目的 探索土木香内酯体内外的抗病毒作用，并基于I型干扰素（type I interferon，IFN-Ⅰ）通路探究其抗病毒作用机制。方法 CCK-8法检测土木香内酯对A549细胞活力的影响；利用表达绿色荧光蛋白的水疱性口炎病毒（vesicular stomatitis virus expressing green fluorescent protein，VSV-GFP）感染细胞模型，结合流式细胞术检测土木香内酯对病毒复制的影响；qRT-PCR检测土木香内酯对甲型流感病毒（influenza A virus，H1N1）、脑心肌炎病毒（encephalomyocarditis virus，EMCV）复制的影响；Western blotting检测土木香内酯对VSV病毒G蛋白和H1N1病毒NP蛋白表达的影响；利用生物信息学初步分析土木香内酯的抗病毒分子机制；qRT-PCR检测药物处理后干扰素α1（interferon α1，Ifna1）、干扰素β1（interferonβ1，Ifnb1）、干扰素诱导蛋白1（interferon-induced protein with tetratricopeptide repeats 1，Ifit1）、干扰素刺激基因15（interferon stimulated gene 15，Isg15）的mRNA表达变化；流式细胞术和qRT-PCR检测I型干扰素受体1（type I interferon receptor 1，IFNAR1）敲除（Ifnar1^−/−）细胞中，土木香内酯的抗病毒作用与干扰素信号通路的关系；构建小鼠H1N1病毒滴鼻感染模型探究土木香内酯的体内抗病毒作用。结果 土木香内酯体外抑制VSV、H1N1和EMCV病毒复制；土木香内酯激活IFN-I信号通路；在Ifnar1^−/−细胞中土木香内酯抗病毒作用被削弱；土木香内酯提高H1N1感染小鼠的存活率，降低脏器H1N1病毒载量。结论 土木香内酯激活基于IFN-I信号通路的抗病毒天然免疫反应，发挥抵抗多种病毒复制的作用。

Objective To explore the antiviral effects of alantolactone in vitro and in vivo, and investigate its mechanism based on type I interferon (IFN-I) pathway. Methods The CCK-8 assay was used to evaluate the effect of alantolactone on viability of A549 cells. The vesicular stomatitis virus expressing green fluorescent protein (VSV-GFP) was employed to infect cells to establish a model, and flow cytometry was used to assess the influence of alantolactone on virus replication. qRT-PCR was conducted to measure the effects of alantolactone on the replication of influenza A virus (H1N1) and encephalomyocarditis virus (EMCV). Western blotting was performed to determine the effect of alantolactone on the expressions of VSV G protein and H1N1 NP protein. Bioinformatics was used to investigate the antiviral molecular mechanism of alantolactone. qRT-PCR was employed to measure the mRNA expressions of interferon α1 (Ifna1), interferon β1 (Ifnb1), interferon-induced protein with tetratricopeptide repeats 1 (Ifit1) and interferon stimulated gene 15 (Isg15) after alantolactone treatment. Flow cytometry and qRT-PCR were used to investigate the relationship between the antiviral effects of alantolactone and the interferon signaling pathway in type I interferon receptor 1 (IFNAR1) knockout cells (Ifnar1^−/−). To investigate the antiviral effects of alantolactone in vivo, a mouse model of H1N1 virus intranasal infection was established. Results Alantolactone exhibited inhibition of VSV, H1N1 and EMCV virus replication in vitro. Alantolactone activated IFN-I signaling pathway. The antiviral effects of alantolactone were weakened in Ifnar1^−/− cells. Alantolactone increased the survival rate of H1N1-infected mice and reduced the viral load of H1N1 in organs. Conclusion Alantolactone activates the antiviral innate immune response based on IFN-I signaling pathway, exhibiting the ability to resist the replication of various viruses.

北京市科技新星计划课题（20230484342）；中华中医药学会青年人才托举工程项目（CACM-2023-QNRC2-A02）；北京市自然科学基金资助项目(7242239)