Shenling Jiangzhi formula alleviates atherosclerosis in ApoE-/- mice by inhibiting vascular inflammation via the NF-κB/NLRP3 pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Atherosclerosis (AS) falls into the categories of "blood stasis" and "vascular impediment" in traditional Chinese medicine (TCM) theory. Its core pathogenesis is spleen deficiency with excessive dampness, and intermingling of phlegm and stasis. Shenling Jiangzhi Formula (SJF), a clinically common prescription for treating AS, possesses the effects of replenishing qi and invigorating the spleen, tonifying the kidney and resolving turbidity, promoting blood circulation and dredging collaterals, clearing the liver and detoxifying. However, its specific pharmacological mechanism remains unclear.

AIM OF THE STUDY: To characterize the chemical components of SJF and explore its pharmacodynamic substances, observe the regulatory effects of SJF on plaque formation and inflammatory response in the AS model of ApoE-/- mice, and clarify the pharmacodynamic effects and anti-AS mechanism of SJF.

MATERIALS AND METHODS: UPLC-Q-Exactive Orbitrap MS/MS technology was used to analyze the chemical components of SJF and attribute each component to its source medicinal materials. Network pharmacology and molecular docking techniques were utilized to screen the core targets and active components of SJF against AS, followed by molecular docking verification. ApoE-/- mouse AS model was established using a combination of a high-fat diet and an intermittent water environment (sleep deprivation) model and rosuvastatin gavage was used as the positive control. ELISA was used to detect the serum concentrations of blood lipids (TC, TG, LDL-C, HDL-C), inflammatory cytokines (hs-CRP, IL-1β, IL-6, IL-18, NF-κB, MCP-1), and other relevant markers (Hb, TMAO, Orexin A); HE staining and Oil Red O staining were performed to observe cell morphology and aortic plaque area; Western blotting was used to detect the expression levels of IL-1β, IL-18, NF-κB p65, p-NF-κB p65, NLRP3, Caspase-1, cleaved Caspase-1 (p20), and ASC proteins in the aorta.

RESULTS: A total of 313 chemical components were identified from SJF, including terpenoids, flavonoids, anthraquinones, etc. On this basis, network pharmacology combined with molecular docking technology was used for screening, and the active components of SJF for treating AS were found to be luteolin, quercetin, kaempferol, isorhamnetin, β-sitosterol, and arachidonic acid, with key therapeutic targets including IL-1β, MMP9, IL-6, CXCL8, etc. HE staining and Oil Red O staining showed that compared with the model group, the aortic intimal thickness of the administration group was thinner and atherosclerotic plaques were reduced. In addition, SJF could reduce blood lipid levels, down-regulate the expression of inflammatory factors, and increase hemoglobin concentration to improve the hypoxic microenvironment of AS lesions, thereby alleviating vascular endothelial injury, inhibiting macrophage foam cell formation, and plaque lipid deposition. Mechanistic studies suggested that SJF may suppress atherosclerotic plaque progression, possibly through inhibition of the NF-κB/NLRP3 signaling pathway, which could contribute to its anti-AS effects.

CONCLUSIONS: SJF improves AS-related inflammatory responses by inhibiting the activation of the NF-κB/NLRP3 signaling pathway, reduces abnormal blood lipid levels, accelerates lipid metabolism, decreases lipid deposition, inhibits the development of atherosclerotic plaques, and exerts anti-AS effects.