芹黄素对体外人大肠癌Lovo细胞生物学活性的影响及机制研究

目的探讨芹黄素对体外人大肠癌Lovo细胞生物学活性的影响及其可能的作用机制。方法不同浓度的芹黄素作用于体外培养的人大肠癌Lovo细胞24、48或72 h,采用光学显微镜和透射电镜分别观察芹黄素对Lovo细胞形态和超微结构的影响;采用四甲基偶氮唑盐比色法(MTT法)和平板克隆形成实验分别检测芹黄素对Lovo细胞增殖和克隆形成能力的影响;采用PI染色法、Annexin V FITC/PI双染法,流式细胞术分别检测芹黄素对Lovo细胞周期分布和凋亡的影响;采用Western blot法检测芹黄素对Bcl-2、pro-caspase-3、cleaved caspase-3表达的影响。结果芹黄素可浓度-时间依赖性的抑制人大肠癌Lovo细胞的增殖及克隆形成,作用48、72 h时的半数抑制浓度IC50分别为98.05μmol·L-1及33.91μmol·L-1;细胞周期分布发生改变,主要表现在G0/G1期细胞比例减少、G2/M期细胞比例增加,细胞周期阻滞于G2/M期;细胞凋亡率增加;Bcl-2、pro-caspase-3表达下调;cleaved caspase-3表达上调;差异具有统计学意义(P<0.05)。结论芹黄素可通过阻滞细胞周期于G2/M期抑制Lovo细胞增殖,通过下调Bcl-2、pro-caspase-3表达,上调cleaved caspase-3表达,诱导Lovo细胞凋亡,是一种有开发前景的大肠癌化疗药物。

Experimental research of the effect of apigenin on the biological activity of human colon cancer cell line Lovo and its mechanism in vitro

Aim To investigate the effect of apigenin on the biological activity of human colon cancer cell line Lovo and its possible mechanism in vitro.Methods After Lovo cells were cultured and treated with different concentrations of apigenin for 24,48 or 72 h,the morphological and ultrastructural changes were observed through light microscope and transmission electron microscope respectively;cell proliferation was measured by methyl thiazolyl tetrazolium(MTT) colorimetric assay;cell clonogenicity was detected by colony-forming assay;cell cycle distribution and apoptosis were examined by flow cytometry after cells were stained with PI and Annexin V FITC/PI respectively;the expression of Bcl-2,pro-caspase-3 and cleaved caspase-3 in Lovo cells was detected by Western blot analysis.Results Compared with untreated cells,cells treated with apigenin exhibited a marked proliferation and clone-formation inhibition in a dose-time dependent manner.The half maximal inhibitory concentration(IC50) on cell proliferation was approximately 98.05 μmol·L-1 when Lovo cells were treated with apigenin for 48 h and 33.91 μmol·L-1 for 72 h.Accompanying with the decreased growth,apigenin treatment could also cause a decreased percentage of cells at G0/G1 phase,a cell cycle block at G2/M phase and an increased rate of apoptosis.Moreover,apigenin treatment could also dramatically downregulate the expression of Bcl-2,pro-caspase-3 and upregulate the expression of cleaved caspase-3 in Lovo cells.Differences had statistical significance(P<0.05).Conclusions Apigenin partially inhibits the proliferation of Lovo cells by blocking cell cycle at G2/M phase,promotes apoptosis by downregulating the expression of Bcl-2,pro-caspase-3 and upregulating the expression of cleaved caspase-3 in Lovo cells in vitro and has potential for further development as a chemotherapeutic agent for human colon cancer.