Hepatocellular carcinoma (HCC) is the third leading cause of cancer mortality worldwide, causing over 700,000
deaths annually. In the US, HCC rates have tripled in the last three decades. The rise in obesity, non-alcoholic
fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) is increasingly contributing to advanced
liver disease and HCC development. Chronic injury, inflammation and fibrosis are strongly associated with
disease progression and HCC development in patients and sufficient to trigger HCC in mice - suggesting that
HCC truly represents “a wound that does not heal”. Even though HCC may develop in the absence of overt
cirrhosis in NASH, almost all NASH-HCCs still develop in fibrotic livers. Hence, fibrosis is one of the most
prominent but least understood risk factors for HCC and NASH-HCC. Moreover, fibrosis is a main determinant
of outcomes and most therapeutic efforts in NASH aim to reducing fibrosis. Hence, understanding the role of
fibroblasts in HCC development in NASH is highly relevant. We and others have previously shown that hepatic
stellate cells (HSC) are the main source of fibroblasts in the liver. This proposal is based on preliminary data
showing for the first time by reliable and complementary genetic approaches a strong promotion of HCC
development by HSC. However, while the overall effect of HSC in chronic liver disease is tumor-promoting, we
made the striking observation that HSC can both promote and protect from HCC. Based on this, our central
hypothesis is that a cytokine- and growth-factor secreting subpopulation (cyHSC), resembling quiescent HSC in
the healthy liver, protect from NASH-HCC whereas highly activated myfibroblastic HSC (myHSC) promote
NASH-HCC. Linked to this, we also propose the higher percentage of cyHSC in normal liver provide protections
from HCC in the healthy state, whereas the higher percent of myHSC in injured livers contribute to disease-
associated promotion of HCC. Here, we seek to functionally characterize the contribution of HSC-derived
fibroblasts and subpopulation-specific mediators in NASH-associated HCC. We will first determine the role of
HSC-derived fibroblasts by genetic depletion and inhibition approaches, and characterize HSC subpopulations
and their cellular and ligand-receptor interactions and their evolution during disease progression by single cell
RNA-sequencing and CellPhoneDB in murine and human NASH (Aim 1). We will determine the role of myHSC-
secreted mediators, focusing on type I collagen, its receptors and downstream pathways (Aim 2) as well as the
role of cyHSC-secreted mediators, focusing on hepatocyte growth factor and CXCL12 (Aim 3), in NASH-
associated HCC. Together, the proposed studies will reveal novel insights into the pathophysiology of NASH-
HCC and provide evidence for tumor-promoting as well as tumor-suppressive functions of HSC. These studies
may not only open up new therapeutic avenues, targeting tumor-promoting pathways while sparing or restoring
tumor-suppressive pathways, but also shift current paradigms on the role of HSC in HCC development.