[Transfection of pancreatic cancer cells BxPC-3 with recombinant plasmid pSilencer4.1-CMV neo-hTERT-siRNA and its silencing effects].
OBJECTIVE
To study the transfection of pancreatic cancer cells BxPC-3 with recombinant plasmid pSilencer4.1-CMV neo-hTERT-siRNA and its silencing effects.


METHODS
Pancreatic cancer cells BxPC-3 transfected with recombinant plasmid pSilencer4.1-CMV neo-hTERT-siRNA were selected as target and divided into five groups: (1) T1 group (pSilencer4.1CMV neo-hTERT1-siRNA), (2) T2 group (pSilencer4.1CMV neo-hTERT2-siRNA), (3) Lipofectamine (Lipofectamine), (4) mismatch group(pSilence4.1CMV, as negative control), (5) cell control group(without transfection). The expression of hTERT mRNA was detected by RT-PCR. The viability of cells was measured by MTT method. The cell cycle and cell apoptosis was measured by flow cytometry. The expression of telomerase protein was measured by Western blot.


RESULTS
Compared with Lipofectamine group, negative control group and cell control group, the expression of hTERT-mRNA and telomerase protein in cells was downregulated significantly(P<0.05), the viability of BxPC-3 cells was decreased significantly (P<0.05), the ratio of cells in G0/G1 stage was increased, the ratio of cells in S stage and G2/M stage was decreased, and the ratio of apoptotic cells was increased significantly in T1 group and T2 group.


CONCLUSIONS
Recombinant plasmid T1 and T2 can downregulate the expression of hTERT mRNA and telomerase protein in BxPC-3 cells , and has good RNAi silencing effects. T1 and T2 can also inhibit the growth of BxPC-3 cells, block the cell cycle, promote the apoptosis of cells, and has anti-pancreatic cancer effects in vitro.
Tan
Jing
J
Department of Hepatobiliary and Pancreatic Surgery, The 2nd Affiliated Hospital of Kunming Medical University, Kunming 650101, China.
Sun
Min
M
Mo
Xiao-hua
XH
Lu
Qi-yu
QY
Zhou
Xiao-hu
XH
D017209:Apoptosis / Q000235:genetics; D045744:Cell Line, Tumor; D005822:Genetic Vectors; D006801:Humans; D010190:Pancreatic Neoplasms / Q000235:genetics*; D010957:Plasmids; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics*; D019098:Telomerase / Q000235:genetics; D014162:Transfection
D034741:RNA, Small Interfering; D019098:Telomerase