盐酸千金藤碱对HepG2.2.15细胞HBX/NF-κB通路的影响

Effect of cepharanthine hydrochloride on HBX/NF-κB signal pathway of HepG2.2.15 cells

目的探讨盐酸千金藤碱(cepharanthine hydrochloride,CH)对Hep G2.2.15细胞HBX/NF-κB信号通路的影响。方法选用不同浓度的CH(2.5,5,10μmol/L)干预稳定表达野生型乙型肝炎病毒(hepatitis B virus,HBV)的HepG2.2.15细胞后,采用RT-PCR和ELISA法分别检测细胞内X基因mRNA和X蛋白表达变化;采用AnnexinⅤ-FITC/PI法检测细胞的凋亡率,Western blot法检测细胞内NF-κB信号通路成员P65和IκBα蛋白表达。结果与未经CH处理的HepG2.2.15细胞(对照组)相比,CH各剂量组明显抑制细胞内X基因mRNA和X蛋白的表达(P<0.05)。CH干预细胞48 h能够浓度依赖性地诱导Hep G2.2.15细胞凋亡,2.5,5,10μmol/L CH处理细胞后,HepG2.2.15细胞平均凋亡率分别为(11.9±1.21)%,(19.8±2.32)%,(29.3±1.27)%。CH处理组细胞内IκBα蛋白的表达水平明显高于对照组,且P65蛋白的表达水平明显下调(P均<0.05)。结论 CH能够抑制HBX/NF-κB信号通路的活性,并具有促进HepG2.2.15细胞凋亡的作用。

Objective To investigate the effect of cepharanthine hydrochloride（ CH） on HBX/NF-κB signal pathway of HepG2. 2. 15 cells stably expressing wide-type HBV. Methods HepG2. 2. 15 cells were treated with different concentration of CH（ 2. 5,5,10μmol/L）. Then the expression levels of X gene and X protein of HBV were detected by reverse thanscriptase polymerase chain reaction（RT-PCR） and enzyme-linked immunosorbent assay（ELISA）,respectively. Flow cytometry method was used to detect the cell apoptosis rates. The expression levels of IκBα and P65 protein in HepG2. 2.15 cells were detected by Western blot. Results Compared with HepG2. 2. 15 cells without CH treament（ control group）,the expression of X gene mRNA and X protein in HepG2. 2. 15 cells after treatment with CH（2. 5,5,10 μmol/L） were significantly inhibited（ P 〈 0. 05）. The apoptosis retes of HepG2. 2. 15 cells were（11. 9± 1. 21） %,（19. 8 ± 2. 32） %,（29. 3 ± 1. 27） % after treatment with 2. 5,5,10 μmol/L CH for 48 h,respectively. CH increased apoptosis rates of HepG2. 2. 15 cells in a concentration-dependent manner. CH effectively up-regulated the protein expression of IκBα and down-regulated the expression of P65 protein（ both P 〈 0. 05）. Conclusion CH can inhibit the activity of HBx/NF-κB signal pathway and increase cell apoptosis of HepG2. 2. 15 cells.