基于液相色谱-质谱代谢组学方法研究中药定喘汤对呼吸道合胞病毒感染的疗效

Respiratory syncytial virus (RSV) can cause lower respiratory tract infections, such as bronchiolitis in infants. In China, traditional asthma-relieving medicine has numerous clinical applications in the treatment of RSV infections. However, due to the complexity of the traditional Chinese medicine system, its therapeutic mechanism and main pharmacological components remain unclear. Metabolomics can be used to analyze the efficacy of traditional Chinese medicine to provide modern scientific evidence for such treatments. In this study, an animal model experiment was performed with seven groups of three-week-old rats. The model group and five intervention groups were inoculated nasally with RSV for three consecutive days, and the normal group was treated with the same amount of saline for three consecutive days under the same conditions. In parallel, the five intervention groups were treated separately with the following via intragastric administration for seven consecutive days: asthma-relieving traditional Chinese medicine decoction, its three constituent agents (ascending (xuan) therapy, descending (jiang) therapy, pyretic clearing (qing) therapy), and ribavirin. Both normal group and RSV model group were administered with normal saline via intragastric administration as controls for seven consecutive days. The fundus plasma of rats in each group was collected on day 0, day 3, and day 7. Liquid chromatography-mass spectrometry-based untargeted metabolomics analysis was performed to investigate the changes in the metabolome after RSV infection, the effects of the asthma-relieving decoction on the regulation of metabolites related to RSV infection, and the primary source of efficacy. The detected metabolite ions were corrected using internal standards. Multivariate analysis of ions with an RSD value of less than 30% in quality control (QC) samples was used to construct principal component analysis models to monitor the overall metabolic changes of each group. The results showed that, during RSV infection and treatment, the asthma-relieving decoction and the positive control ribavirin had similar effects on the overall metabolic regulation of RSV-infected rats. Among the three asthma-relieving decoction constituent agents, the ascending (xuan) therapy agents which was composed of ephedra and ginkgo had a closer metabolic regulation effect with asthma-relieving decoction, and might be the main source of pharmacological efficacy. Based on the retention time, m/z value and tandem mass spectra in the database established by our laboratory, a total of 150 metabolites were identified. Paired t-tests were performed using data of the identified metabolites before and after RSV infection in each group, and it was found that 83 metabolite levels significantly changed after RSV infection, indicating that RSV infection could lead to disorders of multiple metabolic pathways in rats. The altered pathways included aminoacyl-tRNA biosynthesis, phenylalanine, tyrosine, and tryptophan biosynthesis, primary bile acid biosynthesis, phenylalanine metabolism and sphingomyelin metabolism. On the third day, the asthma-relieving decoction had regulatory effects on several metabolites such as bile acids, amino acids, organic acids, lipids, etc. Among the three asthma-relieving decoction constituent agents, the ascending (xuan) therapy agents had more similar effects on the regulation of metabolites with the asthma-relieving decoction. On the other hand, the descending (jiang) therapy agents and pyretic clearing (qing) therapy agents down-regulated the abnormal increase in acylcarnitine caused by the RSV infection. Additionally, both asthma-relieving decoction and its constituent agents could maintain the stability of the immune system and metabolism of the intestinal flora in rats. This study used metabolomics to evaluate the efficacy of an asthma-relieving decoction and demonstrate the metabolites and the corresponding changes after asthma-relieving decoction-based treatment. It provides theoretical support for research on the therapeutic mechanism and active ingredients of asthma-relieving decoction.