[Experimental study on bladder cancer by the small interfering RNA targeting survivin].
OBJECTIVE
To study the influence of small interfering RNA (siRNA) targeting survivin on the biological behavior of bladder cancer.


METHODS
One pair of survivin target sequence-specific siRNA was designed, then siRNA/liposome complex was used to transfect bladder cancer cell line-T24. The efficiency of transfection and the apoptosis were detected by flow cytometry. The transcriptional level of survivin was analyzed using real time PCR. DNA sequence corresponding to siRNA targeting survivin was cloned into pRNAT-U6.1/Neo to produce plasmid targeting surviving.


RESULTS
The ratio of T24 cells releasing fluorescence in total cells were 92.3%; siRNA-survivin efficiently down-regulated survivin expression (mRNA) in a dose-and time-dependent manner. Its maximal effect was achieved at the concentration of 100 nmol/L, at which survivin expression level was down regulated by 75.91%. Similar results were found in the inhibition ratio of cell growth, which was 55.29%(P< 0.01). Simultaneously the apoptotic rate was markedly increased, which was 45.70%(P< 0.01). After cutting the vector with Bam H I and Hin d III and ligating the vector with the insert by using T4 ligase, the recombinant vector was confirmed by restriction digestion and DNA sequencing.


CONCLUSIONS
The application of siRNA-survivin can markedly inhibit survivin expression in bladder cancer cell line, induce apoptosis and inhibit the growth of the tumor. It may be a new gene therapy tool for bladder cancer. The successful construction of the siRNA expressing plasmid will facilitate the application of RNA interference technique, and lay a foundation for further studies on the function of surviving.
Department of Urology, First Affiliated Hospital of Soochow University, Suzhou, Jiangsu, 215006 P. R. China.
Jian-quan
JQ
Hou
Jun
J
He
Duan-gai
DG
Wen
Xiao-lin
XL
Wang
D045744:Cell Line, Tumor; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins / Q000235:genetics*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics*; D000077022:Survivin; D014162:Transfection; D001749:Urinary Bladder Neoplasms / Q000235:genetics / Q000473:pathology
C107489:BIRC5 protein, human; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins; D034741:RNA, Small Interfering; D000077022:Survivin