Study On The Effects Of HER-2/neu-shRNA On Apoptosis And Sensitivity To Cisplatin Of Endometrial Cancer Cell

PrefaceEndometrial cancer is one of the common malignancy in reproductive system of woman.It is not very sensitive to chemotherapy drugs.The molecular modulation of endometfial cancer is still unknown.Many scholars have proven that there are about 30%～40%of endometfial cancer which show higher amplification and overexpression of HER—2/neu.HER—2/neu can be called "C-erbB2" gene,it can encode a glycoprotein of 185kD and is extremely similar to EGFR.It is relative with the development of human,tumors by gene amplification and the overexpression of protein production.In addition,overexpression of HER-2/neu increase transfer capacity of cancer cells by means of start lots of metastasis-related mechanisms.Besides,we have discovered in transfection studies and clinical observation that overexpression of HER-2/neu changes the chemosensitivity to hormone and chemotherapeutic drugs.In conclusion,the overexpression of HER-2/neu protein plays a important role in the occurrence and development of Endometrial cancer,and might become a index to judge the outcome.Therefore,HER-2/neu can become meaningful therapeutic target in tumor with overexpression of HER-2/neu.Nowadays,the technique of RNA interference becomes a new heated point of gene therapy.RNAi is a new credible method for inhibition of gene expression.The short double strands RNA interferences the target gene expression specifically and the mRNA were degraded.A unique phenotype which lack of a unique gene was induced in the cell.The inhibition of the gene expression is at the RNA grade.RNA interference,higher transfection efficacy, stronger gene block,high stability,easy taking to the cell,was used as a new gene therapy which target on the HER-2/neu. ObjectOur experiment want to use shRNA to interfere the expression of HER—2/neu in Ishikawa cells effectly aims to explore a new method of gene treatment for endometrial cancer.MethodsAccording to gene recombination technology,We built two specific recombinant plasmids pSilener4.1-HER-2/neu-shRNA1,2 which with the DNA sequence corresponding to shRNA and one nonspecific recombinant plasmid,then these three recombinant plasmids were transfected into Ishikawa cell using liposomes.The expression of HER-2 mRNA of three groups cell after transfection was detected by RT-PCR technique.The apoptosis of the three group transfected cells was detected by Annexin V technique.MTT technique was used to detect the chemosensitivity to cisplatin of the three groups cells.Results24h,48h,72h after transfected,RT-PCR results showed that compared with control group,the expression of HER-2 mRNA of transfected HER-2 shRNA1 and HER-2 shRNA2 groups was significantly decrease.24h,48h,72h after transfected,The apoptosis of the two groups cells which transfected HER-2 shRNA1 and HER-2 shRNA2 were significantly increase compared with control group,there were statistical significance difference(p≤0.01).MTT results showed that:24 hours after exposured to cisplatin, the growth inhibition rate of three groups of cell increasing with the increased concentration of cisplatin,but the group which transfected with HER-2/neu shRNA1 increased more significantly,there were statistical significance difference(p≤0.05). The chemosensitivity of transfected cells to cisplatin significantly increased.ConclusionAfter transfected specific recombinantplasmids,the expression of HER-2 mRNA and Cell proliferation rate are Significantly lower.The chemosensitivity to cisplatin is Significantly increased.Finally we had succeededly proven that:transfecting specific recombinant plasmids would effectively inhibite proliferation of Endometrial cancer Ishikawa cell and increase chemosensitivity to cisplatin.