There is extreme inter-individual variation in human platelet reactivity, which likely contributes to occlusion of coronary and cerebral arteries upon atherosclerotic plaque rupture in some individuals, whereas other individuals repair the wound without occluding the vessel. However, there is a lack of understanding of the molecular and genetic mechanisms responsible for this variation in platelet function. Our previous genome wide scan and platelet RNA expression studies have elucidated mechanisms for epinephrine-induced platelet aggregation, identified mRNA expression differences, as well as unexpected evidence that microRNAs (miRNAs) regulate expression of the platelet secretory protein, VAMP8. Additional analyses suggest that alternate exon splicing and genomic copy number variation also contribute to variation in platelet gene expression. Although our platelet RNA profiling study included 29 subjects, it was limited in its ability to identify genes involved in the platelet response to agonists other than epinephrine. We now propose to recall 180 subjects for new platelet phenotyping, preparation of leukocyte-depleted platelet RNA and microarray profiling of mRNA and miRNA. Rigorous bioinformatic and statistical approaches will minimize false positives, and candidate mRNAs and miRNAs will be characterized experimentally. This approach will allow definition of genes regulating the platelet response to specific agonists (ADP, CRP, collagen, TRAP, U46619, ristocetin, CD9 and epinephrine), as well as genes common to multiple platelet stimuli, and permit assessment of the effects of gender and ethnicity (Aim 1).
In Aims 2 -4 we will characterize molecular mechanisms regulating mRNA levels in platelets of differing reactivity. Our analyses will identify associations between platelet reactivity and alternately spliced exons, thus serving as a screen for functional domains in the platelet protein (Aim 2). We will determine platelet miRNA profiles and characterize the function of differentially expressed miRNAs in platelets of differing reactivity (Aim 3). By correlating RNA expression data with our prior GWAS genotyping on these same subjects, we will address the role of copy number variation on platelet gene expression (Aim 4). We will also generate public web tools that provide the relative expression of platelet mRNAs and miRNAs according to gender, race and age. This research will result in new insights into platelet physiology, enhance our understanding of the genetics of platelet gene expression, facilitate the selection of gender and race-specific biomarkers for thrombosis risk, and provide useful tools for other platelet researchers.

Public Health Relevance

Cardiovascular disease is the major cause of mortality in the U.S. The goal of this research is to understand better the mechanisms that contribute to blood clotting in cardiovascular disease in order to develop better targets for drug development and better predictors of disease risk.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL102482-03
Application #
8252243
Study Section
Special Emphasis Panel (ZRG1-VH-E (90))
Program Officer
Sarkar, Rita
Project Start
2010-04-26
Project End
2014-03-31
Budget Start
2012-04-01
Budget End
2013-03-31
Support Year
3
Fiscal Year
2012
Total Cost
$725,979
Indirect Cost
$158,299
Name
Thomas Jefferson University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
053284659
City
Philadelphia
State
PA
Country
United States
Zip Code
19107
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Simon, Lukas M; Chen, Edward S; Edelstein, Leonard C et al. (2016) Integrative Multi-omic Analysis of Human Platelet eQTLs Reveals Alternative Start Site in Mitofusin 2. Am J Hum Genet 98:883-97
Holinstat, M; Bray, P F (2016) Protease receptor antagonism to target blood platelet therapies. Clin Pharmacol Ther 99:72-81
Zhou, Yuhang; Abraham, Shaji; Andre, Pierrette et al. (2015) Anti-miR-148a regulates platelet FcγRIIA signaling and decreases thrombosis in vivo in mice. Blood 126:2871-81
Londin, Eric; Loher, Phillipe; Telonis, Aristeidis G et al. (2015) Analysis of 13 cell types reveals evidence for the expression of numerous novel primate- and tissue-specific microRNAs. Proc Natl Acad Sci U S A 112:E1106-15
Tourdot, Benjamin E; Conaway, Stanley; Niisuke, Katrin et al. (2014) Mechanism of race-dependent platelet activation through the protease-activated receptor-4 and Gq signaling axis. Arterioscler Thromb Vasc Biol 34:2644-50
Londin, Eric R; Hatzimichael, Eleftheria; Loher, Phillipe et al. (2014) The human platelet: strong transcriptome correlations among individuals associate weakly with the platelet proteome. Biol Direct 9:3
Edelstein, Leonard C; Simon, Lukas M; Lindsay, Cory R et al. (2014) Common variants in the human platelet PAR4 thrombin receptor alter platelet function and differ by race. Blood 124:3450-8
Teruel-Montoya, Raul; Kong, Xianguo; Abraham, Shaji et al. (2014) MicroRNA expression differences in human hematopoietic cell lineages enable regulated transgene expression. PLoS One 9:e102259
Simon, Lukas M; Edelstein, Leonard C; Nagalla, Srikanth et al. (2014) Human platelet microRNA-mRNA networks associated with age and gender revealed by integrated plateletomics. Blood 123:e37-45

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