荔枝核总黄酮对活化大鼠肝星状细胞的增殖抑制作用及TLR4表达的影响

The total flavone of litchi chinensis sonn inhibits the proliferation of actived hepatic stellate cells via the TLR4 pathway in rats

目的研究荔枝核总黄酮(TFL)在体外对活化的大鼠肝星状细胞(HSC-T6)的增殖抑制作用及Toll样受体4(TLR4)表达的影响。方法使用转化生长因子β1(TGF-β1)(5 ng/ml)激活HSC-T6,显微镜下观察HSC-T6和激活的HSC-T6形态变化;CCK8试剂盒(CCK8)法观察TFL作用24、48、72 h后活化的HSC-T6的增殖情况;逆转录聚合酶链反应(RT-PCR)检测HSC-T6中TLR4的表达;流式细胞术检测细胞周期和细胞凋亡。结果显微镜下观察HSC-T6和激活的HSC-T6形态有明显变化;CCK8结果显示TFL可以抑制HSC-T6的增殖(P<0.05),并且随着TFL浓度和作用时间增加,HSC-T6增殖明显下降,呈浓度依赖性;RT-PCR结果显示随着TFL浓度增加,HSC-T6中TLR4表达下降,与实验对照组比较差异有统计学意义(P<0.05);流式细胞仪检测结果表明TFL阻滞细胞于S期,G0/G1期细胞减少(P<0.05),S期细胞增加(P<0.05),促进细胞的凋亡(P<0.05),呈浓度依赖性。结论 TFL可以抑制HSC-T6的增殖,其作用机制可能是通过降低TLR4在HSC-T6中的表达,阻滞其细胞增殖和诱导细胞凋亡,达到抗肝纤维化效果。

Objective To investigate the anti-proliferative effect of total flavone of litchi chinensis sonn （ TFL） on the activated hepatic stellate cells （ HSC-T6 ） in vitro and the influence on Toll-like receptors 4 （ TLR4 ） . Methods HSC-T6 were activated with TGF-β（5 ng/ml） and then divided into the following groups： blank control, experi-mental control, five experimental groups. Different morphological changes of HSC-T6 and HSC-T6 were observed under the microscope. CCK8 was used to observe the proliferation of three kinds of the activated HSC-T6 which had been respectively affected by TFL for 24, 48 and 72 h. The expressions of TRL4 were detected by RT-PCR （ for mRNA） . The cell cycle and cell apoptosis were detected by flow cytometry （ FCM） . Results There were obvious morphological changes of HSC-T6 and HSC-T6 under the microscope. The results of CCK8 showed that TFL could inhibit the proliferation of HSC-T6 （ P〈0. 01 ） , and HSC-T6 decreased significantly with the increase of the con-centration and action time of TFL. It could be found in the results of RT-PCR that expressions of TLR4 of HSC-T6 declined with the increase of TFL concentration. There were visible differences compared with the experimental control （P〈0. 01）. The test of FCM showed that TFL retarded the cells during PeriA S, G0/G1cells reduce （P〈0. 01） and then promoted the apoptosis of cells （P〈0. 01）. So it depended on the concentration of TFL. Conclu-sion TFL can inhibit the proliferation of HSC-T6 , whose mechanism is to inhibit the proliferation of cells and in-duce the cell apoptosis by reducing the expressions of TLR4 in HSC-T6 , so as to have the effect of anti-liver fibro-sis.