Hypoxia-inducible factor 1-alpha inhibition mitigates inflammatory responses induced by H9N2 avian influenza virus.

H9N2 avian influenza virus (AIV) poses significant threats to poultry and public health, with the potential to contribute to the emergence of more virulent influenza strains. Hypoxia-inducible factor 1-alpha (HIF-1α) plays a crucial role in viral pathogenesis, immune regulation, and inflammation, making it a promising target for therapeutic intervention. Here, we explored the involvement of HIF-1α in H9N2 AIV pathogenesis and associated inflammatory responses. Our data revealed that HIF-1α exacerbates inflammation induced by H9N2 AIV infection through its regulation of mitochondrial fitness. Additionally, we investigated Sotetsuflavone (Seo), a naturally derived flavonoid, for its potential to modulate inflammatory responses following H9N2 infection. Molecular docking coupled with surface plasmon resonance assays indicated that Seo exhibits robust binding affinity to HIF-1α, implying a direct inhibitory mechanism. Subsequent in vitro studies demonstrated that Seo greatly reduced the production of inflammatory cytokines, including IL-1β, IL-6, and TNF-α, through suppressing the HIF-1α signaling pathway in HD11 cells infected with H9N2 AIV. Correspondingly, in vivo experiments established that Seo administration significantly ameliorated pulmonary inflammation in H9N2-infected mice, while also enhancing alveolar epithelial repair, as evidenced by increased expression of type II alveolar epithelial cell markers Abca3 and Sftpb. Collectively, these findings identify Seo as a potent inhibitor of HIF-1α, highlighting its therapeutic potential in mitigating inflammation and lung damage associated with influenza virus infection.IMPORTANCEH9N2 avian influenza virus triggers inflammatory lung injury via hypoxia-inducible factor-1 alpha (HIF-1α)-mediated dysregulation of mitochondrial function. The natural flavonoid Sotetsuflavone (Seo) directly binds and inhibits HIF-1α, as confirmed by molecular docking and surface plasmon resonance assays. In H9N2-infected HD11 cells, Seo suppresses HIF-1α signaling, reducing IL-1β, IL-6, and TNF-α production. In vivo, Seo treatment attenuates pulmonary inflammation and enhances alveolar epithelial repair in mice, upregulating type II pneumocyte markers (Abca3 and Sftpb). These findings identify Seo as a promising HIF-1α-targeted therapeutic for influenza-associated immunopathology.