HAF prevents hepatocyte apoptosis and progression to MASH and hepatocellular carcinoma through transcriptional regulation of the NF-κB pathway

Background: HCC incidence is increasing worldwide due to the obesity epidemic, which drives metabolic dysfunction-associated steatohepatitis (MASH) that can lead to HCC. However, the molecular pathways driving MASH-HCC are poorly understood. We have previously reported that male mice with haploinsufficiency of hypoxia-associated factor, HAF ( SART1 +/- ) spontaneously develop MASH-HCC. However, the cell type(s) responsible for HCC associated with HAF loss are unclear. Results: We generated SART1 -floxed mice, which were crossed with mice expressing Cre-recombinase within hepatocytes (Alb-Cre; hepS -/- ) or myeloid cells (LysM-Cre, macS -/- ). HepS -/- mice (both male and female) developed HCC associated with profound inflammatory and lipid dysregulation suggesting that HAF protects against HCC primarily within hepatocytes. HAF-deficient hepatocytes showed decreased P-p65 and P-p50 and in many components of the NF-κB pathway, which was recapitulated using HAF siRNA in vitro . HAF depletion also triggered apoptosis, suggesting that HAF protects against HCC by suppressing hepatocyte apoptosis. We show that HAF regulates NF-κB activity by regulating transcription of TRADD and RIPK1 . Mice fed a high-fat diet (HFD) showed marked suppression of HAF, P-p65 and TRADD within their livers after 26 weeks, but showed profound upregulation of these proteins after 40 weeks, implicating deregulation of the HAF-NF-κB axis in the progression to MASH. In humans, HAF was significantly decreased in livers with simple steatosis but significantly increased in HCC compared with normal liver. Conclusions: HAF is novel transcriptional regulator of the NF-κB pathway and is a key determinant of cell fate during progression to MASH and MASH-HCC.