壮药三七姜醇提物及含药血清对脂多糖诱导RAW264.7细胞炎症的抗炎作用及机制

Anti-inflammatory effect and mechanism of the ethanol extract and the drug-containing serum of Zhuang medicine Stahlianthus involucratus on LPS-induced RAW264.7 cell inflammation

目的研究壮药三七姜醇提物及含药血清对脂多糖(LPS)诱导RAW264.7细胞炎症的抗炎作用及机制。方法将三七姜醇提物(75.35 g/kg)或纯净水灌胃大鼠以制备含药血清或空白血清。以RAW264.7细胞为研究对象,将细胞分为正常对照组,LPS组(1μg/mL),三七姜醇提物高、中、低剂量组(50、25、12.5μg/mL),4%或15%空白血清组,4%或15%空白血清+LPS组,4%或15%含药血清组,4%或15%含药血清+LPS组,按相应条件培养24 h后,检测各组细胞活力和细胞中一氧化氮(NO)、肿瘤坏死因子α(TNF-α)、白细胞介素1β(IL-1β)、IL-6含量,以及Toll样受体4(TLR4)、核因子κB(NF-κB)mRNA表达水平,一氧化氮合酶(NOS)、环氧化酶2(COX-2)蛋白表达水平。结果各组细胞培养24 h后,细胞活力差异无统计学意义。与正常对照组比较,LPS组细胞中NO、TNF-α、IL-1β、IL-6含量,TLR4、NF-κB mRNA表达水平和NOS、COX-2蛋白表达水平均显著升高(P<0.05)。与4%或15%空白血清组比较,4%或15%空白血清+LPS组细胞中上述指标水平显著升高(P<0.05)。与LPS组比较,三七姜醇提物各剂量组大鼠上述指标水平均显著降低(P<0.05)。与4%或15%空白血清+LPS组比较,4%或15%含药血清+LPS组细胞中上述指标水平均显著降低(P<0.05)。结论三七姜醇提物及含药血清均可明显减轻LPS诱导的细胞炎症反应,其抗炎机制可能与抑制TLR4/NF-κB信号通路活性,下调COX-2、NOS蛋白表达,减少炎症因子释放有关。

OBJECTIVE To study the anti-inflammatory effect and mechanism of the ethanol extract and the drug-containing serum of Zhuang medicine Stahlianthus involucratus on lipopolysaccharide(LPS)-induced RAW264.7 cell inflammation.METHODS The drug-containing serum or blank serum was obtained by intragastrical administration of ethanol extract of S.involucratus(75.35 g/kg)or purified water.Using RAW264.7 cells as objects,RAW264.7 cells were divided into normal control group,LPS group(1μg/mL),S.involucratus ethanol extract high-dose,medium-dose and low-dose groups(50,25,12.5μg/mL),4%or 15%blank serum groups,4%or 15%blank serum+LPS groups,4%or 15%drug-containing serum groups,4%or 15%drug-containing serum+LPS groups.After culturing for 24 h,cell viability,the contents of nitric oxide(NO),tumor necrosis factorα(TNF-α),interleukin 1β(IL-1β)and IL-6 as well as mRNA expressions of Toll-like receptor 4(TLR4)and nuclear factorκB(NF-κB)and protein expressions of nitric oxide synthase(NOS)and cyclooxygenase 2(COX-2)were all detected in each group.RESULTS After culturing for 24 h,there was no statistical significance in the difference of cell viability.Compared with normal control group,the contents of NO,TNF-α,IL-1βand IL-6,mRNA expressions of TLR4 and NF-κB,and protein expressions of NOS and COX-2 were increased significantly in LPS group(P<0.05).Compared with 4%or 15%blank serum groups,the levels of above indexes were increased significantly in 4%or 15%blank serum+LPS groups(P<0.05).Compared with LPS group,the levels of above indexes were decreased significantly in S.involucratus ethanol extract groups(P<0.05).Compared with 4%or 15%blank serum+LPS groups,the levels of above indexes were decreased significantly in 4%or 15%drug-containing serum+LPS groups(P<0.05).CONCLUSIONS The ethanol extract and the drug-containing serum of S.involucratus can significantly alleviate LPS-induced inflammatory reaction,the mechanism of which may be associated with inhibiting the activity of TLR4/NF-κB signaling pathway,down-regulating the protein expressions of COX-2 and NOS,and reducing the release of inflammatory factors.