Silence Gene ZNF217 Expression In HO-8910 Cells Of Ovarian Cancer

BACKGROUND&OBJECTIVEOvarian cancer is a threat which is one of the common malignancies to women'shealth. The five-year survival rate is about 30% of patients in old age. Ovarian canceris more than 90% of origin of ovarian surface epithelium. Currently epithelial ovariancancer etiology and pathogenesis is not yet very clear. With development of cancergenetics and molecular biology technology, pathogenesis of ovarian cancer have hadextensive and in-depth study.It is recognizated that the abnormal genetic material isthe main reason for tumor formation.Chromosome abnormalities are found widely in ovarian cancer genomic byhybridization technique comparative, gene amplification have higher probability inthe chromosome 20, particularly 20ql 3.2, the site have gene ZNF217.ZNF217 gene is at chromosome 20q13.2 which is newly cloned cancer gene on the20th chromosome, Krupple like transcription factor which coded belongs to zincfinger protein family. An increasing number of studies show that at present, zincfinger protein family members play an important role in the development process of avariety of cancers. It was proved that ZNF217 restrains the role of transcription through mutual reactionbetween PXDLS sequences (At the combination of gullies of the C-terminal-bindingprotein-binding domain),RRT sequences(Combining in domain of the surface of theditch of the C-terminal-binding protein nucleotide-binding domain) and theC-terminal-binding protein. With the increase of the copy number of ZNF217, geneexpression changes accordingly, for example, inhibiting the expression of tumorsuppressor gene promoter can lead to the occurrence of tumor. Peixiang Li etc.thinkthat ZNF217 has the following main role in the occurrence of tumors: Low the levelof expression of certain tumor suppressor, raising the level of expression ofoncogenes. Gene ZNF217 can induce anchorage-independent and serumindependent which means it can activate autocrine of growth-promoting Factor, andfound that the difference expression of ZNF217 in different cells has a relation of thestation of p53 and pRB.ZNF217 gene in esophageal squamous cell carcinoma, gastric cancer, prostate cancer,colorectal cancer study, is believed to be involved in occurrence and developmentprocess of cancer. ZNF217 gene is one of the two promoting factors for breast cancer.Tanner have detected that ZNF217 gene is amplification and higher expression inovarian cancer patients, However, It is limited it's structural change, the papersrelated to its current function are not reported.This study focused on the espression of the ZNF217 gene in ovarian cancer; toconstruct the shRNA expression vector of ZNF217 and inhibit the expression ofZNF217 through RNA interference in ovarian cancer cell lines HO—8910. METHODS1. ZNF217 gene expression in ovarian cystadenocarcinomaThe expression of ZNF217 was examined in specimens of 30 cases of ovariancystadenocarcinoma and matching the seven normal cases and 30 cases of ovariancystadenoma ovarian tissue by immunohistochemistry(S-P) method.2. Silence gene ZNF217 in ovarian carcinoma cell lineDesign ZNF217 gene fragment interference, recombinant Plasmid vector pGenesil/ZNF217shRNA.The pGenesil/ 217shRNA was transfected into epithelial ovariancancer cell line HO-8910 by cationic lipids Lipofectaming^TM2000. After resistancemonoclonal selected with G-418, picked a strong fluorescent clones expandingculture Quantitative, and by RT-PCR and Western blot detect expression of ZNF217gene in the cells after interference.3. STATISTICAL ANALYSISNonparametric Kruskal-Wallis test was used in the immunohistochemicalexperiment.RESULT1. Expression of ZNF217 protein in ovarian Cystadenocarcinoma.Immunohistochemistry results showed ZNF217 in normal ovarian tissue.cystadenoma and cystadenocarcinoma have three different types of tissue expression(x²=15.822, p=0.000). Advanced stage Cystadenocarcinoma ZNF217 expressedstronger than earlier period ones. (x²=5.573, p=0.018).2. Down-regulation of ZNF217 expression by RNAi We constructed PGenesil Plasmid vector system that expressed short hairpin RNAsthat were targeted against gene ZNF217.It is confirmed by sequencing results. ThepGenesil pGenesil/ ZNF2171 shRNA and pGenesil/ ZNF2172 shRNAcells weretransfected intointroduced HO-8910 cells by Liposome technology. The follow-upexperiment will adopt pGenesil/ZNF2171 shRNA which have higher transfectionrate. After transfection, resistance monoclonal was selected with G-418. We foundclone 2 (named HO-8910/ZNF217-)exhibited a dramatic knock down of ZNF217mRNA and proteinexpression (86%) by quantitative RT-PCR and Western Blot.CONCLUSIONGene ZNF217 is closely related to ovarian cancer.It is success that the gene ZNF217was knock down in ovarian cancer cells, to lay the foundation for follow-upexperiments.