Influenza A virus (IAV) infection represents a global health challenge. Excavating antiviral active components from traditional Chinese medicine (TCM) is a promising anti-IAV strategy. Our previous studies have demonstrated that 14-deoxy-11,12-didehydroandrographolide (DAP), a major ingredient of a TCM herb called Andrographis paniculata, shows anti-IAV activity that is mainly effective against A/chicken/Hubei/327/2004 (H5N1), A/duck/Hubei/XN/2007 (H5N1), and A/PR/8/34 (H1N1) in vitro and in vivo. However, the underlying anti-IAV molecular mechanism of DAP needs further investigation. In the present work, we found that DAP can significantly inhibit the apoptosis of human lung epithelial (A549) cells infected with A/chicken/Hubei/327/2004 (H5N1). After DAP treatment, the protein expression levels of cleaved PARP, cleaved caspase-3, and cleaved caspase-9, and the activities of caspase-3 and caspase-9 in H5N1-infected A549 cells were all obviously downregulated. However, DAP had no inhibitory effect on caspase-8 activity and cleaved caspase-8 production. Meanwhile, the efficacy of DAP in reducing the apoptotic cells was lost after using the inhibitor of caspase-3 or caspase-9 but remained intact after the caspase-8 inhibitor treatment. Moreover, DAP efficiently attenuated the dissipation of mitochondrial membrane potential, suppressed cytochrome c release from the mitochondria to the cytosol, and decreased the protein expression ratio of Bax/Bcl-2 in the mitochondrial fraction. Furthermore, the silencing of caspase-9 reduced the yield of nucleoprotein (NP) and disabled the inhibitory ability of DAP in NP production in A549 cells. Overall results suggest that DAP exerts its antiviral effects by inhibiting H5N1-induced apoptosis on the caspase-9-dependent intrinsic/mitochondrial pathway, which may be one of the anti-H5N1 mechanisms of DAP.Keywords: 14-Deoxy-11,12-dehydroandrographolide; Antiviral; Apoptosis; Caspase-9; H5N1.

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乐备实（上海优宁维生物科技股份有限公司旗下全资子公司），是国内专注于提供高质量蛋白检测以及组学分析服务的实验服务专家，自2018年成立以来，乐备实不断寻求突破，公司的服务技术平台已扩展到单细胞测序、空间多组学、流式检测、超敏电化学发光、Luminex多因子检测、抗体芯片、PCR Array、ELISA、Elispot、PLA蛋白互作、多色免疫组化、DSP空间多组学等30多个，建立起了一套涵盖基因、蛋白、细胞以及组织水平实验的完整检测体系。

 
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