No / Bacterial lipoproteins are a diverse and functionally important group of proteins that are amenable to bioinformatic analyses because of their unique signal peptide features. Here we have used a dataset of sequences of experimentally verified lipoproteins of Gram-positive bacteria to refine our previously described lipoprotein recognition pattern (G+LPP). Sequenced bacterial genomes can be screened for putative lipoproteins using the G+LPP pattern. The sequences identified can then be validated using online tools for lipoprotein sequence identification. We have used our protein sequence datasets to evaluate six online tools for efficacy of lipoprotein sequence identification. Our analyses demonstrate that LipoP (http://www.cbs.dtu.dk/services/LipoP/) performs best individually but that a consensus approach, incorporating outputs from predictors of general signal peptide properties, is most informative.

No / Streptococcus equi is the causative agent of strangles, a prevalent and highly contagious disease of horses. Despite the animal suffering and economic burden associated with strangles, little is known about the molecular basis of S. equi virulence. Here we have investigated the contributions of a specific lipoprotein and the general lipoprotein processing pathway to the abilities of S. equi to colonize equine epithelial tissues in vitro and to cause disease in both a mouse model and the natural host in vivo. Colonization of air interface organ cultures after they were inoculated with a mutant strain deficient in the maturase lipoprotein ( prtM138-213, with a deletion of nucleotides 138 to 213) was significantly less than that for cultures infected with wild-type S. equi strain 4047 or a mutant strain that was unable to lipidate preprolipoproteins ( lgt190-685). Moreover, mucus production was significantly greater in both wild-type-infected and lgt190-685-infected organ cultures. Both mutants were significantly attenuated compared with the wild-type strain in a mouse model of strangles, although 2 of 30 mice infected with the lgt190-685 mutant did still exhibit signs of disease. In contrast, only the prtM138-213 mutant was significantly attenuated in a pony infection study, with 0 of 5 infected ponies exhibiting pathological signs of strangles compared with 4 of 4 infected with the wild-type and 3 of 5 infected with the lgt190-685 mutant. We believe that this is the first study to evaluate the contribution of lipoproteins to the virulence of a gram-positive pathogen in its natural host. These data suggest that the PrtM lipoprotein is a potential vaccine candidate, and further investigation of its activity and its substrate(s) are warranted.

No / Streptococcus equi and Streptococcus zooepidemicus are major etiological agents of upper and lower airway disease in horses. Despite the considerable animal suffering and economic burden associated with these diseases, the factors that contribute to the virulence of these equine pathogens have not been extensively investigated. Here we demonstrate the presence of a homologue of the Streptococcus pneumoniae PsaA protein in both of these equine pathogens. Inhibition of signal peptide processing by the antibiotic globomycin confirmed the lipoprotein nature of the mature proteins, and surface exposure was confirmed by their release from intact cells by mild trypsinolysis. / Project grant 056042 from The Wellcome Trust.

Thesis (PhD) -- University of Stellenbosch, 2001. / ENGLISH ABSTRACT: The advent of the new millennium saw the complete sequencing of the entire human genome. Only approximately 30 000 genes, much less than was initially predicted, have been identified to be responsible for the genetic diversity in humans. This discovery has prompted a shift in the approach to disease research, since one gene can be involved in numerous diseases. This phenomenon seems to be especially true for the low-density lipoprotein receptor (LDLR) gene. Various substances beside sterols can induce transcription of the LDLR gene. Non-communicable diseases (e.g. hypertension) are common in the developing world and contribute significantly to mortality rates. The fmding that a promoter variant (-175 g~t) in the LDLR gene is associated with elevated diastolic blood pressure may explain the phenomenon of high LDL-cholesterollevels in hypertensive individuals. Studies have demonstrated that the lowering of cholesterol, especially LDL-cholesterol, can reduce the incidence of hypertension. The -175 g~t variant is located in a newly described cis-acting regulatory element which contains a putative binding site for Yin Yang (YY)-l and also demonstrates great homology to the cAMP response element (CRE) which bind the Ca2+- dependent transcription factor, CRE binding protein (CREB). The fact that Ca2+ can induce transcription of the LDLR gene may, at least in part, explain the association between the - 175g~t variant and elevated diastolic blood pressure. Cholesterol is important for various processes, such as apoptosis, maintenance of cellular membranes and immune function. The -59 c-ot mutation in repeat 2 of the LDLR gene abolishes binding of the sterol regulatory element binding protein(SREBP) to the SRE-l site. SREBP is proteolytically activated during apoptosis by two caspases (CPP32 and Mch3) to induce cholesterol levels. Our results imply that the -59C/T mutation, in repeat 2 of the LDLR gene promoter, may inhibit apoptosis under normal immunological conditions. Atherosclerosis can be considered an immunological disease, since various humoral and cellular immune processes can be detected throughout the course of the disease. The fmding that certain lipoproteins can protect against infection by binding and lysing of pathogens, or competing with pathogens for cellular receptors, prompted the investigation into the potential role of variation in the LDLR gene promoter in immune function. A significant difference in allelic distribution was detected between asymptomatic HIY -infected subjects and fast progressors for the -124 c-ot variant (P=O.006), shown to increase (~160%) transcriptional activity of the LDLR gene. Of relevance to this particular study is the fact that human herpesvirus (HHV) 6 can transactivate CD4 promoters through a partial CRE site. It has been shown that the CREB and YYl can regulate viral and cellular promoters, and these transcription factors can potentially bind to the LDLR promoter at the FP2 site. The mutation enrichment in the LDLR gene promoter seen in the South African Black and Coloured population groups can possibly provide insight into the pathogenesis of various diseases. This could also potentially, provide novel targets for treatment, since manipulation of cholesterol levels may affect the pathogenesis of various diseases. / AFRIKAANSE OPSOMMING: Die volledige DNA volgorde bepaling van die mensgenoom is voltooi vroeg in die nuwe millennium. Slegs ongeveer 30 000 gene is geidentifiseer, heelwat minder as wat in die verlede voorspel is, wat verantwoordelik is vir die genetiese diversiteit in die mens. Hierdie ontdekking het gelei tot 'n verandering in die benadering van navorsing ten opsigte van siektes, aangesien een geen 'n rol by verskeie siektes kan speel. Hierdie gewaarwording blyk veral waar te wees vir die lae digtheids lipoproteien reseptor (LDLR) geen. Verskeie stimuli, buiten sterole, kan transkripie van die LDLR geen inisieer. Verskeie siektes soos hipertensie is algemeen in die ontwikkelende wereld, en dra by tot die hoe mortaliteit syfers. Die bevinding dat 'n promoter variant in die LDLR geen (-175g-H) geassosieer is met verhoogde diastoliese bloeddruk, kan moontlik verhoogde lipiedvlakke in hipertensiewe individue verklaar. Studies het aangetoon dat die verlaging van cholesterol, veral LDL-cholesterol, die voorkorns van hipertensie kan verlaag. Die -175 g~t variant is gelee in 'n cis-regulerende element wat na bewering 'n bindingsetel vir die Yin Yang (YY)-l transkripsie faktor bevat asook sterk homologie met die cAMP respons element (CRE) toon, wat bind aan die Ca2 +_ afhanklike transkripie faktor, CRE bindings proteiene (CREB). Die feit dat Ca2+ transkripsie van die LDLR geen kan inisieer, kan dalk tot 'n mate, 'n verklaring bied vir die assosiasie tussen die -175 (g~t) variant en verhoogde diastoliese bloeddruk. Cholesterol is noodsaaklik vir verskeie prosesse soos apoptose, die instandhouding van selmembrane sowel as immuun funksies. Die -59 c-ot mutasie in die sterol regulerende element 1 (SRE-l) van die LDLR geen vernietig binding van die sterol regulerende element bindingsprotei'en (SREBP) aan SRE-l. SREBP word proteolities geaktiveer tydens apoptose deur twee kaspases (CPP32 en Mch3) om cholesterolvlakke te induseer. Ons resultate impliseer dat die -59C/T mutasie, in herhaling-2 van die LDLR-geen promoter, apoptose kan inhibeer onder normale immunologiese toestande. Aterosklerose kan beskou word as 'n immunologiese siekte, aangesien verskeie humorale en sellulere immuun prosesse deur die verloop van die siekte waargeneem kan word. Die feit dat Iipoproteiene beskermend kan wees teen infeksies, deur binding en lisering van virusse of kompeteer met patogene vir sellulere reseptore, het aanleiding gegee tot 'n ondersoek na die potensiele rol van variasies in die promoter area van die LDLR geen in immuun funksie. Betekenisvolle verskille in alleel verspreiding vir die -124c~t variant (P=0.006) is waargeneem tussen asimptomatiese MIV -geinfekteerde pasiente en individue met vinnige siekte progressie. In vitro studies het voorheen getoon dat die -124c~t 'n verhoging in LDLR geen transkripsie (160%) tot gevolg het. Dit is noemenswaardig dat 'n vroee studie getoon het dat die mens like herpesvirus-6 (MHV6) transaktivering van die CD4 promoters deur 'n gedeeltelike CRE bindingsetel kan bewerkstellig. Beide CREB en YYl kan virus en sellulere promotors reguleer, en hierdie transkripsie faktore toon bindingshomologie met die FP2 element van die LDLR promotor Die mutasie verryking van die LDLR geen promoter soos waargeneem in Suid Afrikaanse Swart en Kleurling populasies, kan moontlik lig werp op die patogenese van verskeie siektetoestande. Hierdie bevindinge kan potensieel nuwe teikens vir behandeling identifiseer, aangesien manipulasie van cholesterolvlakke 'n effek mag he op die patogenese van verskeie siektes.

M.Phil. / Numerous studies have demonstrated the favourable effects of aerobic training on blood lipid profiles. However, few studies have generated conclusive data on the effects of dynamic resistance training (DRT) on blood lipid profiles. In order to evaluate the effect of DRT on lipoprotein-lipid profiles, a group of 28 sedentary but healthy males (mean age 28 years and 7 months) were matched and randomly assigned into a control/non-exercising (n = 15) or an experimental (n = 13) group. To control for variations in lipoprotein-lipid profiles, the present investigation recorded dietary intake and smoking behaviour in an attempt to account for any changes in lipoprotein-lipid profiles over the eight-week period. The experimental group (EG) exercised using DRT for a period of eight weeks and was monitored for changes in lipoprotein-lipid profiles. The control group (CG) took part in no structured exercise throughout the eight-week period. The experimental training programme consisted of nine exercises (dumbbell (D/B) shoulder shrugs, D/B lateral shoulder raises, seated chest press, latissimus dorsi pulldowns, seated pulley rows, biceps curls, triceps extensions, crunchies and unilateral leg press). These exercises were performed at 60% of one repetition maximum (1-RM) and were performed three times per week on non-consecutive days. Serum was analyzed for total cholesterol (TC), triglycerides (TG), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C). In addition to this, the TC: HDL-C and LDL-C: HDL-C ratios were calculated. The Independent t-Test and the Paired t-Test were utilized to determine the significance (at a 95% confidence level (p ¡Ü 0.05)) of the lipoprotein-lipid profile changes from pre- to post-test. These student t-Tests demonstrated no statistically significant changes in TC, TG, LDL-C, HDL-C, TC: HDL-C ratios and LDL-C: HDL-C ratios in the EG. However, the present investigation did demonstrate the following changes: a 0.50% decrease in TC, a 1.74% increase in TG, a 2.95% decrease in LDL-C, a 4.61% increase in HDL-C, a 4.12% decrease in the TC: HDL-C ratio and a 5.96% decrease in the LDL-C: HDL-C ratio. The lack of statistically significant changes in the individual lipoprotein-lipid parameters could not have been affected by diet, cigarettes smoked daily, aerobic fitness and/or body mass, since these parameters did not change significantly from pre- to post-test. Specifically, both the EG and CG demonstrated no statistically significant changes in intake in total calories consumed, carbohydrates, proteins, fats (monounsaturated, polyunsaturated and saturated fatty acids), cholesterol and fibre. Although the present investigation findings suggest that this study¡¯s eight-week combination of dose, workload, number of repetitions and order and number of exercises may not have been sufficient to elicit significant improvements in lipoprotein-lipid parameters in this population of sedentary but healthy males, it is the opinion of the author that DRT should be included with aerobic modes of exercise. DRT should be used in conjunction with aerobic modes of exercise for its additional benefits. Such additional benefits include inter alia: increased strength, increased lean tissue mass, increased maintenance of metabolically active tissue in the elderly and increased muscle control. / Prof. J.M. Loots Mr. L. Lategan

by Lo Hoi Ka Elka. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2001. / Includes bibliographical references (leaves 172-181). / Abstracts in English and Chinese. / ABSTRACT --- p.i / 摘要 --- p.iv / LIST OF TABLES AND FIGURES --- p.viii / ABBREVIATIONS --- p.xiv / Chapter CHAPTER 1 : --- INTRODUCTION / Chapter 1.1. --- DIFFERENT TREATMENTS OF THE CANCER THERAPY --- p.1 / Chapter 1.2. --- THE SIDE EFFECTS OF CANCER TREATMENT / Chapter 1.2.1. --- Surgery --- p.1 / Chapter 1.2.2. --- Radiotherapy --- p.2 / Chapter 1.2.3. --- Chemotherapy --- p.2 / Chapter 1.3. --- THE CHARACTERISTICS OF DOXORUBICIN (DOX) / Chapter 1.3.1. --- The structure of Dox --- p.6 / Chapter 1.3.2. --- The actions of Dox --- p.8 / Chapter 1.3.3. --- The adverse side effect of Dox --- p.8 / Chapter 1.4. --- THE RATIONALE OF USING LOW DENSITY LIPOPROTEIN (LDL) AS A TARGET CARRIER IN CANCER THERAPY / Chapter 1.4.1. --- The correlation between cholesterol and cancer --- p.9 / Chapter 1.4.2. --- Low density lipoprotein (LDL) as a target carrier --- p.11 / Chapter 1.4.3. --- The down and up regulation of LDL receptors --- p.14 / Chapter 1.4.4. --- The characteristics of Fuctus Craegus (FC) --- p.15 / Chapter 1.5. --- DIFFERENT METHODS OF THE PREPARATION OF THE LOW DENSITY LIPOPROTEIN-DRUG (LDL- DRUG) --- p.18 / Chapter 1.6. --- THE CHARACTERISTICS OF LOW DENSITY LIPOPROTEIN (LDL) / Chapter 1.6.1. --- The structure of LDL --- p.20 / Chapter 1.6.2. --- The metabolic pathway of LDL in human bodies --- p.23 / Chapter 1.7. --- THE MULTIDRUGS RESISTANCE IN TUMOR CELLS --- p.25 / Chapter 1.7.1. --- The mechanism of multidrug resistance --- p.27 / Chapter 1.7.2. --- The structure of P-glycoprotein --- p.27 / Chapter 1.7.3. --- The mechanism of P-glycoprotein --- p.30 / Chapter 1.8. --- COMBINED TREATMENT WITH HYPERTHERMIA --- p.31 / Chapter 1.9. --- AIM OF THE STUDY --- p.33 / Chapter CHAPTER 2 : --- MATERIALS AND METHODS / Chapter 2.1. --- MATERIALS / Chapter 2.1.1. --- Animals --- p.34 / Chapter 2.1.2. --- Buffers --- p.34 / Chapter 2.1.3. --- Cell culture reagents --- p.36 / Chapter 2.1.4. --- Chemicals --- p.38 / Chapter 2.1.5. --- Culture of cells --- p.40 / Chapter 2.2. --- METHODS / Chapter 2.2.1. --- In vitro studies / Chapter 2.2.1.1. --- "LDL, doxorubicin complex formation" --- p.41 / Chapter 2.2.1.2. --- Determination of the concentration of LDL-Dox --- p.42 / Chapter 2.2.1.3. --- In vitro cytotoxicity --- p.43 / Chapter 2.2.1.4. --- The cytotoxicity of the combined treatment with anticancer drugs --- p.44 / Chapter 2.2.1.5. --- The preparation of Fructus Crataegus (FC) --- p.46 / Chapter 2.2.1.6. --- Western blot --- p.47 / Chapter 2.2.1.7. --- Flow cytometry --- p.49 / Chapter 2.2.1.8. --- Confocal laser scanning microscopy --- p.52 / Chapter 2.2.2. --- In vivo studies / Chapter 2.2.2.1. --- Subcutaneous injection of R-HepG2 cells in nude mouse --- p.55 / Chapter 2.2.2.2. --- Treatment schedules --- p.55 / Chapter 2.2.2.3. --- Assay of investigating of the myocardial injury --- p.56 / Chapter 2.2.2.4. --- Tissue preparation procedure for light microscope (LM) --- p.57 / Chapter 2.2.3. --- Statistical analysis in our research --- p.59 / Chapter CHAPTER 3 : --- RESULTS / Chapter 3.1. --- in vitro STUDIES / Chapter 3.1.1. --- The preparation of low density lipoprotein-doxorubicin (LDL-Dox) --- p.60 / Chapter 3.1.2. --- Studies on human hepatoma cells line (HepG2 cells) / Chapter 3.1.2.1. --- The comparison of Dox and LDL-Dox accumulated in HepG2 cells --- p.63 / Chapter 3.1.2.2. --- Confocal laser scanning microscopic (CLSM) studies on the accumulation of Dox and LDL-Dox in HepG2 cells --- p.65 / Chapter 3.1.2.3. --- The comparsion of the cytotoxicity of Dox and LDL-Dox on HepG2 cells --- p.67 / Chapter 3.1.2.4. --- The comparison of the cytotoxicty of Dox and LDL-Dox with and without hyperthermia on HepG2 cells --- p.73 / Chapter 3.1.2.5. --- The comparison of accumulation of Dox and LDL-Dox in HepG2 cells treated with and without combination of with hyperthermia --- p.77 / Chapter 3.1.2.6. --- Confocal laser scanning microscopic (CLSM) studies on the accumulation of Dox and LDL-Dox in HepG2 treated cells with and without hyperthermia --- p.80 / Chapter 3.1.2.7. --- Modulation of LDL receptors on HepG2 cells------Up- regulation of LDL receptors by Fructus Craegtus (FC) / Chapter 3.1.2.7.1. --- The comparsion of LDL receptor expression on HepG2 cells after Fructus Craegtus (FC) pre-treatment --- p.83 / Chapter 3.1.2.7.2. --- The comparison of accumulation of LDL-Dox accumulated in HepG2 cells pre-treated with and without Fructus Craegtus (FC) --- p.85 / Chapter 3.1.2.7.3. --- Confocal laser scanning microscopic (CLSM) studies on the accumulation of LDL-Doxin HepG2 cells after Fructus Craegtus (FC) pre- treatment --- p.88 / Chapter 3.1.2.7.4. --- Cytotoxicity of combined treatment with LDL-Dox and Fructus Craegtus (FC) --- p.91 / Chapter 3.1.3. --- Studies on multidrug human resistant hepatoma cell line (R-HepG2 cells) / Chapter 3.1.3.1. --- The overexpression level of P-glycoprotein in resistant cell line R-HepG2 --- p.93 / Chapter 3.1.3.2. --- The comparison of Dox and LDL-Dox accumulated in R- HepG2 cells --- p.95 / Chapter 3.1.3.3. --- Confocal laser scanning microscopic (CLSM) studies on the accumulation of Dox and LDL-Dox in R-HepG2 cells --- p.97 / Chapter 3.1.3.4. --- The comparsion of the cytotoxicity of Dox and LDL-Dox on R-HepG2 cells --- p.99 / Chapter 3.1.3.5. --- The comparison of the cytotoxicty of Dox and LDL-Dox with and without hyperthermia on R-HepG2 cells --- p.109 / Chapter 3.1.3.6. --- The comparison of the accumulation of Dox and LDL- Dox in R-HepG2 cells treated in combination with hyperthermia --- p.113 / Chapter 3.1.3.7. --- Confocal laser scanning microscopic (CLSM) studies on the accumulation of Dox and LDL-Dox in R-HepG2 cells with and without hyperthermia --- p.117 / Chapter 3.1.3.8. --- Modulation of LDL receptors on R-HepG2 cells ------ Up-regulation of LDL receptors by Fructus Craegtus (FC) / Chapter 3.1.3.8.1. --- The comparsion of LDL receptor expression on R-HepG2 cells after Fructus Craegtus (FC) pre-treatment --- p.120 / Chapter 3.1.3.8.2. --- The comparsion of the accumulation of LDL- Dox in R-HepG2 cells after Fructus Craegtus (FC) pre-treatment --- p.122 / Chapter 3.1.3.8.3. --- Confocal laser scanning microscopic (CLSM) studies in the accumulation of LDL-Dox by Fructus Craegtus pre-treatment in R-HepG2 cells --- p.125 / Chapter 3.1.3.8.4. --- The comparison of cytotoxicity of combined treatment with LDL-Dox and Fructus Craegtus (FC) in R-HepG2 cells --- p.128 / Chapter 3.2. --- in vivo STUDIES / Chapter 3.2.1. --- The comparison of Dox and LDL-Dox on reducing the tumor sizes and weight in nude mice bearing R-HepG2 cells / Chapter 3.2.1.1. --- The comparison of Dox and LDL-Dox on reducing the tumor size in nude mice bearing R-HepG2 cells --- p.130 / Chapter 3.2.1.2. --- The comparison of Dox and LDL-Dox on reducing the tumor weight in nude mice bearing R-HepG2 cells --- p.138 / Chapter 3.2.2. --- Myocardial injury measured by Lactate dehydrogenase (LDH) activity in nude mice bearing R-HepG2 cells treated with Dox and LDL-Dox --- p.140 / Chapter 3.2.3. --- Myocardial injury measured by Creatine kinase (CK) activity in nude mice bearing R-HepG2 cells treated with Dox and LDL-Dox --- p.143 / Chapter 3.2.4. --- Histological studies of heart of nude mice bearing R-HepG2 cells treated with Dox and LDL-Dox / Chapter 3.2.4.1. --- Heart section of nude mice --- p.146 / Chapter 3.2.4.2. --- Heart section of nude mice bearing R-HepG2 cells --- p.148 / Chapter 3.2.4.3. --- Heart section of lmg/kg Dox treated nude mice bearing R- HepG2 cells --- p.150 / Chapter 3.2.4.4. --- Heart section of 2mg/kg Dox treated nude mice bearing R- HepG2 cells --- p.152 / Chapter 3.2.4.5. --- Heart section of lmg/kg LDL-Dox treated nude mice bearing R-HepG2 cells --- p.154 / Chapter CHAPTER 4 --- : DISCUSSION / Chapter 4.1. --- in vitro STUDIES / Chapter 4.1.1. --- The cytotoxicity of Dox and LDL-Dox on HepG2 cells and R- HepG2 cells --- p.156 / Chapter 4.1.2. --- The combined treatment on HepG2 cells and R-HepG2 cells --- p.157 / Chapter 4.1.3. --- The modulation of LDL-R expression --- p.159 / Chapter 4.2. --- in vivo STUDIES --- p.162 / Chapter CHAPTER 5 --- : CONCLUSION / Chapter 5.1. --- CONCLUSION / Chapter 5.1.1. --- In vitro studies --- p.167 / Chapter 5.1.2. --- In vivo studies --- p.169 / Chapter 5.2. --- FUTURE PROSPECTIVE --- p.170 / REFERENCES --- p.172

Les macrophages jouent un rôle clé dans l'athérosclérose. Après la captation massive des LDL oxydées (oxLDL), les macrophages sous-endothéliaux sont chargés en cholestérol et se transforment ainsi en cellules spumeuses qui contribuent à la formation de la plaque d'athérome. Les oxystérols, produits d'oxydation du cholestérol, sont retrouvés en quantité importante dans les oxLDL. Au niveau cellulaire, ils sont impliqués dans la régulation de l'homéostasie du cholestérol, l'induction du stress oxydatif cellulaire et de la cytotoxicité. Notre travail montre que le cholestérol, associé aux LDL et d'origine cellulaire, est fortement oxydé par les macrophages lors d'une exposition aux oxLDL. Les principaux produits d'oxydation sont le 7-cétocholesterol et 7α/β-hydroxycholestérol. De plus, nous démontrons que ces oxystérols sont exportés hors des macrophages via les HDL, mais pas l'apoA1. Nous avons aussi caractérisé les oxystérols formés dans les HDL suite à des modifications oxydatives et les HDL issues de patients diabétiques. Nous avons en outre montré que ces modifications sont associées à une diminution de la capacité des HDL à exporter les oxystérols. / Macrophages play a key role in atherosclerosis. After massive uptake of oxidized LDL (oxLDL), subendothelial macrophages are overloaded with cholesterol thereby leading to the formation of foam cells, which is one characteristic of atherogenesis. Oxysterols, the oxidation products of cholesterol, are one of major components of oxLDL; they are involved in the regulation of cholesterol homeostasis, induction of cellular oxidative stress and cytotoxicity. Our works show that both LDL derived-cholesterol and cellular cholesterol can be strongly oxidized in human THP1 and murine RAW macrophages, especially during exposure of oxLDL. The major oxidative products are 7-ketocholesterol and 7α/β-hydroxycholesterol. Moreover, we demonstrate that both oxysterols derived from LDL cholesterol and cellular cholesterol can be exported to HDL, whereas not to apoA1. Then, we studied the functionality of modified HDL and diabetic HDL on oxysterols efflux. A decrease of oxysterols efflux was observed with oxidized and glycoxidized HDL. Compared to the HDL of healthy controls, the HDL of diabetic subjects are less efficient to efflux oxysterols. Taken together, the increased production of oxysterols in presence of oxLDL and their lower efflux by modified HDL support the detrimental role of these oxidative compounds in pathophysiological conditions like diabetes.

高膽固醇血症是產生動脈粥樣硬化的危險因素，本研究旨在探討甾醇的氧化產物和植物化學物質在餵食高膽固醇食金黃地鼠模型中對脂蛋白代謝的影響及其相關機制。 / 本研究包含四個部分。食物中同時含有植物甾醇及其氧化產物。第一部分旨在研究β-穀甾醇（Si）、甾醇(St)、β-穀甾醇氧化產物(SiOP)和豆甾醇氧化產物(StOP)對金黃地鼠血脂的影響。本研究顯示，Si和St組能有效降低血總膽固醇（TC）、非高密度脂蛋白膽固醇（non-HDL-C）和甘油三酯（TAG）的水平，而SiOP和StOP則失去此能力。RT-PCR分析表明，Si和St而非SiOP和StOP，能下調腸道醯基輔酶A：膽固醇醯基轉移酶2（ACAT2）和微粒體甘油三酯轉移蛋白（MTP）的mRNA表達。Si和St而非SiOP和StOP能有效防止動脈粥樣硬，Si和St的動脈弓舒張能力強於對照組和SiOP、StOP組。 / 辣椒鹼是辣椒中的活性成分。本研究第二部分表明，辣椒鹼能降低TC，NON-HDL-C，TAG，而不影響高密度脂蛋白膽固醇。餵養辣椒鹼能增加糞便中總酸性固醇的排泄，此作用有可能是通過上調膽固醇7α-羥化酶（CYP7A1）和下調肝X受體α（LXRα）的基因表達來實驗。辣椒鹼可通過抑制COX-2基因表達來改善內皮依賴性收縮。 / 藍莓含有豐富的抗炎抗氧化劑，例如花青素。本研究第三部分表明，食物中添加0.5和1.0藍莓花青素能導致TC呈劑量效益地降低6-12%，其中還伴隨22-29的中性固醇和41-74%的膽汁酸排泄的增加。RT-PCR分析表明食物中添加的藍莓花青素能下調腸道Niemann-Pick C1 Like 1 (NPC1L1)，ACAT2，MTP， 腺苷三磷酸結合盒轉運體G8(ABCG8)和肝臟3-羥基-3-甲基戊二醯輔酶A還原酶(HMG-CoA　Reductase)的基因表達。 / 芝麻素是芝麻種子中含有抗氧化活性的木脂素類化合物。本研究第四部分表明，在食物中添加芝麻素可有效調控TC和non-HDL-C，同時不影響TAG，並導致非高密度脂蛋白膽固醇與高密度脂蛋白膽固醇比例的下降。這有可能與膽汁酸排泄增加、CYP7A1基因的上調，LXR的下調有關。 / 綜上所述，本研究證實了植物甾醇、辣椒鹼、藍莓花青素和芝麻素降低血膽固醇的能力。與此同時，本研究還表明植物甾醇被氧化後將失去其降低膽固醇的能力。 / Hypercholesterolemia is a major risk factor in the development of atherosclerosis. Functional foods that can lower or regulate cholesterol concentration are of interest to both public and scientific communities. The present study was to investigate the effects of phytosterols, phytosterol oxidation products (POPs), capsaicinoids, blueberry anthocyanins and sesamin on plasma cholesterol concentration using hamsters as a model. / The whole project consisted of four parts. Human diets contain both phytosterols and POPs. Part I was to examine the effect of β-sitosterol (Si), stigmasterol (St), β-sitosterol oxidation products (SiOP) and stigmasterol oxidation products (StOP) on plasma cholesterol concentration. Results showed both Si and St could reduce while SiOP and StOP lost the capacity of lowering plasma total cholesterol (TC), non-high density lipoprotein cholesterol (non-HDL-C) and triacylglycerols (TAG). Real-Time PCR analysis demonstrated Si and St but not SiOP and StOP down-regulated mRNA levels of intestinal acyl CoA: cholesterol acyltransferase 2 (ACAT2) and microsomal triglyceride protein (MTP). In addition, aortas from hamsters given diets containing Si and St relaxed better than those from the control and their corresponding SiOP- and StOP-treated hamsters, suggesting that Si and St not SiOP and StOP were beneficial in improving lipoprotein profile and aortic function. / Capsaicinoids refer to a group of pungent compounds that are the active components found in chili peppers. Part II was to investigate the cholesterol-lowering activity of capsaicinoids and the associated molecular mechanisms. Results demonstrated that capsaicinoids reduced plasma TC, non-HDL-C and TAG with high-density lipoprotein cholesterol (HDL-C) being unaffected. This was accompanied by an increase in the fecal excretion of total acidic sterols, possibly mediated by up-regulation of cholesterol 7α-hydroxylase (CYP7A1) and down-regulation of liver X receptor alpha (LXRα). Capsaicinoids could also improve the endothelium-dependent relaxations and reduce the endothelium-dependent contractions by inhibiting the gene expression of COX-2. / Blueberries are rich in anthocyanins. Results from Part III experiments demonstrated that dietary supplementation with 0.5 and 1.0 % blueberry anthocyanins for 6 weeks decreased plasma TC concentration by 6-12% in a dose-dependent manner. This was accompanied by increasing the excretion of fecal neutral and acidic sterols by 2229% and 4174%, respectively. Real-time PCR analyses demonstrated that incorporation of blueberry anthocyanins into diet down-regulated the genes of intestinal Niemann-Pick C1-like 1 (NPC1L1), ACAT2, MTP, ABCG 8 and hepatic 3-hydroxy-3-methylglutaryl CoA (HMG-CoA) reductase. / Sesamin is a major lignan in sesame seed and is known to exhibit antioxidative activity. Part IV was to investigate the mechanism by which sesamin decreased plasma cholesterol concentration. Results clearly demonstrated supplementation of sesamin into diets could favorably reduce serum TC and non-HDL-C with TAG being unaffected. In addition, dietary supplementation of 0.2 or 0.5% of sesamin could cause a significant decrease in the ratio of non-HDL-C to HDL-C. This was accompanied by a marked increase in bile acid excretion and up-regulation of CYP7A1 and down-regulation of LXRα. / In conclusion, phytosterols, capsaicinoids, blueberry anthocyanins and sesamin were beneficial in improving lipoprotein profile in hamsters fed a high-cholesterol diet. However, phytosterols lose the cholesterol-lowering capacity when they are oxidized. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Liang, Yintong. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 112-123). / Abstracts also in Chinese. / Chapter Chapter 1 --- General Introduction / Chapter 1.1 --- Cardiovascular diseases --- p.1 / Chapter 1.2 --- Cholesterol --- p.2 / Chapter 1.3 --- Lipoproteins --- p.4 / Chapter 1.4 --- Cholesterol homeostasis --- p.6 / Chapter 1.4.1 --- HMG-CoA reductase --- p.7 / Chapter 1.4.2 --- LDL receptor --- p.9 / Chapter 1.4.3 --- Intestine ACAT2 --- p.10 / Chapter 1.4.4 --- NPC1L1 --- p.11 / Chapter 1.4.5 --- CYP7A1 and LXRα --- p.12 / Chapter 1.4.6 --- SREBP2 --- p.14 / Chapter 1.4.7 --- ABCG5 and ABCG8 --- p.15 / Chapter 1.5 --- Phytochemicals --- p.16 / Chapter 1.5.1 --- Phytosterols --- p.16 / Chapter 1.5.2 --- Capsaicinoids --- p.17 / Chapter 1.5.3 --- Blueberry anthocyanins --- p.19 / Chapter 1.5.4 --- Sesamin --- p.20 / Chapter 1.6 --- Animal model --- p.22 / Chapter Chapter 2 --- Effect of Phytosterols and their Oxidation Products on Lipoprotein Profiles and Vascular Function / Chapter 2.1 --- Introduction --- p.23 / Chapter 2.2 --- Objective --- p.24 / Chapter 2.3 --- Materials and methods --- p.24 / Chapter 2.3.1 --- Preparation of sitosterol oxidation products (SiOP) and stigmasterol oxidation products (StOP) / Chapter 2.3.2 --- Diets --- p.25 / Chapter 2.3.3 --- Hamsters --- p.25 / Chapter 2.3.4 --- Analysis of individual SiOP and StOP in serum and liver --- p.26 / Chapter 2.3.5 --- Analysis of plasma lipoproteins --- p.28 / Chapter 2.3.6 --- Measurement of atherosclerotic plaque --- p.28 / Chapter 2.3.7 --- Analysis of cholesterol in the liver and aorta --- p.28 / Chapter 2.3.8 --- Determination of fecal neutral and acidic sterols --- p.29 / Chapter 2.3.9 --- Real-time PCR analysis of mRNA of liver SREBP2, LDL receptor, HMG-CoA reductase, CYP7A1, LXRα, and small intestine NPC1L1, ABCG5, ABCG8, ACAT2, MTP. --- p.29 / Chapter 2.3.10 --- Western blotting analysis of hepatic SREBP2, LDL receptor, HMG-CoA reductase, LXRα and CYP7A1 --- p.32 / Chapter 2.3.11 --- Vascular reactivity --- p.32 / Chapter 2.4 --- Results --- p.34 / Chapter 2.4.1 --- Composition of SiOP and StOP --- p.34 / Chapter 2.4.2 --- Food intake, body and organ weights --- p.34 / Chapter 2.4.3 --- Plasma TC, HDL, non-HDL ,TAG, Non-HDL-C/HDL-C --- p.34 / Chapter 2.4.4 --- Aortic cholesterol and atherosclerotic plaque --- p.35 / Chapter 2.4.5 --- Liver cholesterol, SiOP and StOP --- p.35 / Chapter 2.4.6 --- Fecal neutral, acidic sterols and cholesterol balance --- p.35 / Chapter 2.4.7 --- Immunoblot and mRNA analysis --- p.36 / Chapter 2.4.8 --- Vascular reactivity --- p.36 / Chapter 2.4.9 --- Role of COX in endothelium-dependent contractions --- p.37 / Chapter 2.5 --- Discussion --- p.50 / Chapter Chapter 3 --- Cholesterol-Lowering Activity of Capsaicinoids Is Mediated by Increasing Sterol Excretion in Hamsters Fed a High Cholesterol Diet / Chapter 3.1 --- Introduction --- p.54 / Chapter 3.2 --- Objective --- p.55 / Chapter 3.3 --- Materials and methods --- p.55 / Chapter 3.3.1 --- Diets --- p.55 / Chapter 3.3.2 --- Hamsters --- p.57 / Chapter 3.3.3 --- Analysis of plasma lipoproteins --- p.57 / Chapter 3.3.4 --- Measurement of atherosclerotic plaque --- p.57 / Chapter 3.3.5 --- Analysis of cholesterol in the liver and aorta --- p.57 / Chapter 3.3.6 --- Determination of fecal neutral and acidic sterols --- p.57 / Chapter 3.3.7 --- Real-time PCR analysis of mRNA of liver SREBP2, LDL receptor, HMG-CoA reductase, CYP7A1, LXRα, and small intestine NPC1L1, ABCG5, ABCG8, ACAT2, MTP --- p.57 / Chapter 3.3.8 --- Western blotting analysis of hepatic SREBP2, LDL receptor, HMG-CoA reductase, LXRα and CYP7A1 --- p.58 / Chapter 3.3.9 --- Vascular reactivity --- p.58 / Chapter 3.4 --- Results --- p.59 / Chapter 3.4.1 --- Food intake, body and organ weights --- p.59 / Chapter 3.4.2 --- Plasma TC, HDL, non-HDL,TAG, Non-HDL-C/HDL-C --- p.59 / Chapter 3.4.3 --- Aortic cholesterol and atherosclerotic plaque --- p.59 / Chapter 3.4.4 --- Fecal neutral, acidic sterols and cholesterol balance --- p.59 / Chapter 3.4.5 --- Immunoblot and mRNA analysis --- p.60 / Chapter 3.4.6 --- Vascular reactivity --- p.60 / Chapter 3.4.7 --- Role of COX in endothelium-dependent contractions --- p.61 / Chapter 3.5 --- Discussion --- p.74 / Chapter Chapter 4 --- Effect of Blueberry Anthocyanins on Lipoprotein Profiles in Hamsters Fed a Cholesterol Diet / Chapter 4.1 --- Introduction --- p.77 / Chapter 4.2 --- Objective --- p.78 / Chapter 4.3 --- Materials and methods --- p.78 / Chapter 4.3.1 --- HPLC analysis of blueberry anthocyanins --- p.78 / Chapter 4.3.2 --- Diet --- p.79 / Chapter 4.3.3 --- Hamsters --- p.80 / Chapter 4.3.4 --- Analysis of plasma lipoproteins --- p.80 / Chapter 4.3.5 --- Analysis of cholesterol in the liver --- p.80 / Chapter 4.3.6 --- Determination of fecal neutral and acidic sterols --- p.81 / Chapter 4.3.7 --- Real-time PCR analysis of mRNA of liver SREBP2, LDL Receptor, HMG-CoA Reductase, CYP7A1, LXRα, and small intestine NPC1L1, ABCG5, ABCG8, ACAT2, MTP --- p.81 / Chapter 4.3.8 --- Western blotting analysis of hepatic SREBP2, LDL Receptor, HMG-CoA reductase, LXRα and CYP7A1 --- p.81 / Chapter 4.4 --- Results --- p.82 / Chapter 4.4.1 --- Food intake, body, and organ weights --- p.82 / Chapter 4.4.2 --- Plasma TC, HDL-C, non-HDL-C, and TAG --- p.82 / Chapter 4.4.3 --- Liver cholesterol concentration --- p.82 / Chapter 4.4.4 --- Fecal total sterols and apparent sterol retention --- p.82 / Chapter 4.4.5 --- Immunoblot and mRNA analysis --- p.83 / Chapter 4.5 --- Discussion --- p.92 / Chapter Chapter 5 --- Effect of Sesamin on Lipoprotein Profiles in Hamsters Fed a high Cholesterol Diet / Chapter 5.1 --- Introduction --- p.95 / Chapter 5.2 --- Objective --- p.95 / Chapter 5.3 --- Materials and methods --- p.96 / Chapter 5.3.1 --- Diets --- p.96 / Chapter 5.3.2 --- Hamsters --- p.96 / Chapter 5.3.3 --- Methods --- p.97 / Chapter 5.4 --- Results --- p.98 / Chapter 5.4.1 --- Food intake, body and organ weights --- p.98 / Chapter 5.4.2 --- Plasma TC, HDL-C, non-HDL-C ,TAG, Non-HDL-C/HDL-C --- p.98 / Chapter 5.4.3 --- Liver cholesterol --- p.98 / Chapter 5.4.4 --- Fecal neutral, acidic sterols and cholesterol balance --- p.98 / Chapter 5.4.5 --- Immunoblot and mRNA analysis --- p.99 / Chapter 5.5 --- Discussion --- p.109 / References --- p.112

Thesis (M.Sc.), Memorial University of Newfoundland, 2000. / Restricted until June 2003. Bibliography: leaves 98-117.

Thesis (Ph. D.)--University of Missouri--Columbia, 2000. / Typescript. Vita. Includes bibliographical references (leaves 120-131). Also available on the Internet.