Thesis (Ph. D.)--Columbia University, 1934. / Vita. Published also as Child development monographs, Monograph no. 16. Bibliography: p. 55-56.

Thesis (M.S.)--University of Wisconsin--Madison, 1983. / Typescript. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Thesis (Maters Diploma (Medical Technology)--Cape Technikon, Cape Town, 1993 / The object of the present study was to determine whether there was a difference in the flux through two different metabolic pathways, the purine salvage pathway and the urea cycle, in skin fibroblasts from three species. A double label approach was adopted to measure the incorporation of the purine bases, hypoxanthine and adenine, and the amino acids, citrulline and arginine in acid precipitable material. Before examining the between species variation it was necessary to examine different levels of possible variation such as the variation between individuals from each species, the variation between separate experiments and the replication error within experiments. Eight individuals from each of three species, bUffalo, human, and rhinoceros were examined with the labelled purine bases. Skin fibroblasts from four humans and eight rhinoceros individuals were also examined over varying passage numbers until the fibroblasts senesced, to determine the effect of ageing on the uptake of hypoxanthine and adenine. The same four human fibroblast cultures were transformed with a transforming virus and examined to see the effects of transformation on the uptake of the purine bases, these transformed fibroblasts were compared with previously transformed rhinoceros fibroblasts. The uptake of labelled citrulline and arginine was also examined in three individuals from each of the three species. The major part of variation throughout the study was found to be at the between experiment level, despite stringently controlled conditions. This between experiment variation obscured any variation found within individuals from each species. In spite of this major between - experiment variation, the results showed that there was significant variation between the three species in the uptake of hypoxanthine. Adenine uptake was similar in the buffalo and human, but was significantly different between both these species and the rhinoceros. citrulline uptake showed no variation between the three species, whereas arginine showed a significant variation between the rhinoceros and the other two species. Buffalo and human showed no significant variation in arginine uptake. There was a significant increase in the uptake of hypoxanthine and adenine in transformed fibroblasts relative to untransformed fibroblasts. As a consequence of the significant between-experiment variation demonstrated in this study, it is apparent that great care must be taken to standardize the conditions when using a double label approach, especially if the assay is to be used for the diagnosis of inborn errors of metabolism.

Marine organisms are known to produce secondary metabolites which have novel structures and are often biologically active. Chemical studies of biologically active metabolites from three different marine organisms led to the discovery of six new compounds and six previously known compounds. The brown alga Dictyota binghamiae is fairly abundant in British Columbia coastal waters. A chemical study of this alga yielded ten diterpenoids of which four are new compounds. All the new compounds, dictyoxide A (66), dictyol G acetate (68), dictyotriol A diacetate (69), and epidictyol B acetate (70) contain a perhydroazulene carbon skeleton first encountered in the algal metabolite pachydictyol A (29). Dictyoxide A (66) appears to be an artifact of isolation. The acetates 68, 69, 70 were found to be antibacterial and antifungal. Six previously known compounds pachydictyol A (29), dictyol C (32), dictyoxide (35), acetyldictyolal (49) and the acetals 61a and 61b were also isolated from this alga. Chemical studies on an Agelas sp. of sponge collected in Sri Lanka yielded the antimicrobial metabolite desbromooroidin (79). An interesting interaction between the starfish Dermasterias imbricata and the sea anemone Stompia coccinea was observed a long time ago. When contacted by the starfish, the anemone displays an unusual "swimming" response which was, according to other subsequent studies, caused by a single chemical substance in the starfish. A study conducted to elucidate the structure of this starfish metabolite led to the isolation of imbricatine (91), a unique benzyltetrahydroisoquinoline alkaloid. Imbricatine (91) induced S. coccinea swimming response at a very low concentration and also exhibited antitumor activity. Structures of all the new metabolites were determined by spectral analysis, and chemical degradations and chemical interconversions. / Science, Faculty of / Chemistry, Department of / Graduate

Human cancer intervention trials have found beta-carotene to be effective in reducing the genotoxic damage to oral mucosa cells that resulted from carcinogen exposure. Design of intervention trials using beta-carotene has so far lacked an important component, knowledge of the accumulation and retention of this putative chemopreventive agent in the tissues of interest. The factors of dose, timing of administration of supplements, and the effects of confounding factors are aspects of trial design that demand an understanding of the kinetics of beta-carotene in human tissues. The oral mucosa is the only tissue so far that has been investigated for beta-carotene levels in exfoliated cells. A non-invasive technique for sample collection, suitable for sampling populations, in combination with a highly sensitive assay for beta-carotene, is appropriate for use with other epithelial sites that may be targets of intervention trials; one such site is the uro-genital tract. This thesis describes preliminary investigations towards design of an intervention trial with beta-carotene directed at cells of the uro-genital tract. Initial studies established the feasibility of measuring beta-carotene in uro-genital tract cells, and verified that the technical variation in the assays of oral mucosa cells and urogenital tract cells was less than the variation between individuals in the study population. A short-term (four-day) supplementation trial compared the effects of three doses of beta-carotene, given orally, on the beta-carotene content of exfoliated oral mucosa cells. Ingestion of 360 mg, 180 mg, and 90 mg of beta-carotene resulted in a rise in median cell beta-carotene levels from 1.8, 1.5, and 1.0 ng/10⁶ cells to 9.2, 7.7, and 3.9 ng/10⁶ cells, respectively, one week after the loading. Due to variation in response, the effects of the three doses were not significantly different from one another. The elevation in beta-carotene levels persisted for 2 weeks after the loading. The kinetic profile of the accumulation and retention of beta-carotene in uro-genital tract cells differed markedly from that of oral mucosa cells, in individuals who were supplemented for four weeks with a field trial dosage of beta-carotene. Median beta-carotene levels in uro-genital tract cells of beta-carotene-supplemented individuals increased from 0.6 ng/10⁶ cells to 2.8 ng/10⁶ cells during the loading period, a level significantly higher than that of the placebo group, but declined immediately after the end of the loading to levels that were not significantly elevated. In contrast, beta-carotene in exfoliated oral mucosa cells remained at significantly elevated levels until four weeks after the cessation of loading. Tissue-specific features of beta-carotene accumulation in response to its administration need to be taken into consideration when designing intervention strategies that use beta-carotene. / Medicine, Faculty of / Pathology and Laboratory Medicine, Department of / Graduate

Osteoblasts are bone-forming cells and therefore they are responsible of the synthesis of type I collagen, the main component of bone matrix. However, there is an apparent disconnect between the regulation of osteoblast differentiation and bone formation since the synthesis of Type I collagen precedes the expression of Runx2, the earliest determinant of osteoblast differentiation. Recently, genetic experiments in the mouse have revealed the existence of an unexpected cross-regulation between bone and other organs. In particular this body of work has highlighted the importance of osteoblasts as endocrine cells to regulate whole-body glucose homeostasis by secretion of a hormone, osteocalcin. However, the fundamental question of why bone regulates glucose homeostasis remained to be answered. Therefore, in my thesis, considering that bone is a metabolically demanding organ that constantly renews itself, I hypothesized that characterizing the connection between the need of glucose as a main nutrient in osteoblasts and bone development will provide a key to deeper understanding of why bone regulates glucose homeostasis. My work shows here that glucose uptake through GLUT1 in osteoblasts is needed for osteoblast differentiation by suppressing the AMPK-dependent activation by phosphorylation at S148 of Smurf1 that targets Runx2 for degradation. I also uncovered the mechanism of action of Smurf1 in this setting. In a distinct but synergetic way, glucose uptake promotes bone formation by inhibiting a distinct function of AMPK. In turn, Runx2 favors Glut1 expression, and this feedforward regulation between Runx2 and Glut1 determines the onset of osteoblast differentiation during development and the extent of bone formation throughout life. Furthermore, I also identified that Smurf1 not only regulates osteoblast differentiation by targeting Runx2 for degradation but also contributes to whole-body glucose homeostasis by regulating the activation of osteocalcin by targeting the insulin receptor for degradation in vivo. These results identify Smurf1 as a determinant of osteoblast differentiation during development, of bone formation and glucose homeostasis post-natally. Most importantly, we show that these Smurf1 functions required AMPK-phosphorylation site S148 in vivo. Altogether, these results revealed the absolute necessity of glucose as a regulator of Runx2 accumulation during osteoblast differentiation and bone formation in vivo and highlight the fundamental importance of the intricate cross-talk between bone and whole-body glucose metabolism.

published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

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