Construction of antisense RNA expression plasmid for u-PAR and its transfection to highly invasive PC-3M cell subclones.
To evaluate the specific inhibition of antisense u-PAR on the u-PAR expressions in highly invasive cell subclones and to determine its blocking function in the invasion by those cells, a cDNA fragment of u-PAR obtained by RT-PCR was inserted into a plasmid vector named pcDNA3 in antisense orientation. Then the antisense u-PAR recombinant was transfected into highly invasive cell subclones. The u-PAR expression in neo-resistant cells was examined by RT-PCR and immunohistochemical assay. Compared to the control cells, the content of mRNA and protein of u-PAR in transfected cells decreased sharply, and the rate of inhibition was 53% and 73%, respectively, indicating that an antisense u-PAR might have played a specific inhibitory role in its expression in the cells, which may provide a good cell model for making further investigation of the inhibitory effects of the antisense u-PAR on invasion in highly invasive cell subclones of human prostate carcinoma.
Liao
Guoning
G
Department of Biochemistry and Molecular Biology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030.
Li
Qingfen
Q
Feng
Youmei
Y
Deng
Yaozu
Y
Li
Zhuoya
Z
Gong
Feili
F
Ma
Ding
D
D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D006801:Humans; D008297:Male; D009361:Neoplasm Invasiveness; D010957:Plasmids; D011471:Prostatic Neoplasms / Q000378:metabolism / Q000473:pathology*; D016372:RNA, Antisense / Q000096:biosynthesis / Q000235:genetics; D011956:Receptors, Cell Surface / Q000096:biosynthesis / Q000235:genetics*; D055293:Receptors, Urokinase Plasminogen Activator; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014568:Urokinase-Type Plasminogen Activator / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism
C524898:PLAUR protein, human; D016372:RNA, Antisense; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D014568:Urokinase-Type Plasminogen Activator