红花多糖对人结肠癌 LoVo 细胞增殖、凋亡、侵袭作用机制研究

摘要

Objective To study the mechanism of safflower polysaccharide (SPS)on apoptosis of LoVo cell.Methods The LoVo cell was treated with different concentrations of SPS (i.e.0.5、1.0、1.5 g·L -1 ).At 24、48、72 h after intervention,the morphology and state of cell were observed by inverted microscope,the inhibitions of cell proliferation were analysed by MTT assay,and the change of cell cycle phase and cell apoptosis were detected by flow cytometry in Annexin V-FITC /PI.The expressions of Bcl-2,Bax and Caspase-3 were detected by RT-PCR,and the expressions of Bax,Bcl-2,Caspase-3 protein were detected by Western-blot.Transwell cell invasion assay was performed to detect the effect of safflower polysaccharide on the invasion ability of colon cancer LoVo cells.Results Com-pared with normal cell,the cells treated with SPS showed typical apoptosis detected by MTT assay.Many cells were floated in medium by inverted microscope,the cell cycle was blocked in phase G1 by flow cytometry,and RT-PCR and Western blot both indicated SPS can restrain the Bcl-2 and enhance Bax and Caspase-3.Conclusions SPS can significantly induce apoptosis,inhibit growth of the LoVo cell,decrease its ability of invasion,and regulate the cycle of cells.The effect may be related to pathways of Bax,Bcl-2,and Caspase-3.%目的：红花多糖对人结肠癌 LoVo 细胞增殖、凋亡、侵袭作用机制研究。方法体外培养人结肠癌 LoVo 细胞，在不同浓度（0．5、1．0、1．5 g·L －1）的红花多糖处理后，分别在干预24、48、72 h 后采用 MTT 比色法测定红花多糖对 LoVo 细胞的增殖抑制作用，利用倒置显微镜观察细胞的形态，采用 PI 单标流式细胞仪测定细胞周期分布，Annexin V-FITC ／PI 双标法检测细胞凋亡率；采用 RT-PCR 检测 Bax、Bcl-2、Caspase-3 mRNA 表达水平及检测 Caspase-3活性，Western-blot 检测 Bax、Bcl-2、Caspase-3蛋白表达情况；应用 Transwell 细胞侵袭试验检测红花多糖对结肠癌 LoVo 细胞侵袭能力的影响。结果MTT 检测显示红花多糖能显著抑制人结肠癌 LoVo 细胞的生长、增殖，呈现浓度和时间效应关系；倒置显微镜显示红花多糖能使结肠癌 LoVo 细胞出现典型凋亡特征；流式细胞仪分析结果提示红花多糖具有调节人结肠癌 LoVo 细胞周期的功能。RT-PCR 实验和 Western-blot实验结果显示随着红花多糖浓度增加可促进 LoVo 细胞中 Bax、Caspase-3 mRNA 表达，并抑制 Bcl-2 mRNA 表达。Transwell 侵袭实验示随着红花多糖浓度增加，LoVo 细胞侵袭能力降低。结论红花多糖能显著促进人结肠癌 LoVo 细胞凋亡，可使结肠癌 LoVo 细胞增殖受到抑制，侵袭能力减弱，调节细胞周期，红花多糖的抗肿瘤的生物学效应可能与 Bax、Bcl-2、Caspase-3的信号通路有关。