Zou D.-L.,Chongqing Institute of Oncology |
Zhou Q.,Chongqing Institute of Oncology |
Wang D.,Chongqing Institute of Oncology
Tumor | Year: 2014
Objective: To establish an adenovirus expression vector containing human telomerase reverse transcriptase (hTERT) promoter-driven p53 up-regulated modulator of apoptosis (Puma), and further to achieve the directional expression of Puma in ovarian carcinoma COC1 cells. Methods: The full-length open reading frame (ORF) of Puma and the promoter of hTERT were amplified by RT-PCR and cloned into the adenovirus shuttle vector pDC316. Recombinant adenovirus pDC316-hTERT-Puma was obtained and infected into human normal ovarian epithelial HUM-CELL-0088 cells and ovarian carcinoma COC1 cells. The expression levels of Puma mRNA and protein were detected by real-time fluorogenic quantitative-PCR (RFQ-PCR) and Western blotting, respectively. The cell proliferation was detected by cell counting kit-8 (CCK-8) assay. The apoptosis was detected by flow cytometry (FCM). Results: The adenovirus expression vector pDC316-hTERT-Puma containing hTERT promoter-driven Puma was successfully constructed, and the corresponding adenovirus could achieve the specific and high expression of Puma in ovarian carcinoma cells but not in the normal ovarian cells. The successful expression of Puma could inhibit the proliferation of COC1 cells evidently (P 〈 0.05) and induce apoptosis (P 〈 0.05). Conclusion: The tumor-targeted expression of Puma gene in ovarian carcinoma cells is achieved through adenovirus vector, and it will be beneficial for the use of Puma in targated therapy of ovarian carcinoma. Copyright© 2014 by TUMOR.