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研究山柰酚对脂多糖(lipopolysaccharide,LPS)诱导的HMC-1肥大细胞炎症反应的抑制作用。利用MTT法检测山柰酚对HMC-1细胞的细胞毒性作用,同时确定山柰酚的给药浓度;ELISA检测活化的HMC-1细胞经不同浓度的山柰酚(10、20和40μmol·L-1)作用后组胺、IL-6、IL-8、IL-1β和TNF-α的释放量;Western blot检测经不同浓度的山柰酚作用于活化的HMC-1细胞后,胞质中p-IKKβ、IκBα、p-IκBα和胞核中NF-κB(p65)蛋白水平。根据MTT法的实验结果,确定了山柰酚的给药浓度范围为5~40μmol·L-1。山柰酚能够使活化的HMC-1肥大细胞中组胺、IL-6、IL-8、IL-1β和TNF-α的释放量显著下降(P<0.01)。山柰酚作用于LPS诱导的HMC-1细胞后,p-IKKβ、p-IκBα和胞核中NF-κB(p65)蛋白水平会显著下降(P<0.01)。结果表明,山柰酚对于肥大细胞炎症反应具有显著的抑制效应,它能够抑制IKKβ的活化,抑制IκBα的磷酸化,阻止NF-κB(p65)进入细胞核内,进而影响相关炎症介质的释放。
To investigate the inhibitory effect of kaempferol on the inflammatory response of HMC-1 mast cells induced by lipopolysaccharide (LPS). MTT assay was used to determine the cytotoxicity of kaempferol to HMC-1 cells and the concentration of kaempferol was determined. The activity of kaempferol was detected by ELISA with different concentrations of kaempferol (10, 20 and 40 μmol·L -1), and the release of histamine, IL-6, IL-8, IL-1β and TNF-α after treated with different concentrations of kaempferol were detected by Western blot. p-IKKβ, IκBα, p-IκBα and nuclear NF-κB (p65) protein levels. According to the results of MTT assay, the concentration of kaempferol was determined to be in the range of 5 ~ 40μmol·L-1. The kaempferol significantly decreased the release of histamine, IL-6, IL-8, IL-1β and TNF-α in activated HMC-1 mast cells (P <0.01). The expression of p-IKKβ, p-IκBαand NF-κB (p65) in the cells of LPS-induced HMC-1 cells were significantly decreased by kaempferol (P <0.01). The results showed that kaempferol had a significant inhibitory effect on the inflammatory response of mast cells. It inhibited the activation of IKKβ, inhibited the phosphorylation of IκBα, and prevented the entry of NF-κB (p65) into the nucleus, thereby affecting the release of relevant inflammatory mediators.