姜黄素对脂多糖诱导大鼠肝脏Kupffer细胞NF-κB及下游炎症因子的影响

The effect of curcumin on expression of NF-κB and TNF-α,IL-1β and IL-6 in rat liver Kupffer cells induced by lipopolysaccharides

目的研究姜黄素对脂多糖(LPS)诱导大鼠肝脏Kupffer细胞NF-κB活化的影响。方法将分离的Kupffer细胞按1×105/ml接种于24孔板内培养48 h后,对照组:常规培养;LPS组:给予终浓度为100 ng/ml的LPS培养8 h;姜黄素组:给予25μmol/L姜黄素干预2 h后,其余处理同LPS组。细胞免疫荧光技术检测NF-κB核转位水平,Western blotting检测胞浆IκBα、p-IκBα及胞核NF-κB p65蛋白表达水平,ELISA法检测上清TNF-α、IL-1β和IL-6的水平。结果①LPS能明显诱导NF-κB核转位,姜黄素能明显逆转这一效应。②对照组胞浆蛋白IκBα表达显著高于LPS组和姜黄素组(P<0.05),但LPS组明显低于姜黄素组(P<0.05);LPS组胞核NF-κB p65和p-IκBα表达显著高于对照组和姜黄素组(P<0.05),但姜黄素组表达显著高于对照组(P<0.05)。③LPS组TNF-α、IL-1β和IL-6表达显著高于对照组(P<0.05),姜黄素组TNF-α、IL-1β和IL-6显著低于LPS组(P<0.05)。结论姜黄素对LPS诱导的Kupffer细胞炎症因子分泌有明显的抑制作用。

Objective To investigate the effect of curcumin on NF-KB activation in rat liver Kupffer cells （KCs） in- duced by lipopolysaccharides （LPS）. Methods The isolated KCs were inoculated into culture plate with the density of 1 ×105/ml, then were cultured for 48 hours. KCs were divided into control group, LPS group and curcumin group. KCs were normally cultured in control group, while KCs treated with 100 ng/ml LPS for 8 hours in LPS group, and KCs trea- ted with 25 μmol/L curcumin for 2 hours and subsequently treated with 100 ng/ml LPS in curcumin group. The nuclear factor KB （NF-KB） in nuclear and cytosolic fraction was assayed by immunofluorescence staining. Nuclear extracts were prepared and analyzed by Western blotting with NF-KB p65. Cytosolic extracts were analyzed by Western blotting with anti-IKBoc and phosphorylated- IKBcα antibody. The expression of TNF-oc, IL-1β and IL-6 in the culture supernatant were assayed by ELISA. Results ① Compared with control group, LPS induced the activation of NF-KB markedly, which could be inhibited by curcumin markedly. ② LPS significantly facilitated the phosphorylation of IKBcα and curcu- min obviously inhibited the process （P 〈 O. 05）. The level of nuclear NF-KBp65 expression in LPS group was signifi- cantly higher than that in control group （P 〈 0.05） , the nuclear NF-KBp65 expression decreased significantly in curcu- min group than that in LPS group （P 〈 O. 05） , but the level was higher significantly than that in control group （P 〈 0.05 ）. ③ Compared with control group, LPS stimulation increased significantly the levels of TNF-α, IL-1β and IL-6 in LPS group （P 〈 O. 05 ） , and the levels of TNF-α, IL-1β and IL-6 significantly decreased after curcumin administration （ P 〈 O. 05 ）. Conclusion Curcumin can obviously inhibit LPS-induced the expression of inflammatory cytokines signifi- cantly in KCs.