MicroRNA-9Expression Analysis And Functions In Esophageal Squamous Carcinoma Cell Lines

BackgroundWorldwide, esophageal carcinoma is the sixth most common cause of tumor-related death. This disease is endemic in Asia, Southern and Eastern Africa. Although esophageal carcinoma has a low incidence in North America and many Western European countries, there has been an increasing incidence of esophageal adenocarcinoma (AC) over the past three decades in these countries and the corresponding increase in mortality is remarkable. Although surgery is the mainstay of treatment for patients with resectable esophageal cancer, the effectiveness of surgery alone has been unsatisfactory. Early metastasis and postoperative metastasis is the main reason affecting the prognosis of patients with postoperative five-year survival rate of only10-20%. Decrease the morbidity and mortality of patients with esophageal cancer study is very important.Esophageal disease process is accumulation of more than one gene variant, a variety of molecules involved in the complex process, these mechanisms are still poorly understood.MicroRNA(miRNA) are small RNAmolecules around20-25nucleotides in length, which play an important roles in affecting the activity of post transcriptional level, encording by the endogenous genes. They constitute a class of non-protein encoding RNA molecules which have now emerged as key players in regulating the activity of target mRNA, resulting in preventing its translation into protein or directly degrading it.Each miRNA can have a plurality of target genes and several of the miRNA can jointly regulate the same gene.According to the studies,miRNA regulation of about1/3of the humen gene.There are more and more evidence to support that miRNA plays an import role in tumor metastasis,some of microRNA expression in cells can inhibit tumor cell metastasis,some can promote tumor cell metastasis.miRNAs as a class of highly conserved non-coding RNA in cell proliferation,differentiation and apoptosis process plays an important role.It has been well established that the miR-9expression profile in cancer relies on tissue distribution. In primary brain tumors, miR-9is elevated and functions as an essential factor in neural carcinogenesis.miR-9is also over-expressed in cervical cancer,suggesting its tumor promoter role in tumor development and progression. However, miR-9is down-regulated in multiple human cancers, including pancreatic cancer,ovarian cancer,and colorectal cancer. MiR-9is also associated with the malignant progression of breast cancer,and colorectal cancer.Experiments have confirmed that miR-9in gastric cancer was significantly lower expression.However the inhibition of tumor cell invasion and metastasis in esophageal remains unclear.The purpose of this study is to reveal miR-9expression in esophageal and holds it as an candidate biomarker for the early detection and management of cancer.ObjectivemiR-9expression and function in esophageal squamous carcinoma cell lines KYSE30, KYSE70, KYSE140and KYSE150.MethodsDetection miR-9expression levels in4esophageal cancer cell line (KYSE30, KYSE70, KYSE140, KYSE150) and HEEpiC by real-time RT-PCR analysis of experimental methods. MiR-9precursor molecules (pre-miR-9) were transfected in KYSE30cells, change the expression levels of miR-9, then implementing the following function tests. By CCK.8assay cell growth, draw out the growth curve after three days.Analysis of miR-9on cell proliferation. PI staining using flow cytometry analysis of its role in regulating the cell cycle, to use AnnexinV-FITC/PI double staining by flow cytometry apoptosis. Transwell method to detect cell migrationResultsThe expression of miR-9levels decreased,expression level in four esophageal cancer cell line were different detected by real-time RT-PCR analysis of experimental methods. In the function of miR-9study,transfected with Pre-miR-9, the miR-9in KYSE30cell lines were overexpression.Detected cell growth curve compared with the control, the overexpression of cell growth significantly reduced, low expression of cell growth rate increased, miR-9inhibition in cell proliferation. Over-expression of miR-9in KYSE30,compared with the control group,found that miR-9overexpression group proportion of the G1phase of the cell cycle were increased detected by flow cytometry. The AnnexinV FITC/PI double staining by flow cytometry apoptosis, no significant difference between the three groups of apoptosis. Use the Transwell method to detect cell migration,found that overexpression of miR-9can inhibit cell migration ability.ConclusionmiR-9expression reduced in esophageal cancer cell line, overexpression of miR-9can block tumor cells in G1phase, inhibiting the proliferation of esophageal cancer cells, inhibition of tumor cell migration to play its tumor suppressor role.