HIF-1α-ASS1 axis drives reprogramming of arginine metabolism controls hepatic stellate cell activation in hepatic fibrosis.

Abstract

Dysregulated arginine metabolism promotes hepatic stellate cell (HSC) activation and liver fibrosis. However, the mechanisms underlying arginine hosmeostasis in hepatic fibrogenesis and therapeutic targeting remain unclear. Using Multicellular hepatic spheroid (MCHS) models recapitulating in vivo liver fibrosis, we investigated the effects of arginine deprivation or supplementation. Genetic and pharmacological (ABRL8301, a 2-methoxyestradiol derivative) inhibition of hypoxia-inducible factor 1 alpha (HIF-1α) were assessed for effects on argininosuccinate synthetase 1 (ASS1) expression and antifibrotic efficacy in vitro and in vivo. Holotomography and quantitative imaging approaches characterized cellular and molecular fibrogenic changes. Arginine deprivation increased MCHS compactness and α-smooth muscle actin (α-SMA) expression, whereas l-arginine supplementation reversed effects. Considering pivotal roles of ASS1 and arginase 1 (Arg1) in arginine biosynthesis and catabolism, respectively, ASS1 overexpression suppressed mesenchymal markers, whereas Arg1 overexpression aggravated fibrotic signatures. HIF-1α knockdown and ABRL8301 treatment upregulated ASS1, reduced epithelial-mesenchymal transition (EMT) and fibrotic markers, and enhanced arginine biosynthesis in MCHSs. Holotomography analysis confirmed ABRL8301 attenuated TGFβ1-induced cell elongation and α-SMA fiber accumulation in HSCs and fibroblasts. In CCl₄-mouse liver fibrosis models, ABRL8301 significantly reduced collagen deposition, necrosis, and fibrosis. Combination sorafenib and ABRL8301 therapy decreased MCHS compactness, increased apoptosis, and achieved superior tumor control in HCC xenografts. Restoring arginine metabolic homeostasis through HIF-1α inhibition-mediated ASS1 upregulation suppresses HSC activation and liver fibrosis. ABRL8301 acts as a pharmacologic inducer of ASS1 and arginine biosynthesis, inhibiting EMT and fibrotic progression while enhancing anticancer therapy. These findings support ABRL8301 as a promising antifibrotic and tumor-sensitizing agent in hepatic fibrosis.