The Expression Detection Of EMT Related Proteins And Screening Of Its Differential Expression MicroRNAs In Hepatocellular Carcinoma

Hepatocellular carcinoma (HCC) is the most common maglinant tumor and the second cause of death from cancer in China. The deaths of HCC each year in China accounted for 53% of globals'. Metastasis is the main reason why HCC patients miss the best treatment strategy and have poor prognosis. Therefore, investigating the molecular mechanisms of HCC development and metastasis and finding new prevention and treatment targets of HCC are significant.Metastasis is a complex process that invoves multiple alterations, the specific mechanism remains unclear. Epithelial-mesenchymal transition (EMT) plays an important role in tumor metastasis. EMT is a phenomenon that polarized epithelial cells are converted to motile mesenchymal cells in specific microenvironment, during the process the epithelial proteins E-cadherin is down-regulated and mesenchymal proteins vimentin is up-regulated. The phenotype transition allows tumor cells to overcome the cell-cell connecting and more easily to invasion and metastasis.MicroRNA (miRNA) is a small, noncoding, endogenous RNA gene products (17-25 nucleotides). MiRNAs exist in many organisms and play key regulatory roles in mRNA translation and degradation by base pairing to partially complementary sites of the mRNA, predominantly in the 3'-untranslated region. Recent studies indicate that as important endogenous gene regulator, miRNAs play an important role in tumor metastasis by regulating EMT related genes. In summary, investigating the relationship between EMT and HCC metastasis, finding the miRNAs which may regulate EMT in HCC metastasis are significant. In this dissertation we detect the EMT related proteins E-cadherin and vimentin in 32 HCC tissues using immunohistochemistry. To screen and identify the microRNAs differential expression profile in the metastasis HCC which was E-cadherin(-)/vimentin(+) by the microRNA array technique, for further study on functions of microRNAs in metastasis of HCC.1. Expression of E-cadherin and vimentin in 32 cases of HCCs tissues were examined with immunohistochemistry, and the relationship between the expression and clinicalpathology was analyzed.2. MicroRNA array was used to investigate the differentially expressed microRNAs between E-cadherin(-)/vimentin(+) metastasis HCC and E-cadherin(+)/vimentin(-) no metastasis HCC.3. Real time PCR was applied to verify the reliability of microRNA array results.4. Predicte target genes of the four microRNAs by combination anticipated algorithms.1. The result of immunohistochemistry indicated that low expression of E-cadherin and high expression of vimentin in HCC was significant correlated with metastasis and poor differential of HCC (p<0.05).2. MicroRNA array showed that 8 microRNAs were up-regulated and 28 microRNAs were down-regulated in E-cadherin(-)/vimentin(+) metastasis HCC.3. Compared to E-cadherin(+)/vimentin(-) no metastasis HCC, miR-21,miR-221 were up-regulated and miR-126,miR-199a were down-regulated in E-cadherin(-)/vimentin(+) metastasis HCC.4. The potential targets of miR-21,miR-221,miR-126,miR-199a involves many tumor metastasis biological process such as cell differentiation,oncogene activation, transcription regulation,cell signaling pathway.1. EMT exist in the process of HCC development and metastasis and is closely related with HCC metastasis.2. Commercial miRNA microarray is an effective method for high-throughput screening differential expression miRNAs in tumors. MiRNA expression profile of HCC EMT and metastasis was established.3. MiR-21,miR-221,miR-126,miR-199a have potential regulatory effect in HCC EMT and metastasis.