The Inducible Of IEX-1by UVB And The Relationship Between Other Genes In Ovarian Cancer

Background and ObjectiveOvarian cancer is a common malignancy of the female reproductive system, because of specific symptoms and effective detection tool in the early stage, most patients were treated in the advanced stage. The mortality rate was the first of gynecologic oncology. Therefore, it poses a serious threat to the woman’s life. In recent years, along with the raising level of medical technology, the five-year survival rate of ovarian cancer was improved, but still only25%-30%. The main treatment of ovarian cancer is the surgical treatment, and combined with chemotherapy, radiation therapy, immunotherapy, and so on. But only received little efficient in advanced ovarian cancer patients. Gene therapy and tumor gene-radiation therapy which was based on it are new strategy for cancer treatment in recent years. Immediate early response genes (immediate early response gene X-1, IEX-1) is one of the family member of immediate early genes (immediate early gene, IEG), the gene located on5q31, the regulatory sequences of IEX-1are highly conserved. It will start a rapid, short-term expression after some physical and chemical stimulation such as ionizing radiation, mitogen, and radicals. IEX-1is closely related to cell growth, differentiation and apoptosis, and participate in a variety of tumor development. Recent studies have shown that the low expression of IEX-1in ovarian cancer tissue, the low expression of IEX-1is one of the factors which caused the ovarian benign transformed to malignant, and is associated with the low survival rate of patients in ovarian cancer. Transfected the recombinant plasmid pEGFP-N1-IEX-1in human ovarian cancer cells, S phase+G2/M phase cell ratio was significantly increased and speed up the cell proliferation. Internal tests and external tests were both revealed that IEX-1participated in ovarian cancer development.In our research, UVB was used to irradiate SKOV3cell which was in low expression of IEX-1, at the same time, we transfected the recombinant plasmid pEGFP-N1-IEX-1into SKOV3cells, then observe the radiation-induced effection and sensitivity to cisplatin of IEX-1. Then we detected the expression of IEX-1and NF-κB, Mcl-1, p53in ovarian cancer tissue and the relationship of those proteins were analysed. The purpose is to explore the IEX-1function and the specific mechanism in ovarian cancer and to provide valuable gene therapy of ovarian cancer.Methods1. Different doses and different time of UVB was used to irradiate on SKOV3cells, the expressions of IEX-1mRNA were examined by RT-PCR after irradiated.2. The experiment was divided into three groups:SKOV3, SKOV3-EGFP group and SKOV3-IEX-1group. SKOV3was transfected with recombinant plasmid pEGFP-N1-IEX-1and empty plasmid pEGFP-N1by lipofectamine, observed the transfection efficiency by fluorescence microscopy, and the expression of IEX-1protein was analysised by Western blot.3. The cell apoptisis rate was determined by PI/AnnexinV double staining, to detect the effects of IEX-1gene transfection on sensitivity to cisplatin of ovarian cancer cell line SKOV3.4. The expression of IEX-1and NF-κB, Mcl-1, p53in ovarian cancer tissue was determined by immunohistochemistry, and their relationship was analysised.5. Statistical software SPSS17.0was used to analyze data results. One-way analysis of variance and bonferroni test were used to measure mean differences between three groups. Bilateralα=0.05was considered as test standards. Results1. After different doses and different time of UVB irradiated on SKOV3cells, the expression IEX-1mRNA in SKOV3was significantly increased (P<0.05), with a dose-dependent manner (P<0.05), the abundance of IEX-1mRNA increased rapidly after exposure of the cells to40mJ/cm2of UVB, reaching a maximum by1or2hours after irradiated (P<0.05).2. After transfected SKOV3cells, fluorescence microscopy revealed a large number of fluorescence. The expression of IEX-1protein in SKOV3-IEX-1cells was significantly higher than SKOV3group and SKOV3-EGFP group (P<0.05), The difference between SKOV3group and SKOV3-EGFP group was not statistically significant (P>0.05).3. PI/AnnexinV double staining showed that the apoptosis rate of SKOV3-IEX-1group was significantly higher than that of SKOV3group and SKOV3-EGFP group under the effect of cisplatin (P<0.05), with a concentration-dependent and time-dependent manner (P<0.05), the remaining between the two groups showed no significant difference (P>0.05).4. The positive expression of IEX-1、NF-κB、p53、Mcl-1in ovarian cancer were36.6%(15/41)、73.2%(30/41)、61.0%(25/41)、56.1%(23/41) respectively (P<0.05). The expression of IEX-1was negatively correlated with Mcl-1、 NF-κB in ovarian cancer,and had no correlation with p53(P>0.05)Conclusion1. UVB can induce the expression of IEX-1in SKOV3cells, with a time-dependent and dose-dependent manner.2. IEX-1can increase the sensitivity to cisplatin of SKOV3cells, with a time-dependent and concentration-dependent.3. In the ovarian cancer, IEX-1may be playing a pro-apoptosis function by counteracting to its upstream regulator of NF-κB and inhibiting anti-apoptosis effection of Mcl-1.