| RNA interference(RNAi) refers to the introduction of homologous double stranded RNA(dsRNA) to specifically target a gene's product, resulting in null or hypomorphic phenotypes. This powerful technology has been widely employed to manipulate gene expression in mammalian and human cells, elucidate signal pathways, identify gene functions in a whole-genome scale and develop RNAi-based drugs for various diseases, especially in cancers. A number of experimental studies have made it clear that different types of mRNAs can be efficiently, specifically and rapidly inactivated by RNAi. Over-expression of oncogenes is involved in uncontrolled growth and proliferation of cells, the specific suppression of oncogene activities can reverse the phenotype of tumor cells and control their growth and proliferation. This notion has been tested in HeLa cells, lung adenocarcinoma cells, hepatoma cells, ovarian carcinoma cells, and melanoma cells by using single siRNA(short interfering RNA) or combination of siRNAs with cationic lipidcomplexes. It has been reported that RNAi-based approach is a very useful tool for inhibitiing MDA-MB-231 human breast cancer cell migration in vitro and suppressing the invasion and metastasis of gliomas in vivo for almost 4 month. It is optimism when the whole process of tumor invasion and metastasis was delineated by RNAi and other methods, thecure of cancer seems not far from us.Adenoid cystic carcinoma (ACC), accounting for approximately 30% of salivary glands carcinoma, has a high metastatic ability. Clinical statistics shows that distant metastasis often defeat treatment of patients with ACC, and is associated with a low long-term survival rate.The incidence of distant metastasis in ACC, most often the lung, ranges from 35% to 50% of the cases. In this study, we used salivary gland adenoid cystic carcinoma cell line ACC-2 and its high metastatic ACC-M as model system to identify the genes with differential expression levels by cDNA microarray. The results show that matrix metalloproteinasel(MMP-1) is significantly up-regulated in ACC-M cells.MMP-1, a memeber of MMPs family, is also called fibroblast collagenase. It was the first vertebrate collagenase purified as a protein and cloned as a cDNA, and is considered the prototype for all the interstitial collagenases. Studies show that MMP-1 is up-regulated in a wide variety of advanced cancers, and in nearly all instances, has a significant negative correlation with survival. However, there are few studies on the relationship between MMP-1 and SACC. Using shRNA (short hairpin RNA) expression system, we showed that lnducible knockdown of MMP-1 gene expression in ACC-M cells with high metastatic potential resulted in significant inhibition of cell migration and invision in vitro and lung metastasis in vivo. The study included three parts as follows:1. 26 differential expression genes between ACC-2 and ACC-M cells were identified by cDNA microarray BioStarH-I representing 1152 genes, of which 19 genes were up-regulated and 7 down-regulated. The results were validated by RT-PCR. MMP-1, one of the upregulated genes, was chosen to be the target for RNAi.2. The plasmid expressing shRNA homologous to MMP-1 mRNA was constructed using shRNA expressing vector pWH1, and named pWH1-MMP1. The combinant plasmid was transfected into ACC-M cells with cationic lipid complexes Lipofectamine2000 and then stable transfectant of ACC-M cells aquired using G418 agent. Finally, blocked expression of MMP-1 in the stable transfectant was...
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