目的 探究清肺解毒颗粒通过肠道菌群对呼吸道合胞病毒（respiratory syncytial virus，RSV）感染小鼠固有免疫的影响。方法 将BALB/c雌性小鼠随机分为对照组、模型组、清肺解毒颗粒组、抗生素组、抗生素＋清肺解毒颗粒组。在滴鼻RSV造模前，抗生素组及抗生素＋清肺解毒颗粒组小鼠ig四联抗生素4周（0.15 mL），构建伪无肠道菌表型。除对照组外其余各组滴鼻RSV，清肺解毒颗粒组、抗生素＋清肺解毒颗粒组小鼠ig清肺解毒颗粒（120 mg/kg），1次/d，3 d后处死小鼠。苏木素-伊红（hematoxylin-eosin，HE）染色法观察肺组织的病理改变，q-PCR检测小鼠肺组织呼吸道合胞病毒融合蛋白（respiratory syncytial virus-fusion protein，RSV-F）、呼吸道合胞病毒附着蛋白（respiratory syncytial virus-glycoprotein，RSV-G）、呼吸道合胞病毒非结构蛋白1（respiratory syncytial virus-non-structural protein 1，RSV-NS1）、白细胞介素-1β（interleukin-1β，IL-1β）、IL-6、肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）、干扰素-α（interferon-α，IFN-α）、IFN-β、人I型黏病毒对抗蛋白（myxovirus resistance proteins I，MX1）、MX2、干扰素刺激基因15（interferon stimulated gene 15，ISG15）、三重基序蛋白5（tripartite motif-containing protein 5，TRIM5）、2'-5'寡聚腺苷酸合成酶-1（2'-5'-oligoadenylate synthetase-2，OAS1）、OAS2、蛋白激酶R（protein kinases R，PKR）mRNA表达水平。采用高效液相色谱-质谱法检测小鼠粪便中对羟基苯丙酸（desaminotyrosine，DAT）含量，ELISA方法检测血清IFN-α、IFN-β水平。免疫组化、免疫荧光、Western blotting方法检测维甲酸诱导基因-I（retinoic acid inducible gene-I，RIG-I）、线粒体信号蛋白（mitochondrial antiviral-signaling protein，MAVS）、MX1、OAS1表达水平。结果 与模型组比较，清肺解毒颗粒组药效明显，即小鼠肺组织病理性损伤得以显著改善、RSV病毒水平、炎性因子水平显著降低（P＜0.01），与清肺解毒颗粒组比较，抗生素＋清肺解毒颗粒组小鼠药效减弱（P＜0.05、0.01）。与模型组比较，清肺解毒颗粒组小鼠固有免疫抗病毒效应中的RIG-I、MAVS、IFN-α、IFN-β、MX1、MX2、OAS1、OAS2、ISG15、TRIM5、PKR mRNA表达水平升高（P＜0.01），与清肺解毒颗粒组比较，抗生素＋清肺解毒颗粒组小鼠上述mRNA表达水平显著降低（P＜0.05、0.01）。与模型组比较，清肺解毒颗粒组DAT含量显著增加（P＜0.01），与清肺解毒颗粒组比较，抗生素＋清肺解毒颗粒组DAT含量显著降低（P＜0.05）。结论 清肺解毒颗粒可能通过调控RIG-I/MAVS/I-IFN信号通路，活化固有免疫抗病毒反应，发挥抗RSV效应，清除肠道菌群则影响其药效。

Objective To investigate the effects of Qingfei Jiedu Granules (清肺解毒颗粒) on the innate immunity of respiratory syncytial virus (RSV) infected mice through intestinal flora. Methods BALB/c female mice were randomly divided into control group, model group, Qingfei Jiedu Granules group, antibiotic group, antibiotic + Qingfei Jiedu Granules group. Before nasal RSV modeling, mice in the antibiotic group were administrated with a quadruple antibiotic (0.15mL) for four weeks to establish a pseudopteral bacterial phenotype. Except control group, mice in the other groups were given nasal RSV, Qingfei Jiedu Granules group and antibiotic+Qingfei Jiedu Granules group by intragastric administration (120mg/kg), and the mice were killed 3 days later. Hematoxylin-eosin (HE) was used to observe the pathological changes of lung tissue. Respiratory syncytial virus-fusion protein (RSV-F), glycoprotein (RSV-G), non-structural protein 1 (RSV-NS1), interleukin-1β (IL-1β), IL-6, tumor necrosis factor-α (TNF-α), interferon-α (IFN-α), IFN-β, myxovirus resistance proteins I (MX1), MX2, interferon stimulated gene 15 (ISG15), tripartite motif-containing protein 5 (TRIM5), 2'-5'-oligoadenylate synthetase-2 (OAS2), protein kinases R (PKR) of mice lung tissue were detected by q-PCR. The content of desaminotyrosine (DAT) in feces was detected by HPLC/MS, and the serum IFN-α, IFN-β levels were detected by ELISA. The expression levels of retinoic acid inducible gene I (RIG-I), mitochondrial antiviral-signaling protein (MAVS), MX1 and OAS1 were detected by immunohistochemistry, immunofluorescence and Western blotting. Results Compared with the model group, the efficacy of Qingfei Jiedu Granules group was obvious, that is, the pathological injury of lung tissue was significantly improved, and the levels of RSV virus and inflammatory factors were significantly decreased (P < 0.01). Compared with Qingfei Jiedu Granules group, the efficacy of antibiotic + Qingfei Jiedu Granules group was weakened (P < 0.05, 0.01). Compared with model group, levels of RIG-I, MAVS, IFN-α, IFN-β, MX1, MX2, OAS1, OAS2, ISG15, TRIM5, PKR in Qingfei Jiedu Granules group were increased (P < 0.01). Compared with Qingfei Jiedu Granules group, the expression levels of MX1, MX2, OAS1, OAS2, ISG15, TRIM5 and PKR in antibiotic + Qingfei Jiedu Granules group were significantly decreased (P < 0.05, 0.01). Compared with the model group, the DAT content in Qingfei Jiedu Granules group was significantly increased (P < 0.01), while the DAT content in antibiotic + Qingfei Jiedu Granules group was significantly decreased (P < 0.05). Conclusion Qingfei Jiedu Granules may activate the innate immune antiviral response by regulating RIG-I/MAVS/I-IFN signaling pathway, and exert anti-RSV effect, while clearing intestinal flora affects its efficacy.

R285.5

河南省重点研发与推广专项（科技攻关）项目（242102310508）;2023年度河南省“双一流”创建学科中医学科学研究专项（HSRP-DFCTCM-2023）;河南中医药大学博士科研启动项目（00104311-2024-2-33）