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Expression profiling and functional analysis on bladder tumor suppressor candidate genes, ANXA10 and CDK2AP1Wong, Chui-wei 16 July 2004 (has links)
Bladder cancer is a common malignancy affecting the genitourinary system. Although a large number of studies have been carried out on these areas for a long time, little is know about the molecular events which may involve in tumorigenesis. Until now, no profound immunohistological or molecular markers have been identified to define clinically relevant subsets of bladder cancer. The purpose of this thesis is to identify a novel bladder cancer carcinogenesis related genes. Chapter 1 attempts to illustrate the background, molecular markers, chromosomal abnormalities and genetic instability related to bladder cancer. In Chapter 2, various bioinformatics methodologies were used to annotate and identify candidate genes. Twenty-one genes were identified 1.5-fold up- or down-regulated in mRNA expression from RT4, TSGH8301 and J82, three different stages of bladder cancer cell lines by microarray chips (Dr. Liu, personal communications). Another eight candidate tumor suppressor genes were preliminarily identified from suppression subtractive hybridization (SSH) cDNA library of RT4 cell line based on an isoflavones-treated minus non-treated and further subjected to quantitative RT-PCR analyses to confirm the mRNA expression level in different stages of bladder cancer cell lines. Chapter 3 studies on the ANXA10 gene with special emphasis on its cloning, protein expression, subcellular localization and the preparation of polyclonal antibody. The result suggests that ANXA10 is a cytoplasmic protein in N18 cells. Chapter 4 analyzes the CDK2AP1 gene in mRNA and protein level at different bladder cancer cell lines and various specimens. In our preliminary observations, there are lost of CDK2AP1 expressions at invasive TCCs specimens when compared to noninvasive TCCs specimens. The mechanism of the tumor-associated loss of the CDK2AP1 expression is currently not clear. In Chapter 5, bladder cancer cell lines TSGH8301, UB37, TCCSUP and J82 in SCID mice xenograft model were established for further in vivo studies.
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