Demethylzeylasteral Targets the Sp1/AGAP2 Axis to Impair Autophagic Flux and Induce Apoptosis in Hepatic Stellate Cells.

Background

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Aim

Chronic liver injury-induced liver fibrosis, if left untreated, can progress to cirrhosis or hepatocellular carcinoma, posing a serious threat to human health. Our previous study demonstrated that the natural product demethylzeylasteral (T-96) alleviates liver fibrosis by suppressing ArfGAP with GTPase domain, ankyrin repeat and PH domain 2 (AGAP2) expression. However, the molecular mechanism underlying T-96-mediated regulation of AGAP2 remains unclear. This study aimed to clarify this mechanism and explore T-96's role in liver fibrogenesis.

Methods

A series of molecular biological experiments (e.g., promoter activity assay, co-immunoprecipitation, Western blot, and immunofluorescence) were performed to identify the transcription factor regulating AGAP2. The interaction between T-96 and the transcription factor, as well as the degradation pathway of the latter, were investigated. Additionally, in vitro experiments were conducted to evaluate the effects of T-96 on autophagy, apoptosis, and activation of hepatic stellate cells (HSCs).

Results

We identified Sp1 as the transcription factor mediating AGAP2 promoter activity. Mechanistically, T-96 directly bound to Sp1 and promoted its degradation via the ubiquitin-proteasome pathway, thereby downregulating AGAP2 expression. Furthermore, T-96 impairs autophagic flux and induces apoptosis in activated HSCs through specific targeting of Sp1.

Conclusions

This study delineates a novel regulatory axis centered on Sp1 (T-96/Sp1/AGAP2) in liver fibrosis, wherein T-96 binds to Sp1 to promote its ubiquitination and degradation, subsequently inhibiting AGAP2 expression and inducing autophagic flux impairment/apoptosis in activated HSCs to alleviate liver fibrosis. These findings provide a theoretical foundation for developing targeted therapeutics and clinical strategies for fibrotic liver disease.