Si-Ni Decoction as a Potential Treatment for Ulcerative Colitis: Modulation of Gut Microbiota and AKT1 Inhibition Through Network Pharmacology and in vivo Validation

Abstract

Background: Sini Decoction (SND), a time-honored formulation in traditional Chinese medicine, consists of three key ingredients: aconite, licorice, and ginger rhizome. It has been used for more than a thousand years to relieve symptoms associated with acute gastroenteritis, dyspepsia, and abdominal discomfort, but its therapeutic efficacy in ulcerative colitis (UC) and the mechanisms involved have not been validated. In this study, a comprehensive approach integrating network pharmacology, molecular docking, molecular dynamics simulation and experimentation was used to assess the efficacy of SND in the treatment of UC and to explore its molecular mechanisms.

Methods: The bioactive compounds associated with ulcerative colitis (UC) were identified using the TCMSP database, with potential targets predicted via the Swiss Target Prediction database. Protein-protein interaction networks were constructed using the STRING database and Cytoscape and the most important genes were identified. Subsequently, molecular docking was combined with molecular dynamics simulations using molecular docking to assess the binding affinity of the main active ingredient of SND to AKT1. To evaluate the therapeutic effects of SND, we utilized a dextran sodium sulfate-induced UC mouse model. Additionally, fecal samples were collected for analysis of the intestinal microbiota to explore the influence of SND on gut flora composition.

Results: Fifteen bioactive components from SND were identified, and their activities were validated. The results indicated that AKT serine/threonine kinase 1 is a core target of SND for the treatment of UC. The anti-inflammatory, intestinal barrier-protective, and microbiota-regulating effects of SND were confirmed in animal models, alongside evidence of its inhibitory effect on AKT1.

Conclusion: The active ingredients of SND were screened, with a focus on AKT1 inhibition, to reduce inflammation in UC, protect the intestinal barrier, and regulate the intestinal microbiota, demonstrating significant therapeutic potential.

Keywords: gut microbiota; intestinal barrier; molecular docking; network pharmacology; sini decoction; ulcerative colitis.

Figure 1

Screening of SND targets and key active ingredients for the treatment of UC. (A) Venn diagram, illustrating a total of 193 overlapping genes associated with ulcerative colitis in SND treatment. (B) Herbs- Compounds-Target Network Construction of SND in the treatment of UC. (C) All active ingredients were ranked according to “degree” and the top 15 active compounds were selected.

Figure 2

Specific chemical formulae for the first 15 active compounds. From (A–O) are Sexangularetin, DFV, Jaranol.naringenin.6-prenylated eriodictyol, Quercetin der., Glabridin, Sigmoidin-B, licoisoflavanone, glyuranolide, 7,2’,4’-trihydroxy–5-methoxy-3–arylcoumarin, isorhamnetin, kanzonols W, Karanjin, Delphin_qt. Protein-protein interaction (PPI) network depicting the interactions of SND-related proteins involved in UC treatment.

Figure 3

PPI network of SND targets in UC therapy and core targets. (A) PPI network of 193 targets in the STRING database. (B) 32 core targets were screened out of the 193 individual targets, where the core targets were labelled in yellow. (C) The PPI network was analysed using the Cytoscape software to analyse the PPI network to show the key targets affected by CR. The higher the degree value, the closer the colour is to red.

Figure 4

GO and KEGG enrichment analysis of the target of OA treated by CR. (A) GO analysis was performed to predict biological processes/cellular components/ molecular functions affected by SND. The horizontal axis as well as the large of the dots indicate the number of small, enriched genes, the closer the colour to red means the closer the P-value is to zero, and the vertical axis indicates the name of the term. (B) KEGG analysis was performed to predict pathways affected by snd. The horizontal axis is the number of genes, and the vertical axis is the term name.

Figure 5

Molecular docking of key components of SND and AKT1. From (A–J) are Q Quercetin der., Delphin_qt, licoisoflavanone, Jaranol, 6-prenylated eriodictyol, 7,2’,4’-trihydroxy–5-methoxy-3–arylcoumarin, Glabridin, kanzonols W, Sexangularetin, DFV.

Figure 6

Molecular dynamics modelling of three herbal components of SND (Quercetin der, Delphin_gt and Sexangularetin). A The root mean square deviation (RMSD) of AKT1-Delphin complex system, AKT1-Quercetin complex system, and AKT1-Sexangularetin complex system, the smaller the deviation, the better the conformational stability. B-C Radius of Gyration (Rg) and solvent-accessible surface area (SASA) of AKT1-Delphin, AKT1-Quercetin and AKT1-Sexangularetin composite systems, the slight fluctuation of the complex during movement indicates that the complex system has undergone conformational changes during movement. D The number of hydrogen bonds between small molecules and target proteins in the kinetic process, and the number of hydrogen bonds between AKT1-Delphin complex systems ranged from 0 to 9. In most cases, the complex has about six hydrogen bonds. The number of hydrogen bonds between the AKT1-Quercetin der complex systems ranges from 0 to 4. In most cases, the complex has about three hydrogen bonds. The number of hydrogen bonds between the AKT1-Sexangularetin complex systems ranges from 0 to 6. In most cases, the complex has about four hydrogen bonds. E Root mean square fluctuation (RMSF) of AKT1-Delphin, AKT1-Quercetin der and AKT1-Sexangularetin complex Systems.

Figure 7

SND suppresses the progression of experimental colitis. To induce colitis in mice, mice were first treated with 2% DSS for 7 days, while SND was gavage for 7 days, and mice received 0.9% saline as a negative control. (A) Representative mouse gross morphology of the colon images was captured. (B) Colon length was measured. (C) DAI of mice was calculated daily during the period of observation. (D) The changes of body weight of mice were expressed as a percentage of the initial weight at the start of the experiment. (E) Representative photomicrographs of colon sections stained with haematoxylin and eosin (H&E) were examined. **P < 0.01, ***P < 0.001.

Figure 8

SND treatment inhibits the expression of pro-inflammatory genes and promotes the expression of anti-inflammatory genes in DSS-induced colitis in mice. (A) SND inhibited the increase in IL-1β levels in colon tissue homogenates of DSS mice and (B) suppressed the decrease in IL-10 levels. (C–F) SND reduced the mRNA expression levels of IL-Iβ, IL-6, IL-17A, and TNF-α and (G) suppressed the decrease in IL-10 mRNA expression levels in colon samples from mice with colitis by qRT-PCR. Gene expression was normalized to β-actin in each group. (H) The expression of p-AKT was significantly increased in the DSS group. *P < 0.05, **P < 0.01, ***P < 0.001.

Figure 9

SND alleviates the impaired intestinal barrier associated with colitis. (A) The expression of tight junction proteins Occludin, E-cadherin, CLaudin-1, ZO-1 was increased in the SND DSS group compared to the DSS group. (B) Four groups of mice were gavage with FITC- dextrose and fluorescence levels in plasma were measured. **P < 0.01.

Figure 10

β-Diversity of fecal microbiota in mice. (A) UPGMA Sample Hierarchical Clustering Map of fecal microbiota in four groups of mice. (B and C) β-Diversity of fecal microbiota in colitis mice Represented by PCA and PcoA.

Figure 11

SND increased beneficial bacteria and decreased harmful bacteria. (A) A histogram of the top 15 species in terms of abundance at the phylum level. (B) Relative abundance values of Desulfobacterota at the phylum level.(C) Relative abundance values of Bacteroidota at the phylum level. (D) A histogram of the top 15 species in terms of abundance at the order level. (E) Relative abundance values of Desulfovibrionales at the order level. (F) Relative abundance values of Lactobacillales at the order level. (G) A histogram of the top 15 species in terms of abundance at the family level. (H) Relative abundance values of Desulfovibrionaceae at the family level. (I) Relative abundance values of Peptococcaceae at the family level. *P < 0.05, **P < 0.01, ***P < 0.001.