Therapeutic potential of Isatidis Radix alkaloids in RSV pneumonia: A focus on AhR-mediated pulmonary barrier repair.

ETHNOPHARMACOLOGICAL RELEVANCE: The traditional use of Isatis tinctoria L. (Isatidis Radix) in East Asian medicine centers on managing febrile diseases and various respiratory ailments. Classical medical texts document its function of "clearing heat and detoxifying". These traditional applications are highly consistent with its reported therapeutic effects against viral pneumonia. However, its clinical efficacy in treating respiratory syncytial virus (RSV) pneumonia and the underlying regulatory mechanisms involved have not been systematically elucidated.

AIM OF THE STUDY: This research sought to evaluate the therapeutic efficacy and prospects of Isatidis Radix alkaloids (IRA) in RSV pneumonia and to elucidate the molecular mechanisms underlying their role in restoring pulmonary barrier integrity.

MATERIALS AND METHODS: By employing bioinformatics in conjunction with molecular docking, the initial step was to screen for the bioactive constituents within Isatidis Radix and their corresponding key targets, and to predict their potential mechanisms of action. We next constructed an RSV pneumonia mouse model for evaluating IRA-mediated changes in immune organ parameters, cytokine production, and lung histology. The impact of IRA on immunomodulation was then assessed using flow cytometry, qPCR analysis, and various other methods. Following this, epithelial barrier repair was evaluated by Western blotting and the Evans blue assay. Ultimately, we verified that the barrier-protective effect of IRA is specifically mediated by the aryl hydrocarbon receptor (AhR) using techniques such as gene silencing and Western blotting, along with other assays. Finally, an AhR knockout (AhR KO) mouse model of RSV pneumonia was utilized to verify the specificity of the IRA-mediated barrier protection for AhR.

RESULTS: Alkaloids were identified as the principal active constituents of Isatidis Radix mediating its anti-RSV pneumonia effect, based on bioinformatics analysis and molecular docking results, and AhR was identified as a key target. In the RSV pneumonia mouse model, IRA significantly restored immune organ function, improved inflammatory cytokine levels, upregulated the expression of tight junction proteins, and enhanced pulmonary barrier integrity. Subsequent mechanistic investigation revealed that the activation of AhR served as the primary pathway mediating the effects of IRA. Most importantly, AhR knockout completely abolished the protective effect of IRA on barrier function, confirming that this effect is dependent on the AhR pathway.

CONCLUSION: Our study demonstrates that IRA exerts its therapeutic effect against RSV pneumonia and repairs the lung epithelial barrier by modulating AhR activity. These findings indicate that IRA possesses potential value in the treatment of RSV pneumonia.