Mapping the platelet profile for functional genomic studies and demonstration of the effect size of the GP6 locus.
BACKGROUND
Evidence suggests the wide variation in platelet response within the population is genetically controlled. Unraveling the complex relationship between sequence variation and platelet phenotype requires accurate and reproducible measurement of platelet response.


OBJECTIVE
To develop a methodology suitable for measuring signaling pathway-specific platelet phenotype, to use this to measure platelet response in a large cohort, and to demonstrate the effect size of sequence variation in a relevant model gene.


METHODS
Three established platelet assays were evaluated: mobilization of [Ca(2+)](i), aggregometry and flow cytometry, each in response to adenosine 5'-diphosphate (ADP) or the glycoprotein (GP) VI-specific crosslinked collagen-related peptide (CRP). Flow cytometric measurement of fibrinogen binding and P-selectin expression in response to a single, intermediate dose of each agonist gave the best combination of reproducibility and inter-individual variability and was used to measure the platelet response in 506 healthy volunteers. Pathway specificity was ensured by blocking the main subsidiary signaling pathways.


RESULTS
Individuals were identified who were hypo- or hyper-responders for both pathways, or who had differential responses to the two agonists, or between outcomes. 89 individuals, retested three months later using the same methodology, showed high concordance between the two visits in all four assays (r(2) = 0.872, 0.868, 0.766 and 0.549); all subjects retaining their phenotype at recall. The effect of sequence variation at the GP6 locus accounted for approximately 35% of the variation in the CRP-XL response.


CONCLUSIONS
Genotyping-phenotype association studies in a well-characterized, large cohort provides a powerful strategy to measure the effect of sequence variation in genes regulating the platelet response.
Department of Cardiovascular Sciences, University of Leicester, Leicester, UK.
C I
CI
Jones
S F
SF
Garner
W
W
Angenent
A
A
Bernard
C
C
Berzuini
P
P
Burns
R W
RW
Farndale
J
J
Hogwood
A
A
Rankin
J C
JC
Stephens
B D
BD
Tom
J
J
Walton
F
F
Dudbridge
W H
WH
Ouwehand
A H
AH
Goodall
D000328:Adult; D001792:Blood Platelets / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry; D005260:Female; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D023281:Genomics / Q000379:methods; D006801:Humans; D008297:Male; D008875:Middle Aged; D010455:Peptides / Q000737:chemistry; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins / Q000235:genetics*; D015398:Signal Transduction
D002352:Carrier Proteins; D010455:Peptides; D010975:Platelet Aggregation Inhibitors; D010980:Platelet Membrane Glycoproteins; C435908:collagen-related peptide; C098341:platelet membrane glycoprotein VI