This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. All individuals with sickle cell anemia (SS) have the identical genetic defect (homozygous beta 6 glu to val) but there is a wide variation in clinical severity. In this study we focus on one of the key risk factors for severity identified by the CSSCD: Baseline white blood cell count (baseline WBC). This initially unanticipated risk factor is becoming more obviously relevant as new investigations into the pathophysiology of the disease increasingly emphasize the importance of white cells and inflammation. At the same time, baseline WBC and other markers of inflammation are emerging as risk factors for mortality in the general population, making the exploration of genetic determinants of baseline WBC of interest not only to the SS population, but also to the population at large. Our strategy for locating the genes that are responsible for the variability in baseline WBC involves three unique populations: inbred strains of mice (Jackson Labs, Bar Harbor), baboon pedigrees (Southwest Foundation for Biomedical Research, San Antonio), and nuclear and extended families of ~300 probands with SS (Boston, Creteil). The animals will help us find quantitative trait loci (QTLs) and ultimately individual genes that influence baseline WBC. The present study in baboons is a continuation of earlier NIH supported research (R01 HL054141) to detect, characterize, and localize the effects of genes on normal quantitative variation in platelet biology phenotypes related to the bioavailability and activity of PDGF in pedigreed baboons. In addition to WBC number, we have collected data on other standard indicators of hematologic status (e.g., erythrocyte counts, platelet number and mean platelet volume); concentrations of PDGF, platelet activating factor, thromboxane B2, insulin-like growth factor-1, total serum cholesterol and high-density lipoprotein cholesterol; and inflammation. Whole Genome Gene Expression Profiling in Baboon Lymphocytes (pilot studies). In order to improve our ability to objectively prioritize candidate genes near sites of peak statistical genetic evidence for QTLs that influence variation in traits that influence quantitative variation in lipoprotein metabolism, oxidative stress and inflammation, we have begun whole genome (or transcriptome ) gene expression profiling in stored lymphocytes from 500 of the pedigreed baboons from this project. To date, with RNA isolated from lymphocytes from 122 pedigreed baboons we used the Illumina BeadStation 500 GX genome-wide expression platform and Illumina's Sentrix Human-6 Bead chip to profile the expression of more than 46,000 human transcripts (list of human genes on chip: www.illumina.com/products/arraysreagents/gwehuman6.ilmn). Included among the list of unambiguously detected genes from this initial pilot study were many that map to regions of the human genome that are orthologous to those in which we have localized QTLs for cellular and cytokine markers of inflammation in baboons. Within two weeks, when all 500 transcriptional profiles are completed, we will begin intensive statistical genetic analyses to localize QTLs for significantly heritable transcripts, identify correlations between transcripts and our focal traits, and conduct bivariate linkage genetic analyses between cis-regulated positional expression candidates and focal traits with QTLs the same regions of the baboon genome.
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