Chaihu Guizhi decoction alleviates H1N1 influenza virus and Staphylococcus aureus co-infection in mice by reducing inflammatory cytokines and regulating Th17/Treg balance.

ETHNOPHARMACOLOGICAL RELEVANCE: Chaihu Guizhi Decoction (CGD), a classical formula from Treatise on Cold Damage, is traditionally used to "harmonize Shaoyang" and "regulate Ying and Wei" in traditional Chinese medicine. Clinically, it has been applied in treating influenza and bacterial infections. However, its therapeutic efficacy and pharmacological mechanisms in influenza and Staphylococcus aureus (S. aureus) co-infection remain incompletely clarified.

OBJECTIVE

This study aimed to investigate the intervention effect and underlying mechanism of CGD in influenza and S. aureus co-infection.

METHOD: Male BALB/c mice were randomly divided into 8 groups. On day 1, mice received a nasal injection of 20 μL influenza virus suspension (0.2 TCID50) diluted in normal saline; on day 3, a nasal injection of 20 μL S. aureus suspension (2.5 × 107 CFU) was administered to establish an influenza virus and S. aureus co-infection model. Six days after viral infection, all mice were anesthetized with 1.5 % isoflurane in 98.5 %O2, followed by sample collection for subsequent analysis. The relative expression level of the influenza virus M gene was quantified by RT-qPCR. Bacterial load was determined using the smear-plate method. Lung histopathological changes were evaluated via hematoxylin-eosin (H&E) staining. Flow cytometry was employed to analyze the populations of Th17 and Treg lymphocytes, as well as the levels of 13 inflammatory cytokines.

RESULTS

CGD exerted a therapeutic effect on influenza virus and S. aureus co-infection. Specifically, CGD alleviated clinical manifestations, including improving body weight, spontaneous activity, and food consumption in mice. It increased the thymus index while decreasing the spleen and lung indexes. Additionally, CGD reduced the bacterial load and influenza virus M gene expression in lung tissues, and mitigated inflammatory pathological damage. It also decreased the Th17/Treg cell ratio and the levels of multiple inflammatory cytokines, including IL-4, IL-12p70, IL-6, IL-17A, IL-13, IL-2, IL-10, TNF-α, IFN-γ, BCMA, sCD40L, and TGF-β1.

CONCLUSION

CGD protects mice against influenza and S. aureus co-infection by suppressing viral replication, mitigating the inflammatory cytokine storm, and restoring Th17/Treg immune balance.