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地榆皂苷Ⅱ抑制结肠癌细胞增殖、迁移、侵袭和诱导凋亡的体外实验研究

Experimental study in vitro of ziyuglycoside Ⅱ in inhibition of proliferation, migration, invasion and induction of apoptosis of colon cancer cells

    摘要
  • 摘要:
    目的 探讨地榆皂苷Ⅱ对结肠癌细胞HT-29增殖、迁移、侵袭和凋亡的作用并探讨其作用机制。
    方法 采用CCK-8法检测地榆皂苷Ⅱ对细胞增殖的影响, 采用划痕试验检测地榆皂苷Ⅱ对细胞迁移能力的影响，采用Transwell小室实验检测地榆皂苷Ⅱ对细胞侵袭能力的影响，采用流式细胞术测定地榆皂苷Ⅱ对细胞凋亡的影响，分别采用实时定量反转录聚合酶链反应(qRT-PCR)和Western blot法测定地榆皂苷Ⅱ对细胞中蛋白激酶B (AKT)/磷脂酰肌醇-3-激酶(PI3K)信号通路mRNA和蛋白表达的影响。
    结果 地榆皂苷Ⅱ(0、1、5、10、20、40、60和80 μmol/mL)可剂量依赖性抑制结肠癌细胞HT-29的增殖; 地榆皂苷Ⅱ(5、10和20 μmol/mL)可剂量依赖性抑制结肠癌细胞HT-29的迁移能力; 地榆皂苷Ⅱ(5、10和20 μmol/mL)可剂量依赖性抑制结肠癌细胞HT-29的侵袭能力; 地榆皂苷Ⅱ(5、10和20 μmol/mL)可剂量依赖性促进结肠癌细胞HT-29的凋亡; 地榆皂苷Ⅱ(5、10和20 μmol/mL)可剂量依赖性降低结肠癌细胞HT-29中AKT和PI3K mRNA的表达，增加Caspase-3和Caspase-9 mRNA的表达; 地榆皂苷Ⅱ(5、10和20 μmol/mL)可剂量依赖性地降低结肠癌细胞HT-29中磷酸化蛋白激酶B(p-AKT)和磷酸化磷脂酰肌醇-3-激酶(p-PI3K)蛋白的表达，增加Cleaved-Caspase-3和Cleaved-Caspase-9蛋白的表达，差异有统计学意义(P＜0.05)。
    结论 地榆皂苷Ⅱ具有抑制结肠癌细胞HT-29的增殖、迁移、侵袭，促进细胞凋亡作用，可能与其促进AKT和PI3K蛋白磷酸化、Caspase-3和Caspase-9蛋白活化而调节AKT/PI3K信号通路有关。

     

    Abstract:
    Objective To investigate the effect and its mechanism of ziyuglycoside Ⅱ on proliferation, migration, invasion and apoptosis of colon cancer cells HT-29.
    Methods The effect of ziyuglycoside Ⅱ on cell proliferation of colon cancer cells HT-29 was determined by CCK-8 method; the effect of ziyuglycoside Ⅱ on cell migrative capacity of colon cancer cells HT-29 was determined by scratch assay; the effect of ziyuglycoside Ⅱ on cell invasive capacity of colon cancer cells HT-29 was determined by transwell assay; the effects of ziyuglycoside Ⅱ on cell apoptosis of colon cancer cells HT-29 was determined by flow cytometry; the effects of ziyuglycoside Ⅱ on mRNA and protein expression of protein kinase B (AKT)/phosphatidylinositol-3-kinase (PI3K) signal pathway were determined by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western-blot, respectively.
    Results Ziyuglycoside Ⅱ (0, 1, 5, 10, 20, 40, 60 and 80 μmol/mL) inhibited proliferation of colon cancer cells HT-29 in a dose-dependent manner. Ziyuglycoside Ⅱ (5, 10 and 20 μmol/mL) inhibited migration of colon cancer cells HT-29 in a dose-dependent manner. Ziyuglycoside Ⅱ (5, 10 and 20 μmol/mL) inhibited invasion of colon cancer cells HT-29 in a dose-dependent manner. Ziyuglycoside Ⅱ (5, 10 and 20 μmol/mL) promoted apoptosis of colon cancer cells HT-29 in a dose-dependent manner. Ziyuglycoside Ⅱ (5, 10 and 20 μmol/mL) increased mRNA expression of AKT and PI3K, decreased mRNA expression of Caspase-3 and Caspase-9. Ziyuglycoside Ⅱ (5, 10 and 20 μmol/mL) increased protein expression of phosphorylated protein kinase B(p-AKT) and phosphorylphosphatidylinositol-3-kinase (p-PI3K), increased the expression of Cleaved-Caspase-3 and Cleaved-Caspase-9 protein.
    Conclusion Ziyuglycoside Ⅱ can inhibit proliferation, migration and invasion of colon cancer cells HT-29, and promote the apoptosis of colon cancer cells HT-29. Its mechanism may be related to regulating the signal pathway of AKT/PI3K via promoting phosphorylation of AKT and PI3K protein, activation of Caspase-3 and Caspase-9 protein.

     

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