猕猴桃水提物诱导二相酶产生及抗氧化的机制研究

Study on the Antioxidant Mechanism of Actinidia chinensis Aqueous Extract via Inducing the Produce of Phase 2 Enzymes

目的:本实验以人胃腺癌细胞株SGC-7901为模型,用猕猴桃水提物含药血清进行干预,探讨猕猴桃水提物诱导二相酶产生及抗氧化的机制。方法:取正常SD大鼠随机分为猕猴桃水提物高、中、低剂量组(用猕猴桃水提物溶液46.8,31.2,15.6 g.kg-1ig)和正常对照组(ig给予生理盐水)。连续14 d,每天ig 2次,于末次给药1 h,腹主动脉取血制备猕猴桃水提物含药血清。取对数生长期的SGC-7901细胞,在不同时间段分别给予相应的含药血清进行干预。用免疫组化法检测SGC-7901细胞内活化状态的核因子相关因子2(Nrf2)的细胞定位;RT-PCR法检测猕猴桃水提物高剂量组含药血清对SGC-7901细胞内二相酶mRNA水平的影响。结果:免疫组化法染色观察结果显示与正常对照组相比,猕猴桃水提物含药血清低、中、高剂量组均可提高Nrf2细胞核内的表达水平,颜色从浅黄色变为棕黄色。经积分统计学分析差异有显著性(P<0.05或P<0.01)。RT-PCR检测结果显示与正常对照组相比较,猕猴桃水提物可明显诱导二相酶谷氨酰半胱氨酸合成酶催化亚单位(GCLC)基因的表达(P<0.05),且诱导谷胱甘肽-S-转移酶(GST)-P1和GST-T1基因表达作用差异有高度显著性(P<0.01),但对二相酶GST-M1和醌氧化还原酶(NQO1)基因水平影响不大。结论:猕猴桃水提物可促使Nrf2进入核内,导致二相酶的表达增加,具有诱导细胞解毒抗氧化作用。

Objective： In this study, Human gastric cancer cell line SGC-7901 cells were used as a model and treated with the aqueous extract of actinidia chinensis medicated sera, to investigate the antioxidant mechanism via inducing the produce of phase 2 enzymes. Method ： The normal SD rats were randomly divided into 4 groups： blank control group, high, middle and low dose groups. The actinidia chinensis aqueous extract was gavaged in different doses, respectively and water was gavaged in the blank control group and twice a day for 14 days. The blood was collected from abdominal aorta to prepare serum containing drugs, then which were given to the SGC-7901 ceils separately at different time. Immunohistochemistry staining was used to detect the localization of nuclear factor-E2 related factor2 （Nrf2） in the SGC-7901cells, RT-PCR was used to detect the mRNA level of phase 2 enzymes. Result： The immunohistochemistry detecting showed that the expression of the nuclear Nrf2 was increased and the area broadened significantly （P 〈 0.05） in th groups after administration with different doses of serum containing drugs in the SGC-7901 cells. The results of RT-PCR showed that the mRNA expression levels of the phase 2 enzymes glutamate cysteine ligase catalytic subunit （GCLC） were induced （P 〈 0.05 ）, and the mRNA expression levels of glutathione-S-transferase （GST）-P1 and GST-T1 were enhanced significantly （P 〈 0.01 ） , which was no obvious effect on GST-M1 and heme oxygenase-1 （NQO1）. Conclusion： The Actinidia chinensis aqueous extract can increase nuclear translocation of Nrf2 and increase the phase II enzyme mRNA expression. The Actinidia chinensis aqueous extract has the function of antioxidant.