| Objective: The aim of this study is to establish a simple and efficient evaluation model of Chinese herbal medicine against chronic fatigue syndrome using the model organism zebrafish,and to use metabolomics combined with transcriptomics technology to perform multidimensional evaluation of the model from endogenous metabolites,genes and related metabolic pathways,and to use the model combined with a web-based pharmacology-driven strategy to screen and discover key anti-chronic fatigue active substances in American ginseng.Methods: A model of chronic fatigue syndrome in zebrafish was induced using a combination of swim fatigue training and disturbed sleep.The three key modeling conditions,namely the length of swimming training,the number of days of swimming training and the duration of light-disrupted insomnia,were optimized using three behavioral experiments:the exhaustive swimming experiment,the autonomous behavior experiment and the T-maze.The endogenous metabolites of zebrafish model brain and muscle tissues were analyzed separately using non-targeted metabolomics techniques,and the obtained differential metabolites were analyzed for KEGG enrichment pathways.Transcriptomic techniques were used to identify differential genes in zebrafish brain and muscle tissues,and the obtained differential genes were subjected to GO functional analysis,KEGG enrichment analysis and protein interaction network and other biomimetic analyses to analyse the key genes and metabolic pathways affecting central fatigue and physical fatigue.UPLC/Q-TOF/MS was used to identify the major saponins in American ginseng,and a zebrafish model combined with a network pharmacology-driven strategy was used for the screening of key potent substances in American ginseng.Twenty-nine saponins from American ginseng were identified using UPLC/Q-TOF/MS,and a network pharmacology-driven strategy was applied to screen for the possible key component of ginsenoside F11,which was found to have significant anti-chronic fatigue activity in combination with a new zebrafish model.Results: By optimizing the swimming training time,light interference time and the number of modeling days,the modeling method of zebrafish was finally determined as the optimal modeling method with 120 min of swimming training and 24 h of light interference for a total of 14 days.After the modeling treatment,the zebrafish showed a significant reduction in exhaustive swimming time,autonomous behaviour and memory capacity.Significant increases in lactate,urea nitrogen and phosphokinase levels in muscle tissue and significant changes in DA/5-HT levels in the brain.Metabolomic and transcriptomic analyses revealed that the zebrafish model caused alterations in the brain mainly in proteins such as stat1 b,socs3a,hsd17b8 and ardc3 b,and affected steroid hormone biosynthesis,neuroactive ligand receptor interactions,glutathione metabolism,tryptophan metabolic pathway and iron death metabolic pathway through acetylcholine and 6-hydroxymelatonin metabolites Muscle fatigue in the zebrafish model was associated with the expression of ccnb1,ccnb2 and kpna7,as well as changes in adenosine diphosphate,deoxyribose 5-phosphate,2-methylbutyrylcarnitine and choline phosphate metabolites,which in turn affected valine,leucine and isoleucine degradation,oxidative phosphorylation,fatty acid metabolism and pyruvate metabolic pathways.Conclusions: This study shows that a zebrafish model of chronic fatigue syndrome can be induced using a combination of swimming fatigue training and disturbed sleep.Behavioural and in vivo biochemical parameters as well as brain and muscle metabolomics and transcriptomes show a more pronounced chronic fatigue syndrome profile,with a much shorter time period and reduced sample size compared to the traditional rat and mouse models of the same type,reflecting certain advantages.The new zebrafish model combined with a web-based pharmacology-driven strategy revealed significant anti-chronic fatigue activity of Pseudoginsenoside F11 from American ginseng. |
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