title: Patient-independent variables affecting the assessment of aspirin responsiveness by serum thromboxane measurement.
abstract: The serum TXB2(sTXB2) assay reflects the pharmacodynamics of platelet inhibition by low-dose aspirin. However, different studies reported variable sTXB2values. sTXB2assay requires whole blood incubation at 37 °C as a condition for optimal thrombin generation, arachidonic acid release and its metabolism by platelet cyclooxygenase-1 to form TXA2. Access to 37 °C incubation may be variably delayed, and different methods to quantitate sTXB2may contribute to variable results between different Centers. We investigated whether delaying 37 °C incubation and/or analytical issues affect sTXB2concentrations, biasing the assessment of aspirin responsiveness. Sixty-eight samples from 54 volunteers, on- and off-aspirin, were incubated at 37 °C immediately after sampling (reference sample) or after 5, 10, 15, 20, 30 or 60 minutes at room temperature (RT); 8 samples remained at RT 60 minutes, without subsequent incubation; 314 sera were measured by enzyme immunoassay (EIA) and liquid chromatography-tandem mass-spectrometry (LC/MS-MS) methods. sTXB2concentrations decreased exponentially as a function of the delay before 37 °C incubation, ranging from 94 ± 11 % at 5 minutes to 23 ± 22 % of the reference sample after 60 minutes at RT. There was high agreement between EIA and LC/MS-MS. Moreover, we simulated the influence of a 15- or 30-minute delayed incubation on 300 sTXB2 measurements from previously-studied, aspirin-treated patients. Delayed incubation reduced the percentage of aspirin 'non-responders' by 22 % to 52 %, depending on the response threshold. In conclusion, a variable delay in the 37 °C incubation of blood samples may affect the assessment of platelet cyclooxygenase-1 inhibition by aspirin and confound the characterization of the determinants of aspirin responsiveness.
authors.forename: Giovanna
authors.initials: G
authors.lastname: Petrucci
authors.forename: Alessandro
authors.initials: A
authors.lastname: Rizzi
authors.forename: Viviana
authors.initials: V
authors.lastname: Cavalca
authors.forename: Aida
authors.initials: A
authors.lastname: Habib
authors.forename: Dario
authors.initials: D
authors.lastname: Pitocco
authors.forename: Fabrizio
authors.initials: F
authors.lastname: Veglia
authors.forename: Paola
authors.initials: P
authors.lastname: Ranalli
authors.forename: Francesco
authors.initials: F
authors.lastname: Zaccardi
authors.forename: Francesca
authors.initials: F
authors.lastname: Pagliaccia
authors.forename: Elena
authors.initials: E
authors.lastname: Tremoli
authors.forename: Carlo
authors.initials: C
authors.lastname: Patrono
authors.affiliation: Bianca Rocca, MD, PhD, Istituto di Farmacologia, Università Cattolica, Largo F. Vito 1, 00168 Rome, Italy, Tel.: +39 06 30154253, E-mail: b.rocca@tiscali.it; bianca.rocca@unicatt.it.
authors.forename: Bianca
authors.initials: B
authors.lastname: Rocca
mesh_terms: D000328:Adult; D000368:Aged; D001241:Aspirin / Q000494:pharmacology*; D001792:Blood Platelets / Q000187:drug effects; D051545:Cyclooxygenase 1 / Q000378:metabolism; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology*; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D013929:Thromboxane B2 / Q000097:blood*
chemical_list: D016861:Cyclooxygenase Inhibitors; D013929:Thromboxane B2; D051545:Cyclooxygenase 1; C496511:PTGS1 protein, human; D001241:Aspirin
keywords: Thromboxane B2; aspirin; cyclooxygenase-1; enzyme immunoassay; liquid chromatography-tandem mass spectrometry