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目的:观察知母皂苷是否能抑制脂多糖引起的小鼠腹腔巨噬细胞炎症因子TNF-α和NO释放并探讨PI3K/Akt/p70S6K信号转导通路的影响。方法:小鼠腹腔巨噬细胞培养24 h,加入不同浓度知母皂苷(1、10和100μmol/L)或加入不同的阻断剂(PI3K特异性阻断剂LY294002、Akt特异性阻断剂TCBN),之后加入脂多糖(10 mg/L)继续培养。ELISA法和Griess法分别测定巨噬细胞培养上清液中TNF-α和NO的含量;免疫化学荧光染色法观察巨噬细胞磷酸化p70S6K表达水平的改变。结果:加入脂多糖作用2 h巨噬细胞上清液中TNF-α开始含量明显增加,24 h达高峰。知母皂苷(1、10和100μmol/L)、LY294002(20μmol/L)和TCBN(10μmol/L)均可不同程度的抑制脂多糖引起的巨噬细胞TNF-和NO产生增加。知母皂苷(100μmol/L)可减少脂多糖引起的小鼠腹腔巨噬细胞磷酸化p70S6K表达增加。结论:知母皂苷显著抑制脂多糖引起的巨噬细胞炎症因子TNF-α和NO释放,其作用机制与知母皂苷下调PI3K/Akt/p70S6K信号转导通路表达有关。
OBJECTIVE: To observe whether timosaponin could inhibit the release of TNF-α and NO from mouse peritoneal macrophages induced by lipopolysaccharide and to explore the effect of PI3K / Akt / p70S6K signal transduction pathway. METHODS: Mouse peritoneal macrophages were cultured for 24 h. Different concentrations of timosaponins (1, 10 and 100 μmol / L) or different blockers (PI3K-specific blockers LY294002, Akt-specific blockers TCBN ), Then added lipopolysaccharide (10 mg / L) to continue the culture. The levels of TNF-α and NO in macrophage culture supernatants were measured by ELISA and Griess respectively. The expression of phosphorylated p70S6K in macrophages was detected by immunochemical staining. Results: The content of TNF-α in supernatant of macrophages treated with lipopolysaccharide increased significantly at 24 h and peaked at 24 h. Tiama saponin (1, 10 and 100 μmol / L), LY294002 (20 μmol / L) and TCBN (10 μmol / L) all inhibited the increase of TNF- and NO production by LPS in different degree. Timosaponins (100μmol / L) can reduce lipopolysaccharide-induced mouse peritoneal macrophages phosphorylation of p70S6K increased expression. Conclusion: Tim saponin significantly inhibits the release of TNF-α and NO from macrophages induced by lipopolysaccharide, and its mechanism is related to the down-regulation of PI3K / Akt / p70S6K signal transduction pathway by timosaponins.