The Expression Of Miz-1 In Esophageal Cancer Tissue And Cell Line And Its Biological Behavior

Background:Esophageal cancer is one of the most common cancers which originates in the esophageal mucosal epithelium. Esophageal cancer ranks globally as fifth to eighth for morbidity and fourth to sixth for mortality [29–31] in China. Two-hundred-thousand people die from esophageal cancer every year.Tumorigenesis results from various kinds of dysregulation of oncogenes and tumor suppressors that influence cellular proliferation, differentiation, apoptosis and senescence. The transcription factor Miz-1(Myc-interacting zinc finger protein 1; Zbtb17)can either activate or repress gene expression in concert with binding partners including the Myc oncoprotein in several tumor types. Known target genes of these complexes encode the cyclin-dependent kinase inhibitors such as cdkn2b(p15) and cdkn1a(p21). Here we show that deletion of Miz-1 expression, through sh RNA in a lentiviral vector, influences various biological processes in two kinds of esophageal carcinoma cell lines to discuss its behavior and function in signal pathway in esophageal cancer.It is possible for Miz-1 to be a alternative target of gene therapy or provide a new treatment method for clinical treatment of esophageal cancer.Objective:We built that two kinds of esophageal carcinoma cell lines deletion of Miz-1 expression through sh RNA in a lentiviral vector to discuss its biological behavior such as cellular proliferation, apoptosis, cycle and migration.Through testing related the proteins and c DNA expression,we know more about the mechanism of biological functions and role of signal pathway which Miz-1particaptes in.This is for the certain theoretical basis of Miz-1 to be a target gene in clinical treatment in the further research.We commission Shanghai Innovation Biotechnology Co.Ltd.to build one special lentivirus of Miz-1 target gene sh RNA.Choosing two esophageal cancer cell lines to be the target gene transfection group in seven samples.Dividing them into three groups of Miz-1 transfecteion group(Miz-1),empty vector transfection group(vector) and non-treat control group(control). Methods:Detecting different expression of Miz-1 in esophageal cancer tissue and juxtacancerous tissues through RT-PCR and immunohistochemical technique.Using immunofluorescence,RT-PCR and Western blot technique for validation of lower expression of target gene Miz-1 and testing related proteins and c DNA expression. CCK8 technology and clonogeicity test were for testing cell proliferation ability. Flow cytometry instrument detection by using the grouping cell cycle and cell apoptosis. Using transwell Chambers to detect cell migration and invasion ability of groups. Use SPSS17.0 statistical software for the data from the experiment method for statistical analysis. Results:1, Miz-1 expressed differences exist in the carcinoma tissue and juxtacancerous tissue, and expressed higher in carcinoma tissue compared with juxtacancerous.2, Eca109 and Kys-150 were the selected two kinds of cell lines in seven alterative esophageal cancer cell lines.3, Lower expression of Miz-1 esophageal cancer cell lines were build through lentivirus contain Miz-1 sh RNA.4, Without Miz-1, p21 was expressed at high levels and reduced downstream cyclin D1, leading to cell cycle arrest.Apoptosis related proteins Caspase 3 and PARP are expressed higher compared with control groups.5, CCK8 and clone formation assay result indicates that without Miz-1 cell proliferation was limited. Flow cytometry results showed that the cell cycle was arrested in G0 / G1 phase and increased apoptosis in lower expression Miz 1 transfection group.Transwell migration and invasion experiment results indicate: the migration and invasion ability of treatment group were weaker than empty vector group and control group. Conclusion:The research confirmed that the Miz-1 express higher in esophageal cancer tissue than in juxtacancerous tissue. This indirect illustrates the Miz-1 may play a positive role in esophageal cancer. We knocked down the expression of Miz-1 by sh RNA through lentivirus in Eca109 and Kys-150. Experiments confirmed that the lower expression of Miz-1 can suppress the esophageal cancer cell proliferation by limiting the cell cycle. Apoptosis related proteins expression promote apoptosis in the absence state of Miz-1.At the same time migration and invasion ability also declined. Miz-1 combines or not with Myc oncoprotein to inhibit downstream p21 tumor-suppressor protein expression, which liberate the arrested of cyclin D1. Miz-1 whether combined with Myc play a role in esophageal cancer was not verified in our researches.The further research is needed in exploration for mechanism in esophageal cancer of Miz-1as a cancer gene. This can provide the clinical diagnosis and treatment of esophageal cancer with more options.