Identification of key anti-pulmonary fibrosis components in Hyssopus cuspidatus Boriss. and its mechanism exploration

Background: Hyssopus cuspidatus Boriss. (HC) is a renowned herbal medicine commonly used in treating respiratory diseases. However, its potential in alleviating pulmonary fibrosis (PF) remains unexplored.

Purpose: This study aims to investigate the anti-PF mechanisms of HC and identify activity-guided biomarkers to improve its quality standards.

Methods: The anti-fibrotic effects of HC were evaluated using TGF-β1-induced MRC-5 cell fibrosis and bleomycin-induced mouse PF models. In vitro, GSK-3β was silenced using shRNA or the inhibitor lithium chloride (LiCl), while active GSK-3β was overexpressed to validate the underlying mechanisms. Chemical profiling and spectrum-effect relationship analysis were conducted to identify key anti-fibrotic constituents.

Results: HC alleviates PF by ameliorating inflammatory injury, reducing extracellular matrix (ECM) deposition, and suppressing epithelial-mesenchymal transition (EMT). The underlying mechanism involves modulation of the GSK-3β/β-catenin pathway, specifically through decreased phosphorylation of p-GSK-3β at Ser9, which enhances phosphorylation of β-catenin at Ser33/37/Thr41, thereby promoting its ubiquitination and subsequent degradation. Chemical analysis tentatively identified 88 compounds, including 38 newly reported in HC, with rosmarinic acid (RA) identified as a key activity-guided quality marker for the first time.

Conclusions: The therapeutic potential of HC in pulmonary fibrosis is mainly attributed to its regulation of the GSK-3β/β-catenin pathway, with RA serving as a key bioactive component.