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目的研究染料木黄酮(genistein,Gen)对脂多糖(LPS)诱导的巨噬细胞炎症因子产生和腺苷酸激活蛋白激酶(AMPK)磷酸化的影响。方法体外培养RAW 264.7小鼠单核巨噬细胞,加入1、5、10、50、100 mol/L的Gen共同培养,MTT法检测其对细胞活性的影响。将RAW264.7细胞随机分为4组:空白对照组不加任何药物,模型组加入终浓度为1 g/ml的LPS进行刺激,Gen高、低剂量组分别加入终浓度为10、5 mol/L的Gen预处理1h后,再加入终浓度为1 g/ml的LPS刺激,24h后收取细胞上清用酶联免疫(ELISA)方法检测TNF-α、IL-6含量,Westernblot检测AMPKα蛋白及其磷酸化的表达水平。结果 100 mol/L的Gen对体外培养RAW264.7细胞的增殖有影响,而其他浓度则无。LPS处理的RAW 264.7细胞与对照组比较,TNF-α、IL-6生成显著增多(P<0.01)、AMPKα磷酸化水平显著降低(P<0.01),而AMPKα总蛋白表达水平无差异(P>0.05)。Gen干预可减少LPS诱导的TNF-α、IL-6生成,上调AMPKα磷酸化水平的表达,与LPS组相比,差异均有统计学意义(P<0.01)。结论 Gen能抑制LPS诱导巨噬细胞的炎症反应,减少TNF-α、IL-6生成,这可能与Gen促进AMPKα磷酸化,从而激活AMPKα有关。
Objective To investigate the effects of genistein on the production of lipopolysaccharide (LPS) -induced macrophage inflammatory factor production and phosphorylation of adenylate activated protein kinase (AMPK). Methods RAW 264.7 mouse monocyte-macrophage cells were cultured in vitro and cultured in Genistein (1, 5, 10, 50 and 100 mol / L). The cell viability was measured by MTT assay. RAW264.7 cells were randomly divided into 4 groups: blank control group without any drug, the model group was added to a final concentration of 1 g / ml of LPS stimulation, Gen high and low dose groups were added to a final concentration of 10,5 mol / L for 1 h and then added LPS at a final concentration of 1 g / ml. After 24 h, the cell supernatants were harvested for enzyme-linked immunosorbent assay (ELISA) to detect the contents of TNF-α and IL-6. Its phosphorylation level of expression. Results The proliferation of RAW264.7 cells cultured in vitro with 100 mol / L Gen had an effect, but no other concentrations. Compared with the control group, the production of TNF-α and IL-6 in LPS-treated RAW 264.7 cells was significantly increased (P <0.01), while the phosphorylation of AMPKα was significantly decreased (P <0.01) 0.05). Gen interference could decrease the production of TNF-α and IL-6 induced by LPS, and up-regulate the phosphorylation of AMPKα. Compared with LPS group, the difference was statistically significant (P <0.01). Conclusion Gen inhibits the inflammatory response induced by LPS in macrophages and decreases the production of TNF-α and IL-6, which may be related to the effects of Gen on AMPKα phosphorylation and AMPKα activation.