The Effect And Mechanism Of APC Expression In The Growth And Development Of Epithelial Ovarian Carcinoma

Part1The relationship of APC methylation and expression and thegrowth and development of epithelial ovarian carcinomaObjective: To detect the level of APC expression and methylation in thenormal ovarian tissue, benign epithelial ovarian tumor, borderlineepithelial ovarian tumor and epithelial ovarian carcinoma, to analysis therelationship of APC expression and promotor methylation anddevelopment of epithelial ovarian carcinoma.Methods1. The expression of APC in the normal ovarian tissue, benign epithelialovarian tumor, borderline epithelial ovarian tumor and epithelial ovariancarcinoma was examined by immunohistochemical staining.2. The protein expression of APC in the normal ovarian tissue, benignepithelial ovarian tumor, borderline epithelial ovarian tumor and epithelial ovarian carcinoma was examined by western blot analysis.3. The methylation of APC gene in the normal ovarian tissue, benignepithelial ovarian tumor, borderline epithelial ovarian tumor and epithelialovarian carcinoma was examined by MSP.4. Analysis the association of APC protein expression withclinicopathological characteristics in epithelial ovarian carcinoma.Results1. Immunohistochemical staining showed that APC was not expressed or ata low level in EOC compared with samples from normal ovary, benignepithelial ovarian tumor and borderline epithelial ovarian tumor（P<0.05）.2. Western blot analysis showed that APC protein was low expressed inEOC compared with samples from normal ovary, benign epithelial ovariantumor and borderline epithelial ovarian tumor（P<0.05）.3. MSP showed that Promoter methylation of APC genes was high in EOCcompared with samples from normal ovary, benign epithelial ovariantumor and borderline epithelial ovarian tumor.4. Expression of APC was significantly associated with advanced FIGOstage and poor grade of EOC Conclusion1. Low expression of APC is associated with the development of epithelialovarian carcinoma.2. The abnormal promoter methylation possibly leads to low expression ofAPC, and furthermore result in the development of epithelial ovariancarcinoma. Objective: To observe the expression and promoter methylation ofAPC in ovarian cancer cell lines treated with5-Aza-2′-deoxycytidine;at same time, observe the effect on the cell cycle, apoptosis and invasion and the expression of relevant protein in Wnt signal pathway.Methods1. Screening tests: the protein expression of APC in ovarian cancercells was examined by western blot analysis to screen two cells wit h low expression of APC protein.2. Determination of5-Aza-2′-deoxycytidine concentration: select suitableconcentration of5-Aza-2′-deoxycytidine by MTT analysis.3. After ovarian cancer cell lines treated with5-Aza-2′-deoxycytidine for24hours, the protein expression of APC was examined by western blotanalysis, and mRNA expression of APC was examined by RT-PCR, thepromoter methylation of APC was examined by MSP.4. After ovarian cancer cell lines treated with5-Aza-2′-deoxycytidine, thecell cycle and apoptosis was observed by flow cytometry technology; theability of invasion were examined by transwell.5. After ovarian cancer cell lines treated with5-Aza-2′-deoxycytidine, theexpression of relevant protein in Wnt signal pathway was examined bywestern blot analysis.Results1. Western blot analysis showed that APC protein was low expressed inovarian cancer cells, and SKOV3, OVCAR-3was screened for followingexperiment.2.1.0uM was determined as suitable concentration of5-Aza-2′-deoxycytidine by MTT analysis. 3. After ovarian cancer cell lines treated with5-Aza-2′-deoxycytidine,compared with contral, the protein and mRNA expression of APC wasincreased（P <0.05）, and the promoter methylation of APC was downregulated.4. After ovarian cancer cell lines treated with5-Aza-2′-deoxycytidine,compared with contral, the apoptosis rate was increased obviously（P <0.05）; the proportion of the cells in G2/M was reduced obviously（P <0.05）; the numbers of invasion were reduced obviously（P <0.05）.5. After ovarian cancer cell lines treated with5-Aza-2′-deoxycytidine,compared with contral, the expression of β-catenin and cyclinD1were notvaried obviously（P>0.05）, while the expression of p-β-catenin wereincreased and C-myc were decreased obviously（P <0.05）.Conclusion1. The abnormal promoter methylation leads to low expression of APC,and furthermore results in the development and progress of epithelialovarian carcinoma.2. The abnormal promoter methylation of APC and low protein andmRNA expression were related with multiplication and invasion of ovarian caner cells.3. The effect of APC in inhibiting the development and metastasis ofepithelial ovarian carcinoma may possiblely via Wnt signal pathway.4. APC is a novel and crucial predictor in inhibiting EOC developmentand metastasis.