Pan, Tak Yin. / Thesis submitted in: November 2007. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references. / Abstracts in English and Chinese. / Acknowledgements --- p.i / Abstract (English) --- p.ii / Abstract (Chinese) --- p.iv / Content --- p.v / Abbreviations --- p.x / List of Figures --- p.xi / List of Tables --- p.xii / Chapter Chapter 1: --- Introduction --- p.1 / Chapter 1.1 --- Alzheimer's Disease --- p.1 / Chapter 1.1.1 --- Cholinergic hypothesis --- p.2 / Chapter 1.1.2 --- p-amyloid hypothesis --- p.2 / Chapter 1.1.3 --- Taupathy hypothesis --- p.3 / Chapter 1.1.4 --- Current therapies --- p.4 / Chapter 1.2 --- Proteins Involved in Alzhemer's Disease --- p.5 / Chapter 1.2.1 --- Acetylcholinesterase (AChE) --- p.5 / Chapter 1.2.2 --- p-amyloid --- p.6 / Chapter 1.2.3 --- Paired helical filaments (PHF) --- p.7 / Chapter 1.2.4 --- Protein kinases --- p.8 / Chapter 1.2.4.1 --- Glycogen synthase kinase-3 (GSK-3) --- p.9 / Chapter 1.2.4.2 --- Cyclin-dependent kinase-5 (CDK-5) --- p.9 / Chapter 1.2.5 --- Protein phosphatase (PP) --- p.10 / Chapter 1.2.5.1 --- Protein phosphatase 1 (PP-1) --- p.11 / Chapter 1.2.5.2 --- Protein phosphatise 2A (PP-2A) --- p.12 / Chapter 1.2.5.3 --- Protein phosphatise 2B (PP-2B) --- p.13 / Chapter 1.2.6 --- Apoptotic and Anti-apoptotic proteins --- p.14 / Chapter 1.2.6.1 --- Caspase-3 --- p.15 / Chapter 1.2.6.2 --- Bcl-2 --- p.15 / Chapter 1.3 --- Flavonoids --- p.16 / Chapter 1.3.1 --- Biosynthesis of flavonoids --- p.17 / Chapter 1.3.2 --- Biological functions of flavonoids in plants --- p.18 / Chapter 1.3.3 --- Beneficial effects of flavonoids on human health --- p.19 / Chapter Chapter 2: --- Materials and Methods --- p.20 / Chapter 2.1 --- Differentiation of SHSY-5Y cells --- p.20 / Chapter 2.1.1 --- SHSY-5Y cell culture --- p.20 / Chapter 2.1.2 --- Counting cells --- p.20 / Chapter 2.1.3 --- Retinoic acid differentiation --- p.21 / Chapter 2.2 --- Western blot analysis --- p.21 / Chapter 2.2.1 --- Extraction of proteins from mammalian cells --- p.21 / Chapter 2.2.2 --- Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) --- p.22 / Chapter 2.2.3 --- Semi-dry protein transfer to nitrocellulose membrane --- p.23 / Chapter 2.2.4. --- Membrane blocking and immunostaining --- p.24 / Chapter 2.3 --- MTT assay --- p.25 / Chapter 2.4 --- Hoechst 33342 Nuclei staining --- p.25 / Chapter 2.5 --- Cell cycle analysis --- p.25 / Chapter 2.5.1 --- Ethanol fixation --- p.25 / Chapter 2.5.2 --- Propidium iodide staining --- p.26 / Chapter 2.6 --- Annexin V-FITC & Propidium iodide staining --- p.26 / Chapter 2.7 --- DNA fragmentation analysis --- p.26 / Chapter 2.7.1 --- Phenol/Chloroform extraction of DNA --- p.26 / Chapter 2.7.2 --- Ethanol precipitation of DNA --- p.27 / Chapter 2.7.3 --- Agarose gel electrophoresis of DNA --- p.27 / Chapter 2.8 --- Proteomic analysis --- p.28 / Chapter 2.8.1 --- First dimension: isoelectric focusing --- p.28 / Chapter 2.8.2 --- Second dimension: SDS PAGE --- p.29 / Chapter 2.8.3 --- Gel staining --- p.30 / Chapter 2.8.3.1 --- Silver staining --- p.30 / Chapter 2.8.3.2 --- SYBRO Ruby staining --- p.31 / Chapter 2.8.4 --- Gel scanning and image analysis --- p.31 / Chapter 2.8.5 --- ln-gel digestion --- p.32 / Chapter 2.8.6 --- Zip Tip for desalting the digested sample --- p.33 / Chapter 2.8.7 --- Protein identification with mass spectrometry and database search --- p.33 / Chapter Chapter 3: --- Characterization of Okadaic acid-induced tail hyperphosphorylation in SHSY-5Y cells --- p.35 / Chapter 3.1 --- Introduction --- p.35 / Chapter 3.2 --- Objectives --- p.37 / Chapter 3.3 --- Results --- p.38 / Chapter 3.3.1 --- Differentiation of SH-SY5Y cell --- p.38 / Chapter 3.3.2 --- Changes of protein expression after okadaic acid treatment --- p.40 / Chapter 3.3.3 --- Neurite Retraction Induced by okadaic acid --- p.42 / Chapter 3.3.4 --- Okadaic acid-induced Cell Death measured by MTT assay --- p.44 / Chapter 3.3.5 --- Hoechst 33342 Nuclei Staining --- p.44 / Chapter 3.3.6 --- Cell cycle analysis by propidium iodide staining --- p.47 / Chapter 3.3.7 --- Early Apoptotic cells detection by Annexin V/PI staini --- p.49 / Chapter 3.3.8 --- DNA fragmentation --- p.51 / Chapter 3.4 --- Discussion --- p.53 / Chapter Chapter 4: --- Flavonoids screening for protecting neuronal death by preventing tau hyperphosphorylation --- p.57 / Chapter 4.1 --- Introduction --- p.57 / Chapter 4.2 --- Objectives --- p.58 / Chapter 4.3 --- Tested flavonoids --- p.59 / Chapter 4.4 --- Results --- p.60 / Chapter 4.4.1 --- Toxicity of flavonoids --- p.60 / Chapter 4.4.2 --- Effects of flavonoid pre-treatment on OA-induced neu retractions and cell death --- p.62 / Chapter 4.4.3 --- Western blot analysis --- p.65 / Chapter 4.4.4 --- The effect of different concentrations of hesperidin or OA treatment --- p.70 / Chapter 4.4.5 --- Proteomic analysis --- p.74 / Chapter 4.5 --- Discussion --- p.78 / Chapter Chapter 5: --- General Discussion --- p.82 / References
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_326208 |
| Date | January 2008 |
| Contributors | Pan, Tak Yin., Chinese University of Hong Kong Graduate School. Division of Biochemistry. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, bibliography |
| Format | print, xiii, 86 leaves : ill. ; 30 cm. |
| Rights | Use 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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