中药药理与临床

目的：考察诃子抗呼吸道病毒作用，探索其具有抗病毒活性的单体成分及可能的作用机制。方法：通过H1N1和H1N1-RSV共感染两种小鼠模型，体内评价诃子的抗病毒作用；通过转录组学预测作用通路及靶点；qRT-PCR法检测感染小鼠肺组织中病毒载量及宿主相关因子表达变化；WB法检测造模及给药前后对cGAS-STING通路关键蛋白的调控作用。通过HPLC和对照品分析诃子中的单体成分，对检出的成分进行体外抗病毒活性检测。结果：与正常对照组比较，H1N1及H1N1-RSV感染模型对照组小鼠体质量显著降低(P<0.01)，肺指数、肺组织病毒载量及TNF-α、IL-6含量显著升高(P<0.01)，肺组织切片显示明显炎症病理改变。转录组测序结果表明，模型对照组中cGAS-STING-TBK1-IRF7信号轴相关基因表达上调(P<0.05)；qRT-PCR验证显示，模型对照组肺组织中cGAS、STING、IRF7、ZBP1等关键因子的mRNA水平较正常对照组显著升高(P<0.01)；Westernblot结果亦证实模型对照组cGAS-STING通路关键蛋白(STING、TBK1、IRF7)表达显著上调。与模型对照组比较，诃子0.8、1.6 g/kg组小鼠体质量显著升高 (P<0.01)，肺指数、病毒载量及TNF-α、IL-6含量降低(P<0.05或P<0.01)，肺组织炎症病理明显减轻。转录组及qRT-PCR结果显示，诃子能显著下调cGAS、STING、IRF7、ZBP1等因子的表达(P<0.01)；Western blot法进一步验证了诃子可显著抑制病毒感染诱导的cGAS-STING通路过度激活。体外实验中，与病毒感染细胞模型对照组比较，诃子中的诃子鞣酸、没食子酸、安石榴苷等活性成分显著抑制H1N1及RSV的复制，并发挥细胞保护作用。结论：诃子具有较好的体内抗H1N1、H1N1-RSV等呼吸道病毒共感染的活性，作用机制为抑制病毒激活的cGAS-STING通路，从而降低炎症反应，保护受损肺组织。抗病毒活性单体为诃子鞣酸、没食子酸、安石榴苷等。该研究为诃子在抗呼吸道病毒研究和临床应用方面提供一定参考，也为抗病毒药物的研发提供了新的思路。

Objective: To investigate the anti-respiratory virus effects of CHEBULAE FRUCTUS， and to explore its bioactive monomer components with antiviral activity and their potential mechanisms of action. Methods: The antiviral effects of CHEBULAE FRUCTUS were evaluated in vivo using two mouse models of H1N1 infection and H1N1-RSV co-infection. Transcriptomic analysis was performed to predict relevant pathways and targets. Quantitative real-time PCR (qRT-PCR) was used to detect viral load and changes in host-related factor expression in lung tissues of infected mice. Western blot (WB) was used to assess the regulatory effects of CHEBULAE FRUCTUS on key proteins in the cGAS-STING pathway before and after treatment. The monomer components in CHEBULAE FRUCTUS were analyzed by HPLC with reference standards， and the antiviral activities of the identified components were further evaluated in vitro. Results: Compared with the normal control group， mice in both the H1N1 and H1N1-RSV model groups showed significantly decreased body weight (P < 0.01)， significantly increased lung index， lung viral load， and levels of TNF-α and IL-6 (P < 0.01)， and obvious inflammatory pathological changes in lung tissue sections. Transcriptomic sequencing revealed that genes related to the cGAS-STING-TBK1-IRF7 signaling axis were significantly upregulated in the model groups (P < 0.05). The qRT-PCR validation demonstrated that mRNA expression levels of Cgas， Sting， Irf7， and Zbp1 in lung tissues of the model groups were significantly higher than those in the normal control group (P < 0.01). WB results further confirmed that key proteins in the cGAS-STING pathway (STING， TBK1， IRF7) were significantly upregulated in the model groups. Compared with the model control group， mice treated with CHEBULAE FRUCTUS (0.8 and 1.6 g/kg) showed significantly increased body weight (P < 0.01)， significantly decreased lung index， viral load， and TNF-α and IL-6 levels (P < 0.05 or P < 0.01)， and markedly alleviated inflammatory pathological changes in lung tissue. Transcriptomic and qRT-PCR results showed that CHEBULAE FRUCTUS significantly downregulated the expression of Cgas， Sting， Irf7， and Zbp1 (P < 0.01). WB analysis further confirmed that CHEBULAE FRUCTUS significantly inhibited the excessive activation of the virus-induced cGAS-STING pathway. In vitro experiments showed that， compared with the virus-infected cell model control group， active components of CHEBULAE FRUCTUS， including chebulinic acid， gallic acid， and punicalagin， significantly inhibited the replication of H1N1 and RSV and exerted cytoprotective effects. Conclusion: CHEBULAE FRUCTUS exhibits good in vivo antiviral activity against respiratory virus co-infection models such as H1N1 and H1N1-RSV. Its mechanism may be related to inhibition of virus-induced activation of the cGAS-STING pathway， thereby reducing inflammatory responses and protecting damaged lung tissue. The main antiviral active monomers include chebulinic acid， gallic acid， and punicalagin. This study provides a reference for the research and clinical application of CHEBULAE FRUCTUS against respiratory viral infections， and offers new insights for antiviral drug development.