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目的:探讨二十二碳六烯酸(DHA)对5-FU诱导乳腺癌细胞MDA-MB-231增殖与凋亡的作用及对Wnt/β-Catenin信号传导通路的影响。方法:采用四甲基偶氮唑蓝(MTT),测定DHA对5-FU诱导乳腺癌细胞MDA-MB-231增殖活性的作用;应用流式细胞仪检测细胞周期和凋亡率;RT-PCR检测MDA-MB-231细胞β-catenin、GSK-3β基因的表达情况;Western blot检测MDA-MB-231细胞β-catenin、GSK-3β、磷酸化GSK-3β(Ser9)蛋白的表达情况;细胞免疫组化染色观察DHA及DHA联合Licl对β-Catenin蛋白细胞内表达及定位的影响。结果:20μg/ml、40μg/ml的DHA分别与5-FU联合应用,可增强5-FU对MDA-MB-231细胞增殖抑制作用。DHA(20μg/ml)能促进5-FU增强MDA-MB-231细胞G0/G1期阻滞作用、降低细胞增殖指数及诱导细胞的凋亡。DHA作用于MDA-MB-231细胞后,β-catenin基因及蛋白表达水平下降(P<0.05)、GSK-3β的基因及蛋白表达水平无明显变化(P>0.05),磷酸化GSK-3β(Ser9)蛋白表达水平下降(P<0.05)。结论:DHA能增强5-FU对MDA-MB-231细胞增殖抑制和诱导凋亡,起化疗增敏作用,可能与其通过抑制GSK-3β磷酸化来阻断Wnt/β-Catenin信号通路有关。
AIM: To investigate the effects of docosahexaenoic acid (DHA) on proliferation and apoptosis of breast cancer cell line MDA-MB-231 induced by 5-FU and its effect on Wnt / β-Catenin signaling pathway. Methods: MTT assay was used to determine the effect of DHA on 5-FU-induced proliferation of breast cancer cell line MDA-MB-231. Flow cytometry was used to detect cell cycle and apoptosis rate. RT-PCR The expression of β-catenin and GSK-3β in MDA-MB-231 cells was detected by Western blot. The expressions of β-catenin, GSK-3β and phosphorylated GSK-3β (Ser9) The effect of DHA and DHA combined with Licl on the expression and localization of β-Catenin protein was observed by immunohistochemistry. Results: The combination of 20μg / ml and 40μg / ml DHA with 5-FU could enhance the inhibitory effect of 5-FU on the proliferation of MDA-MB-231 cells. DHA (20μg / ml) can promote the proliferation of MDA-MB-231 cells induced by 5-FU at G0 / G1 phase, decrease the cell proliferation index and induce the apoptosis of MDA-MB-231 cells. The expression of β-catenin gene and protein in MDA-MB-231 cells was decreased after DHA treatment (P <0.05), while the expression of GSK-3β mRNA and protein did not change significantly (P> 0.05) Ser9) protein expression decreased (P <0.05). CONCLUSION: DHA can enhance the inhibition and induce the apoptosis of MDA-MB-231 cells induced by 5-FU, which may be related to the block of Wnt / β-Catenin signaling pathway by inhibiting the phosphorylation of GSK-3β.