降低COX-2表达与乳腺癌细胞迁移和侵袭关系的研究

The Effect of Disruption of Cyclooxygenase-2 on Invasiveness and Migration of Breast Cancer Cells

目的 探讨利用小RNA干扰技术降低COX-2表达对高度恶性乳腺癌MDA-MB-231细胞趋化和侵袭能力的影响.方法 应用合成的小RNA干扰质粒转染MDA-MB-231细胞株,采用逆转录-聚合酶链反应（RT-PCR）检测COX-2mRNA的表达.通过划痕实验检测细胞的运动能力;体外侵袭实验检测细胞的侵袭能力;应用细胞黏附实验检测细胞的黏附能力.结果 限制性内切酶的酶切结果显示成功构建了干扰质粒pSUPER-siCOX-2,转染后的细胞株分别命名为MDA-MB-231/pSuPER-basic和MDA-MB-231/pSuPER-siCOX-2.转染后48h,与MDA-MB-231/pSUPER-basic细胞相比,MDA-MB-231/pSUPER-siCOX-2细胞的COX-2 mRNA表达水平下降（P＜0.01）;COX-2减低的乳腺癌细胞单位时间内向划痕移动的距离比对照组细胞减少（P＜0.05）;黏附实验结果显示：黏附5min,15min后,COX-2减低的乳腺癌细胞比对照组的黏附细胞数量均减少（P＜0.01）;COX-2减低的乳腺癌细胞侵袭并穿透Matrigel的细胞数量比对照组细胞少（P＜0.01）.结论 利用siRNA干扰技术降低COX-2表达对乳腺癌MDA-MB-231细胞株的迁移和侵袭能力具有明显的抑制作用.

Objective To investigate the role of COX-2 in invasiveness and chemotaxis of tile highly malignant breast cancer cell line MDA-MB-231 l）y using RNA interference. Methods MDA-MB-231 cells were transfeeted with small RNA interierence plasmids to disrupt COX-2 expression. COX-2 mRNA levels were detected using RT-PCR. Scratch assay was performed to detect the migration ability of MDA-MB-231 ceils. The in vitro invasion ability of MDA-MB-231 cells was examined using matrigel invasion assay. The adhering ability of MDA-MB-231 cells was examined using adhesion assay. Results The resuhs of ,＇estriction endonnclease digestion electrophoresis showed that the interenced plasmid pSUPER-siCOX-2 was construeted successfully. The cell strains transfected were named as MDA-MB-23 1/pSUPER-basic and MDA-MB-231/pSUPER-siCOX-2,respeetively. The results of RT-PCR showed that the levels of COX-2 mRNA of MDA-MB-231/ pSUPER-siCOX-2 were obviously reduced at 48 ilours alter transfection,compared to MI）A-MB-231/pSUPER-basic cells. When a seratch was created in the fluent monolayer cells,it took the COX-2-reduced MDA-MB-231 cells a longer time to fill the gap（P 〈 0.05）. Adhesion assay showed the numt）er of the adhering cells was decreased for reduction of COX-2 within 5 and 15 minutes （ P 〈 0.01 ）. The invasion assay showed prominent differences between the COX-2-reduced MDA-MB-231 cells and the control cells（P 〈0.01 ）. Conclusion Using RNA interference,the reduction of COX-2 expression can obviously inhibit the invasion and migration of the highly malignant breast cancer cell line MDA-MB-231.