School of Pharmaceutical Sciences, Jilin University, Changchun 130021, PR China.
Mol Med Rep. 2011 Nov-Dec;4(6):1201-9. doi: 10.3892/mmr.2011.540. Epub 2011 Jul 26.
Matrix metalloproteinase 26 (MMP-26) is a novel member of the matrix metalloproteinase (MMP) family and is widely expressed in cancer cells of epithelial origin. MMP-26 has been shown to contribute to tumor development and to the restoration of tissue injury. In this study, in order to identify the functions of MMP-26 that contribute to the biological phenotype and behavior of human lung carcinoma A549 cells, we established an MMP-26 low-expressing tumor cell model using RNA interference (RNAi) transfection. These cells were used to investigate the role of MMP-26 in tumor progression. The MTT, colony forming, adhere-nce and spreading, wound-healing and Transwell chamber invasion assays were performed to analyze the invasion ability of pshRNA-MMP26-transfected A549 cells. Semi-quantitative reverse transcription polymerase chain reaction, Western blotting and double immunofluorescent staining were employed to detect the relationship between MMP-26 and MMP-9. Results showed that the adhesive rate was down-regulated in pshRNA-MMP26-transfected cells, compared to the controls. Silencing of the MMP-26 gene significantly retarded the invasiveness of A549 cells in Transwell insert invasion assays. The cells proliferated in the three-dimensional (3D) culture system. A549 cells transfected with the pshRNA-MMP26-C plasmid mainly developed reticular structures in morphology, and formed few clones with clear and smooth edges as well as tight intercellular junctions. The mRNA and protein expression of MMP-9 in pshRNA-MMP26-transfected cells were significantly lower than those of the controls. Double immunofluorescence labeling and focal laser scanning microscopy showed that MMP-26 was colocalized with MMP-9 in the controls. In conclusion, we successfully established an MMP-26 low-expressing cell model and confirmed that MMP-26 contributed to A549 cell invasion and migration in vitro. We also demonstrated that MMP-26 plays an important role in local invasion at least in part through coordination with MMP-9.
基质金属蛋白酶 26（MMP-26）是基质金属蛋白酶（MMP）家族的一个新成员，广泛表达于上皮来源的癌细胞中。MMP-26 被证明有助于肿瘤的发展和组织损伤的恢复。在这项研究中，为了鉴定 MMP-26 的功能，这些功能有助于人肺癌 A549 细胞的生物学表型和行为，我们使用 RNA 干扰（RNAi）转染建立了 MMP-26 低表达肿瘤细胞模型。使用这些细胞来研究 MMP-26 在肿瘤进展中的作用。通过 MTT、集落形成、黏附和铺展、划痕愈合和 Transwell 室侵袭实验分析 pshRNA-MMP26 转染的 A549 细胞的侵袭能力。采用半定量逆转录聚合酶链反应、Western blot 和双免疫荧光染色检测 MMP-26 与 MMP-9 之间的关系。结果显示，与对照组相比，pshRNA-MMP26 转染细胞的黏附率下调。沉默 MMP-26 基因显著抑制了 A549 细胞在 Transwell 插入侵袭实验中的侵袭能力。细胞在三维（3D）培养系统中增殖。转染 pshRNA-MMP26-C 质粒的 A549 细胞主要在形态上发育为网状结构，并且形成的克隆数较少，边缘清晰、光滑，细胞间连接紧密。pshRNA-MMP26 转染细胞的 MMP-9 mRNA 和蛋白表达明显低于对照组。双免疫荧光标记和共聚焦激光扫描显微镜显示，在对照组中 MMP-26 与 MMP-9 共定位。总之，我们成功建立了 MMP-26 低表达细胞模型，并证实 MMP-26 有助于 A549 细胞体外侵袭和迁移。我们还表明，MMP-26 通过与 MMP-9 的协调至少部分发挥作用，在局部侵袭中发挥重要作用。
