鳖甲煎丸对大鼠肝星状细胞中NF-κB信号通路的影响

Effects of Biejiajian Pills on NF-κB signaling pathway in hepatic stellate cells in rats

目的研究鳖甲煎丸(鳖甲胶、阿胶、蜂房等)对大鼠肝星状细胞(HSC-T6)中核转录因子-κB(NF-κB)信号通路中NF-κB、p65、p50、IκB及靶基因表达的影响。方法使用鳖甲煎丸药物血清培养HSC-T6细胞24 h后,采用q PCR检测p65、p50、血管内皮生长因子(VEGF)、基质金属蛋白酶抑制因子-1(TIMP-1)m RNA的表达,免疫荧光法检测p65的表达,Western blot检测NF-κB抑制蛋白α(IκBα)、NF-κB抑制蛋白β(IκBβ)及α平滑肌肌动蛋白(α-SMA)的表达。结果与空白对照组及阴性对照组相比,鳖甲煎丸的中、高剂量组及阳性对照组p65、VEGF、TIMP-1 m RNA的表达显著降低,各组间p50 m RNA的表达无显著差异,但免疫荧光显示p65在细胞质中的表达降低。同时,鳖甲煎丸能显著上调IκBα的蛋白表达,并能明显下调α-SMA的表达,具有剂量依赖性,但对IκBβ的表达无显著影响。结论鳖甲煎丸对肝纤维化的治疗作用可能与影响NF-κB信号通路、抑制下游靶基因的表达有关。

AIM To study the effects of Biejiajian Pills （Colla Carapacis Trionycis, Asini Corii Colla, Nidus Vespae, etc. ） on NF-κB, p65, p50 and IκB in NF-κB signaling pathway and target gene expression in HSC-T6 cells of rats. METHODS HSC-T6 cells were cultured with Biejiajian Pills drug serum for 24 hours, the expres- sions of p65, p50, VEGF and TIMP-1 mRNA were determined by qPCR; the expression of p65 was measured by immunofluorescence; the expressions of IκBα, IκBβ and α-SMA were determined by Western blot. RESULTS The Biejiajian Pills middle-, high-dose and positive control groups showed significantly lower expressions of p65, VEGF and TIMP-1 mRNA as compared with the blank control group and negative control group, the expressions ofp50 mRNA among various groups showed no significant differences. But immunofluoreseence showed that the ex- pression of p65 in cytoplasm was decreased. Meanwhile, Biejiajian Pills showed significantly higher IκBα protein expression and obvious down-regulation of α-SMA expression in a dose-dependent manner, but had no significant influence on the expression of 1κBβ. CONCLUSION Biejiajian Pills＇ therapeutic effects on hepatic fibrosis may be related to influencing NF-κB signaling pathway and inhibiting the expression of down-stream target gene.