Knockdown of cyclin A2 expression by small interfering RNA in MG-63 cells / 中华肿瘤杂志
Chinese Journal of Oncology
;
(12): 670-675, 2007.
Article
in Chinese
| WPRIM
| ID: wpr-298521
ABSTRACT
<p><b>OBJECTIVE</b>To study the inhibitory effect of small interference RNA (siRNA) targeting cyclin A2 gene on the growth of osteosarcoma MG-63 and human normal skin fibroblast HSF cells and to explore whether cyclin A2 siRNAs could become a useful tool in the treatment of osteosarcoma.</p><p><b>METHODS</b>Three pairs of siRNAs targeting cyclin A2 mRNA and a pair of nonsense siRNA were designed according to the current criteria. SiRNAs were chemically synthesized and purified. The siRNAs were transfected into MG-63 cells and HSF cells via oligofectamine. The cells transfected with nonsense siRNA served as negative control group and those only treated with PBS as blank control group. Quantitative fluorescence RT-PCR, Western-blot, MTT assay, reverse transcriptase (RT)-PCR, flow cytometry and clone forming test were employed to evaluate the efficacy of RNA interference. At the same time, the mRNA expression of PCNA and cyclin B1 in siRNA-treated MG-63 cells were examined.</p><p><b>RESULTS</b>Although all three siRNAs could reduce the cyclin A2 expression, siRNA, appeared to be the most effective. After 48 h treatment with siRNA1, cyclin A2 mRNA and protein expression in MG-63 cells was significantly reduced by nearly 80% as compared with that of the blank control group, whereas the negative and blank control groups had similar expression levels. MG-63 cells treated with siRNA1 were arrested at G0/G1 phase by 80.1% and the proliferation of these tumor cells was suppressed 48 h after transfection. Furthermore, MG-63 cells showed a decreased colony forming ability after siRNA1 treatment. In addition, the cyclin A2-depleted MG-63 cells showed decreased levels of PCNA and cyclin B1. In contrast, although cyclin A2 expression in HSF reduced by nearly 60% after treatment by siRNA1 for 48h, these cells exhibited only a slight change in cell cycling, and neither clear inhibition of proliferation nor impaired colony forming ability was observed.</p><p><b>CONCLUSION</b>Cyclin A2 is critical for proliferation of MG-63 cells. Cyclin A2-siRNAs can induce obvious inhibition of cyclin A2 mRNA and protein expression in MG-63 and HSF cells, which consequently down-regulate the proliferation of MG-63 cells. There is little effect on the proliferation of siRNA-treated HSF cells. Those results indicate that siRNAs against cyclin A2 may become a potential antiproliferative tool in future antitumor therapy.</p>
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Pathology
/
Skin
/
Bone Neoplasms
/
RNA, Messenger
/
Transfection
/
Osteosarcoma
/
Cell Cycle
/
Proliferating Cell Nuclear Antigen
/
Cell Biology
/
RNA, Small Interfering
Limits:
Humans
Language:
Chinese
Journal:
Chinese Journal of Oncology
Year:
2007
Type:
Article
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