Novel ultrasound-targeted microbubble destruction mediated short hairpin RNA plasmid transfection targeting survivin inhibits gene expression and induces apoptosis of HeLa cells.
Survivin is an attractive target for tumor growth inhibition and represents a significant approach to anticancer therapy. RNA interference is an important tool for specifically down-regulating the expression of cellular genes. However, the efficiency of short hairpin RNA (shRNA) on the expression of survivin gene and the influence on the cell apoptosis transfected by the non-viral gene transfer system of ultrasound-targeted microbubble destruction was not explored. In this work, recombinant expression plasmid of shRNA targeting survivin gene was constructed and added to cultured cervical cancer cells followed by ultrasound exposure and SonoVue((R)) microbubble. Expression of survivin mRNA and protein were assessed by RT-PCR and western blot analysis. Apoptosis ratio was quantified by flow cytometry marked with annexin V and 7-AAD. After transfected for 48 h, the expression of survivin mRNA and protein were (16.67 +/- 2.73)% and (21.33 +/- 3.55)%, respectively. The apoptosis rate was (45.41 +/- 1.47)%. The differences were significant as compared with other groups (P < 0.01). In conclusion, we suggested that survivin could be regarded as an ideal anticancer target of cervical cancer. Recombinant expression plasmid of shRNA targeting survivin gene mediated by ultrasound-targeted microbubble destruction technique could effectively inhibit the expression of target gene and induce cell apoptosis. This novel method for RNA interference represents a powerful, promising non-viral technology that can be used in the tumor gene therapy and research.
Chen
Zhiyi
Z
Department of Medical Ultrasound, The Third Affiliated Hospital of Guangzhou Medical College, Guangzhou 510150, China. winchen1408@126.com
Liang
Kun
K
Xie
Mingxing
M
Wang
Xinfang
X
Lü
Qing
Q
Zhang
Jing
J
D000704:Analysis of Variance; D017209:Apoptosis; D005434:Flow Cytometry; D015870:Gene Expression; D018014:Gene Transfer Techniques; D006367:HeLa Cells; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D045423:Microbubbles; D008869:Microtubule-Associated Proteins / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D034622:RNA Interference; D012333:RNA, Messenger; D034741:RNA, Small Interfering / Q000235:genetics* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000077022:Survivin; D014162:Transfection / Q000379:methods; D014463:Ultrasonography / Q000379:methods*; D000097573:Red Fluorescent Protein
D051034:Inhibitor of Apoptosis Proteins; D008164:Luminescent Proteins; D008869:Microtubule-Associated Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D000077022:Survivin; D000097573:Red Fluorescent Protein; C107489:BIRC5 protein, human