木犀草素对肥大细胞脱颗粒影响及机制的研究

摘要

目的：探讨木犀草素（ Luteolin）的抗I型变态反应的作用机制。方法：通过建立DNP－BSA－IgE激发致敏的大鼠RBL－2H3细胞模型，分别采用MTT法检测不同浓度（5、15、25μmol／L）Luteolin对RBL－2H3肥大细胞活性的影响；ELISA法检测不同浓度Luteolin对RBL－2H3细胞分泌β－hexosa minidase（β－HEX）及细胞因子TNF－α影响；Flou－4AM荧光探针检测细胞内Ca2＋浓度变化；Western blot检测AKT，P－AKT的表达。结果：成功建立致敏的细胞模型，低浓度的Luteolin对RBL－2H3细胞活性无明显影响；不同浓度Luteolin刺激RBL－2H3细胞后，对β－HEX和TNF－α的释放抑制作用呈线性相关，且细胞内Ca2＋明显减少；Western blot结果显示随着Luteolin浓度的增加AKT磷酸化水平明显下降。结论：Luteolin呈剂量依赖性抑制RBL－2H3肥大细胞脱颗粒，且通过调节细胞内Ca2＋浓度与AKT活性参与其中。%Objective:To investigate the effect of Luteolin on the degranulation function of RBL-2H3 mast cells and its mechanism of action.Methods:DNP-BSA-IgE was used to establish the sensitized cell model in RBL-2H3 cells.Cell viability was exa mined by MTT assay at different concentrations (5,15,25 μmol/L) of Luteolin pretreated RBL-2H3 cells.The effect of Luteolin on β-hexosa minidase(β-hex) and TNF-αwas evaluated by ELISA.The change of Ca2+influx was detected by Flou-4AM calcium ion fluorescent probe.The expression of AKT and P-AKT were detected by Western blot.Results:The sensitized cell model was established successfully .Cell viability had not significantly changes stimulated by different concentrations of Luteolin ,and a significantly inhibition role showed the release of β-hex and TNF-αfrom RBL-2H3 cells.The concentration of intracellular Ca 2+ was significantly decreased.However,the Western blot results showed that the level of Phosphorylation AKT was decreased .Conclusion: Luteolin suppress RBL-2H3 cell degranulation in dose dependence ,which may via impacting the Ca2+influx and the activity of AKT.