Antiviral effect of fufang yinhua jiedu (FFYH) granules against influenza A virus through regulating the inflammatory responses by TLR7/MyD88 signaling pathway

Ethnopharmacological relevance: Fufang-Yinhua-Jiedu Granules (FFYH) optimized from a Yin-Qiao-San, as traditional Chinese medicine (TCM), was used to treat influenza and upper respiratory tract infection and was recommended for the prevention and treatment of SARS in 2003 and current COVID-19 in Anhui Province in 2020. Aim of study: In the clinical studies, FFYH was very effective for the treatment of influenza, but the mechanism of action against influenza A virus remains unclear. In the present study, we investigated the antiviral effect of FFYH against influenza A virus in vitro and vivo. Moreover, the potential mechanism of FFYH against influenza A virus in vivo was investigated for the first time. Materials and methods: CPE inhibition assay and HA assay were used to evaluate the in vitro antiviral effects of FFYH against influenza A virus H1N1, H3N2, H5N1, H7N9 and H9N2. Mice were used to evaluate the antiviral effect of FFYH in vivo with ribavirin and lianhuaqingwen as positive controls. RT-PCR was used to quantify the mRNA transcription of TNF-alpha, IL-6, IFN-gamma, IP10, and IL-1 beta mRNA. ELISA was used to examine the expression of inflammatory factors such as TNF-alpha, IL-6, IFN-gamma, IP10, and IL-1 beta in sera. The blood parameters were analyzed with auto hematology analyzer. Moreover, the potential mechanism of FFYH against influenza A virus in vivo was also investigated. Results: FFYH showed a broad-spectrum of antiviral activity against H1N1, H3N2, H5N1, H7N9, and H9N2 influenza A viruses. Furthermore, FFYH dose-dependently increased the survival rate, significantly prolonged the median survival time of mice, and markedly reduced lung injury caused by influenza A virus. Also, FFYH significantly improve the sick signs, food taken, weight loss, blood parameters, lung index, and lung pathological changes. Moreover, FFYH could markedly inhibit the inflammatory cytokine expression of TNF-alpha, IL-6, IFN-gamma, IP10, IL-10, and IL-1 beta mRNA or protein via inhibition of the TLR7/MyD88/NF-kappa B signaling pathway in vivo. Conclusion: FFYH not only showed a broad-spectrum of anti-influenza virus activity in vitro, but also exhibited a significant protective effect against lethal influenza virus infection in vivo. Furthermore, our results indicated that the in vivo antiviral effect of FFYH against influenza virus may be attributed to suppressing the expression of inflammatory cytokines via regulating the TLR7/MyD88/NF-kappa B signaling pathway. These findings provide evidence for the clinical treatment of influenza A virus infection with FFYH.