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目的探讨可溶性活化的matriptase-2对乳腺癌细胞的侵袭作用及其可能机制。方法构建表达matriptase-2胞外区片段且与C末端V5标签形成融合表达的真核表达质粒,转染CHO卵巢细胞,潮霉素筛选稳定表达重组蛋白的细胞克隆,收集上清液过镍琼脂糖凝胶柱。采用Western blot法鉴定matriptase-2活化,Transwell侵袭实验观察重组可溶性活化的matriptase-2对MDA-MB-231乳腺癌细胞侵袭能力的影响,Western blot法检测MMP-9的表达。结果建立了稳定分泌matritpase-2蛋白的CHO细胞克隆,可溶性matritpase-2蛋白过镍柱后自身活化。可溶性活化的matripase-2蛋白能显著抑制MDA-MB-231细胞的侵袭能力以及MMP-9的表达。结论可溶性活化的matriptase-2可能通过降低MMP-9的表达,从而抑制乳腺癌细胞的体外侵袭作用。
Objective To investigate the invasion and metastasis of breast cancer cells induced by soluble activated matriptase-2 and its possible mechanism. Methods Eukaryotic expression plasmids expressing the extracellular domain of matriptase-2 and fused with the C-terminal V5 tag were constructed and transfected into CHO ovarian cells. The cells stably expressing the recombinant protein were screened by hygromycin, Sugar gel column. Western blot was used to detect the activation of matriptase-2. Transwell invasion assay was used to investigate the effect of recombinant soluble matriptase-2 on the invasiveness of MDA-MB-231 breast cancer cells. Western blot was used to detect the expression of MMP-9. Results CHO cell clones stably secreting matritpase-2 protein were established. Soluble matritpase-2 protein activated itself through nickel column. Soluble activated matripase-2 protein significantly inhibited the invasive ability of MDA-MB-231 cells and the expression of MMP-9. Conclusion Soluble activated matriptase-2 may inhibit the invasion of breast cancer cells in vitro by decreasing the expression of MMP-9.