Investigating the mechanism of Tanyu Tongzhi decoction against atherosclerosis by regulating bile acid metabolism based on network pharmacology and experimental validation

Objective: This case is aimed to look into and analysis in prophylactic and therapeutic effect of Tanyu tongzhi decoction (TYTZD) in atherosclerosis (AS), and to explore its possible regulating mechanism, including that of bile acid metabolism, using the integrated methodology in combination of network pharmacology and experiment validation. Methods: ApoE -/- mice were fed with a high-fat diet and administered trimethylamine N-oxide (TMAO) for 12 weeks to establish an atherosclerosis model. At the end of the eighth week, the murine group was divided into four groups: high fat diet (HFD), HFD plus trimethylamine N-oxide (TMAO), Tanyu Tongzhi decoction (TYTZD), and TMAO inhibitor (DMB), one of which was a regular chow diet group as control. Following a 4-week intervention period, TMAO concentrations across all experimental groups were quantified using HPLC-MS methodology. TYTZD’s therapeutic benefits on lipid profiles and atherosclerotic lesion development in murine models were assessed via serum biochemical tests, HE staining, and Oil Red O staining techniques. The TCMSP and ETCM databases facilitated prediction of molecular targets for TYTZD bioactive constituents. AS-related disease targets were retrieved from GEO database repositories. Network pharmacology approaches enabled identification of differentially expressed genes linking “TYTZD-AS-bile acid metabolism” pathways. Enrichment analyses combined with PPI network construction revealed critical regulatory targets. TYTZD’s influence on bile acid metabolic processes in atherosclerotic mice was examined through targeted metabolomic profiling of serum and fecal samples. Finally, we validated the expression of proteins in core signaling pathways by western blotting. Results: In vivo experiments demonstrated that a high-fat diet combined with TMAO intervention significantly promoted AS progression in ApoE -/- mice, manifested primarily as a significant increase in aortic plaque burden, markedly elevated serum lipid levels, and disordered serum bile acid metabolism. Both DMB and TYTD treatments effectively reduced the in vivo TMAO levels, regulated blood lipids, alleviated aortic lipid deposition, and inhibited the formation of AS plaques. Network pharmacology analysis indicated that the potential target of TYTZD for dealing with AS was being mainly enriched by cholesterol conversion and primary bile acid synthesis pathways. CYP7A1 and CYP27A1 were identified as key genes through which TYTZD regulates bile acid metabolism. Biochemical and metabolomic analyses demonstrated that DMB and TYTZD treatments substantially elevated total circulating bile acids, augmented bile acid reserves, and restored homeostatic balance in serum and fecal bile acid profiles. Specifically, both interventions decreased blood concentrations of CA, CDCA, DCA, and LCA while concurrently increasing TUDCA levels in both serum and feces. Immunoblot analysis indicated that DMB and TYTZD suppressed hepatic FXR along with its negative regulator SHP, enhanced liver CYP7A1 expression, and reduced ileal FXR and FGF15 protein abundance. Collectively, these data indicate that TYTZD counteracts TMAO-mediated inhibition of CYP7A1, the rate-limiting enzyme governing bile acid biosynthesis, thus facilitating cholesterol catabolism into bile acids. Conclusion: TYTZD prevents the events of atherosclerosis progression by regulating the bile acid metabolism. It does this by controlling liver FXR-SHP and intestinal FXR-FGF15 signaling pathways together. Thus, bile acid synthesis facilitated by CYP7A1 increases.