Title: Construction of recombinant secreting tumor necrosis factor related apoptosis inducing ligand adenoviral vector and the effects of secreting-trail adenovirus on the prostate cancer PC-3 cells’ proliferation
Abstract: Objective
To construct a recombinant secreting-trail adenoviral vector and to investigate the effect of tumor necrosis factor related apoptosis inducing ligand (TRAIL) gene on the prostate cancer PC-3 cells’ proliferation.
Methods
The TRAIL gene with a secreting signal peptide was obtained by gene synthesis, and cloned into GV315 plasmid vector. Then plasmids were constructed and amplified in 293A cells. The adenoviral vector were infected into prostate cancer cell line PC-3 in vitro. The up-regulation of TRAIL protein was determined by indirect immunofluorescence assay. The protein expressions of cysteinyl aspartate-specific protease (Caspase)-3 and Caspase-8 in the cells infected with Ad-sTRAIL and Ad-LacZ were determined by Western blotting respectively. Methyl thiazol tetrazolium (MTT) assay and plate colony formation were used to determine the effect of cells proliferation.
Results
Construct the Ad-sTRAIL successfully. After infection, it had been verified that the protein expression of TRAIL in the cells by indirect immunofluorescence assay. Through the Western-blot, the expressions of active-Caspase-3 and active-Caspase-8 in Ad-sTRAIL infected PC-3 cells were significantly higher than the Control group and Ad-LacZ group. The colony formation rate of the PC-3 cells was [(30.6±2.10)%] in Ad-sTRAIL group significantly lower than (46.6±4.86)% in Control group (q=7.56, P<0.01) and (45.9±3.56)% in Ad-LacZ group (q=7.19, P<0.01). MTT assay showed that Ad-sTRAIL significantly inhibited the growth of the PC-3 cells(20.15%)at 48 h (q=6.82, P<0.01).
Conclusion
The secreting-trail adenoviral vector could up regulate the expression of TRAIL, active-Caspase-3 and active-Caspase-8, furthermore it could inhibit the proliferation of PC-3.
Key words:
Prostate cancer; Tumor necrosis factor related apoptosis inducing ligand gene; Cell proliferation; Secreting-trail adenoviral vector
Publication Year: 2016
Publication Date: 2016-12-08
Language: en
Type: article
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