Dissertations / Theses on the topic 'Hypercholesterolaemia'

Familial Hypercholesterolaemia (FH) is an autosomal dominant disorder with a frequency of 1 in 250 to 500 in most European populations. It is characterised by a raised low density lipoprotein-cholesterol (LDL-C) and a high incidence of premature coronary heart disease (CHD). There are three genes where mutations are known to cause FH: the low-density lipoprotein receptor (LDLR) gene, the apolipoprotein B (APOB) gene and the pro-protein convertase subtilisin/kexin type 9 (PCSK9) gene. An FH-causing mutation can be found in around 40% of patients with a possible diagnosis of FH. It has been suggested that the patients with a clinical diagnosis of FH where no mutation were found might have a polygenic cause for their raised LDL-C. FH disorder is an under-diagnosed condition in many countries such as Poland. An analysis of a Polish FH cohort in this thesis, demonstrated the heterogeneous aetiology of FH. We found 39 different pathogenic mutations in the LDLR gene with 10 of them being novel and an overall detection rate of 43.4%. The aim of this thesis was to compare preclinical atherosclerosis between patients with monogenic FH and subjects with polygenic hypercholesterolaemia by means of a neck ultrasound to measure carotid Intima Media Thickness and a cardiac CT scan to assess coronary artery calcification. This study showed that preclinical atherosclerosis was greater in patients with monogenic FH. Lipoprotein(a) [Lp(a)] is a well-known biomarker for CHD risk prediction. The Lp(a) concentration and its association with two LPA single nucleotide polymorphisms (rs3798220 and rs6919346) were assessed in FH patients participating in the Simon Broome registry and a group of the general population participating in the Northwick Park Heart Study II. The results showed that the Lp(a) concentration and the frequency of rs3798220 was significantly higher in the FH patients compared to the general population.

Cholesterol is an integral part of the cell membrane and influences both its fluidity and activity. Furthermore membrane lipids are influenced by plasma lipid levels. In familial hypercholesterolaemia (FH) red cell rheology and platelet aggregation is abnormal. These effects could be due to changes in plasma lipids since they are improved by cholesterol reduction. Section A Fasting lipid profile, membrane cholesterol, red cell ghost and platelet fluidity [using diphenyl hexatriene (DPH) and trimethylamino-diphenyl hexatriene (TMA-DPH) fluorescence anisotropy] and Ca2+ -Mg2+ -ATPase activity (using [32P] ATP hydrolysis) in 30 patients with heterozygous FH and 19 controls were compared before and after treatment with colestipol (10g) , simvastatin (10mg) and maxepa (10g). The two groups were generally comparable with respect to age, sex, BMI and blood pressure. In FH plasma cholesterol, membrane cholesterol, TMA-DPH anisotropy and red cell Ca2+ -Mg2+ -ATPase were increased whilst platelet Ca2+ -Mg2+ -ATPase was reduced (p<0.05). Colestipol and simvastatin reversed these changes toward controls. Maxepa increased DPH anisotropy and reduced Ca2+-Mg2+ -ATPase. In FH membrane fluidity and activity is altered and normalised by cholesterol lowering. This may account for the abnormal cell function in this condition. Section B A double blind, randomized, placebo controlled trial in hypercholesterolaemia (cholesterol 6.5-10 mmol/l, triglycerides <3 mmol/l). After 8 weeks of an AHA step I diet, patients were randomized to A: placebo; B: 5g colestipol + 10mg simvastatin; or C: 10g colestipol + 10mg simvastatin for 8 weeks. Patients were assessed 4 weekly. 44 patients were screened, 32 completed the study. Groups were generally comparable (group C patients were older p<0.02). Active treatment resulted in a 38% reduction in LDL cholesterol (p0.8). Mild gastro-intestinal upset was the commonest adverse event. Low dose combinations are effective and well tolerated. There is no apparent advantage of 10g over 5g colestipol when combined with l0mg simvastatin. HDL cholesterol and triglycerides were unchanged.

Familial Hypercholesterolaemia (FH) is a common autosomal dominant disorder of the defective plasma clearance of LDL-cholesterol. Mutations in three genes, LDLR/APOB/PCSK9, can be detected in 60-90% of definite FH patients. DNA-based testing for FH mutations has important clinical utility and is recommended by the UK and European guidelines to identify affected relatives. This thesis aimed to determine the frequency and spectrum of FH mutations in two independent cohorts of FH patients (from one Oxford lipid clinic, and of Indian background). The FH mutation spectrum was shown to be highly heterogeneous and the mutation detection rate was significantly dependent on the pre-treatment total cholesterol and triglyceride levels. This project also validated the findings that a proportion of clinically diagnosed FH patients have a polygenic cause of hypercholesterolaemia due to an accumulation of common mild LDL-C-raising alleles by analysing LDL-C gene score in 88 mutation negative and 21 mutation positive FH patients, and by replicating the results in further 231 FH patients. A high-throughput DNA sequencing method was assessed as a novel diagnostic tool for detection of FH mutations, and compared it with the currently used methods. This highlighted the need for updating the current FH mutation screening methods as well as the need for more efficient bioinformatics for the next generation sequencing data analysis. Lastly, whole exome sequencing of 125 definite FH patients with no mutations detected in known genes was performed to identify novel monogenic causes of FH. Variants in two genes, CH25H and INSIG2, were identified as potential novel FH mutations. Overall, the results of this thesis demonstrate the heterogeneous FH aetiology and help to understand the genetic architecture of the disease.

Familial hypercholesterolaemia (FH) is an autosomal dominant genetic disorder characterised by elevated levels of plasma low density lipoprotein (LDL) cholesterol. Loss-of-function mutations in the Low-Density Lipoprotein Receptor (LDLR) gene are responsible for ~85% of all FH cases. Heterozygous FH (HeFH) affects 1 in 200-500 individuals world-wide, while homozygous FH (HoFH) has a prevalence of one in a 160000-1000000. Elevated levels of LDL-cholesterol in the blood lead to an accelerated development of atherosclerosis and progression to coronary heart disease. The work presented in this thesis investigates two novel molecular therapies for the treatment of FH. A small molecule approach for the treatment of HeFH and a non-viral gene therapy for the treatment of HoFH. Small molecule: Through a compound screen, we identified a series of small molecules able to upregulate the LDLR at nanomolar potencies, via a mechanism distinct from statins. We then went on to elucidate the mechanism of action as inhibitors of squalene synthase and demonstrated, that when used in combination with statins, these compounds give a much greater increase in LDLR expression than can be achieved with statins alone. By carrying out extensive structure-activity relationship studies we improved the potency and pharmacokinetics of our lead compound and demonstrated in vivo efficacy. Non-viral gene therapy: We have previously generated mini-gene vectors carrying the human LDLR cDNA, driven by 10 kb of genomic DNA from the native human LDLR locus, encompassing the promoter region. To further enhance LDLR transgene expression, a miRNA, targeting Hmgcr (miR82) was cloned into the same vector, generating a combinatorial RNAi-LDLR vector. In vivo, we show the RNAi-LDLR vector is able to significantly reduce total and LDL-cholesterol in Ldlr^-/- mice fed a high cholesterol diet twelve weeks post-delivery. The LDL-cholesterol lowering resulted in reduced atherosclerosis in the RNAi-LDLR vector treated mice. Here we demonstrate for the first time, that an episomal non-viral vector is able to significantly reduce LDL-cholesterol and slow the progression of atherosclerosis in a mouse model of FH.

Includes bibliographical references.
The primary objective was to substitute phytosterol and regula margarine into the diets of familial hypercholesterolaemia (FH) patients and to investigate whether the phytosterol containing margarine had a cholesterol lowering effect. Another objective was to investigate immune responses to phytosterols. A randomized double-blind cross-over study in which subjects with FH consumed 20g of regular or phytosterol margarine for a period of 6 weeks on each margarine, after which a washout period of 4 weeks was followed. There were 5 visits in total, visit 1 being the collection of baseline data, visit 2 the randomisation to either of the regular of phytosterol margarine periods, visit 3 for the cross-ver to the alternative margarine form, visit 4 marked the onset of washout period and visit 5, was the last evaluation visit. Margarine use was in addition to current, albeit limite, pharmacological treatment with statins.

The expression of two phenotypically-contrasting LDL receptor mutations was characterized in cultured fibroblasts from the genetically-homozygous Afrikaner subjects, FH1a and lb, and FH3a and 3b, respectively. Surface receptor expression and functional activity were studied by ligand (¹²⁵I-LDL) and monoclonal antibody (¹²⁵I-IgG-C7) binding, and c35s]-methionine pulse-chase experiments were used to analyze biosynthesis, processing and degradation of IgG-C7- immunoprecipitable mutant receptors. Cells from the "receptor-negative" subjects, FH3a and 3b exhibited reduced, but significant (40-60% of normal) LDL receptor synthesis rates. Newly-synthesized precursors were processed slowly (t½ 1.5 hours versus normal t½ of approximately 15 minutes) to mature receptors which reached the cell-surface, but were rapidly degraded thereafter with a half-life of approximately 1.7 hours (normal value 12.6 hours) thus representing a new type of LDL receptor defect. Lysosomotropic weak bases such as ammonium chloride partially inhibited rapid degradation of the mutant receptors, suggesting the involvement of proteolysis in acidic compartments such as lysosomes or endosomes. Fibroblasts from FH1a and lb exhibited normal synthesis rates of LDL receptor precursors that were processed at a severely reduced rate (t½ approximately 5 hours) to functionally heterogeneous mature surface receptors. Onethird of the receptors (20% of normal levels) bound ¹²⁵I-LDL with normal affinity at 4°C and 37°C, whereas the majority were able to recognize only ¹²⁵I-IgG-C7, and apparently showed defective internalisation and subsequent degradation of the bound IgG-C7 at 37°C. The existence of the two receptor populations was further supported by selective intracellular trapping and degradation of only the active, LDL-binding population, in the presence of ammonium chloride and LOL. The abnormal form predominated even in newly-synthesized receptors and reached a maximum of 50-70% of normal levels after 48 hours of upregulation. Upregulation kinetics and degradation rates (t½ = 10-11 hours) of both functionally-active and abnormal receptor populations were similar to normal. A progressive increase in apparent molecular weight of the slowly-processed precursor receptors suggested a possible role for abnormal glycosylation in the formation of both "normal" and abnormal conformations of the same receptor molecule.

Essential hypertension and hyperlipidaemia co-segregate in the population and act synergistically in increasing coronary heart disease. Using invasive procedures such as brachial artery infusion of drugs, functional abnormalities of the vascular endothelium have been observed in forearm resistance vessels in essential hypertension, and in hypercholesterolaemia. This 'endothelial dysfunction' has been attributed, at least in part, to impaired generation of nitric oxide. However, it is unclear whether this dysfunction is confined to nitric oxide in the forearm vascular bed, or whether other mediators and other microvessels are involved. Hypertension is characterised by increased peripheral vascular resistance, which may be due to functional or structural abnormalities in the microcirculation, particularly affecting pre-capillary vessels which make the greatest contribution to vascular resistance. However, from studies in established hypertension it is not possible to ascertain whether microvascular abnormalities are a cause or consequence of the rise in blood pressure (see below). This thesis, therefore, aims (i) to examine the importance of nitric oxide in maintaining basal vascular tone and in the control of blood pressure, and to investigate the physiological role of nitric oxide in the human skin microcirculation; (ii) to establish the non-invasive technique of drug iontophoresis combined with laser Doppler fluximetry for assessing endothelial function in dermal microvessels in health, examining the principle mediators involved; (iii) to recruit patients with endothelial dysfunction, to determine whether microvascular abnormalities are present and measurable in the dermis using iontophoresis; (iv) to examine whether functional and/or structural abnormalities in the microcirculation are a cause or consequence of hypertension. Inhibition of nitric oxide synthase with systemic administration of l-A^-monomethyl-arginine (l-NMMA) increased blood pressure and substantially increased peripheral vascular resistance, emphasising the importance of nitric oxide in the maintenance of basal vascular tone. This led to an investigation of the role of nitric oxide at the microvascular level - the seat of greatest contribution to peripheral resistance. Using local brachial artery infusion of l-NMMA, tissue-specific differences in response suggested a selective role for nitric oxide in microvessels. To investigate further the role of nitric oxide and other mediators involved in microvascular function in vivo, the non-invasive technique of drug iontophoresis was used in combination with brachial artery infusion of l-NMMA and noradrenaline. Despite substantial reductions in forearm blood flow caused by these drugs, no differences were observed in dermal blood flow. However, aspirin, which blocks prostanoid synthesis via the cyclo-oxygenase pathway, inhibited the dermal response to acetylcholine (ACh). Thus, iontophoresis measured prostanoid-mediated cholinergic vasodilatation in human dermal vessels. With the pharmacological mechanisms of iontophoretic drug delivery emerging, the technique was extended to investigate endothelial function in 10 hypercholesterolaemic patients compared with 10 matched controls. In the patients, forearm vasodilatation was impaired with ACh but not with the endotheliumindependent vasodilator, sodium nitroprusside (SNP). In contrast, dermal vasodilatation was impaired with SNP and not with ACh. In the presence of aspirin, in hypercholesterolaemic patients, forearm vasodilatation to ACh was partially restored, but dermal vasodilatation to ACh was impaired. Aspirin did not affect SNP responses. Therefore, impaired stimulated nitric oxide generation was confirmed in forearm resistance vessels in hypercholesterolaemic patients. In dermal microvessels in these patients, reduced smooth muscle sensitivity to nitric oxide was observed, and an enhanced, possibly compensatory, role for prostanoid vasodilators. To identify the likely importance of microvascular abnormalities in determining the familial risk of essential hypertension, subjects were recruited from an epidemiological model which identifies young people with contrasting predisposition to high blood pressure. In this model, subjects widi a predisposition to high blood pressure which was familial were found to have structural rarefaction with a substantial increase in minimum vascular resistance in dermal microvessels. These structural abnormalities suggest that capillary rarefaction is a cause rather than a consequence of essential hypertension. As hypertension and hypercholesterolaemia are cofactors which increase the risk of cardiovascular, cerebrovascular and renal diseases, an investigative method which would recognise individuals at risk would be a useful adjunct to their clinical management. Functional abnormalities are present in established essential hypertension, and in hypercholesterolaemia. The non-invasive technique of drug iontophoresis provides important new information about differences between dermal microvessels and forearm resistance vessels. However, further investigation is required if this technique is to become established in assessing endothelial function in large groups of patients.

This thesis presents a study of the incidence of hypercholesterolaemia in a 1988 sample of male personnel from the Royal Australian Air Force (RAAF). The thesis defines hypercholesterolaemia as a plasma total cholesterol level in excess of the risk threshold promulgated by the National Heart Foundation of Australia (NHF). Since 1981, RAAF personnel have been subjected to a series of regular physical and physiological examinations, including a comprehensive blood lipd profile, as part of a preventive approach to reducing the incidence of coronary heart disease in the Service. The study presents data on the extent of hypercholesterolaemia in the RAAF and illustrates the spread of the condition through various age cohorts. In excess of 40% of the RAAF personnel studied had total cholesterol levels in excess of the NHF "increased risk" threshold with the prevalence varying from 11.8% of personnel aged 20-24 years to some 64% of personnel older than 49 years. The increase in total cholesterol level with increasing age is statistically significant. The study compares these data with previously published observations on certain motivational characteristics of military communities, with a view to providing an evaluation of the structure of the RAAF Health Promotion Programme as a lifestyle based educational initiative. The data are further compared with data from two conceptually similar studies of male personnel in the United States armed forces. These comparisons suggest that the age related frequency of educational intervention which characterized the 1988 RAAF programme may well have been inappropriate for optimal effect. The study further utilizes a tabular (matrix) analysis of a number of educational strategies to identify several preferred option(s) for programme design in the RAAF community. The analysis suggests that the present mode of educational intervention may also be inappropriate for optimal effect (when addressing the 1988 RAAF population at least). The study concludes with the presentation of a series of recommendations aimed fundamentally at a redesign of the cholesterol monitoring component of the RAAF Health Promotion Programme. The study recommends, however, that given the multifactorial nature of coronary heart disease risk, the cholesterol monitoring component of the programme should not be redesigned in isolation. Instead, the findings of this study suggest that a more wide-ranging study of the educational basis for the client interface to the RAAF Health Promotion Programme is warranted.

Introdução: Hábitos alimentares (HAs) inadequados têm importante relação com hipercolesterolemia, que constitui fator de risco para doenças cardiovasculares. Objetivo: Analisar HAs de idosos hipercolesterolêmicos, atendidos em ambulatório da cidade de São Paulo. Métodos: Participaram do estudo idosos ( 60 anos), de ambos os sexos, voluntários, com prontuários ativos no Centro de Referência do Idoso José Ermírio de Moraes (CRIJEM), com diagnóstico de hipercolesterolemia ou prescrição de medicamento hipolipemiante. O método de inquérito utilizado foi história alimentar. As variáveis de estudo foram: número de refeições diárias, ingestão habitual de alimentos, incluindo água e bebidas alcoólicas, necessidade de ajuda para se alimentar, companhia às refeições e local (fora e dentro da residência) das mesmas. O critério adotado, para considerar hábito alimentar do grupo, foi a frequência de ingestão 50 por cento . Resultados: Foram analisados os HAs de 106 idosos, com idade entre 60 e 84a (média de 69,3 anos - DP=5,9), 86 por cento do sexo feminino, 58 por cento moravam com familiares, e 93 por cento apresentaram prescrição de medicamento hipolipemiante. O número médio de refeições diárias foi 5,0 (DP=1,2), um terço dos idosos referiu ingestão diária 6 a 8 copos de água, e a maioria (78 por cento ) negou ingestão de bebidas alcoólicas. Quanto às respostas com múltiplas alternativas, 71 por cento das citações mostraram não haver necessidade de ajuda, 89 por cento indicaram realizar as refeições na própria residência, e cerca da metade (49 por cento ) informaram não ter companhia. Foram considerados como HAs os seguintes alimentos e preparações: café, leite, pão francês, arroz, feijão, carne bovina e de frango, alface crua, banana, óleos de oliva e de soja. Conclusões: Nesse grupo, constatou-se pequena variedade de alimentos, especialmente em relação a hortaliças e frutas, e ausência de hábito alimentar para ingestão de água
Introduction: Inadequate eating habits are strongly related to hypercholesterolaemia, a risk factor for cardiovascular diseases. Objective: This study aims the analysis of food habits of hypercholesterolaemic elderly patients attending to an ambulatory unit in São Paulo city. Methods: Among the patients who were attending to Centro de Referência do Idoso José Ermírio de Moraes (CRIJEM), 106 elders ( 60y) volunteered to be interviewed about their eating habits; all of them were hypercholesterolaemic or were on hypolipaemic drugs. It was used an inquiry on dietetic history. Data was collected on the number of daily meals, their habitual ingestion of food (water and alcoholic beverages included), the need for help for feeding themselves, if they eat accompanied by anyone, and where do they have their meals. Any food was included as part of their customary diet only if it was mentioned by at least 50 per cent of the group. Results: The eating habits of 106 elders were evaluated. Their age ranged from 60 to 84 years old, mean age was 69,3 years (±5,9), 86 per cent of them were female, 58 per cent were living with family members, and 93 per cent were prescribed hypolipaemic drugs. The average frequency of meals was 5,0 (±1,2), one third drank 6 to 8 glasses of water a day, and 78 per cent denied alcohol consumption. Besides, 71 per cent of the answers indicated no need for any help in feeding themselves, 89 per cent had their meals at their own house, and almost half of them ate alone. The following foods were part of the dietary habits of this group: coffee, milk, white bread, rice, beans, red meat and poultry, lettuce, banana, olive and soy oils. Conclusions: This group demonstrated a small variety of foods, specially fruit and vegetables, and absence of water

This thesis provides an empirical investigation of the geneticisation thesis. Geneticisation is one of the most prominent critiques of the social and cultural implications of developments in genetics. It incorporates a set of claims and expectations about the way genetic knowledge and technologies are transforming or will transform ideas about health and illness, and health care practices. This research aims to explore the empirical basis of these claims, by looking at the place of genetic discourses and practices in one specific area. The thesis focuses on familial hypercholesterolaemia (FH), a treatable hereditary cholesterol condition associated with high rates of coronary heart disease (CHD). It asks how much and in what ways patients with FH and professionals involved with the condition construct FH and CHD as genetic conditions. The thesis draws on three main areas of data - biomedical literature concerning CHD and FH; ethnographic work concerning the activities of HEART UK, the main UK health charity involved with inherited lipid disorders and cholesterol; and interviews with patients with FH and with staff and members of HEART UK. The analysis suggests that FH is not understood or managed within a strong genetic frame, and that neither professionals involved in HEART UK, nor patients with FH, provided or contributed to radically new or geneticised accounts of CHD. In short, the research suggests that geneticisation overstates the transformatory potential of genetics, and that factors such as the availability of effective therapeutics, the sites where care takes place, the disciplines involved, and existing lay and professional models of disease are important for the construction of a particular field. Furthermore, in arguing that FH is not associated with a strong specific disease identity or community, the analysis questions the notion of biosociality, suggesting that is may be less relevant to some biological states or conditions than to others.

Ingestion of vegetables rich in inorganic nitrate (NO3-) content has emerged as an effective method, via the formation of a nitrite (NO2-) intermediate, for acutely elevating vascular nitric oxide (NO) levels. As such a number of beneficial effects of NO3- ingestion have been demonstrated including the suggestion that platelet reactivity is reduced. I initially investigated whether inorganic NO3- supplementation might also reduce platelet reactivity in healthy volunteers and have determined the mechanisms involved in the effects seen. I conducted a randomised crossover study in 24 (12 of each sex) healthy subjects assessing the acute effects of potassium nitrate capsules (KNO3, 8 mmol) vs placebo (KCl) control capsule ingestion on platelet reactivity. Inorganic NO3- ingested via supplementation raised circulating NO3- and NO2- levels in both sexes and attenuated ex vivo platelet aggregation responses to adenosine diphosphate (ADP) and, albeit to a lesser extent, collagen but not epinephrine in male but not female volunteers. These inhibitory effects were associated with a reduced platelet P-selectin expression and elevated platelet cyclic guanosine monophosphate (cGMP) levels. In addition, I have shown that NO2- reduction to NO occurs at the level of the erythrocyte and not the platelet. These results demonstrate that inorganic NO3- ingestion, whether via the diet or through supplementation, results in a modest decrease in platelet reactivity in healthy males. I then sought to examine the effects of 6 weeks daily intake of NO3--rich beetroot juice versus a placebo NO3--deplete juice on endothelial and platelet function in a cohort of otherwise healthy non-diabetic untreated hypercholesterolaemics. In this randomised double blind placebo controlled parallel study 69 subjects were recruited. The primary end point was change in endothelial function determined using ultrasound flow-mediated dilatation (FMD). Secondary endpoints included change in pulse wave analysis (PWA), aortic pulse wave velocity (aPWV), platelet P-selectin and platelet monocyte aggregate (PMA) expression and plasma, urine and salivary NO3- and NO2- levels. Baseline characteristics, including lipid levels, were similar between the groups. Dietary NO3- caused an improvement in FMD of ~24% from 4.6%±2.2% to 5.7%±2.6% in the treatment group (p<0.001) not seen in the placebo group (4.5%±1.9% versus 4.3%±1.8% p=0.07). This improvement in FMD was also noted following acute administration of dietary NO3-. Small but significant improvements also occurred in aPWV and PWA augmentation index (p=0.04). The % of platelet monocyte aggregates was significantly reduced in the NO3- limb by 7.6% versus an increase of 10.1% in the placebo group (p=0.004). No adverse effects of dietary NO3- were detected. In this study population, chronic dietary NO3- ingestion improves endothelial function, vascular stiffness and platelet markers of atherogenesis in a cohort of hypercholesterolaemics who are otherwise at increased risk of cardiovascular disease (CVD). This thesis provides strong support for assessment of the potential of dietary NO3- as a primary prevention strategy to prevent atherothrombotic and atherogenic complications in larger cohorts.

Thesis (MNutrition (ITE))--Stellenbosch University, 2012.
ENGLISH ABSTRACT: Background Statins are drugs of known efficacy in the treatment of hypercholesterolaemia. However, statin-induced myopathy, an adverse effect of statins in up to 15% of its users, has warranted a reduction in the prescription dose or discontinuation of the drug. The exact mechanism of statin-induced myopathy is unknown, but the potential of Coenzyme Q10 (CoQ10) as treatment has been recognized due to decreased human plasma CoQ10 levels found after statin use and the concomitant role of CoQ10 in muscle function. Objectives This systematic review assessed the effect of CoQ10 supplementation on: the severity of statin-induced myopathic symptoms, levels of plasma creatine kinase, intramuscular and plasma CoQ10, as well as whether any adverse effects of CoQ10 supplementation such as abdominal pain, nausea and vomiting or headaches were experienced. Search methods Two searches for studies were conducted in The Cochrane Central Register of Controlled Trials (inception to March 2011 and inception to November 2011), MEDLINE (inception to March 2011 and inception to November 2011), Web of Science (inception to March 2011 and inception to November 2011), Science Direct (inception to March 2011 and inception to February 2012), Wiley Online Library (inception to March 2011 and inception to February 2012), Springerlink (inception to April 2011 and inception to February 2012), EBSCOhost [Academic Search Premier and CAB abstracts (inception to March 2011 and inception to February 2012), CINAHL (inception to March 2011 and inception to November 2011)], Scopus (inception to March 2011 and inception to November 2011) and Google Scholar (inception to March 2011 and inception to February 2012). Reference lists of articles were hand searched for relevant clinical trials. Only trials with a full text were included in the review. Selection criteria Randomised controlled trials (RCTs) were included with adult participants (mean of 18-64.99 years) of all race/ethnic groups and gender on statin therapy with reported myopathic symptoms from an unknown cause. The intervention was in the form of a pure oral supplement of CoQ10 irrespective of dose, duration and frequency, and the control in the form of a placebo, a similar antioxidant, or no intervention. Outcomes included the severity of myopathic symptoms, levels of plasma creatine kinase (U/L), intramuscular CoQ10 (μmol/kg) and plasma CoQ10 (μmol/L), as well as adverse effects of CoQ10. Data collection and analysis The principle investigator and one independent reviewer selected the studies, extracted data and assessed for risk of bias using the Cochrane Collaboration‘s tool for assessing risk of bias. Authors of relevant clinical trials were contacted for additional information. Results Two RCTs were included in the review, totaling 76 participants. A meta-analysis could not be performed, thus the review is narrative. There were an insufficient number of RCTs to confirm whether routine supplementation of CoQ10 improves statin-induced myopathic symptoms. Conclusions More and larger RCTs are required to determine the efficacy of CoQ10 supplementation in statin-induced myopathy. Consensus needs to be reached regarding the definition and measurement instrument/s of myopathy so that results of future studies can easily be compared and synthesized.
AFRIKAANSE OPSOMMING: Agtergrond Statiene is medikasie bekend vir die effektiewe behandeling van hipercholesterolemie. Statien-geïnduseerde miopatie is egter 'n newe-effek wat voorkom in tot 15% van gebruikers, wat 'n vermindering in die voorgeskrewe dosis of staking van die medikasie tot gevolg het. Die presiese meganisme van statien-geïnduseerde miopatie is onbekend, maar die potensiaal van Koënsiem Q10 (CoQ10) is geïdentifiseer as 'n moontlike behandeling aangesien menslike plasma CoQ10 vlakke verlaag na die gebruik van statiene en as gevolg van die rol van CoQ10 in spierfunksie. Doelwitte Hierdie sistematiese literatuuroorsig het die effek van CoQ10 supplementasie bepaal op: die graad van statien-geïnduseerde miopatiese simptome, plasma kreatien kinase vlakke, intra-muskulêre en plasma CoQ10 vlakke, asook die teenwoordigheid van enige newe-effekte van CoQ10 supplementasie soos abdominale pyn, naarheid en braking of hoofpyne. Soektogstrategie Twee soektogte vir studies is uitgevoer in The Cochrane Central Register of Controlled Trials (ontstaan tot Maart 2011 en ontstaan tot November 2011), MEDLINE (ontstaan tot Maart 2011 en ontstaan tot November 2011), Web of Science (ontstaan tot Maart 2011 en ontstaan tot November 2011), Science Direct (ontstaan tot Maart 2011 en ontstaan tot Februarie 2012), Wiley Online Library (ontstaan tot Maart 2011 en ontstaan tot Februarie 2012), Springerlink (ontstaan tot April 2011 en ontstaan tot Februarie 2012), EBSCOhost [Academic Search Premier en CAB abstracts (ontstaan tot Maart 2011 en ontstaan tot Februarie 2012), CINAHL (ontstaan tot Maart 2011 en ontstaan tot November 2011)], Scopus (ontstaan tot Maart 2011 en ontstaan tot November 2011) en Google Scholar (ontstaan tot Maart 2011 en ontstaan tot Februarie 2012). Verwysingslyste van artikels is ook met die hand nagegaan vir relevante kliniese proewe. Slegs kliniese proewe waarvan die volteks beskikbaar was, is ingesluit in die oorsig. Seleksiekriteria Ewekansige gekontroleerde proewe (EGP) is ingesluit met volwasse deelnemers (gemiddeld 18-64.99 jaar) van alle rasse/etniese groepe en geslag op statien-terapie met gerapporteerde miopatie simptome van onbekende oorsaak. Die intervensie was 'n suiwer orale supplement van CoQ10 ongeag die dosis, duurte en frekwensie, en die kontrole 'n plasebo, soortgelyke antioksidant, of geen intervensie. Uitkomste het ingesluit: die graad van miopatie simptome, vlakke van plasma kreatien kinase (U/L), intra-muskulêre CoQ10 (μmol/kg) en plasma CoQ10 (μmol/L), sowel as newe-effekte van CoQ10. Dataversameling en -analise Die hoof ondersoeker en een onafhanklike hersiener het die seleksie van studies en data-ekstraksie onderneem en die risiko vir sydigheid geassesseer deur gebruik te maak van die Cochrane Collaboration’s tool for assessing risk of bias. Outeurs van relevante kliniese proewe is geraadpleeg vir addisionele inligting Resultate Twee EGP is ingesluit in die oorsig met 'n totaal van 76 deelnemers. 'n Meta-analise kon nie uitgevoer word nie, dus is die oorsig beskrywend. Daar was te min EGP om te bewys dat roetine supplementasie van CoQ10 statien-geïnduseerde miopatiese simptome verbeter. Gevolgtrekkings Meer en groter EGP is nodig om die effektiwiteit van CoQ10 supplementasie in statien-geïnduseerde miopatie te bepaal. Konsensus moet bereik word ten opsigte van die definisie en metingsinstrument/e van miopatie sodat die resultate van toekomstige studies makliker vergelyk en verwerk kan word.

Cette thèse tente d’apporter, en la sociologisant, des éléments de réponse à une question émanant du monde médical : pourquoi des individus atteints d’hypercholestérolémie et donc surexposés à des risques cardio-vasculaires ne mettent-ils pas en application les conseils diététiques formulés par leur médecin ? Trois phases d’enquête ont été menées : 1) des entretiens d’experts, dans le but d’identifier les controverses qui ont transformé l’hypercholestérolémie en un problème de santé publique majeur ; 2) des entretiens individuels et collectifs auprès de « mangeurs à risques », afin de saisir les principaux freins au suivi des règles hygiéno-diététiques prescrites et les formes de régulation sociale de la pathologie ; et 3) une enquête quantitative, avec passation de questionnaires auprès de 800 individus, ayant pour but la description socio-démographique de la population d’étude et la validation des résultats qualitatifs. Si les rapports à l’alimentation et à la santé apparaissent comme étant en partie surdéterminés par des variables sociales et culturelles, les résultats invitent surtout à resituer les individus dans leurs contextes interactionnels et familiaux. Les décisions alimentaires et les manières de faire face à cette pathologie chronique ne sont pas construites dans une perspective individuelle. Au final, l’articulation entre la sociologie de l’alimentation et les champs de la santé, du risque, de la famille et du genre a permis de renouveler la problématique de l’observance et d’ouvrir un vaste territoire de recherches sur les effets du « manger ensemble » et du « vivre ensemble ». En cela, la thèse entend contribuer à une sociologie de la décision alimentaire
This thesis attends answering a question originated from the medical world, by framing it through a sociological perspective: why do some hypercholesterolaemic individuals not comply with their doctor’s dietetic advice although being overexposed to cardiovascular risks? Three surveys were conducted: 1) some experts’ interviews, so as to identify the controversies that have led hypercholesterolaemia to be considered as a major public health problem; some in-depth interviews and a focus group with “at risk eaters”, in order to grasp the main obstacles to the dietary compliance and the forms of social regulation for the pathology; and 3) a quantitative survey, carried out by questionnaires among 800 people and developed with the aim of collecting sociodemographic data on the population, and validating the qualitative results. Notwithstanding that the relations with food and health are partly over-determined by social and cultural variables, results underlined the need to situate individuals in their interactional and family environment. Neither eating choices nor the ways to cope with this chronic pathology are constructed within an individual perspective. Connecting the sociology of food with other works from the health, risk, family and gender domains eventually enabled revitalizing the compliance issue and initiating a vast field of researches on the effects of “eating together” and “living together”. This thesis therefore intends contributing to a sociology of eating choices

A homeostase do colesterol é mediada por proteínas envolvidas na absorção (NPC1L1), regulação (SREBP1, SREBP2, SCAP), síntese (HMGCR) e remoção plasmática (LDLR). Os fármacos inibidores da síntese (vastatinas) e absorção (ezetimiba) do colesterol são potentes agentes hipocolesterolemiantes. Alterações em vários genes têm sido associadas a diferenças na resposta a diversos agentes terapêuticos. Com a finalidade de estudar os efeitos de hipolipemiantes e polimorfismos sobre a expressão dos genes HMGCR, LDLR, SREBF1a, SREBF2, SCAP e NPC1L1, foram selecionados 25 indivíduos com hipercolesterolemia familial (HF), 72 com hipercolesterolemia não familial (HNF) e 125 indivíduos normolipidêmicos e sem doença cardiovascular (NL). Os indivíduos HF foram tratados com sinvastatina (40 mg/dia/4 sem) combinada ou não com ezetimiba (10 mg/dia/4sem) e os HNF foram tratados com atorvastatina (10 mg/dia/4sem). Amostras de sangue foram obtidas antes e após o tratamento para a extração de DNA e RNA e analise do perfil lipídico sérico. A expressão de mRNA dos genes SREBF1a, SREBF2, SCAP, HMGCR, LDLR e NPC1L1 em células mononucleares do sangue periférico (CMSP) foi determinada por RT-PCR em tempo real empregando-se o gene da GAPD como controle endógeno. Os polimorfismos SREBF1a 36delG, SREBF2 G1784C e SCAP A2386G foram determinados por PCR-RFLP. Os indivíduos HF apresentaram maior expressão de mRNA dos genes NPC1L1, HMGCR e LDLR que os grupos HNF e NL (p<0,05). O efeito da atorvastatina sobre a expressão dos genes estudados parece depender da expressão basal nos indivíduos HNF. A variação da expressão após o tratamento com atorvastatina nos pacientes do grupo HNF esteve correlacionada nos genes: SREBF1a e SREBF2; SREBF1a e SCAP; SREBF1a e LDLR; SREBF2 e SCAP; SREBF2 e LDLR; HMGCR e LDLR. O tratamento com sinvastatina e ezetimiba não modificou o padrão de expressão dos genes estudados no grupo HF. Os polimorfismos SREBF2 G1784C e SCAP A2386G parecem estar relacionados com diminuição da expressão de mRNA após o tratamento com atorvastatina. Foi observado que os portadores do genótipo GG do polimorfismo SREBF2 G1784C apresentaram maiores concentrações séricas de colesterol total e LDL-C após o tratamento com atorvastatina. O polimorfismo SCAP A2386G parece estar associado com maiores concentrações de apoB em pacientes do grupo HNF antes do tratamento com atorvastatina. Os resultados são sugestivos que os genes HMGCR, LDLR e NPC1L1 são regulados diferentemente de acordo com o estado metabólico do indivíduo e a taxa de expressão de mRNA é influenciada pelos polimorfismos SREBF2 G1784C e SCAP A2386G após o tratamento com atorvastatina
The regulation of cholesterol is mediated by proteins involved in the absorption (NPC1L1), regulation (SREBP1, SREBP2, SCAP), synthesis (HMGCR) and removal of plasma cholesterol (LDLR). Potent hypocholesterolemic agents inhibit cholesterol synthesis (statins) and its absortion (ezetimibe). Changes in several genes have been associated to different responses to various therapeutic agents. In order to evaluate the association between genes involved in the metabolism of cholesterol and their response to lipid lowering drugs, patients with familial (FH, n = 25) and non familial hypercholesterolemia (NHF, n = 72) were selected. Additionally, 125 normolipidemic individuals and without cardiovascular disease were selected (NL). The HF group were treated with simvastatin (40 mg/day/4 weeks) combined or not with ezetimibe (10 mg/day/4weeks). The NHF group were treated with atorvastatin (10 mg/day/4weeks). Blood samples were obtained prior to and following treatment for extraction of DNA and RNA, and serum lipid profile analysis. The mRNA expression of SREBF1a, SREBF2, SCAP, HMGCR, LDLR, and NPC1L1 genes was determined by real time RT-PCR using the GAPD gene as endogenous control. The polymorphisms SREBF1a-36delG, SREBF2 G1784C, and SCAP A2386G were determined by PCR-RFLP. Individuals with HF showed higher expression of mRNA of genes NPC1L1, HMGCR and LDLR when compared with HNF and NL groups (p <0.05). The effect of atorvastatin on the gene expression seems to depend on the baseline expression in HNF subjects. The change of expression after treatment with atorvastatin in group HNF was correlated as followed: SREBF1a and SREBF2; SREBF1a and SCAP; SREBF1a and LDLR; SREBF2 and SCAP; SREBF2 and LDLR; HMGCR and LDLR. Treatment with simvastatin and ezetimibe did not change the gene-expression profile in HF group. The polymorphisms SREBF2 G1784C, and SCAP A2386G appear to be related to a decreased expression of mRNA after treatment with atorvastatin. HNF group Carriers of GG genotype of SREBF2 G1784C polymorphism had higher serum concentrations of total cholesterol and LDL-C after therapy. The SCAP A2386G polymorphism seems to be associated with higher concentrations of apoB in patients from HNF group prior to treatment with atorvastatin. The results suggest that the HMGCR, LDLR and NPC1L1 genes are regulated according to the metabolic status of the individual, and the expression rate of mRNA is influenced by SREBF2 G1784C and SCAP A2386G polymorphisms after atorvastatin therapy.

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Obesity represents a global public health problem, being a disorder influenced by genetic and environmental factors. It is an important cause of morbidity and mortality worldwide and the disease characteristics, has been understood as a chronic disorder of inflammatory nature. Several studies of polymorphisms associated with the metabolic syndrome, has been performed in obese children and adolescents, demonstrating significant relationship with this pathology. The several inflammatory cytokines exert significant influence in the development and maintenance of obesity for her role in the regulation of energy homeostasis and metabolism. The aim of this study was to evaluate the association of the polymorphism of SNP 1082G / A in the interleukin-10 gene promoter with the Children and Youth obesity in 48 cases and 45 controls, associated with the lipid profile. In this study case-control was used to ARMS-PCR methodology for allelic determination. Of obese individuals, 27 (56%) had the genotype A / G, followed by 13 (27%) with genotype A / A and 8 (17%) with genotype G / G. Of the analyzed controls, 28 (62%) had Genotype A / G, followed by 9 (20%) with genotype A / A and 8 (18%) with genotype G / G. In the analysis it was found that the presence of the SNP-1082G / A polymorphism was similar in both groups, no representing risk factor to obesity. Also there was a significant difference in HDL cholesterol levels in both groups, linking obesity to low levels of HDL in adolescents and reforcing the relevance of this parameter.
A obesidade corresponde a um problema mundial de saúde pública, sendo uma desordem influenciada por fatores genéticos e ambientais. Consiste em uma importante causa de morbidade e mortalidade no mundo todo e, pelas características da doença, tem sido entendida como uma desordem crônica de natureza inflamatória. Diversos estudos de polimorfismos associados com a síndrome metabólica, tem sido realizados em crianças e adolescentes obesos, demonstrando significativa relação com essa doença. As diversas citocinas inflamatórias exercem influência significativa no desenvolvimento e na manutenção da obesidade por seu papel na regulação da homeostase energética e no metabolismo. O presente estudo teve como finalidade associar o polimorfismo da citocina anti-inflamatória IL-10 e a possível relação desta com a obesidade. Foi avaliado a associação do polimorfismo do SNP 1082G/A no promotor do gene da interleucina-10 com a obesidade infanto juvenil em 48 casos e 45 controles, associando com o perfil lipídico. Nesse estudo tipo caso-controle foi utilizada a metodologia de ARMS-PCR para a determinação alélica. Dos indivíduos com obesidade, 27 (56%) apresentaram o genótipo A/G, seguidos de 13 (27%) com o genótipo A/A e 8 (17%) com o genótipo G/G. Dos controles analisados, 28 (62%) apresentaram genótipo A/G, seguidos de 9 (20%) com genótipo A/A e 8 (18%) com o genótipo G/G. Nas análises foi verificado que a presença do polimorfismo SNP- 1082G/A foi semelhante nos dois grupos, apresentando maior prevalência de heterozigotos para o loco da IL-10. Conclui-se com o presente estudo que as frequências alélicas e genotípicas foram semelhantes nos dois grupos analisados não sendo possível sugerir tal alteração como fator de risco para obesidade. Foi evidenciada uma significativa diferença dos níveis séricos de colesterol HDL, reforçando-se a importância da avaliação desse parâmetro que se mostrou em níveis inferiores nos indivíduos obesos.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior
In mammals, the liver plays a central role in whole-body lipid metabolism. Unfortunately, dysregulation of these pathways has been implicated in hyperlipidemias. In recent years, a significant association between the trace element selenium and hypercholesterolaemia in human and animals has been reported. This study was designed to investigate a potential hypolipidaemic effect of diphenyl diselenide ((PhSe)2) in Triton WR-1339-induced hyperlipidaemia in mice. Triton was administered intraperitoneally (400 mg/kg) to overnight-fasted mice to develop acute hyperlipidaemia. (PhSe)2 was administered orally (10 mg/kg) 30 min before Triton. At 24 h after Triton injection, blood samples were collected to measure plasma lipid levels. The hepatic thiobarbituric acid reactive substances and ascorbic acid levels as well as catalase and glutathione peroxidase activity were recorded. (PhSe)2 administration significantly lowered total cholesterol, non-high- density lipoprotein-cholesterol and triglycerides,whilst it increased high-density lipoprotein-cholesterol levels in plasma of hyperlipidaemic mice. Neither oxidative stress nor the antioxidant effect of (PhSe)2 was observed in the mouse liver in this experimental protocol. These findings indicated that (PhSe)2 was able to lower plasma lipid concentrations. Further studies are needed to elucidate the exact mechanism by which (PhSe)2 exerted its hypolipidaemic effect in the management of hyperlipidaemia.
Nos mamíferos, o fígado desempenha um papel extremamente importante na manutenção da homeostase do metabolismo dos lipídios plasmáticos. Entretanto, problemas na regulação desses mecanismos têm sido implicados na ocorrência de dislipidemias (alterações na concentração adequada de lipídios no plasma). Nos últimos anos, tem sido evidenciada a existência de uma relação entre os níveis de selênio (Se) e o aumento nas concentrações plasmáticas de lipídios em humanos e animais. Dessa forma, o presente trabalho teve por objetivo investigar um possível efeito hipolipidêmico do (PhSe)2, um composto orgânico de Se, no modelo de hiperlipidemia aguda induzida por Triton WR-1339 em camundongos, bem como investigar se a hiperlipidemia aguda induzida pela administração intraperitonial (i.p.) de Triton WR-1339 altera parâmetros relacionados à ocorrência de dano oxidativo no tecido hepático de camundongos e determinar se o efeito antioxidante do (PhSe)2 está presente nesse processo. Para isso, o Triton WR-1339 foi injetado i.p. (400 mg/kg) em camundongos que estavam em jejum de 12 horas. O (PhSe)2 foi administrado oralmente (10 mg/kg) 30 minutos antes do Triton WR-1339. 24 horas após a injeção do Triton WR-1339, amostras de sangue foram coletadas para dosagem das concentrações de lipídios plasmáticos. Os níveis de espécies reativas ao ácido tiobarbitúrico (TBARS) e ácido ascórbico, bem como a atividade das enzimas catalase e glutationa peroxidase foram determinados no tecido hepático. A administração de (PhSe)2 foi capaz de prevenir o aumento nos níveis de colesterol total, colesterol não-HDL e triglicerídeos bem como aumentar os níveis de HDL-colesterol no plasma de camundongos hiperlipidêmicos. No protocolo experimental utilizado nesse estudo não foram observadas alterações nos parâmetros hepáticos de estresse oxidativo analisados. Conseqüentemente, o efeito antioxidante do (PhSe)2 não pode ser verificado. Os resultados obtidos nesse trabalho encorajam a realização de estudos posteriores no intuito de elucidar o exato mecanismo através do qual o (PhSe)2 exerce seu efeito hipolipidêmico.

Projeto de Pós-Graduação/Dissertação apresentado à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Ciências Farmacêuticas
As doenças cardiovasculares são uma das principais causas de morbilidade e mortalidade a nível mundial. Neste enquadramento, um dos principais fatores de risco associado às doenças cardiovasculares é a hipercolesterolemia. As opções farmacológicas existentes para o tratamento e prevenção desta dislipidemia centram-se, sobretudo, no uso de fármacos como as estatinas, a ezetimiba, os fibratos, o ácido nicotínico e as resinas sequestradoras de ácidos biliares. Porém, esta abordagem farmacológica no combate da hipercolesterolemia caracteriza-se pelo prolongado período terapêutico decorrendo daí possíveis efeitos laterais a longo prazo, pela intolerância a grupos terapêuticos observada em alguns doentes (como acontece com as estatinas) ou, ainda, pela eficácia limitada de alguns grupos terapêuticos (como é o caso, dos fibratos), o que suscita preocupação. Os avanços científicos no conhecimento dos processos que envolvem a hipocolesterolemia e a incessante procura de fármacos mais seguros e eficazes impulsionou o desenvolvimento dos inibidores da pró-proteína convertase subtilisina/kexina tipo 9 (PCSK9), afirmando-se como uma nova e promissora estratégia terapêutica. Os níveis plasmáticos elevados de colesterol proveniente das lipoproteínas de baixa densidade (C-LDL) são um fator de risco, no desenvolvimento de aterosclerose e de doença vascular isquémica. O recetor LDL (R-LDL) é essencial no metabolismo do colesterol, uma vez que ao se ligar ao C-LDL é capaz de eliminá-lo da circulação. É aqui, que reside o principal foco de interesse desta nova estratégia terapêutica, uma vez que a PCSK9 promove a degradação do recetor R-LDL, conduzindo a uma redução da depuração de LDL, aumentando os níveis de colesterol LDL. Desta forma, a inibição da atividade da PCSK9, veio revelar-se como uma nova abordagem potencialmente interessante para o desenvolvimento de novos fármacos destinados à redução do C-LDL. Os anticorpos monoclonais humanos contra PCSK9 estão em desenvolvimento clínico e são, neste momento, a aposta mais promissora de inibição da PCSK9. Até ao momento, os resultados dos ensaios clínicos demonstraram a eficácia destas moléculas com redução do C-LDL na ordem dos 60%. Adicionalmente, os seus efeitos na redução do C-LDL são aditivos aos das estatinas e até à data, não mostraram qualquer efeito tóxico a nível muscular, como acontece com estas últimas, sendo fármacos bem tolerados e aparentemente seguros.
Cardiovascular diseases are a major cause of morbidity and mortality worldwide. In this context, one of the main risk factor associated with cardiovascular disease is hypercholesterolemia. The treatment and prevention of this dyslipidemia is mainly focused on the use of drugs such as statins, ezetimibe, fibrates, nicotinic acid and bile acid sequestrants. However, the pharmacological approach in hypercholesterolemia treatment is characterized by prolonged therapeutic period elapsing possible long-term side effects, by the intolerance to treatment in some patients (as is the case of statins), or by the limited efficacy of various drugs/pharmaceuticals (as for example of fibrates), which raise concerns. Scientific advances in the understanding of hypercholesterolemia and the constant need for safer and more effective drugs prompted the development of the convertase pro-protein subtilisin inhibitor/kexin type 9 (PCSK9), as a promising new therapeutic strategy. Elevated plasma LDL cholesterol (LDL-C) levels are a risk factor for atherosclerosis and ischemic vascular disease. The LDL receptor (LDL-R) has an essential role in the cholesterol metabolism, since it binds to LDL-C removing it from circulation. Here, lies the main focus of interest of this novel therapeutic strategy, since PCSK9 promotes LDL-R the degradation, leading to a reduction of LDL clearance, increasing levels of LDL cholesterol. Therefore, inhibition of PCSK9 activity is a potentially interesting new approach for the development of new drugs to reduce LDL-C. Human monoclonal antibodies against PCSK9 are in clinical development and are presently the most promising strategy for the inhibition of PCSK9. At the moment, results of clinical trials show the efficacy of these molecules with reductions efficiencies of LDL-C in the order of 60%. Furthermore, this LDL-C reduction are additive to those of statins and until now have not shown any toxic effect in muscle, as observed with statins, and have a good record for safety and tolerability.

Includes bibliographical references (21 leaves). Previous studies have found that green tea and its antitoxidant constituents, the catechins, are hypocholesterolaemic in both epidemiological and animal intervetion studies. The main objectives of the present study were to investigate the mechanism by which green tea and its most abundant catechin constituent epigallocatechin gallate increase the low-density lipoprotein (LDL) receptor of HepG2 cells. In addition, it was hoped to determine if a crude catechin extract from green tea could lower plasma cholesterol levels in the hypercholesterolaemic rabbit and ascertain if this effect was due to an increase in the LDL receptor. The study provides evidence that green tea and its catechins exhibit hypocholesterolaemic properties and may therefore provide protection against heart disease.

Statins, the standard treatment for hypercholesterolaemia, have been associated with side effects, including statin intolerance. This study determined the prevalence of SLCO1B1 single nucleotide variations (SNVs) and possible associations between SLCO1B1 SNVs, statin intolerance and creatine kinase (CK) in hypercholesterolemic patients on statin therapy. One hundred and eighty one healthy controls and 100 hypercholesterolaemic patients receiving either simvastatin or atorvastatin were recruited. A questionnaire was used to assess the risk of statin intolerance. Polymerase Chain Reaction - Restriction Fragment Length Polymorphism (PCR-RFLP) was used to identify the presence of SLCO1B1 SNVs (rs4149056, rs2306283 and rs4363657) and enzyme-linked immunosorbent assay (ELISA) was used to quantify serum creatine kinase (CK) levels. Of the 100 hypercholesterolaemic patients, 15% presented with high risk, 49% with moderate risk and 36% with low risk to statin intolerance. The prevalence of the rs4149056 variant was 16% for the control group and 20% for the test group, while the rs2306283 variant present in 31.5% of the control group compared to only 10.5% in the test group. The prevalence of rs4363657 variant was similar in each group. A comparison of genotype frequencies based on calculated statin intolerance risk, i.e. low risk versus moderate to high risk, showed no significant association between any of the SNVs and the either low risk or moderate to high risk statin intolerant presentation. CK levels in patients on simvastatin were significantly higher compared to patients on atorvastatin. The prevalence of the SLCO1B1 SNVs in this population is a novel finding. No association between the presence of any one of the SNVs and the statin intolerance severity risk score or CK elevation was found.
Dissertation (MSc)--University of Pretoria, 2020.
Pharmacology
MSc
Unrestricted

La transplantation d’hépatocytes représente une alternative à la transplantation hépatique pour le traitement de certaines maladies métaboliques dont l’hypercholestérolémie familiale. Les cellules souches embryonnaires (ES) et les cellules souches pluripotentes induites (iPS) humaines représentent de nouvelles sources de cellules hépatiques. Nous avons mis au point une approche de différenciation des cellules souches humaines en cellules hépatiques et généré ainsi des cellules dérivées de cellules iPS de patients atteints d’hypercholestérolémie familiale
Hepatocyte transplantation represents an alternative to liver for the treatment of metabolic diseases including familial hypercholesterolaemia. Embryonic stem cells (ES) and induced pluripotent stem cells (iPS) represent new sources of hepatic cells. We have developed an approach to differentiate human stem cells into hepatic cells and thus we have generated hepatic cells derived from iPS of familial hypercholesterolaemia patients

Cardiovascular disease remains one of the leading causes of death globally. Many pharmacotherapeutic strategies are constantly being developed in order to effectively reduce the prevalence, as well as the morbidity and mortality associated with cardiovascular disease. In recent years, studies in our laboratory were able to demonstrate that dietary red palm oil (RPO) supplementation could offer protection against myocardial ischaemia/reperfusion injury. Several possible mechanisms for this protection were proposed, including: 1) upregulation of nitric oxide (NO), 2) cyclic guanylyl-mono-phosphate (cGMP) signaling, 3) scavenging of harmful free radicals associated with ischaemia/reperfusion injury and 4) the modulation of mitogen activated protein kinase (MAPK) signaling pathways which inhibit myocyte apoptosis. These findings created opportunities for further investigations to be conducted in order to elucidate RPO mediated pathways involved in cardiac protection. Matrix metalloproteinase 2 (MMP2) is an endogenous protease which is normally associated with the digestion of gelatin. This protease has recently become the focus of many cardiovascular studies, as it was found to be a mediator of ischaemia/reperfusion injury. Activation of MMP2 by free radicals leads to induction of myocyte death through degradation of intracellular targets. Research has shown that dietary RPO supplementation is able to increase activity of glutathione peroxidase (GPX) which is an endogenous antioxidant expressed in most mammalian tissue, including the heart. Increased antioxidant activity in the heart may lead to a reduction in ischaemia/reperfusion injury. However, the mechanism by which RPO supplementation was able to increase GPX activity is not known. The aims of the study were: 1) To investigate whether dietary RPO supplementation can reduce myocardial infarct size of rats fed a standard rat chow diet and rats fed a cholesterol supplemented diet 2) To investigate the whether inhibition of MMP2 plays a role in RPO mediated protection against ischaemia/reperfusion injury 3) To determine whether dietary RPO supplementation regulates GPX activity through gene transcription. Three study designs are described in this thesis where use was made of a male Wistar rat model being fed RPO supplemented diets for a 5 to 9 week period. Study design 1 compared rats placed on RPO supplemented diets with sunflower oil (SFO) fed rats and standard rat chow (SRC) fed rats while Study design 2 compared rats fed cholesterol supplemented diets with rats placed on a cholesterol+RPO supplemented diet. Study design 3 makes use of similar groups as Study design 1. After the supplementation period, rats were sacrificed and the excised hearts perfused on either a Langendorff perfusion apparatus or a working heart perfusion apparatus. Myocardial infarct size and aortic output recovery were measured in order to determine whether RPO does offer protection against ischaemia/reperfusion injury. MMP2 activity was measured in coronary effluent samples of the hearts, in order to determine its level of activity in RPO supplemented hearts. GPX1, -3 and -4 gene transcription were also measured by quantitative real time polymerase chain reaction (qrtPCR), in hearts of RPO supplemented rats, and compared to controls after supplementation. Other assays performed include ELISA (enzyme linked immunosorbent assay) for the determination of serum cholesterol, serum triglyceride and myocardial 3-nitrotyrosine levels as well as western blots to determine protein kinase B (also known as Akt (PKB/Akt)) and extracellular signal-regulated kinases (ERK) in myocardial tissue. Our results showed that dietary RPO supplementation was able to reduce myocardial infarct size in SRC fed rats (9.17 ± 1.03% in the RPO group, versus 30.20 ± 3.97% in the SRC group), as well as rats fed a cholesterol supplemented diet 26,87 ± 2,96% in the HCRPO group, versus 37.16 ± 3.58% in rats fed a cholesterol supplemented diet). Dietary RPO supplementation was also able to increase aortic output recovery in SRC fed rats (47.16 ± 5.46 % versus 13.44 ± 6.34 %). The decreased infarct size was associated with decreased MMP2 activity during reperfusion in the SRC fed rats (1389.27 ± 124.34 arbitrary units versus 1724.42 ± 69.77 arbitrary units; 72kDa isoform: 2635.03 ± 163.02 arbitrary units versus 3201.63 ± 104.97 arbitrary units). Dietary SFO supplementation reduced MMP activity at the same time point, with no reduction in infarct size. Dietary supplementation of RPO and cholesterol to rats, led to decreased MMP2 activity before ischaemia (228.43 ± 28.06 arbitrary units versus 450.83 ± 33.62 arbitrary units). The MMP2 activity was significantly increased in the cholesterol+RPO supplemented group when compared to the cholesterol fed rats after ischaemia (2107.06 ± 50.99 arbitrary units versus 1821.90 ± 56.92 arbitrary units). RPO supplementation did not show any significant differences in GPX transcription. We therefore conclude that dietary RPO supplementation reduced myocardial infarct size in SRC fed rats and rats fed a cholesterol supplemented diet. Inhibition of MMP2 activity was also shown not to be a major pathway of protection involved in RPO mediated protection against ischaemia/reperfusion injury. Furthermore, our results are in agreement with previous studies which show that dietary RPO supplementation is able to improve aortic output recovery. RPO supplementation does however, not increase GPX activity through upregulation of GPX gene transcription.

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

Dissertation submitted in partial compliance with the requirements for the Master's Diploma in Technology in the Department of Homoeopathy at Technikon Natal, 1994.
The object of the present research trial was to evaluate the efficacy of a single homoeopathic medication, Cholesterinum, in the ninth atcenuation (9CH) in the treatment of hypercholesterolaemia. Special attention was paid to its effect on total choles~erol (TC) levels and the high-density lipoprotein cholesterol/ low-density lipoprotein cholesterol (HDL-C/LDL-C) ratio.
M

A research report submitted to the Faculty of Health Sciences, University of the Witwatersrand in fulfilment of the requirements for the Degree of Master of Medicine in the branch of Internal Medicine
Untreated patients with heterozygous familial hypercholesterolaemia (hFH) are at increased risk for atherosclerosis. Surrogate markers to predict atheroma include the presence of endothelial dysfunction (ED) as well as increased levels of inflammatory markers
IT2018

A research report submitted to the Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, in partial fulfilment of the requirements for the degree of Master of Medicine in Ophthalmology Johannesburg, 2017.
Homozygous familial hypercholesterolaemia is a fatal disease if untreated and has a high prevalence of premature coronary artery disease. Ocular findings may help with earlier identification and coronary artery disease risk stratification. Objectives: The primary objective was to determine ocular findings in patients with homozygous familial hypercholesterolaemia. The secondary objective was to correlate ocular findings with clinical and biochemical data. Design and Method: A cross-sectional study was conducted in 2011. Thirty patients were recruited from the Lipid Clinic at Charlotte Maxeke Johannesburg Academic Hospital. Results: Xanthelasma palpebrarum, corneal arcus, retinal arteriosclerosis and visual field defects were detected. Xanthelasma palpebrarum and corneal arcus were common in patients with overt coronary artery disease. Conclusion: In addition to well-known ocular features of hyperlipidaemia, i.e. xanthelasma palpebrarum, corneal arcus and retinal arteriosclerosis, we detected visual field defects. The assessment of xanthelasma palpebrarum and corneal arcus may help to prognosticate coronary artery disease risk.
LG2018

Tese de mestrado em Biologia Molecular e Genética, apresentada à Universidade de Lisboa, através da Faculdade de Ciências, 2016
As doenças cardiovasculares (DCV) afetam o funcionamento normal do coração e dos vasos sanguíneos. Existem vários tipos de doenças cardiovasculares, sendo as principais a doença das artérias coronárias e a doença das artérias do cérebro. A doença coronária é, maioritariamente, provocada por aterosclerose, uma doença progressiva e inflamatória que resulta na formação de placas ateroscleróticas. Agentes infeciosos ou concentrações elevadas de lipoproteínas de baixa densidade (LDL) no sangue podem levar ao aparecimento destas placas na parede interna das artérias impedindo a circulação sanguínea. As doenças cardiovasculares são uma das causas de morte mais comum no planeta. Fatores de risco como o uso de tabaco, obesidade, prática de exercício físico reduzida, consumo excessivo de álcool, diabetes, hipertensão, stress e dislipidémia aumentam a probabilidade de ocorrência prematura deste tipo de doenças. A dislipidémia é um dos mais importantes fatores de risco da aterosclerose, uma vez que se caracteriza por anomalias quantitativas ou qualitativas dos lípidos no sangue. A identificação de indivíduos em risco e o conhecimento da causa de hipercolesterolemia é de extrema importância para que estes indivíduos possam ser corretamente tratados evitando-se, assim, a morte devida a esta causa. A Hipercolesterolemia Familiar (FH) é uma doença autossómica dominante do metabolismo do colesterol. A FH é hereditária e resulta, maioritariamente, de mutações no gene do recetor das lipoproteínas de baixa densidade (LDLR) cuja função é a remoção do colesterol LDL do plasma, transportando-o para o fígado, onde é processado. Ficando esta função afetada, os níveis de colesterol LDL circulante, aumentam. Mutações noutros genes como o gene da apolipoproteína B-100 (APOB) e da pro-proteína convertase subtilisina/quexina tipo 9 (PCSK9) são também causa, embora menos frequente, desta doença. Alterações no gene da apolipoproteína E (APOE) afetam a afinidade com os recetores de lipoproteínas e, consequentemente, a remoção do colesterol do sangue, podendo também causar dislipidémia. A forma homozigótica da FH é rara e mais severa, mas a heterozigótica é comum embora sub-diagnosticada em muitas populações, nomeadamente na portuguesa. Estima-se que, na maioria dos países europeus, a prevalência destas duas formas seja 1/1000000 e 1/500 indivíduos, respetivamente. O colesterol total, na forma heterozigótica, varia entre 290 e 500 mg/dL (com LDL>190 mg/dl) e na forma homozigótica, habitualmente, encontra-se entre os 600 mg/dL e os 1000 mg/dL. O nível elevado de colesterol no plasma resulta, frequentemente, na formação de depósitos de colesterol nos tecidos extravasculares que, por vezes, podem ser facilmente identificados em indivíduos ainda jovens (abaixo dos 45 anos): xantelasmas, arco corneano e, mais difíceis de reconhecer mas mais específicos, os xantomas nos tendões. A presença de valores altos de colesterol LDL desde o nascimento, característico desta doença, leva a um incremento do risco de doença coronária prematura. Este fenótipo permite diferenciar a FH de outras causas de hipercolesterolemia, nomeadamente da hipercolesterolemia comum ou poligénica, embora nem sempre seja fácil essa diferenciação. Foram descritas, em todo o mundo, mais de 1700 mutações diferentes no gene LDLR. Contudo, grande parte destas não possuem estudos funcionais, o que impede o diagnóstico definitivo destes doentes. Em 1999 iniciou-se no Instituto Nacional de Saúde Dr. Ricardo Jorge o Estudo Português de Hipercolesterolemia Familiar. Este estudo tem como objetivos a pesquisa de alterações genéticas que possam confirmar o diagnóstico clínico de FH em indivíduos da população portuguesa e a determinação da prevalência e distribuição da FH em Portugal. Os casos índex são incluídos no estudo caso cumpram os critérios adaptados de Simon Broome. Estes critérios categorizam a FH como “definitiva” ou “possível”, sendo que a primeira se define, em adultos, por valores de colesterol total acima de 290 mg/dL ou de colesterol LDL acima de 190 mg/dL, e em crianças, até aos 16 anos de idade, por uma concentração de colesterol total cima de 260 mg/dL ou de colesterol LDL acima de 155 mg/dL, com presença de xantomas no doente ou num familiar de primeiro ou segundo grau ou ainda quando existe evidência genética de uma mutação num dos 3 genes que cause FH. O diagnóstico “possível” requer a presença de níveis de colesterol acima destes valores, valores totais de colesterol acima dos 290 mg/dL num familiar de primeiro ou segundo grau e história familiar ou enfarte do miocárdio antes dos 50 anos num familiar de segundo grau ou antes dos 60 anos num familiar de primeiro grau. Sempre que possível, após a identificação da possível alteração causadora de doença é feito o estudo funcional para as variantes de patogenicidade desconhecida, para que o diagnóstico seja o mais completo e definitivo possível, contribuindo para uma abordagem terapêutica mais personalizada. O principal objetivo deste estudo foi a identificação molecular de variantes genéticas nos genes LDLR, APOB e APOE que provoquem dislipidémia. Os doentes referenciados para o Estudo Português de Hipercolesterolemia Familiar em 2015/2016 foram o alvo deste projeto. A realização de estudos funcionais ao nível do RNA para variantes que afetam o splicing foi também objetivo deste estudo. Neste projeto, foram estudados 60 casos índex incluídos no Estudo Português de Hipercolesterolemia Familiar. O estudo molecular foi dividido em várias fases: 1. O DNA genómico é isolado a partir dos linfócitos do sangue periférico; 2. Os 18 exões, regiões adjacentes e o promotor do gene LDLR, parte dos exões 26 e 29 do gene APOB e o exão 4 do gene APOE foram amplificados por PCR e sequenciados pelo método de sequenciação direta de Sanger. As sequências foram analisadas em computador e comparadas com as sequências de referência de forma a detetar variantes que possam ser a causa desta doença, confirmando o diagnóstico clínico; 3. Estudo de grandes rearranjos por Multiplex Ligation-dependent Probe Amplification (MLPA). Foram realizadas predições in silico para todas as alterações encontradas para prever o seu impacto ao nível da proteína. Para as alterações em regiões codificantes foram utilizadas as ferramentas PolyPhen-2, SIFT, PROVEAN e Mutationtaster. As ferramentas HSF, NNSSP e FSPLICE foram usadas para prever o efeito no splicing causado pelas alterações em regiões intrónicas. Quando nenhuma mutação é encontrada nestes três genes e o doente apresenta um fenótipo agressivo, procede-se à pesquisa no gene PCSK9 e em todo o gene APOB, representando a quarta fase do estudo molecular. Quando são encontradas variantes de patogenicidade desconhecida, são realizados estudos funcionais in vitro (fase 5). Embora o estudo completo dos genes PCSK9 e APOB não tenha sido realizado durante este trabalho, foram feitos estudos funcionais ao nível do RNA. No total, foram identificadas 18 variantes diferentes em 24 destes doentes: 16 no gene LDLR, 1 no gene APOB e 1 no gene APOE. Apenas 11 destas apresentam estudo funcional. Sempre que existia amostra disponível, foi feito também o estudo genético dos familiares, o que permitiu a identificação e caracterização genética adicional de 28 indivíduos, num total de 43 doentes com uma alteração possivelmente patogénica. Entre estas mutações encontram-se 3 nonsense, 12 missense e 3 que possivelmente afetam o splicing. Uma destas mutações foi descrita, pela primeira vez, em Portugal neste projeto. A análise de grandes rearranjos não revelou alterações deste tipo no grupo em estudo. A confirmação dos efeitos causados ao nível do splicing foi feita para duas das três alterações de splicing encontradas no gene LDLR durante este projeto. Para tal, recorreu-se ao isolamento das células mononucleares do sangue periférico dos doentes e à extração do RNA utilizando o RNeasy® Mini Kit (Qiagen). Após retrotranscrição para cDNA e amplificação da zona a estudar utilizando primers específicos, as bandas em gel de agarose foram analisadas e os fragmentos foram sequenciados. As duas alterações em causa levam ao skipping de exões e ao aparecimento de codões stop prematuros: no caso da alteração c.1060+1G>A, a inativação do donor site no intrão 7 resulta no skipping do exão 7; a alteração no último nucleótido do exão 16 (c.2389G>A) leva ao aparecimento de um novo acceptor site e, consequentemente, ao skipping do exão 16. A FH é caracterizada por níveis elevados de colesterol plasmático desde a nascença. Por isso, é de extrema importância que o diagnóstico seja feito o mais cedo possível, principalmente em idade pediátrica para que estas crianças recebam acompanhamento médico personalizado durante toda a vida, prevenindo o aparecimento de DCV prematura, permitindo, assim, uma melhor e maior esperança de vida. No entanto, é necessária uma maior divulgação da doença, principalmente junto do corpo clínico dos hospitais e centros de saúde assim como junto do público em geral. Só após a identificação clínica se pode realizar o estudo genético para comprovar a doença e o doente pode então receber acompanhamento e tratamento personalizado.
Cardiovascular disease (CVD) remains the most common cause of death globally. Dyslipidaemia is one of the most important risk factors that leads to CVD. It can be due to a monogenic condition or to polygenic/environmental causes as diabetes, obesity, tobacco use, excess of alcohol or reduced physical activity. The identification of the individuals at risk and the distinction of these two types of dyslipidaemia is important for a correct cardiovascular risk assessment, counselling, and treatment reducing, this way, cardiovascular mortality. Familial hypercholesterolaemia (FH) is an autosomal dominant disorder of cholesterol metabolism. Most commonly, FH results from inherited defects in the Low-Density Lipoprotein Receptor Gene (LDLR) leading to increased levels of circulating LDL cholesterol and lipid accumulation in arteries and tendons. Mutations in other genes as the apolipoprotein B gene (APOB) and proprotein convertase subtilisin/kexin type 9 gene (PCSK9), are also responsible for FH. The distribution pattern of apolipoprotein E gene (APOE) polymorphisms affects the affinity to lipoprotein receptors and, consequently, the clearance of dietary fat from the blood, also causing dyslipidaemia. The homozygous form of FH is rare and more severe, but the heterozygous form is common, with a frequency of 1/500 in most of European countries, although underdiagnosed in several populations, including the portuguese. FH is characterized by increased levels of plasmatic cholesterol since birth, which results in cholesterol deposits in extravascular tissues that can be identified in young patients (below 45 years old): xanthelasma, corneal arcus deposits and tendon xanthomas. This accumulation can cause premature arteriosclerosis and coronary heart disease (CHD). The presence of tendon xanthomas allows the differentiation of FH from other causes of hypercholesterolaemia as polygenic hypercholesterolaemia. More than 1700 different alterations in LDLR gene have been described worldwide. However, the functional studies for the great majority of these variants, have not been performed. For patients carrying these variants, a definitive molecular diagnosis for FH is not possible, representing a serious problem for FH diagnosis. In 1999, the Portuguese FH study was established at the National Institute of Health to identify the genetic cause of hypercholesterolaemia in individuals with a clinical diagnosis of FH. Index patients are included in this study using an adaptation of the Simon Broome (SB) criteria. Nonetheless, FH remains underdiagnosed and undertreated in the portuguese population. The main aim of this project was to perform the molecular identification of genetic variants in LDLR, APOB and APOE genes, causing dyslipidaemia in patients referred to the Portuguese FH Study in 2015/2016 with a clinical diagnosis of FH, in order to improve the identification of individuals at risk. Functional studies in RNA for putative splicing variants were also performed. The molecular diagnosis was performed for 60 index cases. Genomic DNA was isolated from peripheral blood lymphocytes using the salting out method. The 18 exons and promotor region of LDLR, part of exons 26 and 29 of APOB and exon 4 of APOE were amplified by PCR and sequenced by direct Sanger sequencing. A total of 18 variants were identified in 24 of these patients. The cascade screening in relatives of these 24 index patients allowed the identification and genetic characterization of additional 19 FH patients in Portugal. All alterations found have been previously reported, although only 11 had been functionally assessed. The search for large rearrangements was performed by Multiplex Ligation-dependent Probe Amplification (MLPA). In silico analysis was performed for all the variants found. In order to access the effect of splicing mutations, RNA was isolated from patients’ blood with RNeasy® Mini Kit (Qiagen), after isolation of peripheral blood mononuclear cells, and then transcribed to cDNA. Regions of interest were amplified with specific primers designed to evaluate the effect on cDNA of two of the tree putative splicing variants found in LDLR gene. Specific detection of each transcript was accessed by an agarose gel and the fragments were sequenced by Sanger sequencing. Both alterations lead to skipping of an entire exon and create premature stop codons: c.1060+1G>A causes an inactivation of the donor site in intron 7 resulting in skipping of exon 7; the alteration in the last nucleotide of exon 16 (c.2389G>A) creates a new acceptor site causing the skipping of exon 16. The early genetic identification of a mutation, confirming the clinical diagnosis of FH, is very important, especially for young patients, since they can receive appropriate dietary and lifestyle advice and adequate therapeutic measures providing them longer and better lives.

Familial hypercholesterolaeinia (FH) is an inherited disorder caused by mutations in th e low-density lipoprotein (LDL) receptor which lead to diminished clearance of cholesterol from the circulation and, consequently, to markedly elevated LDL-cholesterol (LDL-C) levels. The resultant hypercholesterolaem ia predisposes these patients to severe prem ature atherosclerosis, particularly coronary artery d isease (CAD). T he concentration of LDL-C and lifetime vascular exposure to raised plasm a LDL-C concentrations are major determ inants of atherosclerosis, but there rem ains a considerable variability in the extent of atherosclerosis presen t and in the expression of clinical disease in these patients. Qualitative differences in LDL such a s LDL particle size and susceptibility to lipid oxidation may play a role, as may other biochemical risk factors. The purpose of this thesis was to determine whether such qualitative differences in LDL are important determinants of the extent and severity of atherosclerosis and to determine whether it is mainly the reduction in LDL-C that is of benefit, or whether antioxidant therapy would also be effective in preventing progression of atherosclerosis in FH subjects. FH patients were found to have large, buoyant LDL particles, which are less susceptible to lipid oxidation than smaller, d en ser particles. In the absence of other causes of insulin resistance, patients with FH have normal fasting insulin and triglyceride levels, normal postprandial lipaemia, and do not have microalbuminuria. They, therefore usually show no features of the metabolic syndrome despite severe, accelerated atherosclerosis. Similarly, the role of lipid oxidation in the pathogenesis of atherosclerosis in FH remains uncertain. LDL isolated from FH patients is more resistant to oxidation, and antioxidant therapy appears to be of little or no benefit in preventing progression of atherosclerosis in these hypercholesterolaem subjects. Particularly in severely hypercholesterolaem ic subjects, m ore conclusive proof of a protective effect of antioxidants from large prospective studies presently in progress is needed before antioxidant therapy can be advocated for the treatm ent and prevention of atherosclerosis. In subjects with FH, quantitative rather than qualitative differences in LDL are associated with accelerated atherosclerosis. Therapy in FH should therefore be aimed primarily at reducing LDL-C levels.

Submitted in fulfilment with the requirements for the degree Master in Medicine (MMed)
Background: Homozygous Familial hypercholesterolaemia (HoFH) is a rare genetic disorder affecting approximately one in every million people worldwide. It is characterized by severely elevated LDL-cholesterol (LDL-C) levels usually as a result of mutations in both LDL receptor alleles, and is associated with a markedly increased risk of premature cardiovascular disease with death often occurring in the first 3 decades of life. Standard treatment with statin therapy has been shown to yield suboptimal results with additional therapy required to achieve lower LDL-C levels. As not all centers worldwide have access to newer treatment modalities, cheaper and more accessible therapy needs to be considered. The addition of ezetimibe to statin therapy in HoFH individuals has only been reported in one previous study, but in that study other factors which may have influenced the response to ezetimibe such as body mass index (BMI), gender and the type of LDLR mutation were not evaluated Objectives: Firstly to assess whether the addition of ezetimibe to statin therapy can result in further reduction in LDL-cholesterol in subjects with HoFH. Secondly, to assess whether the reduction in LDL-C (response rate) is dependent on the underlying LDLR mutations, gender and/or BMI. Lastly, to compare HoFH patients which showed higher responses in LDL-C reduction to ezetimibe (“responders”) to those who responded poorly (non-responders), Study design: This was a retrospective study which evaluated HoFH patients known to the Charlotte Maxeke Johannesburg Academic Hospital’s lipid clinic. All patients were confirmed to have HoFH and were already on high intensity statin therapy prior to initiating ezetimibe at a fixed dose of 10mg daily given orally. Their lipograms prior to ezetimibe initiation were recorded and used as a baseline. In addition, their BMI, gender, age, FH genotype and cardiovascular complications were recorded. Follow up lipograms were recorded at 3 and 6 month after ezetimibe initiation. Results: 48 patients who fulfilled the entry criteria were eligible for the study. Of the 48 patients, 24 were males and 24 females. The average BMI in males was 22.7 ± 6.9 kg/m2 and 24 ± 7.1 kg/m2 in females. The two commonest FH genotypes were Afrikaner FH1/FH1 (17 patients) and Afrikaner FH1/FH2 (11 patients). Age ranged between 3 and 48 years with a mean age of 25 years. 65% of patients had documented coronary artery disease or aortic stenosis. 86% of patients were on high intensity statin therapy (atorvastatin 80mg or rosuvastatin 40 mg daily) prior to starting ezetimibe. Despite high intensity statin therapy, mean LDL-C at baseline was 12.1 ± 3.3 mmol/L, decreasing to 10 ± 3.4 mmol/L after 3 months of ezetimibe therapy, and 10.4 ± 3.3 mmol/L at 6 months (p=0.0018). The mean percentage reduction of LDL-C on ezetimibe was -18.9% after 3 months and -17.6% at 6 months. There was no significant change in HDL-C or triglyceride levels with the addition of ezetimibe, p>0.05. Response of LDL-C based on BMI, gender and LDLR mutation was evaluated at 3 months. Overweight patients had an overall better response compared to normal weight patients, with a mean percentage reduction of - 20.5% vs -15.7% (p=0.02). A significant difference in response to ezetimibe was also seen amongst different FH genotypes, with FH1/FH1 having a significant lower mean LDL-C level at baseline (p=0.04), and a greater reduction in LDL-C following 3 months of ezetimibe therapy compared to FH1/FH2 (-17.5% vs -11.5%, p=0.027). Lastly, there was no significant difference in LDL-C at baseline or 3 months between gender. However females tended to show a slightly better mean percentage reduction at 3 months (-20.7% vs -17%; p = 0.49).When patients were divided into those who responded to ezetimibe (mean percentage reduction of > 20%), compared to those with that did not (mean percentage reduction of < 20%), no identifiable factor such as BMI, gender or FH genotype was shown to be significant in identifying those patients who were more likely to respond. Conclusion: Ezetimibe is effective in HoFH and, on top of statin therapy, can reduce LDL-C by a further 18.9%. Ezetimibe should therefore be considered in all HoFH patients in order to lower LDL-C levels further. BMI and FH genotype influenced the response to ezetimibe. However, no single factor was able to predict response in the individual patient.
MT2016

by Ying Tat Mak.
Thesis (Ph.D.)--Chinese University of Hong Kong, 1996.
Includes bibliographical references (leaves 178-198).
Title --- p.1
Abstract --- p.2
Acknowledgments --- p.5
Contents --- p.6
Abbreviations --- p.9
List of Tables --- p.11
List of Figures --- p.13
Chapter Chapter 1: --- Introduction
Chapter 1.1 --- Cholesterol Metabolism and Atherosclerosis --- p.15
Chapter 1.1.1 --- Cholesterol and Cholesterol Metabolism --- p.17
Chapter 1.1.2 --- Cholesterol Transport: Apolipoprotein and Lipoprotein --- p.23
Chapter 1.1.3 --- Cholesterol and Atherosclerosis --- p.26
Chapter 1.2 --- Hyperlipidaemia --- p.30
Chapter 1.2.1 --- Primary and Secondary Hyperlipidaemia --- p.31
Chapter 1.2.2 --- Mutations leading to Primary Hypercholesterolaemia --- p.36
Chapter 1.3 --- Familial Hypercholesterolaemia --- p.38
Chapter 1.3.1 --- Historical Aspects --- p.38
Chapter 1.3.2 --- Clinical Features - Diagnosis and Consequences --- p.39
Chapter 1.3.3 --- Population Prevalence --- p.40
Chapter 1.3.4 --- Mutations in the Low Density Lipoprotein Receptor Gene --- p.41
Chapter 1.4 --- Methods for Detecting Mutations in LDL Receptor Gene --- p.51
Chapter 1.4.1 --- Southern Blotting Based Methods --- p.51
Chapter 1.4.2 --- Polymerase Chain Reaction Based Methods --- p.52
Chapter 1.4.3 --- Screening Methods for Unknown Mutations in LDL Receptor Gene --- p.56
Chapter 1.5 --- Mutations of the LDL receptor gene in Chinese --- p.58
Chapter Chapter 2: --- Objectives --- p.63
Chapter Chapter 3: --- Materials and Methods
Chapter 3.1 --- Subjects
Chapter 3.1.1 --- Familial Hypercholesterolaemia Patients --- p.65
Chapter 3.1.2 --- Normocholesterolaemia Subjects --- p.67
Chapter 3.2 --- Materials
Chapter 3.2.1 --- Enzymes --- p.67
Chapter 3.2.2 --- DNA Markers --- p.68
Chapter 3.2.3 --- Reagents Kits --- p.68
Chapter 3.2.4 --- Primers for PCR --- p.68
Chapter 3.2.5 --- Chemicals and Reagents --- p.69
Chapter 3.2.6 --- Radioisotopes --- p.70
Chapter 3.2.7 --- Solutions and Buffers --- p.70
Chapter 3.3 --- Methods
Chapter 3.3.1 --- Blood Collection --- p.71
Chapter 3.3.2 --- General Biochemistry Tests --- p.72
Chapter 3.3.3 --- DNA Extraction --- p.72
Chapter 3.3.4 --- RNA Extraction --- p.73
Chapter 3.3.5 --- Polymerase Chain Reaction --- p.74
Chapter 3.3.6 --- Agarose Gel Electrophoresis --- p.76
Chapter 3.3.7 --- Polyacrylamide Gel Electrophoresis --- p.78
Chapter 3.3.8 --- Single Strand Conformation Polymorphism --- p.79
Chapter 3.3.9 --- Reverse Transcription - Polymerase Chain Reaction --- p.79
Chapter 3.3.10 --- Direct DNA Sequencing --- p.81
Chapter 3.3.11 --- Haplotyping of the LDL receptor gene --- p.83
Chapter 3.3.12 --- Restriction Enzyme Digestion --- p.84
Chapter Chapter 4: --- Results
Chapter 4.1 --- Patients Investigations --- p.88
Chapter 4.1.1 --- Normal Control Subjects --- p.88
Chapter 4.1.2 --- Patients --- p.88
Chapter 4.2 --- PCR-SSCP Analysis of LDL Receptor Gene --- p.90
Chapter 4.3 --- Summary of Mutations Identified --- p.92
Chapter 4.4 --- Novel Mutations --- p.94
Chapter 4.5 --- Previously Reported Mutations --- p.97
Chapter 4.6 --- Polymorphisms and Silent Mutation --- p.100
Chapter 4.6.1 --- New Polymorphism --- p.100
Chapter 4.6.2 --- New Silent Mutation --- p.102
Chapter 4.6.3 --- Reported Polymorphisms --- p.103
Chapter 4.7 --- Southern Blotting --- p.103
Chapter 4.8 --- Haplotypes --- p.104
(All Figures for Chapter 4) --- p.106
Chapter Chapter 5: --- Discussions
Chapter 5.1 --- Use of SSCP in Screening for Mutations and Polymorphisms --- p.158
Chapter 5.2 --- Novel and Reported Mutations --- p.160
Chapter 5.3 --- Novel Polymorphism and Silent Mutation --- p.170
Chapter 5.4 --- Common Polymorphisms --- p.171
Chapter 5.5 --- Possible Common Mutations of the LDL Receptor Gene in Chinese --- p.172
Chapter 5.6 --- Pattern of LDL Receptor Gene Mutations in Chinese --- p.173
References --- p.178