| Objective: Ovarian cancer is the most lethal tumor of the femal genital tract and is seriously threatening femal health. Lacking of early effective diagnosis methods and treatment, 70 percent of patients with ovarian cancer are diagnosed as late-stage. The conventional principles of treatment are surgery and chemotherapy. 5 –year survival continues to be 28%~35% and long-term one isn't over 15%-30%. It is necessary to find a new treatment to improve the prognosis of the patients with ovarian cancer. Cancer has been considered as a disease of gene alterations, therefore, gene therapy may be an effective measure for treating cancer. EF-1A2 gene has the oncogenic and anti-apoptotic properties and is considered as a putative oncogene. It plays an important role in the process of protein synthesis and regulates cytoskeletonic organization to facilitate cell growth. Experimental evidences demonstrate that amplication and overexpression of the EF-1A2 gene have been observed in 25%~30% of human ovarian cancer and established cell lines. Researches indicate that EF-1A2 gene overexpression characterizes a group of patients with unfavorable tumor biology and poor prognosis and contributes to transformations and tumor progression. The genetic amplications of growth-enhancing genes play a key role in the development of human malignancy. Antisense gene therapy can arrest the expression of targeted gene through blocking its DNA or RNA according to complementary base pairing rule, which is highly selective and specific. Therefore, the EF-1A2 gene represents an attractive target for antisense therapy. This study was designed to investigate the effect of antisense oligonucleotides (AS-ODN) of EF-1A2 gene on SKOV3 ovarian cancer cell line.Methods: 1. 21-mer oligodexynucleotides targeted against the translation initiation of EF-1A2 mRNA and complementary-sense control oligonucleotides were synthesized from shanghai sangon and 3 bases each two ends were phosphorothioate. 2. SKOV3 ovarian cancer cells were seeded in 96-well microtiter plates and culture bottles in the same concentration and maintained in RPMI-1640 containing 10% FBS in 37.0℃ 5% CO2 and allowed to adhere for 24 hours, then cells were cultured with RPMI-1640 not containing 10% FBS for 48 hours. Then cells were divided into three groups: AS-ODN group; S-ODN group; untreated control group. Each group cells were transfected with different concentration of AS-ODN, S-ODN or equalled culture liquid for 24, 48 and 72 hours. Cell proliferation inhibitory was measured by MTT assay. Morphological changes of apoptotic cells were observed by light microscopy and Giemusa staining. DNA fragmentation was analysed by agarose gel electrophoresis. Kinetics of induction of apoptosis, cell cycle and bcl-2 gene expression were analyzed by Flow Cytometry.Results: 1. MTT assay showed that cell proliferation was significantly inhibited by EF-1A2 antisense oligonucleotides in time and concentration dependent manner. 2. Ovarian cancer cells SKOV3 treated by EF-1A2 antisense oligonucleotides showed characteristic morphological changes of apoptosis. 3. A ladder-like pattern of DNA fragments was demonstrated on agarose gel electrophoresis by AS-ODN treatments. 4. The apoptotic rate of SKOV3 cells treated by 5μmol/L ASODN, SODN and untreated control groups were 5.73 %, 3.67%, 3.47% respectively. 9.73%, 3.73%, 3.61% at 10μmol/L group, while 14.73%, 3.81%, 3.52% at 20μmol/L group. The induction of apoptosis by EF-1A2 antisense oligonucleotide was concentration dependent. At the same density condition, the apoptotic rate in ASODN group was significantly higher than that in SODN group and untreated control group (P<0.01). At different density condition there was a significant difference in three ASODN subgroups (P<0.01). No significant difference was observed between SODN group and untreated control group (P>0.05). Cell cycle analyzed by Flow Cytometry showed that the cells of AS-ODN group gradually increased in G0~G1 phase and decreased in both S and G2~M phases with the increa...
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