Deciphering the combination mechanisms of Gualou-Xiebai herb pair against atherosclerosis by network pharmacology and HPLC-Q-TOF-MS technology

Introduction: Gualou (Trichosanthes kirilowii Maxim)-Xiebai (Allium macrostemon Bunge) (GLXB) is a well-known herb pair against atherosclerosis (AS). However, the combination mechanisms of GLXB herb pair against AS remain unclear. Objective: To compare the difference in efficacy between GLXB herb pair and the single herbs and to explore the combination mechanisms of GLXB against AS in terms of compounds, targets, and signaling pathways. Methods: The combined effects of GLXB were evaluated in AS mice. The main compounds of GLXB were identified via quadrupole time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS) and UNIFI informatics platforms. The united mechanisms of GLXB in terms of nodes, key interactions, and functional clusters were realized by network pharmacology. At last, the anti-atherosclerotic mechanisms of GLXB were validated using enzyme-linked immunosorbent assay (ELISA) and Western blot in AS mice. Results: The anti-atherosclerotic effects of the GLXB herb pair (6 g/kg) were more significant than those of Gualou (4 g/kg) and Xiebai (2 g/kg) alone. From the GLXB herb pair, 48 main components were identified. In addition, the GLXB herb pair handled more anti-atherosclerotic targets and more signaling pathways than Gualou or Xiebai alone, whereas 10 key targets of GLXB were found using topological analysis. Furthermore, the GLXB herb pair (6 g/kg) could suppress the inflammatory target levels of IL-6, IL-1 beta, TNF-alpha, ALOX5, PTGS2, and p-p38 in AS mice. GLXB herb pair (6 g/kg) could also ameliorate endothelial growth and function by regulating the levels of VEGFA, eNOS, p-AKT, VCAM-1, and ICAM-1 and reducing macrophage adhesion to vascular wall in AS mice. GLXB herb pair (6 g/kg) could improve the blood lipid levels in AS mice. In addition, the regulating effects of GLXB herb pair (6 g/kg) on levels of IL-1 beta, TNF-alpha, ALOX5, VEGFA, eNOS, VCAM-1, ICAM-1, and blood lipids were more significant than those of Gualou (4 g/kg) or Xiebai alone (2 g/kg). Conclusion: The combination mechanisms of the GLXB herb pair were elucidated in terms of components, targets, and signaling pathways, which may be related to suppressing inflammation, regulating vascular endothelial growth/function, and improving blood lipid levels.