Background: Network pharmacological methods were used to predict the anti-inflammatory targets and related pathways of rhein in the treatment of asthma, and to elucidate its mechanism of action. In addition, we validated the anti-inflammatory effects of rhein in human bronchial epithelial (HBE) cells.Methods: The corresponding targets of rhein were obtained from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform 2.3 (TCMSP), and molecular docking was also performed. A network of predicted rhein targets was established and analysed with Cytoscape 3.7.1. The anti-inflammatory targets in the Therapeutic Target Database 2020 (TTD) were searched to build a protein-protein interaction network (PPI), which was merged with the ingredient-target network to screen anti-inflammatory targets associated with rhein. A network of anti-inflammatory rhein targets during the in vivo treatment of asthma was constructed to screen the anti-inflammatory targets related to asthma. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis was performed with the Enrichr database and Cytoscape 3.7.1. The expression levels of proteins in the mitogen-activated protein kinase / nuclear factor kappa-B (MAPK/NF-κB) signalling pathway were assessed by western blot analysis.Results: Altogether, Eighty-three targets were obtained. Epidermal active growth factor receptor (EGFR), E-selecting (E-SELE), macrophage migration inhibitory factor (MIF), and mitogen-activated protein kinase 14 (MAPK14) might be important anti-inflammatory targets of rhein during asthma treatment. We selected the MAPK signalling pathway to determine the anti-inflammatory effects of rhein.Conclusion: The anti-inflammatory mechanism of the treatment of asthma with rhein may be related to MAPK14, EGFR, E-SELE and MIF as well as their signalling pathways. To prevent the exacerbation of asthma, instead of targeting a single pathway or a single target, all these targets and their signalling pathways should be controlled holistically. Rhein may reduce inflammation by inhibiting the MAPK/NF-κB pathway.