| Background and ObjectiveCholangiocarcinoma, (CCA) is a kind of gastrointestinal tumor originated from epithelial cells of bile duct malignant, in human malignant tumor of digestive tract in total cases accounted for about 2%～3%. In 1840 Durand Fardel first reported in bile duct carcinoma, liver and biliary system malignant tumor is common in a, accounted for hepatobiliary neoplasme 10%～15%. Bile duct cancer incidence in the world rate of about 1.2/10 million, but nearly ten years of bile duct cancer incidence trend for increased year by year, its incidence in China rising velocity is the annual increase of 5%, rising the fastest of the digestive tract tumor. As a result of bile duct cancer specific anatomic location and to the surrounding tissue, blood vessels and nerve infiltration of the special pathological feature and many other reasons, led to the lack of early clinical manifestations in patients with bile duct cancer specific and effective means of diagnosis, many patients have been in the most advanced cancer that has metastasized, and lymph node metastasis or distant metastasis, although in recent years, surgery, chemotherapy, radiotherapy and other treatment methods have made great progress in the treatment of bile duct cancer, but it is still difficult to achieve radical tumor effect. The resection rate is low, is not sensitive to conventional chemotherapy means, high mortality rate is the current status of bile duct cancer treatment, patients in the world scope of bile duct cancer postoperative 3 years survival rate is only 35% to 50%,5 year survival rate is less than 10%, in our country the bile duct cancer postoperative 5 years survival rate is only about 5%. Now the treatment of bile duct cancer is surgical resection for the surgical treatment of choice, surgical resection can not only achieve radical results, recurrence within 2 years and a lot of complete surgical patients after operation, all of these seriously restrict the effect of surgery and prognosis of patients. Therefore, the study of bile duct cancer occurrence, development, invasion and metastasis mechanisms and possible molecular steps to understanding the bile duct cancer progression process, looking for the treatment and prognosis of clinical analysis of bile duct cancer patients to provide suitable target is very necessary.Wild type P53 induced protein phosphatase 1 (Wipl) is a serine/threonine enzyme activity of the newly discovered protein phosphatase, as a proto oncogene, is located on human chromosome 17q22/q24 area, it is highly expressed in a variety of tumors, and can promote tumor growth, and closely related with the prognosis of the patients. But there is no research on the expression of Wip1 gene in human cholangiocarcinoma cells in.RNA interference (RNAi) is a kind of resisting the foreign gene and virus infection conservative evolutionary mechanism widely found in living, is a research method of post transcriptional gene regulation, the nature of its mechanism of action is the intracellular mRNA and siRNA specific binding, mediated by RISC in order to target degradation in combination with mRNA, stop the translation function, leading to gene silencing, with high effectiveness and targeting, is caused by the mechanisms of gene silencing at the mRNA level. Because the RNAi is fast, high efficiency, high specificity and other advantages, each field of the technology has been widely applied in the identification of gene function and so on life science.This study is to design and synthesize can silence Wipl gene expression siRNA, transfection of human bile duct carcinoma cell line QBC939 transfected cholangiocarcinoma, the influence of later generations the biological behavior of QBC939 cells.MethodsDesign and synthesize specific siRNA, which can inhibit the expression of Wipl gene. Transfect QBC939 cells with siRNA using lipofectamine2000 and experiment were divided into experimental group and control group. The protein and mRNA expression level of Wipl gene were studied using Western blot and qRT-PCR. The proliferation of QBC939 cells was detected using CCK-8 and cell migration and invasion capability were tested by scratch test and Transwell assay.ResultsThe expression of Wipl protein and mRNA were significantly decreased in siRNA group compared with the control group (P<0.05). CCK-8 testing results showed that cell proliferation was reduced compared with untransfected group after three days transfection(P<0.05)and significantly reduced after four days transfection (P<0.01). Transwell results show that cell number were significantly less than that of non transfected group (P<0.05)ConclusionSpecific siRNA was successfully designed and can effectively reduce the protein and mRNA expression level of Wipl, which can inhibit the growth of QBC939 cells and reduced cell migration and invasion. |
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