Hastatoside ameliorates alcohol-associated liver disease, potentially involving ATF4-regulated PANoptosis signaling cascades.

Abstract

This study aimed to elucidate the therapeutic efficacy of Hastatoside (Has) against alcohol‑associated liver disease (ALD) and characterize the molecular mechanisms underlying its hepatoprotective effects. For in vivo studies, mice were fed a Lieber‑DeCarli ethanol diet for 28 days to establish ALD models, followed by ATF4 silencing via lentiviral shRNA. In vitro experiments involved exposing HepG2 cells to Has and ethanol, followed by transfection with ATF4 siRNA. The interaction between ATF4 and the ZBP1 promoter was verified using a dual‑luciferase reporter assay. Additionally, to investigate mouse bone marrow‑derived macrophages (BMDMs) and hepatocyte crosstalk in ALD, primary hepatocytes isolated from normal mice were first exposed to conditioned media-the culture supernatants of BMDMs stimulated with LPS/ATP, and were subsequently subjected to LPS/ATP stimulation in the presence or absence of Has. Our data revealed that Has downregulated ATF4 expression as well as levels of PANoptosis‑related receptors in ALD models. Silencing of ATF4 via shRNA notably reduced ZBP1 levels and ameliorated liver injury accompanied by global PANoptosis. Hepatocyte‑specific ATF4 deficiency potentiated Has‑mediated inhibition of PANoptosis pathways. Moreover, Has suppressed pyroptosis in BMDMs, thereby mitigating inflammatory responses. Collectively, these results show that Has may alleviate ALD by modulating the ATF4‑driven PANoptosis axis. In summary, our findings highlight Has as a promising candidate for therapeutic interventions against ALD.