Studies On Cationic Polymer Gene Vector Which Mediatedmirna Transfection

Background:in recent years esophageal cancer is a serious threat to the health of general public. The clinical treatment mainly includes surgery, radiotherapy and chemotherapy nowadays. Esophageal cancer has a very bad prognosis after treatment. Gene therapy has been a hot area of research currently. It is one of tumor therapy that gene vector carry specific gene into target cancer cells efficiently. Firstly of all, we should make sure that specific exogenous genes tightly bound to gene vector effectively. Secondly, the complexes can avoid being degradation by vitro nucleic acid enzyme. Thirdly, the complexes can escape from immune system and enter cancer cell through endocytic pathways. Fourthly, they can avoid be degrading by protease in lysosome. The last and most important, the purpose genes successfully enter cancer cell nucleus and lead to gene silence, and then regulate activity of gene in cells to affect proliferation or migration of cancer cell. As a result, mechanisms of suppressing cancer development involve silencing oncogene, promoting tumor suppressor genes, restraining tumor angiogenesis gene. Gene therapy has some significant advantages, such as high-security, highly cost-effective and time-efficient.With development of biotechnology, new method and thought emerge in endlessly so there were lots of useful results about gene level. And Extensive studies and data analysis in molecular mechanism of tumor etiology promote the application of new technology.MALAT1 is a kind of long chain non-coding RNAand length greater than 200 nucleotides. It is found increase in the metastasis and recurrence of many kinds of cancers. However, miRNA is a kind of non-coding single-stranded RNA measures just about 20-25 nucleotides. For the study, miR-101 and miR-217 have been found down-regulated in many tumors.Objective:our study first unites gene vector PER-PGEA and PER-PGEA-FA (with folic acid ligand) with miRNA to form complex. It is transfected into several kinds of esophageal cancer cells and regulated activity of gene in cells to affect proliferation or migration of cancer cell. By broadening the possibilities of applications in this way, we can provide experimental basis for future clinical use.Methods:we unite gene vectors PER-PGEA and PER-PGEA-FA (with folic acid ligand) with miRNA to form complex. Some of the research contents include enhanced green fluorescent protein experiment and luciferase report gene experiment. They prove that gene vector and DNA are combined well and with effective gene transfection and the low toxicity. The other part result includes conjugation conditions, which let us know strict discipline of the experiment, we have to pay attention and must be careful to the environmental conditions, the equipment, and operational techniques. And then the measurements of the particle size and potential reveal us the optimum condition of conjugation. And then MTT toxicity experiments, which let us understand the toxic effects appearance remains different from one cell type to another. And RT-qPCR make us aware of comparatively accurate influence when complexs entering cells. In general, these results help us understand the ratio of complexes (nitrogen/phosphorus), toxic effects on human cells and the ability of efficiently transfection and expression. And then we use RT-qPCR after transfection of miR-101, miR-217and siRNA-MALAT1 to study influence on target gene relatively. Counting experiments help us understand the complexes affect adversely on proliferation. Invasion experiments help us understand the complexes affect adversely on migration. Scratch experiments let us know the complexes affect adversely on proliferation and migration. Finally, cell clone formation experiments reveal a great deal about proliferation and metastasis of tumor cells.Conclusions:gene vector PER-PGEA and PER-PGEA-FA (with folic acid ligand) combined firmly and transfected into esophageal cancer cell. The primary advantages of compound involve low toxicity, efficient transfection, down-regulating the expression of target lncRNA and inhibition the proliferation, migration and invasion of esophageal cancer cells. Especially, gene vector PER-PGEA-FA is better than PER-PGEA.