小檗碱抑制HCT116细胞在小鼠体内增殖的分子机制研究

Molecular mechanism of berberine inhibiting proliferation of HCT116 cells in mice

目的探究小檗碱抑制人结肠癌HCT116细胞在小鼠体内增殖的分子机制。方法将60只小鼠随机分为空白组、模型组、对照组、给药组(小檗碱低、中及高剂量组)。除空白组以外的各组小鼠接受右前肢皮下注射HCT116细胞悬液100μl,空白组小鼠同部位注射100μl磷酸盐缓冲液。小檗碱低、中及高剂量组分别腹腔注射浓度为1、2和4 mg/ml的小檗碱溶液,给药标准为10 ml/kg,对照组腹腔注射20 mg/kg氟尿嘧啶,空白组和模型组腹腔注射10 ml/kg的生理盐水,各组给药频率均为1次/d。结果小檗碱对结肠癌HCT116细胞有抑制作用,其半数抑制浓度为(0.24±0.02)μmol/L。各组小鼠体重比较,差异有统计学意义(P<0.05)。小檗碱低、中及高剂量组肿瘤长径、短径较模型组短,体积较模型组小,呈剂量依赖性改变,且剂量越大数值越小(P<0.05)。模型组存活率低于空白组(P<0.05)。小檗碱低、中和高剂量组、对照组存活率均高于模型组,且小檗碱给药组呈剂量依赖性改变(P<0.05)。小檗碱低、中及高剂量组肿瘤重量较模型组轻,且小檗碱给药组呈剂量依赖性(P<0.05)。小檗碱低、中及高剂量组均具有较高的抑瘤率,小檗碱高剂量组较小檗碱低、中剂量剂量组高(P<0.05),对照组较小檗碱低、中剂量组高(P<0.05)。小檗碱低、中及高剂量组Caspase-9、Caspase-3 mRNA的相对表达量较模型组高(P<0.05)。小檗碱低、中及高剂量组Cytochrome C、Caspase-9及Caspase-3蛋白相对表达量较模型组高(P<0.05)。结论小檗碱通过破坏线粒体释放Cytochrome C,进而激活凋亡相关蛋白Caspase-9和Caspase-3的表达,诱导肿瘤组织细胞凋亡,发挥抗肿瘤活性。

Objective To explore the molecular mechanism of berberine inhibiting proliferation of HCT116 cells in mice.Methods Sixty mice were randomly divided into blank group,model group,control group,drug group(the drug groups with low,medium and high berberine).The micro syringe was used to absorb the HCT116 cell suspension 100 ul and was subcutaneously injected into the right forelimb of all mice except for blank group.The mice in the blank group were subcutaneously injected with 100 ul PBS,and the treatment drugs were given seven days later.The drug groups with low,medium and high berberine were intraperitoneally injected with concentration of 1,2 and 4 mg/ml berberine saline solution respectively,and the dosage volume standard is 10 ml/kg.The mice in the control group were injected with 20 mg/kg fluorouracil intraperitoneally.In the blank group and model group,10 ml/kg of saline solution was injected intraperitoneally once a day.Results Berberine inhibited HCT116 cells in vitro,and the IC50 of berberine was 0.24±0.02μmol/L.Comparison of body weight of mice in the berberineadministered group(low,medium,and high doses),positive drug group,model group,and control group on days 1,7,14 and 21 after injection,at different time points within the six groups,there was a difference in body weight between the mice(P<0.05).The tumor long diameter,short diameter and volume of the low,medium and high dose berberine group were lower than those of the model group,and showed a dose-dependent change(P<0.05).Compared with the model group,the survival rates of the low,medium and high dose berberine treatment group and the positive drug group were statistically higher(P<0.05,)and the berberine treatment group was dose-dependently altered.The tumor weight of low,medium and high dose berberine groups was lower than that of model group,and the berberine treatment group was dose-dependent(P<0.05).The tumor inhibition rates of the low,middle and high dose berberine group were higher.The high dose berberine group was higher than the low and medium dose berberine group(P<0.05),and the positive control group was higher than the low and medium dose berberine group(P<0.05).The expression of Caspase-9 and Caspase-3 mRNA in berberine group was higher than that in model group(P<0.05).The expression levels of Cytochrome C,Caspase-9 and Caspase-3 protein in berberine group,especially in middle and high dose group,were higher than those in model group(all P<0.05).Conclusions Berberine releases Cytochrome C by destroying mitochondria,and then activates the expression of apoptosis related protein Caspase-9 and Caspase-3,and induces apoptosis of tumor tissue,and then plays anti-tumor activity.