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Functional characterization of GEF-H1 in liver tumorigenesis.

Tsang, Chi Keung. / "November 2011." / Thesis (M.Phil.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 103-116). / Abstracts in English and Chinese. / Abstract --- p.I / 摘要 --- p.III / Acknowledgement --- p.IV / Table of content --- p.V / List of Figures --- p.VIII / List of Tables --- p.XI / Abbreviations --- p.XII / Chapter Chapter 1: --- INTRODUCTION --- p.1 / Chapter 1.1. --- Hepatocellular carcinoma --- p.2 / Chapter 1.1.1. --- Etiological factors --- p.11 / Chapter 1.1.1.1. --- Chronic Hepatitis and Liver Cirrhosis --- p.13 / Chapter 1.1.1.2. --- HBV --- p.13 / Chapter 1.1.1.3. --- HCV --- p.17 / Chapter 1.1.1.4. --- Male gender --- p.20 / Chapter 1.1.1.5. --- Aflatoxin B1 exposure --- p.21 / Chapter 1.2. --- Genomic abnormalities in HCC --- p.23 / Chapter 1.3. --- GEF-H1 --- p.24 / Chapter 1.4. --- RhoA --- p.26 / Chapter 1.5. --- Epithelial-Mesenchymal Transition (EMT) --- p.29 / Chapter 1.6. --- Aims of Thesis --- p.31 / Chapter Chapter 2: --- MATERIALS AND METHODS --- p.32 / Chapter 2.1. --- Materials --- p.33 / Chapter 2.1.1. --- Chemicals and Reagents --- p.33 / Chapter 2.1.2. --- Buffers --- p.35 / Chapter 2.1.3. --- Cell Culture --- p.37 / Chapter 2.1.4. --- Nucleic Acids --- p.38 / Chapter 2.1.5. --- Enzymes --- p.39 / Chapter 2.1.6. --- Equipments --- p.40 / Chapter 2.1.7. --- Kits --- p.41 / Chapter 2.1.8. --- Antibodies --- p.42 / Chapter 2.1.9. --- Software and Web Resources --- p.43 / Chapter 2.2. --- Fluorescence In Situ Hybridization (FISH) --- p.44 / Chapter 2.2.1. --- Probe Preparation --- p.44 / Chapter 2.2.1.1. --- Human Bacterial Artificial Chromosome (BAC) probe preparation --- p.44 / Chapter 2.2.1.2. --- Nick translation --- p.44 / Chapter 2.2.2. --- Hybridization --- p.45 / Chapter 2.3. --- Genomic DNA extraction --- p.47 / Chapter 2.4. --- Copy number analysis --- p.48 / Chapter 2.5. --- Exon Sequencing analysis --- p.49 / Chapter 2.5.1. --- PCR amplification of GEF-H1 exons --- p.49 / Chapter 2.5.2. --- Cycle sequencing --- p.49 / Chapter 2.6. --- Ectopic expression of GEF-H1 in immortalized hepatocyte cell line --- p.52 / Chapter 2.6.1. --- Construction of GEF-H1 expressing vector --- p.52 / Chapter 2.6.2. --- Sub-cloning --- p.52 / Chapter 2.6.3. --- Transfection and clonal selection --- p.53 / Chapter 2.7. --- Gene Expression Analysis by Quantitative RT-PCR --- p.55 / Chapter 2.7.1. --- Total RNA extraction --- p.55 / Chapter 2.7.2. --- qRT-PCR analysis for gene expression --- p.55 / Chapter 2.8. --- Western blot --- p.58 / Chapter 2.9. --- Functional Analysis --- p.60 / Chapter 2.9.1. --- Cell viability (MTT) assay --- p.60 / Chapter 2.9.2. --- Cell proliferation assays (BrdU-incorporation) --- p.60 / Chapter 2.9.3. --- Mitomycin C treatment --- p.61 / Chapter 2.9.4. --- Migration and Invasion assays --- p.63 / Chapter 2.9.5. --- Wound healing assay --- p.65 / Chapter 2.9.6. --- Transient knock-down of RhoA --- p.65 / Chapter 2. --- 10. Immuno-fluorescent imaging --- p.66 / Chapter 2. --- 11. In vivo tumorigenic study of GEF-H1 by subcutaneous injection --- p.68 / Chapter 2. --- 12. Statistical analysis --- p.69 / Chapter Chapter 3: --- RESULTS --- p.70 / Chapter 3.1. --- Verifying copy number gain of GEF-H1 in high GEF-H1 expressing HCC --- p.71 / Chapter 3.2. --- Verifying if there is any GEF-H1 exon point mutation in HCC --- p.75 / Chapter 3.3. --- Functional roles of GEF-H1 in HCC --- p.77 / Chapter 3.4. --- GEF-Hl-induced functions were RhoA independent --- p.83 / Chapter 3.5. --- GEF-H1 Induction of Epithelial-mesenchymal transition in HCC --- p.88 / Chapter 3.6. --- GEF-H1 induced tumorigenicity of MIHA cells --- p.95 / Chapter Chapter 4: --- DISCUSSIONS --- p.96 / Chapter 4.1. --- GEF-H1 in HCC and other cancers --- p.97 / Chapter 4.2. --- GEF-H1 promotes cell motility --- p.98 / Chapter 4.3. --- GEF-H1 induced tumorigenicity --- p.100 / Chapter Chapter 5: --- CONCLUSIONS AND PROPOSED FUTURE INVESTIGATIONS --- p.101 / Chapter Chapter 6: --- REFERENCES --- p.103

Identiferoai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_327615
Date January 2012
ContributorsTsang, Chi Keung., Chinese University of Hong Kong Graduate School. Division of Anatomical and Cellular Pathology.
Source SetsThe Chinese University of Hong Kong
LanguageEnglish, Chinese
Detected LanguageEnglish
TypeText, bibliography
Formatprint, xiv, 116 leaves : ill. (some col.), maps ; 30 cm.
RightsUse of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/)

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