5 research outputs found

    Molecular analysis of gene expression in rat brain

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    Monogenic hypercholesterolemia in South Africans : familial hypercholesterolemia in Indians and familial defective apolipoprotein B-100

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    LDL-receptor mutations and familial defective apolipoprotein B-100 (codon 3500) (FOB), the known causes of monogenic hypercholesterolemia (MH), have similar clinical features. The nature of the mutations responsible for MH in South Africans of Indian origin was previously unknown. Similarly, the mutations in the LDL-receptor gene of a South African Black FH homozygote had also not been characterised. The aim of this thesis was to identify and analyse the LDL-receptor mutations in the Indian homozygotes NS, D, AV and AA and in the Black homozygote JL. In addition, the possible importance of FOB as a cause of MH in South Africans was also assessed. The patient NS was characterized as having two "Null" LDL-receptor alleles. His skin fibroblasts expressed no detectable LDL-receptor protein and very low levels of LDL-receptor mRNA of approximately normal size. Since NS' s LDLreceptor promoter sequence was normal, his alleles are likely to harbour exonic point mutations or minor rearrangements that cause premature stop codons. The patient D was found to be a heteroallelic homozygote. Two new point mutations in the LDL receptor, Asp₆₉ -Tyr and Glu₁₁₉-Lys, were identified. D's fibroblasts expressed about 30% of the normal surf ace complement of receptors that bound LDL poorly. This low number could at least be partially explained by their decreased stability. These mutations were not identified in any other Indian FH or hypercholesterolemic patients. Patients AV and AA were both shown to be homoallelic homozygotes for the Pro₆₆₄ -Leu mutation. This mutation was identified in 4 unrelated Muslim families of Gujerati origin suggesting that the mutation arose from this area in India. Contrary to previous reports (Knight et al. 1990, Soutar et al. 1989), neither LOL nor β-VLDL binding were shown to be affected by this mutation. These mutant receptors were rapidly degraded. Thus the disease FH in these subjects is presumably due to the low steady-state level of mature receptors that are functionally normal but exhibit accelerated turnover. The Pedi FH homozygote, JL, expressed very few LOL receptors due to decreased receptor synthesis associated with low mRNA levels and not due to enhanced degradation. One of JL's LOL receptor alleles has a 3 b.p. deletion in repeat 1 of the promoter (G. Zuilani, H. Hobbs and L.F. de Waal, personal communication). The nature of the defect in his other allele is unknown. The importance of FOB as a cause of monogenic hypercholesterolemia in the South African Indian, "Coloured" and Afrikaner populations was determined by screening hypercholesterolemic subjects with or without xanthomata. The absence of FOB in such patients, in whom the relevant common or founder South African mutations were excluded, suggested that this disorder was rarer in these groups than in North America and Europe. FOB was identified in two different families of mixed British and Afrikaner ancestry. One family contained individuals who were heterozygous for the FOB mutation, as well as the FH Afrikaner-1 and the FH Afrikaner-2 LOL-receptor mutations. In addition, 4 compound heterozygotes, who had both FOB and the FH Afrikaner-1 mutation and one individual whu inherited all 3 defects, were identified. This family allowed us to characterise the compound heterozygotes with one mutant LOLreceptor allele and FOB as having a condition that was probably intermediate in severity between the FH heterozygote and homozygote states

    Effects of common genetic variation on fasting and postprandial lipid metabolism and atherosclerosis

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    Fasting triglyceride (TG) levels are a strong determinant of postprandial lipaemia, which is thought to influence the development of coronary artery disease (CAD) because of the delayed clearance or prolonged presence of atherogenic remnant lipoproteins. Associations were thus sought between common genetic variation in apolipoprotein (apo) B, AI, CIII and E and lipoprotein lipase (LPL) and both levels of the TG-rich lipoproteins and severity of atherosclerosis. The role of the LPL gene was emphasized because of the known metabolic relationship between LPL activity and lipid metabolism. In Swedish healthy individuals, the C-265 allele of the C to T polymorphism in the apoB promoter was associated with higher apoB levels, the X+ allele of the apoB-XbaI polymorphism with higher low density cholesterol (LDL) cholesterol levels, the common allele (V+) of the apoCIII-PvuII polymorphism with higher LDL-TG levels, and both the T allele of the apoCIII C1100 to T polymorphism and the common allele (H+) of the LPL-HindIII polymorphism with higher TG levels. In Swedish myocardial infarction survivors, fasting lipids and lipoproteins were associated with variation at the apoE gene but not at the apoB, apoCIII or LPL genes. However homozygosity for the insertion allele of the apoB signal peptide polymorphism, homozygosity for the C allele of the apoCIII C1100 to T polymorphism and the presence of the rare alleles of the HindIII and Serine-Stop447 polymorphisms in the LPL gene were associated with worse baseline severity of atherosclerosis in the MI survivors. When genotypes associated with severe atherosclerosis were considered together, individuals with all three genotypes had a 6-fold greater severity of atherosclerosis compared to those with none. In a British patient sample with CAD, variation at the apoB and apoAI-CIII-AIV genes were associated with baseline severity and progression, respectively, of atherosclerosis; homozygosity for the X- allele of the apoB-Xbal polymorphism and homozygosity for the G allele of the apoAI-G-75 to A polymorphism being associated with worse atherosclerosis. In neither of these patient samples, did the genotype effect on atherosclerosis appear to be mediated through genotype-associated differences in fasting lipid and lipoprotein traits. To test the hypothesis that genetic variation was influencing atherosclerosis severity via modulation of postprandial lipaemia rather than through fasting lipids, the effects of common genetic variation on levels of postprandial TG-rich lipoproteins and their atherogenic remnants were examined. The apoB signal peptide variants influenced production of postprandial lipoproteins of both intestinal and hepatic origin. Polymorphisms in the apoCIII, LPL and apoB genes exerted their largest effect on lipolysis or catabolism of postprandial lipoproteins, and the H+ allele of the LPL-HindIII polymorphism was associated with higher postprandial LPL activity and levels of free fatty acids. Using Single Strand Conformation Polymorphism analysis and sequencing, an A1127 to G substitution that changes Asparagine-291 to Serine was found in exon 6. The frequency of the Serine-291 variant, found on the haplotype defined by the common alleles (presence of cutting site) of the PvuII and HindIII polymorphisms, was approximately 0.1 in two independent samples. The effects of this substitution on fasting and postprandial lipids and lipoproteins, LPL activity, interrelationships between plasma traits and influence on apoE phenotype-mediated effects have been investigated, but do not explain all the effects associated with genotypes of the LPL-HindIII polymorphism. Common variation in the genes for apoB, apoCIII and LPL influences fasting serum lipid and lipoprotein levels, with LPL having the largest effect on TGs, and modulating postprandial lipaemia and interrelationships between lipid traits; these may explain associations seen between genetic variation and atherosclerosis. Additive effects occur between common genetic polymorphisms, modulated by environmental factors, with implications for future population studies

    American Society of Nephrology

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