川芎嗪可影响肿瘤坏死因子α刺激肝星状细胞结缔组织生长因子、核因子κB及相关基因产物的表达

背景：川芎嗪延缓肝纤维化形成的机制尚不清楚。目的：观察川芎嗪对肿瘤坏死因子α刺激的肝星状细胞增殖及结缔组织生长因子、核因子κB及其相关基因产物白细胞介素6表达的影响。方法：体外培养HSC-T6细胞株,取对数生长期的细胞用于实验。实验分为4组：对照组仅加入细胞;TNF-α组加入10μg/L TNF-α;川芎嗪干预组先加入终浓度为10μg/L的TNF-α作用30min后,分别加入川芎嗪50,100,200,400,600,1000mg/L;PDTC组先加入终浓度为10μg/L的TNF-α作用30min后,再加入终浓度18μmol/L的核因子κB阻断剂PDTC。结果与结论：MTT结果显示100,200,400,600,1000mg/L的川芎嗪均能抑制HSC-T6增殖,且呈剂量依赖性。免疫细胞化学染色及Western blot检测发现10μg/L的肿瘤坏死因子α刺激后,HSC-T6细胞结缔组织生长因子、核因子κB及白细胞介素6的表达明显增多（P〈0.01或P〈0.05）,200,400,600mg/L的川芎嗪及18μmol/L的PDTC均可明显降低肿瘤坏死因子α刺激后HSC-T6细胞结缔组织生长因子、核因子κB及白细胞介素6的表达（P〈0.01）,且随着川芎嗪质量浓度的增加,抑制作用增强,PDTC的抑制作用最明显。相关性分析结果显示HSC-T6细胞结缔组织生长因子和核因子κB的表达呈正相关（r=0.980,P〈0.01）。说明川芎嗪可以抑制肝星状细胞结缔组织生长因子、核因子κB及白细胞介素6的表达,抑制肝星状细胞的增殖。

Effects of tetramethylpyrazine on expression of connective tissue growth factor,nuclear factor kappa B and its related gene products in tumor necrosis factor-alpha stimulated hepatic stellate cells

BACKGROUND：Previous studies have shown that tetramethylpyrazine（TMP） can decrease expression of connective tissue growth factor（CTGF） ,nuclear factor kappa B（NF-κB） and delay the formation of hepatic fibrosis.But the mechanism and the correlation of CTGF and NF-κB was not clear.OBJECTIVE：To investigate the effects of TMP on expression of CTGF,NF-κB and its related gene product interleukin-6（IL-6） in tumor necrosis factor-alpha（TNF-α） stimulated hepatic stellate cells.METHODS：Hepatic stellate cell-T6（HSC-T6） was cultured to logarithmic growth phase for experiments.Cells were divided into four groups：control group：only cells were added;TNF-α group：10 μg/L TNF-α was added;TMP intervention group：pretreated with 10 μg/L TNF-α for 30 minutes and then 50,100,200,400,600,1 000 mg/L TMP was added;PDTC group：pretreated with 10 μg/L TNF-α for 30 minutes,and then stimulated with 18 μmol/L PDTC,a blocker of NF-κB.RESULTS AND CONCLUSIONS：MTT assay results showed that TMP inhibited the proliferation of HSC-T6 at a concentration of 100,200,400,600,and 1 000 mg/L in a concentration dependent manner（P 0.05） .Immunocytochemical staining and western blot analysis showed that 200,400,600 mg/L TMP and 18 μmol/L PDTC could decrease the expression of TNF-α-induced CTGF,NF-κB and IL-6（P〈0.01） in HSC-T6 cells and with increasing concentrations of TMP,the inhibitory effects were enhanced,and PDTC could inhibit their expression effectively.Correlation analysis showed that the expression of cTGF and NF-κB in HSC-T6 cells was positively correlated（r=0.980,P〈0.01） .These data indicated that TMP could inhibit the expression of cTGF,NF-κB and IL-6 in hepatic stellate cells and inhibit the proliferation of hepatic stellate cells.