阿司匹林丁香酚酯对体外脂多糖诱导小鼠巨噬细胞炎症反应的抑制效应

doi:10.11843/j.issn.0366-6964.2022.11.030

Abstract

Abstract: The purpose of this study was to investigate the inhibitory effects and potential mechanism of aspirin eugenol ester (AEE) on lipopolysaccharide (LPS)-induced inflammatory response in mouse macrophage (RAW264.7 cells). Inflammation in RAW264.7 cells was induced by LPS. Cells were divided into control group, LPS group, aspirin group (Asp, 150 μmol·L^-1), eugenol group (Eug, 150 μmol·L^-1), AEE low-(75 μmol·L^-1), medium-(150 μmol·L^-1) and high-dose (300 μmol·L^-1) groups. CCK-8 assay was used to detect the cytotoxicity of AEE in RAW264.7 cells. The levels of IL-1β, IL-6, IL-8 and TNF-α in each group were detected by ELISA. The mRNA expression levels of COX-2, COX-1, PLA2, CYP450 and 5-LOX, the key enzymes in arachidonic acid(AA) metabolism pathway, were detected by RT-PCR. Meanwhile, the interaction of Asp, Eug and AEE with key enzymes related to arachidonic acid metabolism pathway was studied by molecular docking. The results were showed as follows:1) CCK-8 results indicated that AEE with the concentration of 0 to 350 μmol·L^-1 was non-toxic to cells; 2) The results of ELISA showed that, compared with the control group, the expression levels of IL-1β, IL-6, IL-8 and TNF-α in the LPS group were extremely significantly increased (P<0.01); After treated with different doses of AEE extremely significantly reduced the expression levels of IL-1β, IL-8, and TNF-α (P<0.01); Compared with the Asp and Eug groups, no significant difference was observed for the equimolar AEE in inflammatory factors of IL-1β, IL-6, IL-8 (P>0.05), indicating the similar anti-inflammatory effects of AEE with Asp and Eug; And no significant difference of the levels of IL-1β, IL-6 and IL-8 among different doses of AEE was observed(P>0.05); 3) RT-PCR results showed that, compared with the control group, the expression of key enzymes in the AA metabolic pathway in the LPS group was extremely significantly increased (P<0.01); After treated with different doses of AEE significantly reduced the mRNA expression of key enzymes in the AA metabolic pathway (P<0.05); Compared with the Asp and Eug groups, the equimolar amount of AEE significantly reduced the expression of PLA2 (P<0.05); There was no significant difference of the expression levels of COX-1, COX-2, 5-LOX and CYP450 was observed among different doses of AEE (P>0.05); 4) The results of molecular docking showed that the binding energy of AEE to the target proteins was lower than -20.9 kJ, indicating the binding of AEE to target proteins was stable and high quality. Meanwhile, AEE could form hydrogen bonds with PLA2, COX-2, COX-1, CYP450 and 5-LOX. In summary, AEE could inhibit the inflammatory response of RAW264.7 cells induced by LPS, and the anti-inflammatory effects of AEE were similar to the precursor compounds Asp and Eug, which might be associated with the regulation of AA metabolic pathway.

Key words: aspirin eugenol ester, RAW264.7 cells, inflammation, molecular docking

CLC Number:

S859.791

LIU Xu, PAN Yinchuan, YANG Yajun, LIU Xiwang, MA Ning, LI Jianyong. Inhibiting Effect of Aspirin Eugenol Ester on Lipopolysaccharide-induced Inflammatory Response in Mouse Macrophages in vitro[J]. Acta Veterinaria et Zootechnica Sinica, 2022, 53(11): 4048-4057.

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Inhibiting Effect of Aspirin Eugenol Ester on Lipopolysaccharide-induced Inflammatory Response in Mouse Macrophages in vitro

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