Thesis (Ph. D.)--University of Adelaide, Dept. of Animal Sciences, 1986. / Includes bibliographical references (leaves 194-219).

Thesis (Ph. D.)--University of Wisconsin--Madison, 1941. / Typescript. Includes abstract and vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 94-98).

Thesis (Ph. D.)--University of Wisconsin--Madison, 1984. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 106-112).

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 36-38).

Phosphatidylcholine metabolism was investigated in Nb 2 node rat lymphoma, a cell line which is dependant on prolactin for growth in culture. Treatment of stationary cultures with prolactin stimulated the incorporation of [Me-³H] choline into phosphatidylcholine (1.7-fold after 4 h) and its aqueous precursors, mainly phosphocholine (2-fold after 4 h and 3-fold after 10 h). These effects were blocked by cycloheximide. Pulse-chase studies demonstrated that the reaction catalyzed by CTP:phosphocholine cytidylyltransferase (EC 2.7.7.15) was rate-limiting for phosphatidylcholine synthesis in Nb 2 cells. The specific activity of choline kinase (EC 2.7.1.32) increased 1.9-fold after 4 h of prolactin treatment, in correspondence with the increase in choline incorporation mentioned above, and this was also blocked by cycloheximide. The activities of the other enzymes of phosphatidylcholine synthesis were unchanged. These results suggested that phosphatidylcholine biosynthesis was not altered in Nb 2 cells after prolactin treatment. However, phosphatidylcholine levels increased in prolactin treated-cells (1.4-fold after 16 h). Turnover of labeled phosphatidylcholine was reduced 3.4-fold in prolactin treated cells. Calculated turnover rates for phosphatidylcholine averaged 3.2-fold lower in prolactin treated cells whereas the synthetic rates were similar in prolactin treated and stationary cells. Thus, Nb 2 cells utilize a novel mechanism, inhibition of turnover, to regulate the cellular levels of phosphatidylcholine during growth / Medicine, Faculty of / Biochemistry and Molecular Biology, Department of / Graduate

Choline is indispensable for a number of fundamental processes in the body. Besides being the precursor of the acetylcholine, an important neurotransmitter, choline is found in the cell membrane structure combining with fatty acids, phosphate and glycerol. Its deficiency may result in nervous system disorders, fatty acid build up in the liver, along with increased cholesterol levels, high blood pressure and memory loss. Thus, rapid detection methods are required for the determination of choline in biological fluids. In this study a choline oxidase biosensor was constructed for the determination of choline. During construction of the biosensor, glucose oxidase was used as a model enzyme, before choline oxidase used. The Teflon (PTFE) membrane of the oxygen electrode was grafted with 2-hydroxyethyl methacrylate (HEMA, 15%, v/v) in the presence of ferrous ammonium sulphate (FAS, 0.1%, w/v) by gamma irradiation and ethyleneglycol dimethacrylate (EGDMA, 0.15 %, v/v) was used as a crosslinker in a series of membranes. HEMA-grafted membranes were activated with epichlorohydrin or glutaraldehyde to maintain covalent immobilization of enzyme. The enzyme activity was measured with an oxygen electrode unit based on oxygen consumption upon substrate addition. Membranes were characterized in terms of grafting conditions and mechanical properties. Membranes, gamma irradiated in a solution of HEMA (15%) and FAS (0.1%) for 24 h, were found to be suitable for use in the further studies. Mechanical test results revealed that HEMA grafting made Teflon membrane more flexible and the presence of EGDMA made the grafted membrane stiffer. During optimization stage, it was found that the immobilized enzyme amount was not sufficient to obtain enzyme activity. Thus, the membrane preparation stage was modified to obtain thinner membranes. The immobilized glucose oxidase and choline oxidase contents on thin HEMA grafted membranes were determined by Bradford and Lowry methods. The influence of EGDMA presence and the epichlorohydrin activation duration on enzyme activity studies revealed that the membrane should be prepared in the absence of EGDMA and 30 min activation duration is appropriate for epichlorohydrin coupling. The study on the influence of membrane activation procedures revealed that the membranes activated with glutaraldehyde had a higher specific activity than the membranes activated with epichlorohydrin. Upon stretching membrane on the electrode directly rather than placing in the sample unit, the response of the enzyme immobilized sensor improved with high specific activity. The optimum choline oxidase concentration was found to be 2 mg/mL considering the effect of immobilization concentration on enzyme activity. With the choline oxidase biosensor, the linear working range was determined as 0.052-0.348 mM, with a 40 &plusmn / 5 &micro / M minimum detection limit. The response of the sensor decreased linearly upon successive measurements.

Graduation date: 1993

Thesis (M.S.)--University of Toledo, 2008. / Typescript. "Submitted as partial fulfillment of the requirements for the Master of Science degree in Chemistry." "A thesis entitled"--at head of title. Bibliography: leaves 82-85.

Phosphatidylethanolamine (PE) is an important inner membrane phospholipid synthesized de novo by the CDP-ethanolamine pathway and by the decarboxylation of phosphatidylserine. CTP: phosphoethanolamine cytidylyltransferase (Pcyt2) is the main regulatory enzyme in the CDP-ethanolamine pathway and catalyzes the formation of CDP-ethanolamine from phosphoethanolamine. Complete deletion of the mouse Pcyt2 gene is embryonic lethal, and the single allele deficiency leads to development of metabolic syndrome phenotype, including liver steatosis, hypertriglyceridemia, obesity, and insulin resistance. This study aimed to specifically elucidate the effects of dietary methyl group donors betaine and choline supplementation in Pcyt2 heterozygous mice (ETKO). Evidence here shows choline and its oxidized metabolite betaine are responsible for lowering whole body weight, restoring insulin resistance, reducing hypertriglyceridemia, hepatic steatosis, and alleviating adipose and liver tissue inflammation, by restoring hepatic metabolism and gene expression. Collectively, these results establish that the impaired systemic metabolism resulting from Pcyt2 deficiency is a metabolic adaptation that is restored after methyl group supplementation.

Choline is known to be essential for brain development and neural function, but its impact on the retina, as a type of neural tissue, is unknown. This study examined the effects of choline during fetal development on membrane phospholipid (PL) compositions and functions in neural tissues, brain and retina. Pregnant C57 BL/6 mice were fed one of the 4 choline modified diets from gestation to early development: i) deficient (Def, 0g/kg), ii) control (Cont, 2.5g/kg), iii) supplemented with choline chloride (Cho, 10g/kg), iv) supplemented with egg phosphatidylcholine (PC) (PC, 10g/kg). On postnatal day (PD) 7, pups were culled to 4 from each dam, and kept on the same respective diets until 45 PD. On PD 35, memory function was measured by Morris water maze and on PD 45, retina function by an electroretinogram. Brain and retina were obtained for PL analysis by 31P NMR. Animals on the Def and PC diets were lower in body weights on PD 7, in comparison to the other two groups. While the Def group caught up in weights to its Cont counterparts, the PC group’s weight stayed consistently low until PD 45 (P<0.03). As for brain function, Cho and PC supplemented groups showed enhanced cued learning task, and spatial memory abilities, respectively, whereas the Def group showed the poorest memory recollection (P<0.05). The ERG amplitudes of rod driven photoreceptors and inner neural cell functions were significant (P<0.05) in the following order: Cont > Def > PC > Cho, at all light intensities, without reaching statistical significances in cone-driven responses. There were no differences in major PL compositions in the brain and retina. PC enriched group had increased subclasses of ether PL, PEaa and PCaa in the brain. These results indicate that while the addition of choline supplementation is beneficial for fetal brain development and function during early developmental stages, its contributions in the retina were minor. The effect of choline to the membrane PL structure was negligible for the stage of development in the given experimental design.