Thesis (Ph.D.)--Tufts University, 2004. / Adviser: Jose Ordovas. Submitted to the School of Nutrition Science and Policy. Includes bibliographical references (leaves 136-137). Access restricted to members of the Tufts University community. Also available via the World Wide Web;

Thesis (M.A.)--University of North Dakota, 2006. / Includes bibliographical references (leaves 37-40)

This study was performed to determine the rates of in vitro cholesterol synthesis, as measured by ³H incorporation into cholesterol, in gluteal white adipose tissue and the intestine of infant rats during the early postnatal period. The reason for studying these parameters was to hopefully further elucidate the cause of the differences in blood cholesterol levels between rats raised in different litter sizes. Rats were raised in small (4/dam), medium (8/dam) and large (14/dam) litters. The rate of cholesterol synthesis in the proximal and distal region of the small intestine decreased from birth to a low 14 days later and then increased again by day 21. The rate of the decreases in cholesterol synthesis, from birth to the low on day 14, varied amongst the three litter sizes; the rate of synthesis in both regions of the intestine on day 7 was lowest in the larger litter, and was not significantly different from the values seen on day 14. Previous work has shown that plasma cholesterol levels were low in rats from large litters on day 7, thus in the early postnatal period low rates of intestinal cholesterol synthesis correlate with low plasma cholesterol values. After weaning, on day 21, the rate of synthesis in distal intestine was higher than that of proximal intestine in the medium and small litters, whereas the opposite was found in the larger litter. Cholesterol synthesis in gluteal white adipose tissue remained at a very low rate up to 21 days in the small and medium sized litters. In contrast, the rate of synthesis increased continuously in the large litter to nearly threefold the rate of that in the other two litter sizes on day 21. This study has shown that the rates of cholesterol synthesis in intestinal and adipose tissue can be altered during the early postnatal period by raising the rats in different litter sizes. Also, the pattern of development in the rates of cholesterol synthesis as measured by ³H incorporation into cholesterol are in general agreement with results reported for the activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase for this same period in development. / Land and Food Systems, Faculty of / Graduate

A dissertation submitted in partial compliance with the requirements for the Master's Degree in Technology: Homoeopathy, Technikon Natal, 1996. / The purpose of this study was to evaluate the efficacy of Taraxacum 5CH in the treatment of hypercholesterolemic patients, in terms of measuring the extent of change in serum total cholesterol (TC), low density lipoprotein (LOL) and high density lipoprotein levels (HOL). The expected result was a resultant lowering of bath the TC and LOL levels, as well as an increase of the HOL levels. Convenience sampling was employed to draw patients from Technikon Natal, a Durban company and the general public from the greater Durban area. Only persons with raised TC and LOL levels were accepted into the trial. Of these one half constituted the control group and received only placebo, the remaining half made up the trial group and were treated with Taraxacum 5CH. The remedies were prepared and dispensed by a qualified pharmacist. Venous blood was obtained from all subjects before the trial and monthly, for three consecutive months, during the trial. A reputable pathology laboratory was employed to obtain lipogram studies of all blood samples. The participants were asked not to change their then present diets or lifestyles. Performing paired T-tests on the initial versus the final values of the control group revealed no statistically significant changes in the TC, LOL and HOL levels, whereas in the trial group a significant reduction was computed for the TC and LOL levels, as well as an insignificant increase of the HOL levels. Unpaired T-tests showed that the trial and control groups were not significantly different at the beginning ot the trial, but were found to have changed to become significantly different by the end of the trial with respects to TC and LOL levels. The HOL levels were dissimilar initially, but were shown to be significantly similar at the end of the trial. / M

Dissertation submitted in partial compliance with the requirements for the Master's Diploma in Technology: Homoeopathy, Technikon Natal, 1994. / The purpose of this study was to evaluate the effect of Chelidonium 3X in the treatment of hypercholesterolaemia in terms of changes in the blood cholesterol levels in order to determine the extent to which homoeopathic medicine could be used in the treatment of this condition. It was hoped that there would be a drop i n the total cholesterol levels and an increase in the ratio of high density / low density lipoprotein. Participants in the trial were drawn from the staff at Technikon Natal, convenience sampling being used. For acceptance into the trial, 'participants had to have an elevated total cholesterol level together with a raised low density lipoprotein value according to age. Drawing of blood was performed following an overnight fast. Of those participants meeting the above criteTicrthirty were chosen to participate in the trial. Half constituted the control and were given a placebo and the other half made up the experimental group and were treated with Chelidonium 3X. This was a double blind study with the medicine being dispensed on a random basis by a qualified pharmacist. After commencement of the trial venous blood was collected once a month for three consecutive months from both the control and experimental groups. The participants were asked to make no changes to their normal lifestyle. Statistical analysis of the results using paired T-tests revealed no statistical difference between the initial and final total cholesterol or ratio reading of the control group. However, with the experimental group a statistical difference was noted between both the initial and final total cholesterol reading / M

Yang, Nan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (p. 94-117). / Abstracts in English and Chinese. / ACKNOWLEDGMENTS --- p.I / ABSTRACT --- p.II / LIST OF ABBREVIATIONS --- p.VII / TABLE OF CONTENTS --- p.IX / Chapter Chapter 1 --- general introduction / Chapter 1.1 --- Cholesterol and cardiovascular disease --- p.1 / Chapter 1.2 --- Functions of cholesterol and lipoprotein --- p.2 / Chapter 1.3 --- Cholesterol metabolism and regulation in the body --- p.5 / Chapter 1.3.1 --- General process of cholesterol metabolism --- p.5 / Chapter 1.3.2 --- Cholesterol metabolism in liver --- p.6 / Chapter 1.3.2.1 --- The uptake of LDL cholesterol into the liver --- p.6 / Chapter 1.3.2.2 --- Cholesterol synthesis --- p.7 / Chapter 1.3.2.3 --- Synthesis of bile acids --- p.8 / Chapter 1.3.2.4 --- RCT pathway --- p.9 / Chapter 1.3.3 --- Lipids absorption in the intestine lumen --- p.9 / Chapter 1.3.3.1 --- Niemann-Pick Cl like 1(NPC1L1) --- p.10 / Chapter 1.3.3.2 --- ABCG5/8 --- p.10 / Chapter 1.3.3.3 --- Acyl-CoA:cholesterol acyltransferase (ACAT) 2 --- p.11 / Chapter 1.3.4 --- Cholesterol homeostasis --- p.11 / Chapter 1.3.5 --- The regulation of the cholesterol metabolism --- p.11 / Chapter 1.3.5.1 --- The role of SREBP-2 --- p.11 / Chapter 1.3.5.2 --- The role of LXR --- p.13 / Chapter 1.3.5.3 --- Feedback regulation of cholesterol --- p.13 / Chapter 1.4 --- Bile acid metabolism --- p.13 / Chapter 1.4.1 --- The function of bile acid --- p.13 / Chapter 1.4.2 --- Bile acid synthesis --- p.14 / Chapter 1.4.3 --- Enterohepatic circulation of bile --- p.14 / Chapter 1.5 --- Effect of Dietary composition on the blood cholesterol --- p.15 / Chapter 1.5.1 --- Dietary cholesterol --- p.15 / Chapter 1.5.2 --- Dietary protein --- p.15 / Chapter 1.5.2.1 --- Research history of dietary protein on the cholesterol --- p.15 / Chapter 1.5.2.2 --- Dietary casein --- p.17 / Chapter 1.5.2.3 --- Soy protein --- p.18 / Chapter 1.5.2.4 --- Buckwheat protein --- p.18 / Chapter 1.5.2.5 --- Mechanism of dietary protein on the cholesterol --- p.18 / Chapter 1.5.3 --- Dietary fiber --- p.18 / Chapter 1.5.4 --- Other functional components in the diet --- p.19 / Chapter 1.5.4.1 --- Phytosterol --- p.19 / Chapter 1.5.4.2 --- Dietary flavonoids --- p.21 / Chapter 1.6 --- Chemical composition of Tartary buckwheat --- p.22 / Chapter 1.6.1 --- Buckwheat protein --- p.22 / Chapter 1.6.2 --- Dietary fiber --- p.23 / Chapter 1.6.3 --- Phytosterols --- p.23 / Chapter 1.6.4 --- Flavonoids --- p.23 / Chapter Chapter 2 --- Effect of Tartary Buckwheat Flour on Blood Cholesterol Level in Male Hamsters / Chapter 2.1 --- Introduction --- p.25 / Chapter 2.2 --- Objective --- p.27 / Chapter 2.3 --- Materials and methods --- p.27 / Chapter 2.3.1 --- Hamsters --- p.27 / Chapter 2.3.2 --- Diets --- p.28 / Chapter 2.3.3 --- "Determination of plasma lipid, lipoproteins" --- p.30 / Chapter 2.3.4 --- Determination of cholesterol concentration in organs --- p.31 / Chapter 2.3.5 --- Determination of fecal neutral and acidic sterols output --- p.31 / Chapter 2.3.6 --- "Western blotting of liver SREBP-2, LDLR, HMGR, LXR and CYP7A1 proteins" --- p.36 / Chapter 2.3.7 --- "Real-Time PCR Analysis of mRNA or Liver SREBP-2, LDLR, HMGR, and CYP7A1 and Small Intestine NPC1L1, ABCG5, ABCG8, ACAT2, and MTP" --- p.37 / Chapter 2.3.8 --- Intestinal ACAT2 activity measurement --- p.37 / Chapter 2.3.9 --- Statistics --- p.39 / Chapter 2.4 --- Results --- p.40 / Chapter 2.4.1 --- Nutritional composition of different flours --- p.40 / Chapter 2.4.2 --- "Growth, food intake and relative organ weights" --- p.44 / Chapter 2.4.3 --- Effect of different flour diets on the plasma lipid profile --- p.44 / Chapter 2.4.4 --- Effect of different flour diets on organ cholesterol of hamsters --- p.44 / Chapter 2.4.5 --- Cholesterol balance and excretion of fecal neutral and acidic Sterols --- p.44 / Chapter 2.4.6 --- "Effect of different flour diets on hepatic SREBP-2, HMGR, LDLR and CYP7A1 immunoreactive mass" --- p.51 / Chapter 2.4.7 --- "Effect of different flour diets on intestinal ABCG5, ABCG8, NPC1L1, MTP, and ACAT2 immunoreactive mass" --- p.54 / Chapter 2.4.8 --- Effect of different diet group on intestinal ACAT activity --- p.54 / Chapter 2.5 --- Discussion --- p.57 / Chapter 2.6 --- Summary --- p.61 / Chapter Chapter 3 --- Effect of DefattedTartary Buckwheat Protein Extract on Blood Cholesterol Level in Male Hamsters / Chapter 3.1 --- Introduction --- p.62 / Chapter 3.2 --- Objective --- p.63 / Chapter 3.3 --- Materials and methods --- p.63 / Chapter 3.3.1 --- Hamsters --- p.63 / Chapter 3.3.2 --- Diets --- p.63 / Chapter 3.3.3 --- "Determination of plasma lipid, lipoproteins" --- p.66 / Chapter 3.3.4 --- Determination of cholesterol concentration in organs and fecal neutral and acidic sterols output --- p.66 / Chapter 3.3.5 --- "Western blotting of liver SREBP-2, LDLR, HMGR and CYP7A1 proteins" --- p.66 / Chapter 3.3.6 --- "Real-Time PCR Analysis of mRNA or Liver SREBP-2, LDLR, HMGR, and CYP7A1 and Small Intestine NPC1L1, ABCG5, ABCG8, ACAT2, and MTP" --- p.66 / Chapter 3.3.7 --- Intestinal ACAT2 activity measurement --- p.67 / Chapter 3.3.8 --- Protein digestibility determination --- p.67 / Chapter 3.3.9 --- Statistics --- p.67 / Chapter 3.4 --- Results --- p.68 / Chapter 3.4.1 --- Diet composition --- p.68 / Chapter 3.4.2 --- "Growth, food intake, fecal excretion" --- p.72 / Chapter 3.4.3 --- Relative organ weights and organ cholesterol concentration --- p.72 / Chapter 3.4.4 --- Effect of different defatted protein extracts on the plasma lipid profile --- p.76 / Chapter 3.4.5 --- Cholesterol Balance and Excretion of Fecal Neutral and Acidic Sterols --- p.76 / Chapter 3.4.6 --- "Apparent protein digestibility in casein, TBP, WP and RP diet groups" --- p.77 / Chapter 3.4.7 --- "Effect of different defatted protein extracts on hepatic SREBP-2, HMGR, LDLR and CYP7A1 immunoreactive mass" --- p.83 / Chapter 3.4.8 --- "Effect of different defatted protein extracts on intestinal ABCG5, ABCG8, NPC1L1, MTP, and ACAT2 immunoreactive mass" --- p.83 / Chapter 3.5 --- Discussion --- p.87 / Chapter 3.6 --- Summary --- p.91 / Chapter Chapter 4 --- Conclusion --- p.92 / References --- p.94

acase@tulane.edu / G-protein coupled receptors (GPCRs) represent the largest family of receptor proteins in the living world, having approximately 800 human genes predicted. GPCRs are therapeutically relevant, as approximately 35% of all drugs on the market target them. However, these drugs can be associated with unwanted side effects, due to ubiquitous GPCR expression throughout the body, as well as GPCRs within the same family having common ligands, but varying or contradictory responses. Although there are approximately 800 human GPCRs predicted, the high-resolution crystal structure of only 41 GPCRs are available. The first human GPCR to be crystallized was the β2-adrenergic receptor (β2AR) in 2007. Shortly thereafter an alternate crystal form of the β2AR revealed a specific cholesterol binding site between helices I, II, III and IV. From this work a cholesterol consensus motif (CCM) was established, which defined specific interactions between cholesterol and the receptor. Utilization of this CCM predicted that as many as 25% of all class A GPCRs could have a specific interaction with cholesterol at this site, including the Adenosine A2a receptor. However, the first crystal structure of the human Adenosine A2a receptor (bound to an antagonist) revealed a lipid, not cholesterol bound at the CCM. Simulations have given insight into other potential binding sites for cholesterol on the Adenosine A2a receptor’s helices 5 and 6, based on the crystal structure bound to an antagonist. Cholesterol has been shown to modulate the activity of multiple G Protein-coupled receptors (GPCRs), yet whether cholesterol acts through specific interactions, or indirectly via modifications to the membrane is not well understood. Here we report that the activity of the adenosine A2a receptor (A2aR) is dependent on cholesterol as determined by membrane cholesterol depletion with methyl-beta-cyclodextrin (MβCD). We also tested whether a specific interaction occurs between A2aR and cholesterol, by testing the impact of mutational changes to predicted cholesterol binding sites on functional consequences. Understanding how cholesterol modulates GPCRs could help in the design of superior drugs to target different cell types with varying membrane cholesterol concentrations, or in disease states where cholesterol homeostasis is disrupted. Additionally, the widespread use of cholesterol reducing drugs such as statins, has posed the new question of how plasma membrane cholesterol concentrations and activation of membrane embedded proteins are affected by these drugs. / 1 / Claire McGraw

Cellular cholesterol homeostasis ensures that cells are able to acquire enough cholesterol for cellular needs including membrane renewal, but avoid excess accumulation. Cholesterol homeostasis is maintained by two transcription factors, the Sterol Regulatory Element-Binding Protein-2 (SREBP-2) which regulates the genes involved in cholesterol biosynthesis and uptake; and the Liver X Receptor (LXR), which regulates the genes involved in maintaining Reverse Cholesterol Transport (RCT). Both LXR and SREBP-2 are regulated by oxidized cholesterol derivatives, referred to as oxysterols. 24(S),25-epoxycholesterol (24,25EC) is unique amongst the physiologically produced oxysterols as it is synthesized de novo in the mevalonate pathway. The overall aim of this thesis was to investigate the role of 24,25EC in cellular cholesterol homeostasis. In Chapter 3, using the statin class of HMG-CoA reductase inhibitors, we showed that 24,25EC synthesis correlates with the expression of the LXR-target gene, the ATP-binding cassette transporter, A1 (ABCA1) in human macrophages. In Chapters 4 and 5, we investigated this further showing that the effect of statins on ABCA1 transcription can be modulated by cellular cholesterol status and the extent of macrophage differentiation. Our results also indicated a critical role of SREBP-2 as a positive regulator of ABCA1 transcription by enabling the generation of 24,25EC. In Chapter 6, we showed that 24,25EC synthesis paralleled cholesterol synthesis, suggesting that this oxysterol may protect against the accumulation of newly-synthesized cholesterol. In Chapter 7, we investigated this contention further using a novel strategy of overexpressing the enzyme 2,3-Oxidosqualene Cyclase (OSC) in Chinese Hamster Ovary cells to selectively inhibit 24,25EC synthesis. In cells lacking 24,25EC, fine-tuning of the acute regulation of cholesterol homeostasis was lost, supporting the hypothesis that 24,25EC functions to protect the cell against the accumulation of newly-synthesized cholesterol. In Chapter 8, we focused on 24,25EC in a neurological context since cholesterol is an essential component of the central nervous system. Both astrocytes and neurons had the capacity to synthesize 24,25EC. Furthermore, both added 24,25EC and stimulated cellular production of 24,25EC modulated expression of LXR and SREBP-2 target genes. The evidence present in this thesis demonstrates the importance of 24,25EC as an endogenously synthesized regulator of cholesterol homeostasis.

Oxygen balance is very important and tightly regulated in mammals. Under hypoxia, hypoxia inducible factor 1(HIF-1) dimerizes with hypoxia inducible factor 1± (HIF-) and activates expression of several genes. Using a mammalian two hybrid assay, we found that HIF-1 interacted with sterol response element binding protein 1a (SREBP1a). SREBP1a regulates transcription of HMG-CoA reductase via binding to the sterol response element (SRE) in the promoter region. HMG-CoA reductase is the rate-limiting enzyme in cholesterol biosynthesis. The interaction between SREBP1a and HIF-1suggests that HIF-1 may play an important role in regulation of cholesterol biosynthesis. We tested the effects of hypoxia on the HMG-CoA reductase. We found that hypoxia caused suppression of SRE-driven luciferase reporter gene expression. HMG-CoA reductase mRNA levels decreased under hypoxia in both hepatoma cells and mouse primary hepatocytes. Electrophoretic mobility shift assay showed that HIF-1 blocked binding of SREBP1a to the SRE sequence in vitro. Ectopic expression of HIF-1 suppressed the SRE- driven luciferase reporter gene expression in BPR cells (HIF-1). Our results suggest that hypoxia inhibits cholesterol biosynthesis by suppressing SREBP1a-regulated gene expression and this suppression is caused by the blockage of SREBP1a binding to SRE sequence by HIF-1.

No description available.