Oxymatrine Modulation of TLR3 Signaling Pathway: A Dual-Action Mechanism against H9N2 Avian Influenza Virus and Immune Regulation

Abstract

Abstract Background H9N2 Avian Influenza Virus (AIV) poses a growing public health threat due to its rapid mutation rate and limited vaccine efficacy. Pulmonary Microvascular Endothelial Cells (PMVECs) play a critical role as a gateway for infection, highlighting the need for alternative therapeutic strategies. This study examines the antiviral potential of Oxymatrine (OMT), a traditional Chinese medicine derivative, against H9N2 AIV in PMVECs. Purpose The aim of this study is to explore the efficacy of OMT in modulating antiviral responses and to elucidate its impact on the TLR3 signaling pathway in PMVECs infected with H9N2 AIV. Study Design and Methods: Using an array of in vitro assays such as TCID50, CCK-8, RT-qPCR, ELISA, and Western blot, this study evaluated the viral infectivity, cell viability, gene and protein expression levels, and key cytokine levels in PMVECs. Additionally, RNAi technology was employed to silence TLR3 genes to further understand the mechanisms involved. Results OMT displayed a dose-dependent inhibitory effect on vital antiviral proteins PKR and Mx1 and modulated the expression of Type I interferons and cytokines including IFN-α, IFN-β, IL-6, and TNF-α. It significantly impacted the TLR3 signaling pathways, affecting downstream components such as NF-κB and IRF-3. TLR3 silencing studies indicated that OMT's antiviral efficacy was not solely dependent on the TLR3 pathway. Conclusion Our findings reveal that OMT exhibits a dual-action mechanism by inhibiting H9N2 AIV and modulating immune responses in PMVECs, primarily through the TLR3 signaling pathway. These results lay a promising foundation for the development of OMT as an alternative antiviral therapeutic against H9N2 AIV.