酪醇诱导肝癌细胞NQO_1酶基因表达增加及细胞增殖的抑制

Enhancement of NADPH: quinone oxidoreductase 1 (NQO_1) gene expression and inhibition of cell proliferation induced by β-tyrosol in human hepatoma SMMC-7721 cells

目的 :研究酪醇诱导肝癌细胞Ⅱ相脱毒酶NAD(P)H :醌氧化还原酶 - 1 (NQO1 )基因表达情况和对细胞增殖的影响以及两者之间的关系。方法 :肝癌细胞SMMC - 772 1接种后 2 4h经 β -酪醇处理 2 4h ,分别测定NQO1酶活性 ,mRNA表达和细胞增殖情况。NQO1 酶活性采用微孔板直接测定法 ,诱导结果用NQO1 酶比活性 =NQO1 酶活性 /细胞数 ;mRNA水平的变化采用定量RT -PCR ;细胞增殖采用结晶紫显色法。结果 :NQO1 酶活性诱导上 ,酪醇大于60mg/L时有明显的剂量效应关系 ,且每一浓度点 (60mg/L ,70mg/L ,80mg/L ,90mg/L)与空白组比较均有显著差异 (P <0 0 5) ,80mg/L的酪醇与 80 μmol/L的 β -NF(阳性对照 )诱导的酶比活性相当 ;mRNA表达量存在剂量依赖性增加 (r=0 82 4 ,P <0 0 5) ,且与酶比活性存在明显的相关性 (r=0 .951 ,P <0 0 1 ) ;酪醇在 70mg/L - 1 0 0mg/L范围内 ,其抑制细胞增殖的能力随着浓度的增加而增加。另外 ,细胞增殖与酶比活性呈负相关 (r =- 0 41 0 ,P <0 0 1 )。结论 :酪醇在培养肝癌细胞SMMC - 772 1上能使NQO1 酶活性与mRNA表达量诱导性增加 ,同时酪醇能抑制细胞的增殖 。

AIM: To investigate whether β-tyrosol could induce expression of DT-diaphorase (NQO 1) gene and inhibit proliferation of hepatoma cells and their relationship.METHODS: The DT-diaphorase activity, mRNA expression and cell proliferation were examined using direct measurement of DT-diaphorase from cells cultured in microtiter wells, semi-quantitative reverse transcription-PCR (RT-PCR) technique and crystal violet assay after treatment of SMMC-7721 cells with β-tyrosol for 24 h. RESULTS: Treatment of SMMC-7721 cells with β-tyrosol caused an increase in DT-diaphorase activity and DT-diaphorase mRNA expression in a dose-dependent manner. A statistically significant correlation between DT-diaphorase mRNA levels and enzyme activities was evident. In almost the same concentration range of 70 mg/L to 100 mg/L, β-tyrosol resulted in a dose-dependent inhibition of cell proliferation which well inversely correlated with DT-diaphorase activity.CONCLUSION: β-tyrosol can induce DT-diaphorase activity and enhance expression of its gene, which may contribute to antiproliferation of hepatoma cells caused by the same chemical.