Carvedilol triggers ferroptosis in hepatic stellate cells via the ATF4/SAT1 axis promoting spermidine depletion to ameliorate liver fibrosis

Abstract Liver fibrosis is a major pathological consequence of chronic liver injury, and carvedilol, a drug effective in reducing portal hypertension, has been shown to exert anti-fibrotic effects in recent studies, yet the precise molecular mechanism underlying this activity remains elusive. This study aimed to investigate the core scientific question of whether carvedilol alleviates liver fibrosis by inducing ferroptosis in hepatic stellate cells (HSCs) and to clarify the corresponding regulatory mechanisms. Using a CCl 4 -induced mouse liver fibrosis model and human (LX-2)/rat (HSC-T6) HSC lines treated with carvedilol, mechanistic analyses were conducted via Western blot, quantitative real-time PCR (RT-qPCR), RNA sequencing (RNA-seq), chromatin immunoprecipitation (ChIP), as well as functional assays involving genetic manipulation and pharmacological inhibitors. Carvedilol significantly attenuated HSCs activation and ameliorated liver fibrosis in mice, and it induced ferroptosis in HSCs—characterized by mitochondrial shrinkage, lipid peroxidation, and iron accumulation—an effect that was abrogated by the ferroptosis inhibitor ferrostatin-1. Mechanistically, carvedilol triggered endoplasmic reticulum (ER) stress to activate the PERK/eIF2α/ATF4 signaling axis, and ATF4 transcriptionally upregulated spermidine/spermine N1-acetyltransferase 1 (SAT1). Ectopic overexpression of SAT1 alone was sufficient to induce ferroptosis and suppress HSCs activation, whereas SAT1 knockdown completely abolished the aforementioned effects of carvedilol. Importantly, SAT1 catalyzed spermidine depletion, which led to the post-transcriptional downregulation of GPX4 and xCT proteins, and exogenous spermidine supplementation effectively rescued SAT1-overexpression-induced ferroptosis in HSCs. Collectively, our findings demonstrate that carvedilol ameliorates liver fibrosis by inducing HSCs ferroptosis via the ER stress/ATF4/SAT1/spermidine depletion axis. This study identifies a novel regulatory role of spermidine metabolism in HSCs ferroptosis and liver fibrogenesis, and further establishes spermidine metabolism as a potential and specific therapeutic target for fibrotic liver diseases.