Mechanistic investigation of Liujing Toutong tablet in the treatment of cerebral ischemia–reperfusion injury in rats by the integration of serum pharmacochemistry and metabolomics

Abstract

Background: Liujing Toutong tablet (LTT) is a traditional Chinese patent medicine. Previous studies have demonstrated that LTT exerts a protective effect in permanent cerebral ischemia and cerebral ischemia–reperfusion injury (CIRI). However, the active compounds and underlying mechanisms remain unclear. This investigation aimed to integrate the analyses of serum pharmacochemistry, network pharmacology, and metabolomics to elucidate the therapeutic effects of LTT on CIRI.

Methods: The therapeutic effects of LTT were evaluated in the CIRI rat model, which was induced using the thread embolism method. The prototype active compounds of LTT absorbed into the serum (LPCs) were identified using ultra-performance liquid chromatography-quadrupole/orbitrap high-resolution mass spectrometry (UPLC-Q-Exactive-Orbitrap/MS). Potential targets of LPCs were determined using network pharmacological analysis. A combination of metabolomics and network pharmacology was employed to identify upstream key targets and downstream endogenous metabolites. The predicted mechanisms were then verified by molecular docking. Finally, key targets and signaling pathways were examined using enzyme-linked immunosorbent assay (ELISA) and Western blot.

Results: A total of 27 LPCs were identified as the active ingredients. Furthermore, glycerophospholipid and arachidonic acid (AA) metabolism were also identified, with PTGS2, ALOX5, and AchE as the upstream key targets, and phosphorylcholine, AA, phosphatidate (PA), hosphatidylcholines (PCs), and lysophosphatidylcholines (LysoPCs) as the core endogenous metabolites. Additionally, LPCs binding with these key targets might reduce the inflammatory response via nuclear factor Kappa-B (NF-κB) signaling pathway.

Conclusion: LTT may effectively alleviate symptoms of CIRI by acting on PTGS2, ALOX5, and AchE, improving the glycerophospholipid and AA metabolism and reducing the inflammatory response by inhibiting NF-κB signaling pathway-related proteins.

Keywords

cerebral ischemia–reperfusion injury / Liujing Toutong tablet / metabolomics / network pharmacology / serum pharmacochemistry

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