Thesis (M. Med. Sc.)--University of Hong Kong, 2005.

Thesis (B.A.)--Haverford College, Dept. of Philosophy, 2005. / Includes bibliographical references.

Thesis (Ph. D.)--Drexel University, 2006. / Includes abstract and vita. Includes bibliographical references (leaves 104-112).

No description available.

1. A review of the literature on cartilage canals is given. 2. The formation of cartilage canals commences when the cartilage mass exceeds the maximum thickness which can be nourished by diffusion of fluid from the surface. This maximum thickness or the critical size at which canalization would occur, has been worked out in the distal femoral epiphysis at various developmental stages. It varies with the age of the foetus 0.25 mm. at the fourth month and gradually increasing to about 0.6 mm. at full-term. 3. Because of the restricted area of origin of the cartilaginous epiphysis of long bones, the canals seldom found, to be arranged in a radial fashion to the whole epiphysis, but they arranged so as to distribute the blood evenly through the whole mass. 4. (i) The clear, well-formed communicating canals have been seen in the epiphysis of human long bone as early as the fourth month of foetal life. (ii) As development proceeds, some of the communicating canals appear to become obliterated in, the region of proliferating cartilage adjacent to the metaphysis: this obliteration of canals occurs more rapidly after the onset of epiphyseal ossification so that by the time ossification of the epiphysis is complete, no communications between the diaphysis and the epiphysis remain. (iii) It is suggested that probably the primary cause of the formation of the communicating canal is the chemio-taxio influence in the zone of actively growing cartilage in the region adjacent to metaphysis, which directs the ends of the canals arising from the perichondrium near the end of the shaft to bend towards the diaphyseal end. (iii) The probable function of the communicating canals is that they assist in the supply of nutrition to and in the removal of waste products from the cells in the active juxta-metaphyseal cartilage. The almost invariable absence of osteogenic elements in these canals given no support to the hypothesis that they take part in the formation of the centre of epiphyseal ossification. (v) The vascular connective tissue buds which are identical with the communicating canals in the epiphysis of long bone, have boon observed in the cartilaginous sternal end of the clavicle of a human foetus. 5. It is suggested that the cartilage canals grow by a combination of three methods that is by surface accretion, stretching due to interstitial growth and active invasion of the cartilage by the tip of the canal. 6. The cartilage canal connective tissue contents are of perichondrial origin, and are not formed by back differentiation of the cartilage to an embryonic type of connective tissue. 7. In the long bone of the human, the cartilage canals are probably responsible for the formation of the epiphyseal ossification centre.

Body fat distribution is a strong determinant of human metabolic health but the mechanisms underpinning regional deposition of white adipose tissue (WAT) remain poorly understood. WAT also exhibits striking depot-specific functional properties. The aim of this project was to investigate the potential role of specific candidate genes implicated in regulating WAT development and/or function in a depot-specific manner. Cartilage oligomeric matrix protein (COMP) is an extracellular matrix protein that is highly differentially expressed between subcutaneous abdominal and gluteal WAT but has primarily been studied in the context of bone biology. WAT COMP expression was found to be significantly up-regulated in obesity; COMP expression in preadipocytes was increased by glucocorticoids; and COMP promoted adipogenesis in (immortalised) subcutaneous abdominal and gluteal preadipocytes. Building on a finding during the COMP study, bone morphogenetic protein (BMP)-2 was identified as another candidate. BMP2 exerts positive adipogenic effects in murine models and a recent genome-wide association study meta-analysis identified a significant association between BMP2 and body fat distribution. BMP2 was found to exert a pro-adipogenic effect specifically in subcutaneous abdominal preadipocytes, with this effect requiring activation of SMAD1/5/8 signalling via type 1 BMP receptors. These data identify BMP2 as a novel depot-specific regulator of human adipogenesis. Particularly lipid-laden cells were formed when conventional adipogenic medium was supplemented with fatty acids; these cells were captured, de-differentiated (DFAT) and expanded to generate immortalised abdominal and gluteal DFAT cells. These DFAT cells exhibit a greatly enhanced adipogenic potential compared to the mixed stromovascular (SVF) population from which they derive and retained an intrinsic memory of their anatomical origin. The use of DFAT cells is likely to represent a valid and enhanced model system to study various depot-specific aspects of WAT biology such as adipogenesis. Overall, the data from this thesis emphasise the striking depot-specific biology exhibited by WAT and provide novel insights into the mechanisms governing the regional distribution of WAT in humans.

The study of the role of adipose tissue in the maintenance of the caloric homeostasis of organisms is currently the object of widespread research. In particular, the enzymes of lipid metabolism in adipose tissue are being extensively investigated in both intact fat pads and in broken-cell preparations. Special attention is being paid to factors which control the activity of these enzymes. We have examined some properties of a lipase in epididymal fat pads of rats. The enzyme has been assayed by measuring the free fatty acids liberated when triglycerides are incubated with crude adipose tissue extracts. Quantitative measurements of free fatty acids were performed by (a) titrating the liberated acid with dilute alkali solution, and (b) reacting the free fatty acids with Cu⁺⁺ to form the chloroform-soluble copper soap of long chain fatty acids, then assaying the copper with diethyl-dithiocarbamate spectrophotometrically. It is well known that lipase activity in adipose tissue decreases during incubation in a Krebs-Ringer bicarbonate medium at 37°C. The de-activated enzyme can be activated by briefly exposing the intact tissue to epinephrine. The study of this epinephrine-sensitive lipase in adipose tissue has been the main object of this thesis. When epinephrine was added to media containing intact epididymal fat pads, the dramatic mobilization of free fatty acids from the pads into the media was observed. When epinephrine was added directly to unfractionated homogenates, little, if any, response was elicited, indicating perhaps that some activating factor was destroyed or diluted out during homogenization. When ATP, cyclic 3',5'-AMP and Mg⁺⁺ were added to unfractionated homogenates of adipose tissue, some lipase activation was observed. Similarly, when these nucleotides and Mg⁺⁺ were added to the supernatant fluid obtained from centrifuged homogenates, some activation of the lipase was observed, although the results obtained were not consistent. Other nucleotide 3',5'-cyclic phosphates generally inhibited lipase activity in the supernatant fluid. Our data indicates that epinephrine activates adipose tissue lipase only when added to the intact fat pad before homogenization. Little or no activation occurred when the amine was added to homogenates. Cyclic 3',5'-AMP had some ability to reactivate the lipase, both in unfractionated homogenates and in the supernatant fluid prepared by centrifugation. The effects, however, were not marked. It is concluded that if epinephrine-activation of adipose tissue is mediated through cyclic 3',5'-AMP, precise conditions for showing this have not yet been achieved. Additional experiments were performed on the epinephrine-sensitive lipase. Intact adipose tissue obtained from reserpinized rats was exposed to epinephrine after a 3-hour incubation period. The results indicated that epinephrine does not activate the lipolytic system in adipose tissue of reserpinized rats. Finally, some of the factors regulating the degree of inactivation of the epinephrine-sensitive lipase during incubation were investigated. Fat pads removed from rats which had been either anaesthetized or not anaesthetized prior to sacrifice were incubated for 3 hours. Data collected from a number of experiments indicated that there were virtually no differences in the extent of lipase inactivation between the two groups of rats. / Medicine, Faculty of / Anesthesiology, Pharmacology and Therapeutics, Department of / Graduate

The response of various plant tissues to different conditions of culture was compared. The tissues used were cambium-containing discs from carrot roots, undifferentiated carrot callus, bacteria-free sunflower tumorous (crown-gall) tissue, and segments of sunflower stems. The culture conditions compared, in combination, were agar versus liquid medium, shaken versus non-shaken liquid medium, and continuous light versus continuous dark. The response of the tissues to White's basal nutrient medium with added coconut milk (15 %) and indoleacetic acid (0.1 mg/l) and Hildebrandt's improved sunflower medium was also compared under these different culture conditions. Agitation of the liquid medium was accomplished through the use of a newly designed shaker, which consists basically of a horizontally oscillating bank of shelves. The tissues rested on the bottom of culture flasks (medicine bottles) on these shelves and were alternately exposed to medium and air as the liquid medium washed back and forth. Any horizontally oscillating platform could replace this shaker and almost any type and size of culture flask could be used. Probably any type of plant tissue could be cultured under these shaking conditions. It is not necessary that the tissues adhere to the walls of the culture vessels as in other agitation methods so far used in plant tissue culture. Growth (weight increase) of all tissues in shaken liquid medium (in both light and dark) was markedly superior (two to six times greater average weight in 42 days) to that of tissues on agar and in non-shaken liquid medium. The superiority of growth in shaken liquid medium is probably due to several factors; nutrients and gasses are supplied to the entire surface of the tissues, there is no drying and hardening of the tissue surfaces, resulting in a greatly increased surface area, harmful excretions can not collect at the tissue surface, and diffusion of nutrients is not hindered by adsorptive effects of agar particles. To compare the growth of these cultures with those of other workers using agitation methods is difficult due to the different sources of plant material, different sizes of tissues cultured, and different periods of culture used. In general the stimulatory results of shaking obtained appear to be at least as good as those obtained by Caplin and Steward with the much more elaborate and limited 'auxophyton '. There was no sign of eventual growth stoppage as obtained by White, using roller tubes. Light consistently stimulated tissues grown in liquid medium, particularly those in shaken liquid medium. The effect was especially marked on carrot callus and tumorous sunflower tissues grown in Hildebrandt's medium. It is suggested that light may play a role in the synthesis of growth factors supplied by coconut milk. Light had no significant effect on the growth of tissues on agar medium, indicating that the primary limiting factor in the growth of such tissues may be the rate of diffusion of nutrients from the agar. Carrot tissues showed better overall growth in the enriched White's medium while the sunflower tumorous tissue grew better in Hildebrandt's medium. The effect on carrot was probably primarily through indoleacetic acid and coconut milk. The response of sunflower tissue is difficult to evaluate at present. All carrot tissues developed chlorophyll throughout all of the experiments if cultured in light, while tumorous sunflower tissue remained white until placed in Hildebrandt's medium, when it turned light green. The significance of these differences is not known. One experiment showed that carrot discs derived from different carrots grew at significantly different average rates, indicating that discs to be compared should be derived from the same root. The plane in which the discs were cut did not seem to influence subsequent growth. 'Intra root' variation in disc growth necessitates replication. / Science, Faculty of / Botany, Department of / Graduate

Cultured tissues derived from the coffee plant, Coffea arabica L., were grown in vitro in the form of both callus and suspension cultures. The suspension cultures grew rapidly and appeared healthy. Microscopic examination showed that the cells characteristically grew in long filamentous chains. Suspension cultures were examined for the presence of three components - free amino acids, caffeine and chlorogenic acid. By examining these components the species specificity could be determined. The free amino acids of the coffee bean are thought to be one of the major precursors of coffee aroma on roasting. The coffee suspension cultures were shown to contain a similar pattern of free amino acids although the total content was much higher in the cultures than in the intact green coffee bean. Aspartic acid, glutamic acid, phenylalanine, alanine, valine, threonine, serine, and glycine were the predominant amino acids present in the coffee suspension culture. Threonine, serine, glycine, alanine and phenylalanine were the major free amino acids in the green coffee bean. The free amino acid content in the suspension culture exhibited an initial rise, decreased during active growth, then increased rapidly to the maximum level during the decline of the culture. Roasted coffee bean extracts were investigated to ascertain whether one solvent could in preference extract some of the major precursors of coffee aroma. Methanol was found to extract material from green coffee beans which on roasting produced coffee aroma. Caffeine was detected in the cell suspension cultures. However, problems with the analytical methods gave rise to questionable results. The suspension cultures, at maximum caffeine yield, contained 0.03% caffeine (dry weight) whereas the green coffee bean contained considerably more caffeine (1.15%, dry weight). The caffeine content of the tissues increased during the lag phase, decreased during the rapid phase and then increased again in the stationary phase and ultimately production levelled off during the deline phase of growth. The cell cultures produced chlorogenic acid in low concentrations at the maximum 0.14% dry weight in contrast to the green coffee bean which contains 6.5% dry weight. The production or accumulation of chlorogenic acid followed a similar pattern to that of the cell caffeine production over the growth curve. Caffeic acid was also detected. The cell suspension cultures of Coffea arabica L. were shown to be species specific in their biochemical capabilities. / Land and Food Systems, Faculty of / Graduate

No description available.