Yeung Sau-Man. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2002. / Includes bibliographical references (leaves 139-172). / Abstracts in English and Chinese. / Title page --- p.i / Acknowledgements --- p.ii / Table of Content --- p.iv / List of Figures --- p.viii / List of Graphs --- p.x / List of Tables --- p.xi / Abbreviations --- p.xiv / Abstract --- p.xv / Chinese Abstract --- p.xvii / Chapter Chapter 1 --- General Introduction --- p.1 / Chapter 1.1 --- Diabetes Mellitus --- p.2 / Chapter 1.1.1 --- Type 1 diabetes mellitus --- p.3 / Chapter 1.1.2 --- Type 2 diabetes mellitus --- p.5 / Chapter 1.1.3 --- Maturity onset diabetes of the young (MODY) --- p.6 / Chapter 1.1.4 --- Gestational diabetes --- p.7 / Chapter 1.2 --- Effect of Diabetes on Pregnancy --- p.9 / Chapter 1.3 --- Suggested Causes of Diabetic Embryopathy --- p.10 / Chapter 1.3.1 --- Glucose --- p.10 / Chapter 1.3.2 --- Ketone bodies --- p.11 / Chapter 1.3.3 --- Somatomedin inhibitors --- p.12 / Chapter 1.3.4 --- TNF-α --- p.12 / Chapter 1.3.5 --- Oxidative stress --- p.13 / Chapter 1.4 --- Animal Model of Diabetes --- p.15 / Chapter 1.4.1 --- Chemically-induced --- p.15 / Chapter 1.4.2 --- Mutants --- p.17 / Chapter 1.5 --- Gene-teratogen Interaction under Diabetic Pregnancy --- p.19 / Chapter 1.6 --- Strategy of the Thesis --- p.21 / Chapter Chapter 2 --- General Materials and Methods --- p.24 / Chapter 2.1 --- Mouse Maintenance and Mating Method --- p.25 / Chapter 2.2 --- Induction of Diabetes --- p.25 / Chapter 2.3 --- Preparation of All-trans Retinoic Acid --- p.26 / Chapter 2.4 --- Dissection of Embryos --- p.26 / Chapter 2.5 --- DNA Extraction from Yolk Sac for Genotyping --- p.27 / Chapter 2.6 --- Genotyping of Embryos --- p.28 / Chapter 2.7 --- Preparation of RNA Probes for In Situ Hybridization --- p.29 / Chapter 2.7.1 --- Mini-scale preparation of plasmid DNA --- p.29 / Chapter 2.7.2 --- Linearization of plasmid DNA --- p.30 / Chapter 2.7.3 --- In vitro transcription --- p.31 / Chapter 2.8 --- Whole Mount In Situ Hybridization --- p.33 / Chapter 2.8.1 --- Fixation and dehydration of embryos --- p.33 / Chapter 2.8.2 --- Hybridization --- p.33 / Chapter 2.8.3 --- Post-hybridization wash --- p.34 / Chapter 2.8.4 --- Antibody wash and color development --- p.35 / Chapter 2.8.5 --- Embryo powder preparation --- p.36 / Chapter 2.8.6 --- Pre-absorption of antibody --- p.35 / Chapter 2.9 --- Whole Mount TUNEL Staining --- p.36 / Chapter Chapter 3 --- "Maternal Diabetes, Sp2H and RA Interaction" --- p.39 / Chapter 3.1 --- Introduction --- p.40 / Chapter 3.1.1 --- Neural tube defects --- p.41 / Chapter 3.1.2 --- Retinoic acid as environmental factor --- p.41 / Chapter 3.1.3 --- Sp2H as genetic factor --- p.44 / Chapter 3.1.4 --- Experimental design of this chapter --- p.46 / Chapter 3.2 --- Material and Methods --- p.47 / Chapter 3.2.1 --- Sp2H mice --- p.47 / Chapter 3.2.2 --- Mating and RA injection protocol --- p.47 / Chapter 3.2.3 --- Dissection of fetuses and analysis of neural tube development --- p.48 / Chapter 3.3 --- Results --- p.49 / Chapter 3.3.1 --- Maternal diabetes alone --- p.50 / Chapter 3.3.2 --- Sp2H mutation alone --- p.51 / Chapter 3.3.3 --- RA alone --- p.52 / Chapter 3.3.4 --- Maternal diabetes and RA interaction --- p.53 / Chapter 3.3.5 --- Sp2H mutation and RA interaction --- p.55 / Chapter 3.3.6 --- Sp2H mutation and maternal diabetes interaction --- p.57 / Chapter 3.3.7 --- "Maternal diabetes, Sp2H mutation and RA interaction" --- p.59 / Chapter 3.4 --- Discussion --- p.62 / Chapter 3.4.1 --- Maternal diabetes alone does not cause neural tube defects --- p.62 / Chapter 3.4.2 --- RA induces neural tube defects --- p.63 / Chapter 3.4.3 --- Interaction of maternal diabetes with RA in increasing the susceptibility to neural tube defects --- p.64 / Chapter 3.4.4 --- Embryos with Sp2H allele show increased susceptibility to neural tube defects when triggered by maternal diabetes and RA --- p.67 / Chapter Chapter 4 --- Molecular and Cellular Bases of Interaction --- p.71 / Chapter 4.1 --- Introduction --- p.72 / Chapter 4.1.1 --- Mechanism of diabetic embryopathy --- p.72 / Chapter 4.1.2 --- Mechanism of Sp2H mutation in development of neural tube defects --- p.74 / Chapter 4.1.3 --- Mechanism of RA teratogenicity --- p.75 / Chapter 4.1.4 --- "Possible common pathways shared by maternal diabetes, RA and Sp2H mutation" --- p.76 / Chapter 4.1.5 --- Experimental design of this chapter --- p.78 / Chapter 4.2 --- Materials and Methods --- p.80 / Chapter 4.2.1 --- Sample collection for studying Pax3 expression in Sp2H/+ And +/+ embryos in response to maternal diabetes or RA by whole mount in situ hybridization --- p.80 / Chapter 4.2.2 --- "Sample collection for studying the level of apoptosis in response to the interaction of maternal diabetes, Sp2H mutation and RA by whole mount TUNEL staining" --- p.82 / Chapter 4.3 --- Results --- p.86 / Chapter 4.3.1 --- Expression levels of Pax3 mRNA detected by whole mount in situ hybridization / Chapter 4.3.1.1 --- Expression of Pax3 in Sp2H/+/- and +/+ embryos --- p.86 / Chapter 4.3.1.2 --- Effect of maternal diabetes on Pax3 expression in Sp2H/+ and +/+ embryos --- p.87 / Chapter 4.3.1.3 --- Effect of RA on Pax3 expression in Sp2H /+ and +/+ embryos --- p.88 / Chapter 4.3.2 --- Level of apoptosis detected by whole mount TUNEL --- p.89 / Chapter 4.3.2.1 --- Effect of Sp2H allele on apoptosis --- p.94 / Chapter 4.3.2.2 --- Effect of maternal diabetes on apoptosis in Sp2H/+ and +/+ embryos --- p.95 / Chapter 4.3.2.3 --- Effect of RA on apoptosis in Sp2H/+ and +/+ embryos --- p.96 / Chapter 4.3.2.4 --- Effect of maternal diabetes and RA on apoptosis in Sp2H/+ and +/+ embryos --- p.97 / Chapter 4.4 --- Discussion --- p.99 / Chapter 4.4.1 --- Underexpression of Pax3 and increases in apoptosis under maternal diabetes --- p.99 / Chapter 4.4.2 --- "RA does not down regulate Pαx3, but increases apoptosis" --- p.102 / Chapter 4.4.3 --- Interaction of maternal diabetes and RA in increasing apoptosis --- p.104 / Chapter Chapter 5 --- "Maternal Diabetes, NOD and RA Interaction" --- p.108 / Chapter 5.1 --- Introduction --- p.109 / Chapter 5.1.1 --- Diabetic embryopathy in NOD mice --- p.109 / Chapter 5.1.2 --- Experimental design of this chapter --- p.110 / Chapter 5.2 --- Materials and Methods --- p.112 / Chapter 5.2.1 --- NOD mice --- p.112 / Chapter 5.2.2 --- Mating and RA Injection Protocol --- p.112 / Chapter 5.2.3 --- Sample Collection for the Study of Pax3 Expression --- p.113 / Chapter 5.3 --- Results --- p.115 / Chapter 5.3.1 --- Maternal diabetic alone --- p.116 / Chapter 5.3.2 --- NOD mutation alone --- p.117 / Chapter 5.3.3 --- RA alone --- p.118 / Chapter 5.3.4 --- Maternal diabetes and RA interaction --- p.119 / Chapter 5.3.5 --- NOD mutation and RA interaction --- p.121 / Chapter 5.3.6 --- NOD mutation and maternal diabetes interaction --- p.123 / Chapter 5.3.7 --- "Maternal diabetes, NOD mutation and RA interaction" --- p.125 / Chapter 5.3.8 --- Expression of Pax3 in embryos with different copies of NOD alleles --- p.128 / Chapter 5.4 --- Discussion --- p.130 / Chapter 5.4.1 --- Maternal diabetes interacts with NOD mutation to increase susceptibility to neural tube defects --- p.130 / Chapter 5.4.2 --- Interaction of maternal diabetes with NOD mutation is greatly exacerbated when exposed to RA --- p.131 / Chapter 5.4.3 --- Pax3 is not involved in the interaction --- p.133 / Chapter Chapter 6 --- Conclusion and Future Perspectives --- p.134 / References --- p.139 / Figures / Graphs