ABSTRACT
<p><b>OBJECTIVE</b>To explore the role of endothelin-1 (ET-1) gene in regulating the proliferation, migration and invasion of nasopharyngeal carcinoma cells.</p><p><b>METHODS</b>A lentivirus-mediated shRNA-ET-1 vector was infected into 5-8F cells, and the interference efficiency was examined with Western blotting. MTT assay, cell cycle analysis, plate colony formation assay, Transwell assay, Boyden chamber assay and tumor growth assay were carried out to analyze the changes in cell proliferation, migration and invasion. The expressions of genes related with epithelial-mesenchymal transition (EMT) were examined using Western blotting.</p><p><b>RESULTS</b>shRNA-ET-1 transfection significantly inhibited the expression of ET-1, and suppressed the growth, migration and invasion of 5-8F cells. ET-1 knockdown enhanced the expression of E-cadherin and CK18 and inhibited the expression of N-cadherin and vimentin.</p><p><b>CONCLUSION</b>ET-1 promotes cell growth, migration and invasion by modulating the genes associated with epithelial-mesenchymal transition in nasopharyngeal carcinoma cells.</p>
Subject(s)
Humans , Antigens, CD , Metabolism , Cadherins , Metabolism , Carcinoma , Genetics , Pathology , Cell Cycle , Cell Line, Tumor , Cell Movement , Cell Proliferation , Endothelin-1 , Genetics , Epithelial-Mesenchymal Transition , Gene Silencing , Lentivirus , Nasopharyngeal Neoplasms , Genetics , Pathology , Neoplasm Invasiveness , RNA, Small Interfering , Genetics , Transfection , Vimentin , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To investigate the effects of the enhancer of zeste homolog 2 (EZH2) gene on cell growth and invasion of the nasopharyngeal carcinoma (NPC).</p><p><b>METHODS</b>Recombinant lentivirus vector for shRNA delivery of EZH2 was constructed and transfected into 293FT cells. After collecting the viral particles, the NPC cell line 5-8F cells were transfected. The effects of EZH2 silence on cell proliferation and cell cycle were detected using MTT assay, plate colony formation assay and flow cytometry. The migration and invasion of 5-8F cells were determined by wound healing assay and matrigel invasion assay, respectively. The expressions of EZH2 and epithelial-mesenchymal transition (EMT)-related markers at mRNA and protein levels were examined by real-time PCR and Western blot respectively.</p><p><b>RESULTS</b>The expressions of EZH2 mRNA and protein in the transfected 5-8F cells were obviously reduced. MTT assay showed that EZH2 downregulation significantly inhibited the growth of 5-8F/shEZH2 cells (P < 0.001). Colony formation rate (84.44%) of 5-8F/shEZH2 cells was lower than control (31.56%, P = 0.001). Cell cycle analysis showed that most 5-8F/shEZH2 cells were arrested in G0/G1 phase, with a very low ratio of cells in S phase. Wound healing assay indicated that the migration ability of cells silencing EZH2 decreased significantly, and the 48-hour relative migration distance of 5-8F/ShEZH2 cells and control cells was 0.58 ± 0.05, and 0.81 ± 0.02, respectively (P < 0.000). Matrigel invasion assay, showed the invasive capacity of cells silencing EZH2 was significantly inhibited, with less penetrating cells (72.23 ± 4.08) compared to control (150.95 ± 16.27), P < 0.000. The mRNA expressions of epithelial markers E-cadherin and Keratin 18 in the cells silencing EZH2 increased by 177% and 158% respectively, and the mRNA expressions of mesenchymal markers β-catenin and N-cadherin decreased by 18.04% and 41.18% respectively. Similar results also were obtained with Western blot analysis.</p><p><b>CONCLUSION</b>EZH2 significantly enhanced the proliferation and invasion of nasopharyngeal carcinoma cells in vitro, which might be mediated by inducing EMT.</p>