Thesis (Ph. D.)--Washington State University, May 2009. / Title from PDF title page (viewed on June 1, 2009). "School of Molecular Biosciences." Includes bibliographical references.

Thesis (doctoral)--University of Uppsala, 1984. / Includes bibliographical references (p. 29-37).

Thesis (M.S.)--University of Wisconsin--Madison, 1981. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 30-31).

Lee, Man Yuen. / "March 2008." / Thesis (M.Phil.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 103-128). / Abstracts in English and Chinese. / Title Page --- p.i / Acknowledgements --- p.ii / Table of Content --- p.iii / List of Figures --- p.vii / List of Graphs --- p.viii / List of Tables --- p.x / Abbreviations --- p.xii / Abstract --- p.xiii / Abstract (Chinese) --- p.xv / Chapter Chapter 1: --- General Introduction / 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.4 / Chapter 1.1.3 --- Gestational diabetes --- p.5 / Chapter 1.2 --- Diabetic pregnancy --- p.6 / Chapter 1.3 --- Etiology of diabetes-induced malformations --- p.9 / Chapter 1.3.1 --- Hyperglycaemia --- p.10 / Chapter 1.3.2 --- Hyperketonaemia and somatomedin inhibitors --- p.10 / Chapter 1.3.3 --- Oxidative stress --- p.11 / Chapter 1.3.4 --- Deficiency of myo-inositol and arachidonic acid --- p.12 / Chapter 1.4 --- Vitamin A --- p.13 / Chapter 1.5 --- Retinoic acid --- p.14 / Chapter 1.5.1 --- RA signaling on embryo development --- p.15 / Chapter 1.5.2 --- RA teratogenicity --- p.15 / Chapter 1.5.3 --- RA regulation --- p.17 / Chapter 1.6 --- RA and maternal diabetes-induced caudal regression share similar pathogenic mechanisms --- p.19 / Chapter 1.7 --- Strategy of the thesis --- p.21 / Chapter Chapter 2: --- General Materials and Methods / Chapter 2.1 --- Animal --- p.25 / Chapter 2.2 --- Induction of diabetes --- p.25 / Chapter 2.3 --- Preparation of retinoic acid for mouse injection --- p.26 / Chapter 2.4 --- RA responsive cell line --- p.26 / Chapter 2.4.1 --- Cell culture --- p.27 / Chapter 2.4.2 --- Seeding and adding sample to 96-well plate --- p.28 / Chapter 2.4.3 --- Staining of cells --- p.28 / Chapter 2.5 --- Real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR) --- p.29 / Chapter 2.5.1 --- Collection and storage of tissues --- p.29 / Chapter 2.5.2 --- Total RNA extraction --- p.29 / Chapter 2.5.3 --- Reverse transcription --- p.30 / Chapter 2.5.4 --- Polymerase chain reaction --- p.30 / Chapter 2.5.5 --- Preparation of cDNA standard --- p.31 / Chapter 2.5.6 --- Mini-scale preparation of plasmid DNA --- p.32 / Chapter Chapter 3: --- Effects of Maternal Diabetes on RA Catabolism in the Tail Bud Region / Chapter 3.1 --- Introduction --- p.34 / Chapter 3.2 --- Experimental design --- p.37 / Chapter 3.3 --- Materials and methods --- p.38 / Chapter 3.3.1 --- Preparation of RA stock solution for cell culture --- p.38 / Chapter 3.3.2 --- Preparation of RA standard solutions --- p.38 / Chapter 3.3.3 --- Characterization of RA responsive cell line --- p.39 / Chapter 3.3.3.1 --- Determining optimal culture time for maximum response --- p.39 / Chapter 3.3.3.2 --- Testing toxicity of DMSO --- p.39 / Chapter 3.3.3.3 --- Detection of β-galactosidase activity by β_gal Assay Kit --- p.40 / Chapter 3.3.4 --- In vitro assay of enzymatic degradation of RA --- p.41 / Chapter 3.3.4.1 --- Testing toxicity of enzyme cofactor-reducing agent --- p.41 / Chapter 3.3.4.2 --- Collection of tail bud --- p.42 / Chapter 3.3.4.3 --- In vitro enzymatic reaction --- p.43 / Chapter 3.3.5 --- In vivo assay of enzymatic degradation of RA --- p.44 / Chapter 3.3.5.1 --- Determining the optimal time for maximum release of RA from the tail bud into the medium --- p.44 / Chapter 3.3.5.2 --- Monitoring of RA remained in the tail bud after injection of exogenous RA --- p.45 / Chapter 3.3.6 --- Statistical analysis --- p.45 / Chapter 3.4 --- Results --- p.46 / Chapter 3.4.1 --- Optimization of RA responsive cell line --- p.46 / Chapter 3.4.1.1 --- Effect of incubation time with RA on β-galactosidase expression level --- p.46 / Chapter 3.4.1.2 --- Toxicity of RA and DMSO --- p.47 / Chapter 3.4.1.3 --- Comparison of X-gal staining assay and β_gal Assay Kit for detection of β-galactosidase expression --- p.48 / Chapter 3.4.2 --- In vitro assay of RA catabolic activity in the tail bud --- p.49 / Chapter 3.4.2.1 --- Toxicity of enzyme cofactor-reducing agent --- p.49 / Chapter 3.4.2.2 --- RA catabolic activity in lysed and intact tail buds --- p.50 / Chapter 3.4.2.3 --- Effect of enzyme cofactor and inhibitor --- p.50 / Chapter 3.4.2.4 --- Comparsion of in vitro RA catabolic activity of diabetic and non-diabetic groups --- p.51 / Chapter 3.4.3 --- In vivo assay of RA catabolic activity in the tail bud --- p.52 / Chapter 3.4.3.1 --- Optimal time for maximum release of RA from the tail bud into the medium --- p.53 / Chapter 3.4.3.2 --- Comparison of RA remained in the tail bud of embryos of diabetic and non-diabetic mice --- p.53 / Chapter 3.5 --- Discussion --- p.55 / Chapter Chapter 4: --- Analysis of Cyp26 Expression in the Tail Bud Region / Chapter 4.1 --- Introduction --- p.60 / Chapter 4.2 --- Experimental design --- p.63 / Chapter 4.3 --- Materials and methods --- p.64 / Chapter 4.3.1 --- Sample collection --- p.64 / Chapter 4.3.2 --- Real-time quantitative RT-PCR --- p.64 / Chapter 4.3.3 --- Statistical analysis --- p.66 / Chapter 4.4 --- Results --- p.67 / Chapter 4.4.1 --- "Relative expression levels of Cyp26al, Cyp26bl and Cyp26cl" --- p.67 / Chapter 4.4.2 --- Molecular changes in Cyp26al in tail bud region after maternal RA treatment --- p.68 / Chapter 4.5 --- Discussion --- p.72 / Chapter Chapter 5: --- Comparison of Cyp26al Heterozygous and Wild-type Embryos in Diabetic and Non-diabetic Pregnancies / Chapter 5.1 --- Introduction --- p.76 / Chapter 5.2 --- Experimental design --- p.79 / Chapter 5.3 --- Materials and methods --- p.81 / Chapter 5.3.1 --- Animal --- p.81 / Chapter 5.3.2 --- DNA genotyping --- p.81 / Chapter 5.3.3 --- Measurement of RA remained in the tail bud after RA treatment --- p.82 / Chapter 5.3.4 --- Analysis of extent of caudal regression --- p.83 / Chapter 5.3.5 --- Real-time quantitative RT-PCR --- p.83 / Chapter 5.3.6 --- Statistical analysis --- p.84 / Chapter 5.4 --- Results --- p.85 / Chapter 5.4.1 --- Comparsion of pregnancy outcome of mating with ICR or Cyp26al+/- males --- p.85 / Chapter 5.4.2 --- Expression levels of Cyp26al determined by real-time quantitative RT-PCR --- p.85 / Chapter 5.4.3 --- Determination of RA catabolic activity --- p.86 / Chapter 5.4.4 --- Extent of caudal regression --- p.90 / Chapter 5.5 --- Discussion --- p.93 / Chapter Chapter 6: --- Conclusion and Future Perspectives --- p.96 / References --- p.102 / Figures / Graphs

Graduation date: 2004

Thesis (Ph. D.)--University of Washington, 2006. / Vita. Includes bibliographical references (leaves 153-179).

Not available / text

Vitamin A is an essential dietary component which primarily acts through its active metabolite, retinoic acid (RA), a potent transcriptional regulator that also has non-genomic activities. There is increasing evidence for a role for vitamin A in the regulation of circadian rhythms, and previous studies suggest that it serves important roles in the pineal gland, an integral component of the circadian system due to its function of melatonin production. However very little is currently known about how these effects are mediated, or about RA signalling in the pineal gland. This study aimed to establish whether RA is synthesised in the rat pineal gland and determine its role here. This included investigating whether RA is subject to diurnal changes in synthesis and signalling, and examining its involvement in the key rhythms in this gland: melatonin synthesis, kinase activation and clock gene expression. Organotypic culture of rat pineal glands, qPCR and western blotting were among the techniques employed to do this, as well as RA quantification using a reporter cell line. The rat pineal gland was found to produce RA and robust diurnal changes in synthesis were detected. Furthermore, diurnal changes in expression of RA signalling genes suggested there are corresponding changes in RA activity. RA was not found to rapidly regulate Aanat transcription, melatonin synthesis or clock gene expression in vitro, however it was found to rapidly down-regulate extracellular signal-regulated kinase (ERK) 1/2 phosphorylation. Furthermore, strong cytoplasmic expression of RA receptor α was detected in pinealocytes. These results suggest RA has a non-genomic role in the pineal gland and may be involved in driving the diurnal rhythm in kinase activation.

13-Cis-retinoic acid (13-cis-RA) is a synthetic retinoid and the active ingredient in the oral acne treatment Accutane. The medical literature has suggested that the use of 13-cis-RA for acne treatment can induce depression, but because acne itself can have a negative psychosocial impact on self esteem, whether or not 13-cis-RA can cause depression remains controversial. The purpose of this work was to examine whether chronic 13-cis-RA administration could induce depression-related behaviors in mice and to determine the impact 13-cis-RA has on regions of the brain thought to be associated with mood and depression. We found that chronic treatment of adolescent male mice with 13-cis-RA induced depression-related behaviors, as assessed by immobility in the tail suspension and forced swim tests. Although depression is a multifaceted disease in which many brain regions are involved, the regions that seem particularly vulnerable to the effects of 13-cis-RA are the serotonergic and hippocampal systems. In serotonergic cells in vitro, 13-cis-RA treatment increases protein levels of the serotonergic 5-HT[subscript 1A] autoreceptor and the serotonin reuptake transporter (SERT), two inhibitory components of serotonin (5-HT) signaling. In vivo, the median and dorsal raphe nuclei contain the main 5-HT producing cells. 13-Cis-RA uncoupled the functional connectivity of dorsal raphe nuclei from the hippocampal regions as measured by interregional correlations of cytochrome oxidase (CO) activity, a metabolic marker of neuronal activity. Decreased hippocampal neurogenesis is thought to occur in depression and is decreased by 13-cis-RA. 5-HT is also a known regulator of hippocampal neurogenesis. Uncoupling of the dorsal raphe nuclei from the regions of the hippocampus by 13-cis-RA treatment may be the cause of, or a result from, the decreased neurogenesis. Although retinoids are known regulators of apoptosis, the uncoupling of the dorsal raphe nuclei from the hippocampal regions was not due to serotonergic cell loss. Interestingly, 13-cis-RA treated animals with the lowest CO activity in the dentate gyrus have the highest immobility in the tail suspension and forced swim tests. Ultimately, the effects of 13-cis-RA on the serotonergic and hippocampal systems might be inducing depression-related behaviors. / text

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