Furthermore, a shift in HSC composition to a more activated phenotype exhibiting an increased extracellular matrix production was observed in NASH livers. Using SCENIC, we captured subpopulation-specific transcriptional regulators that may assist as drivers of HSC plasticity at different stages during activation. Lastly, NicheNet-predicted ligand-receptor interactions specified possible cross talk mechanisms between subpopulations of HSCs and MPs that could add to our understanding of intercellular interactions in both healthy and diseased livers.
In summary, we provided insight into the hepatic tissue heterogeneity and cellular plasticity in murine NASH, with a specific focus on HSCs and MPs; important cell types in relation to inflammation and fibrosis. The work gave an increased understanding of liver complexity and better characterization of cell subpopulations, putative cellular trajectories, and transcriptional networks controlling subpopulation-specific cellular identities.