| The N-myc downstream-regulated gene (NDRG) family is composed of NDRG1, NDRG2, NDRG3 and NDRG4, which are important in cell proliferation and differentiation. At amino acid level, the four members share 53-65% identity. The phylogenetic analysis revealed that NDRG1 and NDRG3 belong to one subfamily, whereas NDRG2 and NDRG4 belong to another. Each of the four proteins contains anα/βhydrolase fold as in human lysosomal acid lipase. Deng Y et al first described the human NDRG2 sequence as a protein containing an acyl-carrier protein (ACP)-like domain. Based on UniGene cluster analysis, the gene NDRG2 is located at chromosome 14q11.2, which contains 16 exons and 15 introns. Its mRNA is 2,024 bp and the protein contains 357 amino acid residues. It is known that the expression of NDRG2 is significantly reduced in cancer tissues , such as breast cancer, liver cancer, gastric cancer, oligodendroglial tumors and skin cancer. So, NDRG2 is considered to play an inhibitive role in the tumor. Our previous experimental data showed that in the tissues of colon cancer and high-risk adenomas the expression of NDRG2 was significantly reduced , compared with those in the healthy controls. There was a trend for a decrease in NDRG2 levels with increasing Dukes'stage. Collectively, these data strongly suggest that NDRG2 is an inhibitor during the process of colon cancer, and may be involved in the cell invasion and metastasis. To study the function of NDRG2 and reveal the mechanisms of NDRG2 suppressing cell invasion and metastasis, we performed the following experiments:Experiment 1:Effects of NDRG2 on Migratory and Invasive Ability of Colon Cancer CellsObjective: To observe the influence of NDRG2 on the migratory and invasive ability of human colon cancer cell line SW620. Methods: pcDNA3.1-Ndrg2 and SiRNA-Ndrg2 were transfected transiently respectively into SW620 by Lipofectamine TM 2000. Reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blot were used to investigate the expression of mRNA and protein of NDRG2. Matrigel invasion assays and wound-healing experiment were used to study the movement and invasion of human colon cancer cell line SW620. Results: After transfecting pcDNA3.1-Ndrg2 into SW620 cells, the mRNA and protein levels of NDRG2 increased markedly and the ability of migration and invasion decreased significantly. After transfeting SiRNA-Ndrg2 into SW620 cells, the mRNA and protein levels of NDRG2 declined markedly and the ability of migration and invasion increased significantly. Conclusion: NDRG2 as a candidate tumor suppressor gene can reduce the abilities of Experiment 2:Lentivirus-mediated NDRG2 on the growth of colon cell SW620 Objective: To observe the influence of NDRG2 on the growth and cell cycle regulation of human colon cancer cell line SW620 and to explore the possible mechanism. Methods: Using the second-generation lentivirus vector system, packaging plasmid psPAX2, envelope plasmid pMD2.G and vector plasmid pLenti6.3/V5-DEST-NDRG2 were transfected into the human embryonic kidney epithelial cell line 293T cells. The lentiviral particles carrying NDRG2 gene were harvested. Then SW620 cells were infected by the lentiviral particles and the cells stably transfected NDRG2 were selected. The growth curve and the cell cycles were respectively determined through MTT method and flow cytometry. Results: Growth curve showed that compared with the control, the growth of cells stably transfected NDRG2 was inhibited; the cell cycle determination revealed that cells showed more obvious G1 arrest phenomenon than the control. Conclusion: NDRG2 as a candidate tumor suppressor gene can inhibit the proliferation of colon cancer cells. |
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