In silico and in vivo demonstration of the regulatory mechanism of Qi-Ge decoction in treating NAFLD

Abstract

Background: Nonalcoholic fatty liver disease (NAFLD), a chronic and progressive liver disease, often causes steatosis and steatohepatitis. Qi-Ge decoction (QGD) shows a good effect against NAFLD in the clinic. But the molecular mechanism for QGD in improving NAFLD is unknown.

Purpose: This study explored the molecular mechanism of QGD in NAFLD model rats using comprehensive network pharmacology, molecular docking and in vivo verification strategies.

Methods: Active components and targets of QGD were obtained from public database. The overlapped genes between QGD and NAFLD targets were analyzed by enrichment analysis. Active components and targets were used to predict molecular docking analysis. Finally, seven key targets were screened out and the gene expression were verified in the NAFLD rat's liver tissues after QGD treatment.

Results: Fifty-eight common QGD therapeutic targets were associated with NAFLD. Molecular docking demonstrated that seven targets had strong binding ability for the corresponding active ingredients. GO analysis identified 18 biological process entries, which were mainly related to regulation of lipid storage, lipid localization and peptide transport. KEGG analysis identified multiple signaling pathways, which were mainly associated with tumor necrosis factor signaling and NAFLD. In vivo data confirmed that the effect of QGD in the treatment of NAFLD was mainly exerted through improving liver steatosis and inflammatory cell infiltration. Additionally, QGD upregulated the expression of MAPK8 and ESR1 and downregulated the transcriptional expression of IL6, VEGFA, CASP3, EGFR and MYC. These targets may affect lipid metabolism by regulating lipid storage and inflammation.

Conclusion: The integration of results obtained in silico and in vivo indicated that QGD regulates multiple targets, biological processes and signaling pathways in NAFLD, which may represent a complex molecular mechanism by which QGD improves NAFLD.Key messagesQGD intervention is related to multiple biological processes such as inflammation, oxidation and cell apoptosis in NAFLD.Lipid and atherosclerosis, TNF signaling pathway, IL-17 signaling pathway, non-alcoholic fatty liver disease and AGE-RAGE signaling pathway in diabetic complications are the main pathways for QGD intervention NAFLD.The active components of QGD can form good binding with relevant target proteins through intermolecular forces, exhibiting excellent docking activity.

Keywords: NAFLD; Qi-ge decoction; in silico study; in vivo validation; molecular docking; network pharmacology.

Figure 1.

HPLC of Qi-ge decoction.

Figure 2.

The workflow of study on the mechanism of Qi-ge decoction in the treatment of NAFLD.

Figure 3.

The Venn diagram of the potential targets sharing by Qi-Ge decoction (QGD) in the treatment of NAFLD (A) and QGD-compound-target-NAFLD network (B).①: 1,7-Dihydroxy-3,9-dimethoxy pterocarpene. ②: (6aR,11aR)-9,10-dimethoxy-6a,11a-dihydro-6H-benzofurano[3,2-c]chromen-3-ol.

Figure 4.

PPI network of potential targets (A) and core targets with higher than average degrees (15.7) (B).

Figure 5.

GO enrichment analysis (A), KEGG enrichment analysis (B) and pathway-target network (C).

Figure 6.

Molecular docking diagram of the active compounds of QGD and key targets.

Figure 7.

2D molecular docking diagram of the active compounds of QGD and key targets.

Figure 8.

Effects of the Qi-Ge decoction (QGD) on (A) weight, (B) liver index, serum (C) total cholesterol (TC), (D) triglyceride (TG), (E) low-density lipoprotein cholesterol DL-C), (F) high-density lipoprotein cholesterol (HDL-C) activities and (G) the macroscopic appearance of liver in rats with NAFLD. Data are expressed as means ± SD (n = 10). ∗p < .05 compared with the model group.

Figure 9.

Effects of the Qi-Ge decoction (QGD) on histopathological changes in rats with NAFLD (hematoxylin and eosin staining; 400×).

Figure 10.

Validation of the core genes using real-time PCR. β-actin was used for normalization in real-time PCR (A–G). Data are expressed as means ± SD (n = 3). ∗p < .05 compared with the model group.