Hypercholesterolemia is a major risk factor for atherosclerosis and cardiovascular disease. Most humans and animals respond to increased dietary cholesterol with a moderate to substantial increase in plasma cholesterol levels due to increases in LDL/VLDL cholesterol. The investigators have identified a substrain of the C57BL/6J (J) substrain, which develops hypercholesterolemia and atherosclerotic lesions in response to the atherogenic diet. The Bailey and J substrains exhibit an extremely high degree of genetic identity, with no polymorphisms detected in a panel of 80 microsatellite DNA markers. Thus, these two substrains provide a unique system in which to identify a gene(s) that exerts a major effect on cholesterol response on an otherwise nearly identical genetic background. In a genetic cross between Bailey and the strain A/J, the investigators have detected a locus that segregates with the hypo-response and provides a startin point for identification of a gene that exerts a major effect on cholesterol response. In the proposed studies the investigators will isolate the gene(s) underlying the hypo-response in Bailey mice and characterize the metabolic basis for this phenotype.
Two Specific Aims will be carries out in parallel.
Specific Aim 1 describes determination of the number of variant loci between Bailey and J, genetic mapping of the cholesterol response gene(s) and isolation of the gene(s). If the Bailey x J cross indicates a single locus is involved, we will focus on evaluating the recently mapped locus, first by consideration of candidate genes, and subsequently by high resolution mapping followed by positional cloning strategies employing the techniques of direct selection and representational difference analysis (RDA) to isolate the gene from a physical DNA contig. If the Bailey x J cross indicates more than a single locus is involved, the investigators will perform crosses with additional strains as distinct genetic backgrounds may allow segregation of different loci. Individual cholesterol response loci will then be isolated into congenic strains for further genetic mapping studies.
Specific Aim 2 will complement th genetic studies with characterization of cholesterol metabolism in Bailey compared to J mice, including cholesterol absorption, endogenous cholesterol synthesis, and cholesterol excretion as bile acids. In addition, we compare Bailey and J substrains for susceptibility to aortic lesion development, as a model in which similar HDL levels occur in conjunction with vastly different LDL/VLDL levels.
