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目的:研究下调骨桥蛋白(osteopontin,OPN)对乳腺癌MDA-MB-231细胞生物学行为及基质金属蛋白酶-2(matrix metalloproteinase-2,MMP-2)表达的影响。方法:采用重组的OPN mRNA干扰质粒转染MDA-MB-231细胞,建立稳定转染细胞株。RT-PCR和Western印迹法检测细胞中OPN和MMP-2的表达,MTT法、流式细胞仪检测和软琼脂生长实验观察OPN下调对细胞生长、细胞周期分布和瘤细胞恶性表型的影响,酶联免疫吸附法(ELISA)检测细胞培养液中MMP-2的分泌水平。结果:与MDA-MB-231细胞相比,建立的转染细胞株M-OPNS1和M-OPNS2细胞的OPN mRNA及其蛋白表达均显著下调,细胞生长受到抑制,在细胞周期上表现为S期阻滞;OPN下调的细胞在软琼脂上的生长能力明显下降,MMP-2表达也明显降低。结论:RNA干扰介导的OPN下调能抑制MDA-MB-231细胞增殖、下调细胞恶性程度和MMP-2的表达,这可能为将来乳腺癌治疗提供新的方法。
AIM: To investigate the effects of osteopontin (OPN) on the biological behavior and the expression of matrix metalloproteinase-2 (MMP-2) in human breast cancer cell line MDA-MB-231. Methods: The transfected MDA-MB-231 cells were transfected with recombinant OPN mRNA, and stable transfected cell lines were established. The expression of OPN and MMP-2 were detected by RT-PCR and Western blotting. MTT assay, flow cytometry and soft agar assay were used to observe the effect of OPN down-regulation on cell growth, cell cycle distribution and malignant phenotype of tumor cells. Enzyme-linked immunosorbent assay (ELISA) was used to detect the secretion of MMP-2 in cell culture medium. Results: Compared with MDA-MB-231 cells, OPN mRNA and protein expression of M-OPNS1 and M-OPNS2 cells were significantly down-regulated and the cell growth was inhibited. The expression of S phase Block; OPN down-regulated cells in soft agar significantly decreased the growth capacity, MMP-2 expression was also significantly reduced. Conclusion: Down-regulation of OPN mediated by RNA interference can inhibit the proliferation of MDA-MB-231 cells and down-regulate the expression of MMP-2 and cell malignancy, which may provide a new method for the future treatment of breast cancer.