Identification of Interleukin-1-Beta Inhibitors in Gouty Arthritis Using an Integrated Approach Based on Network Pharmacology, Molecular Docking, and Cell Experiments

Abstract

Background: This study aimed to investigate the molecular mechanism of Tongfengding capsule (TFDC) in treating immune-inflammatory diseases of gouty arthritis (GA) and interleukin-1-beta (IL-1β) inhibitors by using network pharmacology, molecular docking, and cell experiments.

Methods: In this study, the compounds of TFDC and the potential inflammatory targets of GA were obtained from Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), Online Mendelian Inheritance in Man (OMIM), and GeneCards databases. The TFDC-GA-potential targets interaction network was accomplished by the STRING database. The TFDC-active compound-potential target-GA network was constructed using Cytoscape software. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were used to further explore the GA mechanism and therapeutic effects of TFDC. Quantitative real-time PCR (qPCR) was used to verify whether the TFDC inhibited IL-1β in GA. Molecular docking technology was used to analyze the optimal effective compounds from the TFDC for docking with IL-1β.

Result: 133 active compounds and 242 targets were screened from the TFDC, and 25 of the targets intersected with GA inflammatory targets, which were considered as potential therapeutic targets. Network pharmacological analysis showed that the TFDC active compounds such as quercetin, stigmasterol, betavulgarin, rutaecarpine, naringenin, dihydrochelerythrine, and dihydrosanguinarine had better correlation with GA inflammatory targets such as PTGS2, PTGS1, NOS2, SLC6A3, HTR3A, PPARG, MAPK14, RELA, MMP9, and MMP2. The immune-inflammatory signaling pathways of the active compounds for treating GA are IL-17 signaling pathway, TNF signaling pathway, NOD-like receptor signaling pathway, NF-kappa B signaling pathway, Toll-like receptor signaling pathway, HIF-1 signaling pathway, etc. The TFDC reduced IL-1β mRNA expression in GA by qPCR. Molecular docking results suggested that rutaecarpine was the most appropriate natural IL-1β inhibitor.

Conclusion: Our findings provide an essential role and bases for further immune-inflammatory studies on the molecular mechanisms of TFDC and IL-1β inhibitors development in GA.

Figure 1

The workflow of IL-1β inhibitor prediction in GA.

Figure 2

Potential therapeutic targets and PPI network map of TFDC for GA. (a) The Venny results of potential therapeutic targets of TFDC for GA. (b) The PPI network map of 25 targets. (c) Count and list of the above targets of PPI network map.

Figure 3

The TFDC-active compound-potential target-GA network. The green node represents TFDC. The red node represents GA. Purple nodes represent active compounds. Blue nodes represent targets. Gray lines represent interconnections between nodes and nodes.

Figure 4

GO enrichment analysis of potential targets of TFDC in GA.

Figure 5

KEGG pathway enrichment analysis of potential targets of TFDC in GA.

Figure 6

The IL-17 signaling pathway of potential targets of TFDC in GA. Arrows indicate upstream and downstream relationships between targets. The red color represents TFDC-related targets in the network.

Figure 7

The TNF signaling pathway of potential targets of TFDC in GA. Arrows indicate upstream and downstream relationships between targets. The red color represents TFDC-related targets in the network.

Figure 8

Effects of TFDC on MSU-induced THP-1 cell viability. (a) THP-1 cells were exposed to TFDC at various concentrations for 24 h. (b) THP-1 cells were exposed to MSU at various concentrations for 24 h. (c) Protective effects of TFDC on the viabilities of MSU-induced THP-1 cells. Cell viability was assessed by CCK-8 assay and expressed relative to untreated control cells. ^∗∗P < 0.01 versus control group. ^##P < 0.01 versus MSU group.

Figure 9

TFDC protects THP-1 cells against MSU-induced inflammation by affecting the expression of IL-1β. (a) Effects of TFDC on MSU-induced THP-1 cells. (b) Statistical analysis of the effect of TFDC on the mRNA expression level of IL-1β. Data are presented as the mean ± SD (n = 3). ^∗∗∗P < 0.001 versus control group. ^###P < 0.001 versus MSU group.

Figure 10

The diagram of the binding of rutaecarpine (a), dihydrosanguinarine (b), stigmasterol (c), naringenin (d), quercetin (e), dihydrochelerythrine (f), and betavulgarin (g) with IL-1β.