Hepatic stellate cell subpopulations in human MASLD-related hepatocellular carcinoma upregulate YAP via...

Hepatic stellate cells (HSCs) drive cirrhosis and hepatocellular carcinoma (HCC). Statin use decreases cirrhosis and HCC through unclear mechanisms. We aimed to uncover statin-responsive pathways preventing activation of HSC subpopulations in human MASLD-HCC.

We used single-nucleus RNA-sequencing and spatial imaging on matched human MASLD-HCC samples in cirrhotic and non-cirrhotic livers, along with RNA-sequencing of human primary HSCs to identify Yes-Associated Protein (YAP) as a statin-responsive pathway, which was assessed for cellular localization and function, that is, downstream gene expression after statin exposure. We used pharmacologic inhibitors, gene silencing, pharmacological repletion, and direct quantification by UPLC-MS/MS to interrogate the mevalonate pathway as a statin-responsive YAP regulator.

We identified an HCC-HSC subcluster enriched in activated and deactivated marker genes. Spatial resolution of each cell type revealed that myofibroblast-like HSC subpopulations and YAP-effector genes colocalized in the peritumoral pseudocapsule. In fact, although Rho GTPases were differentially expressed across most cell types, YAP-effector genes were largely upregulated in HCC-HSCs by single-nucleus RNA sequencing. Bulk RNA-sequencing of statin-treated activated human primary HSCs showed decreased expression of YAP-related genes and downregulation of Rho GTPase pathways in response to statins. Statins reduced GGPP levels in LX2 HSCs, determined by direct measurement of intracellular GGPP, which was associated with cytoskeletal restructuring, YAP cytosolic retention, and reduced YAP-effector gene expression. Exogenous GGPP repletion reversed these effects. GGPP synthase 1 knockdown and direct Rho inhibition recapitulated the cellular responses of YAP following statin exposure.

Upregulation of YAP-effector genes was largely restricted to HCC-HSCs. Our mechanistic studies support that statins lower GGPP, reduce Rho GTPases prenylation, cause YAP cytosolic retention, and decrease YAP nuclear activity. This study expands on the chemoprotective mechanisms of statins in HCC.