Oxysterol binding protein (OSBP) binds 25-hydroxycholesterol (25OH) and cholesterol, which regulates PH and FFAT domain interaction with the Golgi apparatus and endoplasmic reticulum, respectively. Adjacent to these domains is a phosphorylated serine-rich motif (SRM, T379, S381, S384, S387, S388, S391) that we hypothesize controls sterol transport by OSBP. To test this, OSBP dephospho-mimics or phospho-mimics were expressed in CHO cells. Western blot analysis showed that the S381 is phosphorylated by PKA and is required for phosphorylation of down-stream serine residues. When expressed in OSBP-null CHO cells, there was no difference in the localization of the OSBP mutants, and all mutants restored SM synthesis in response to 25OH. Recombinant OSBP 5S?5E had increased cholesterol binding and extraction, and decreased cholesterol transfer to liposomes compared to OSBP. OSBP 5S?5E also bound VAP more efficiently. A model is proposed wherein SRM phosphorylation facilitates VAP association with the ER and increases cholesterol extraction.

Hydrocarbon autoxidation, a free radical chain reaction, is one of the most important chemical processes, and is ubiquitous in biological systems and industry. While it is vital to maintaining cellular homeostasis and plays central roles in the immune and inflammatory responses, it is also believed to play a role in the onset and development of diseases and degenerative disorders when not kept in check. In vivo, this process is generally initiated by the reduction of O2 to superoxide (O2•-), which can then afford various reactive oxygen species (ROS), such as HOO•, H2O2, HO•, and 1O2. Recently, it was suggested that antibodies, as part of the immune system, produce another ROS: ozone. The evidence for endogenous ozone formation was based largely on the isolation of the known cholesterol ozonolysis products in extracts of arterial plaque and brain tissue. Identification was accomplished by derivatization and subsequent HPLC-MS analysis. Herein, an alternative, more likely explanation for the appearance of these two compounds and their derivatized forms is given, via acid-catalyzed Hock cleavage of cholesterol 5-hydroperoxide. Radical-trapping chain-breaking antioxidants inhibit hydrocarbon autoxidation; in Nature and as additives in industrial materials, formulations, etc. Nature typically employs phenols in this context, and it is well documented that their potency is based largely on the lability of their phenolic O-H bond. While their reactivity can be improved by making the phenol more electron-rich by introducing electron-donating groups on the aromatic ring, this increases their air (oxygen) sensitivity, leading them to decompose in air and generate ROS themselves! To prevent this, nitrogen(s) can be introduced in the aromatic ring to make 3-pyridinols and 5-pyrimidinols; the most effective air-stable radical-trapping antioxidants reported to date. Unfortunately, introduction of nitrogen in the phenolic ring leads to a concomitant increase in the acidity of the O-H bond, leading to stronger interactions with H-bond accepting solvents. This interaction reduces the efficacy of these compounds as antioxidants in polar and heterogeneous media. Herein we describe our efforts to minimize the effect of this interaction, thereby maintaining the strong antioxidant activities of 3-pyridinols and 5-pyrimidinols, by introducing two tert-butyl moieties flanking the reactive hydroxyl group. / Thesis (Master, Chemistry) -- Queen's University, 2008-11-21 14:30:19.24

Sterols serve two important biological functions in animals - they act as cellular membrane components, and as the precursor to steroid hormones. Insects require a dietary source of sterol because they cannot synthesize sterols de novo. Cholesterol is the most common sterol in plant-feeding insects, but because plants contain very little cholesterol, plant-feeding insects must convert plant sterols into cholesterol. In this dissertation I investigate the effect of common and novel plant sterols and steroids found in a transgenic tobacco line on several caterpillar species. I also explore the metabolism of these sterols and steroids, and use a microarray approach to identify genes involved in sterol use and metabolism in plant-feeding insects. I also study cholesterol homeostasis using a grasshopper species. Modified tobacco plants containing a novel sterol profile negatively affected performance three different caterpillar species, especially in the second generation. Insects reared on modified plants contained less total sterols and cholesterol than those on control plants having normal sterol profile. Similar results were found using artificial diets containing atypical steroids, e.g., cholestanol and cholestan-3-one, identified in the tobacco plants that were fed to my experimental caterpillars. More importantly, the sterol/steroid ratio, but not their absolute amount in the diets, determined the negative effects. Caterpillar species could convert stigmasterol, a common plant sterol, into cholesterol. They could also convert cholestan-3-one into cholestanol and epicholestanol, although this ability varied among different species. A microarray study, that focused on gene expression in midgut tissue, indicated that stigmasterol, cholestanol and cholestan-3-one could induce different gene expression level, and that cholestan-3-one caused a the largest pool of genes to be regulated. The genes possibly involved in the metabolism of stigmasterol and cholestan-3-one were reported. These findings are important in directing further research on the potential application of plant sterol modification to control pests in agricultural systems. Insect herbivores could behaviorally regulate the intake of several nutrients, but they could not regulate their sterol intake. They did, however, practice cholesterol homeostasis, by postingestively regulating tissue sterol levels, even when feeding on diets with high cholesterol content. Collectively, the results from this dissertation provide unique insight into cholesterol regulation, which is difficult to achieve in mammals that are capable of synthesizing their own sterols.

1. <i>Cis</i> unsaturated fatty acids were shown to inhibit porcine platelet aggregation in response to both collagen and thrombin. Fatty acids with a <i>trans</i> double bond had an anti-aggregatory effect on collagen-induced aggregation but this was significantly less than that observed with the <i>cis</i> equivalent and was diminished as the dose of agonist increased. Thrombin-induced platelet aggregation was unchanged or slightly potentiated by <i>trans</i> isomers. 2. Both the <i>cis</i> and <i>trans</i> isomeric acids inhibited collagen-induced TXB₂ production. The <i>trans</i> unsaturated fatty acids also inhibited TXB₂ production in response to thrombin, even though they did not inhibit thrombin induced platelet aggregation. 3. Unlike arachidonic acid, the <i>cis</i> and <i>trans</i> mono-unsaturated fatty acids were not rapidly incorporated into membrane phospholipids but modified platelet aggregation whilst in the free acid form. 4. Pre-incubation of platelets with either <i>cis</i> or <i>trans</i> delta 13, 18:1, selectivity inhibited the incorporation of radio-labelled arachidonic acid into membrane PS. 5. <i>Cis</i> and <i>trans</i> unsaturated delta 13, 18:1, inhibited the initial turnover of membrane PI in response to thrombin possibly by an inhibitory effect on PI-specific phospholipase C. After 5 minutes, however, the level of arachidonic acid released from both PI and PE was increased in the presence of the isomeric fatty acids. This may have been via a potentiation of the action of phospholipase A₂. 6. An increased release of arachidonic acid could result in the inhibition of aggregation if metabolised via the 12-lipoxygenase pathway, as the end products of this sytem have direct anti-aggregatory activity and inhibit the cyclo-oxygenase enzymes thus reducing TXA₂ synthesis. 7. <i>Cis</i> unsaturated fatty acids, which produce a greater level of membrane disruption than the <i>trans</i>, may lead to more efficient channelling of the released arachidonic acid in the direction of the lipoxygenase pathway, and thereby produce a greater inhibition of aggregation. The possibility remains that the <i>cis</i> isomers have another, as yet unidentified mechanism by which they inhibit platelet aggregation.

The purpose of the study was to examine the relationship of exercise of self-care agency and serum cholesterol levels. The American Heart Association (1986) and the National Heart, Lung and Blood Institute (1987) have identified the reduction of serum cholesterol as a major factor in decreasing the risk for coronary artery disease. Orem's theory of self-care identified diet and exercise management as activities well being (Orem, 1971). It was hypothesized that high levels of self-care are inversely related to serum cholesterol levels. A retrospective study was conducted. A convenience sample of 176 individuals who have obtained cholesterol screening at a Nursing Center for Family Health was obtained. Written consent was obtained to review individual files for serumcholesterol levels, information related to demographic variables, nutrition and exercise habits; and for the Exercise of Self-Care Agency. Each participant completed the "Exercise of Self-Care Agency" tool. Confidentiality was maintained and responses were coded for analysis only. Correlational statistics were utilized to analyze the data obtained. The hypothesis was tested using Pearson r analysis and demonstrated poor correlation and no significant relationship. Research questions related to diet, exercise, age and the exercise of self-care agency were tested by a 3x2x2 ANOVA and showed no significant difference. Recommendations for future study should include replication of the study with a larger sample and randomization of the sample. / School of Nursing

Thesis (M.Sc. (Human Movement Science))--North-West University, Potchefstroom Campus, 2008.

Thesis (M.Sc. (Human Movement Science))--North-West University, Potchefstroom Campus, 2008.

There is worldwide concern over the possible estrogenic effects of organic chemicals on a variety of wildlife and indeed on humankind. In the U.K., estrogenic compounds in sewage treatment works (STW) effluents have been implicated in causing the increases in egg yolk protein production observed in caged male trout and other fish species. At the initiation of the present study, few of the estrogenic compounds in STW effiuents had been recognised, although circumstantial evidence suggested that steroidal hormones were primary candidates. Cholesterol is abundant in STW effluents and is the precursor of all steroidal hormones biosynthesised in mammalian systems. Thus, the aim of the present study was to examine the hypothesis that cholesterol might undergo A-ring aromatisation, during sewage treatment, producing estrone and 17β-estradiol via, intermediates such as l9-norcholest-l,3,5(10)-trien-3-ol (NCT). To study this hypothesis NCT was first synthesised via a known route and several of its chromatographic and mass spectral properties established for the first time. NCT itself was found to possess some estrogenic potential determined using an established assay but this was rather weak compared to 17β-estradiol - about 200,000 times less active at the minimum concentration needed to invoke a response. NCT also proved to be a much more hydrophobic compound than, for example, 17β-estradiol with a computed log octanol-water partition coefficient (Kow) of over 9 compared with a log Kow of about 4 for l7β-estradiol. The established analytical properties of NCT were then used to investigate possible NCT formation in sewage. Radiolabelled 14C-cholesterol was incubated aerobically and anaerobically in Semi-Continuous Aaivated Sludge (SCAS), Die Away (DA) or simple stand alone STW simulation vessels. The products of incubation in both aqueous and solid fractions were examined by radio-high performance liquid chromatography (r-HPLC), radio-thin layer chromatography (r-TLC) and radio-gas chromatography (r-GC). Aerobic studies showed that side chain cleavage and A-ring rupture of cholesterol occurred rapidly (~25 % of added activity within 24 hrs) as measured by 14C02 evolution. Gaseous evolution was not monitored from the anaerobic experiments. Most remaining activity was associated with the solids fractions in all experiments. In the aqueous experiments both SCAS and DA systems, r-HPLC revealed rapid production of polar products which were not identified further. r-HPLC also revealed non-polar components of which choIest-3,5-diene, an unknown cholestadiene, a cholestadienol (other than 5,7-dienol), cholest-4-en-3-one and possibly NCT were identified by r-GC in the products of DA experiments. Whilst r-HPLC and r-TLC also revealed several products of anaerobic digestion of cholesterol, no compounds were detected by r-GC. STW effluents from two wastewater plants in the North London area were monitored over 7 months for A-ring steroids and other suspected estrogenic chemicals. Both effluents had previously proved estrogenic to caged fish. Liquid and SPM samples were taken, extracted and analysed by GC-MS. The two main estrogens, l7β-estradiol and estrone were identified from all liquid samples but not in SPM extracts. Generally the concentration of estrone (maximum ca 3 ng Lˉ¹) was significantly higher than that for 17β-estradiol (maximum 1 ng fˉ¹). The third natural estrogen, l6a,l7β-estriol, was identified in all Harpenden effluent samples analysed up to a concentration ca 0.5 ng Lˉ¹. However, estriol was not found in Deephams effluent extracts. The phytoestrogen, daidzein, found in soya based products was intermittently found in aqueous effluents samples from both sites >1 ng Lˉ¹. SPM made up <0.001 % of the effluent. Extracts showed that there was a high percentage of steroidal based material with coprostanol>cholesterol=β-sitosterol>>stigmasterol. NCT was twice identified from SPM taken from Deephams with a concentration of 39 and 136 ng 1ˉ¹ but <LOD in Harpenden SPM extracts. In conclusion this study showed that formation of NCT from cholesterol in STW treatment under conditions monitored herein is not a major process. However, estrone, 17β-estradiol and 16a, 17β-estriol are relatively abundant and widespread components of the STW effluents tested. The origins of the components undoubtedly include direct inputs from female urine and faeces but other metabolic sources should not be entirely ruled out.

No description available.

Hypercholesterolaemia (high cholesterol) is a risk factor for atherosclerosis, a major contributor to cardiovascular disease. Much has been written on managing high cholesterol levels including the use of drug therapy and recommendations about beneficial lifestyle changes relating to nutrition and exercise in particular. Despite this, and the need for patients to engage in self management of a condition which may impact invisibly on their health, little is known about how people living with this risk factor actually manage.The purpose of this grounded theory study is to investigate people's process of living with high cholesterol. Eight participants with self reported high cholesterol levels were interviewed and their data analysed using the process of constant comparative analysis, open coding, selective coding and theoretical coding. A substantive theory in the area of patient self-management of high cholesterol levels was generated through the use of this approach. The theory developed was the basic social process of Maintaining Enjoyment of Life, with two supporting categories, Accepting and Getting On With It.Maintaining Enjoyment of Life was central to the participants' process of living with high cholesterol and the recommended health directives. The identification of this basic social process adds to our understanding of the influences on individuals' motivation for behaviour change and adherence to lifestyle changes. The findings of this study provide understanding that issues relating to behaviour change and adherence are not so much about denying that anything is wrong or not taking personal responsibility, but more about trying to hold on to something important, around which the participants' lives revolve. This is Maintaining Enjoyment of Life. By focussing on this basic social process, which is grounded in the participants' data, health professionals could work more effectively towards empowering patients to identify their own individualised strategies for self-management of high cholesterol. In addition, health directives could focus more on process rather than on do(s) and don't(s).