Published in:
01-12-2015 | Research Article
The effect of recombinant lentiviral vector encoding miR-145 on human esophageal cancer cells
Authors:
Tian-Yun Wang, Qing-qing Zhang, Xi Zhang, Qiu-Li Sun, Chun-Peng Zhao, Xiao-Yin Wang
Published in:
Tumor Biology
|
Issue 12/2015
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Abstract
miR-145, a newly identified microRNA molecule, is hypothesized to function as a tumor suppressor, but this activity has not been investigated in esophageal l carcinoma (EC). The aim of this study was to investigate the effect of miR-145 on the biological features of EC cells. miR-145 was obtained using PCR technology and cloned into the lentiviral vector, pLVX-IRES-ZsGreen1, to construct the resulting vector, pLVX-IZ-miR-145. The vector was packaged, the viral titer was tested, and ECA109 cells were infected with the optimal viral titer. Cells that were stably transfected with miR-145 were screened. Flow cytometry was used to analyze enhanced green fluorescence protein gene expression, and to measure cell apoptosis and cell cycle. miR-145 expression was detected by real-time fluorescent quantitative PCR. Furthermore, cell proliferation was assayed using CCK-8 assay. The pLVX-IZ-miR-145 vector was successfully constructed, and the viral titer achieved up to 5.0 × 108 TU/mL. The transfection efficiency was 90 %. Compared to the control group, the expression level of miR-145 in the transfected group was significantly higher (185-fold, P < 0.05). miR-145 overexpression significantly inhibited esophageal cancer cell proliferation (P < 0.05). Moreover, the number of cells at the G2/M stage, as well as the cell apoptotic rate, in the miR-145-transfected group was significantly increased (P < 0.05). Our study reveals that overexpression of miR-145 inhibits cell proliferation, increases apoptosis, and influences the cell cycle progression of EC cell.