Effects of over-expressing UDP-glucuronosyltransferase 1A1 on xenobiotic and therapeutic drug metabolism.

• Other Authors: Leung, Hau Yi.
• Format: Others
• Language: English; Chinese
• Published: 2006
• Subjects: Glucuronosyltransferase > Metabolism > Genetic aspects; Xenobiotics > Metabolism > Genetic aspects; Antineoplastic agents > Metabolism > Genetic aspects; Glucuronosyltransferase > genetics; Glucuronosyltransferase > metabolism; Xenobiotics > metabolism; Antineoplastic Agents > metabolism
• Online Access: http://library.cuhk.edu.hk/record=b5892985; http://repository.lib.cuhk.edu.hk/en/item/cuhk-325706

Leung Hau Yi. === Thesis (M.Phil.)--Chinese University of Hong Kong, 2006. === Includes bibliographical references (leaves 116-131). === Abstracts in English and Chinese. === Thesis Committee --- p.in === Acknowledgement --- p.II === Abstract --- p.III === 摘要 --- p.V === Table of Contents --- p.VII === List of Figures --- p.X === List of Tables --- p.XIII === Appendix Abbreviations --- p.XIV === Chapter Chapter 1 --- Introduction --- p.1 === Chapter 1.1 --- Breast Cancer --- p.1 === Chapter 1.2 --- Development of Breast Cancer --- p.2 === Chapter 1.3 --- Risk Factors of Breast Cancer --- p.3 === Chapter 1.3.1 --- Age --- p.3 === Chapter 1.3.2 --- Genetic Factors --- p.4 === Chapter 1.3.3 --- Hormonal Factors --- p.5 === Chapter 1.3.4 --- Lifestyles --- p.6 === Chapter 1.4 --- Drug Metabolism --- p.6 === Chapter 1.5 --- UGT1A1 --- p.7 === Chapter 1.5.1 --- UDP-glucuronosyltransferase --- p.7 === Chapter 1.5.2 --- UGT1A1 --- p.9 === Chapter 1.6 --- Cytochrome P450 I Enzyme Family --- p.10 === Chapter 1.6.1 --- CYP450 subfamily --- p.10 === Chapter 1.6.2 --- CYP1A1 --- p.11 === Chapter 1.6.3 --- CYP1B1 --- p.12 === Chapter 1.7 --- Reasons why UGT1A1 is being studied --- p.13 === Chapter 1.8 --- Outline of this Study --- p.14 === Chapter 1.8.1 --- Effects of Over-expressing UDP-Glucuronpsyltransferase and Cytochrome P450 1A1 Against Xenobiotic Assault in Breast Cancer Cells --- p.14 === Chapter 1.8.2 --- Effects of Genistein and Resveratrol on Phase I and II Enzymes in a Non-cancerous Breast Cell Line --- p.15 === Chapter 1.8.3 --- Effects of UGT1A1 on Cancer Drug Treatment --- p.15 === Chapter Chapter 2 --- Materials and Methods --- p.16 === Chapter 2.1 --- Chemicals --- p.16 === Chapter 2.2 --- Cell Culture --- p.16 === Chapter 2.2.1 --- Maintenance --- p.16 === Chapter 2.2.2 --- Preparation of Cell Stock --- p.17 === Chapter 2.2.3 --- Cell Recovery from Liquid Nitrogen Stock --- p.17 === Chapter 2.3 --- Cloning and Transfection --- p.18 === Chapter 2.3.1 --- Isolation of RNA from cells and cDNA synthesis --- p.18 === Chapter 2.3.2 --- Amplification of UGTlAl --- p.20 === Chapter 2.3.3 --- Separation and Purification of DNA from Agarose Gel --- p.21 === Chapter 2.3.4 --- Restriction Digestion --- p.22 === Chapter 2.3.5 --- Ligation of DNA Fragment and Vector --- p.22 === Chapter 2.3.6 --- Transformation of DH5a --- p.23 === Chapter 2.3.7 --- Small Scale Plasmid Purification (Miniprep) --- p.24 === Chapter 2.3.8 --- Large Scale Plasmid Purification (Maxiprep) --- p.25 === Chapter 2.3.9 --- Stable Transfection into MCF-7 cells with LipofectAMINE PLUS reagent --- p.26 === Chapter 2.4 --- Analytical Procedures --- p.27 === Chapter 2.4.1 --- Western Blot Analysis --- p.27 === Chapter 2.4.2 --- Measurement of cell proliferation (MTT assay) --- p.28 === Chapter 2.4.3 --- Measurement of DMBA-DNA Adduct Formation --- p.28 === Chapter 2.4.4 --- Comet Assay --- p.29 === Chapter 2.4.5 --- Relative Quantitative Real Time PCR --- p.30 === Chapter 2.4.5.1 --- Real Time PCR Using TaqMan Probe --- p.30 === Chapter 2.4.5.2 --- Statistical Analysis of 2-ΔΔCT Comparative Gene Expression --- p.31 === Chapter 2.4.6 --- Flow Cytometry --- p.31 === Chapter 2.4.7 --- EROD Activity in Intact Cells --- p.31 === Chapter 2.4.8 --- High Performance Liquid Chromatography --- p.32 === Chapter 2.5 --- Statistical Analysis --- p.34 === Chapter Chapter 3 --- Effects of Over-Expressing UDP-GIucuronosyltransferase and Cytochrome P450 1A1 Against Xenobiotic Assault in Breast Cancer Cells --- p.35 === Chapter 3.1 --- Introduction --- p.35 === Chapter 3.2 --- Results --- p.38 === Chapter 3.2.1 --- Effectiveness of Transfection --- p.38 === Chapter 3.2.2 --- Cell Proliferation Experiments --- p.41 === Chapter 3.2.3 --- Regulation of Estrogen Receptor (ER) Expression --- p.43 === Chapter 3.2.4 --- Formation of DMBA-DNA adduct formation --- p.45 === Chapter 3.2.5 --- Single Cell Gel Electrophoresis (Comet Assay) of DMBA-induced DNA Damage in MCF-7UGT1A1 cells --- p.46 === Chapter 3.2.6 --- HPLC for Estradiol-glucuronidation Analysis --- p.49 === Chapter 3.2.7 --- Single Cell Gel Electrophoresis (Comet Assay) of DMBA or E2-induced DNA Damage in MCF-7cyp1A1 cells --- p.51 === Chapter 3.3 --- Discussion --- p.56 === Chapter Chapter 4 --- Effects of Genistein and Resveratrol on Phase I and II Enzymes in a Non-Cancerous Breast Cell Line --- p.61 === Chapter 4.1 --- Introduction --- p.61 === Chapter 4.2 --- Results --- p.66 === Chapter 4.2.1 --- "Genistein and Resveratrol Reduced DMBA-induced UGT1A1, CYP1A1 and CYP1B1 Expression" --- p.66 === Chapter 4.2.2 --- Genistein and Resveratrol Reduced the Formation of DMBA-DNA Adduct in MCF-10A Cells --- p.73 === Chapter 4.2.3 --- Genistein and Resveratrol Reduced the Single Strand DNA Damage Generated by DMBA in MCF-10A Cells --- p.76 === Chapter 4.2.4 --- Genistein and Resveratrol Reduced DMBA-induced EROD Activities --- p.81 === Chapter 4.3 --- Discussion --- p.84 === Chapter Chapter 5 --- Effects of Ugtlal on Cancer Drug Treatment --- p.89 === Chapter 5.1 --- Introduction --- p.89 === Chapter 5.2 --- Results --- p.93 === Chapter 5.2.1 --- Cell Proliferation Experiment --- p.93 === Chapter 5.2.2 --- "Expression of Bcl-2 and Bax proteins in Paclitaxel- or VCR-treated MCF-7, MCF-7control and MCF-7UGt1A1 cells" --- p.98 === Chapter 5.2.3 --- Flow Cytometric Analysis of Cell Cycle Phase Distributionin Paclitaxel- or VCR-treated MCF-7 cells --- p.103 === Chapter 5.3 --- Discussion --- p.110 === Chapter Chapter 6 --- Summary --- p.114 === Bibliography --- p.116