title: Aspirin extrusion from human platelets through multidrug resistance protein-4-mediated transport: evidence of a reduced drug action in patients after coronary artery bypass grafting.
abstract: OBJECTIVE
  In this study we investigate: 1) the role of multidrug resistance protein-4 (MRP4), an organic anion unidirectional transporter, in modulating aspirin action on human platelet cyclooxygenase (COX)-1; and 2) whether the impairment of aspirin-COX-1 interaction, found in coronary artery bypass grafting (CABG) patients, could be dependent on MRP4-mediated transport.
  
  
  BACKGROUND
  Platelets of CABG patients present a reduced sensitivity to aspirin despite in vivo and in vitro drug treatment. Aspirin is an organic anion and could be a substrate for MRP4.
  
  
  METHODS
  Intracellular aspirin concentration and drug COX-1 activity, measured by thrombin-induced thromboxane B2 (TxB2) production, were evaluated in platelets obtained from healthy volunteers (HV) and hematopoietic-progenitor cell cultures reducing or not reducing MRP4-mediated transport. Platelet MRP4 expression was evaluated, in platelets from HV and CABG patients, by dot-blot or by immunogold-electromicrographs or immunofluorescence-microscopy analysis.
  
  
  RESULTS
  Inhibition of MRP4-mediated transport by dipyridamole or Mk-571 increases aspirin entrapment and its in vitro effect on COX-1 activity (142.7 ± 34.6 pg/10(8) cells vs. 343.7 ± 169.3 pg/10⁸ cells TxB2-production). Platelets derived from megakaryocytes transfected with MRP4 small interfering ribonucleic acid have a higher aspirin entrapment and drug COX-1 activity. Platelets from CABG patients showed a high expression of MRP4 whose in vitro inhibition enhanced aspirin effect on COX-1 (349 ± 141 pg/10⁸ cells vs. 1,670 ± 646 pg/10⁸ cells TxB2-production).
  
  
  CONCLUSIONS
  Aspirin is a substrate for MRP4 and can be extruded from platelet through its transportation. Aspirin effect on COX-1 is little-related to MRP4-mediated aspirin transport in HV, but in CABG patients with MRP4 over-expression, its pharmacological inhibition enhances aspirin action in an efficient way.
authors.affiliation: Department of Experimental Medicine, Sapienza University of Rome, Rome, Italy.
authors.forename: Teresa
authors.initials: T
authors.lastname: Mattiello
authors.forename: Raffaella
authors.initials: R
authors.lastname: Guerriero
authors.forename: Lavinia Vittoria
authors.initials: LV
authors.lastname: Lotti
authors.forename: Elisabetta
authors.initials: E
authors.lastname: Trifirò
authors.forename: Maria Pia
authors.initials: MP
authors.lastname: Felli
authors.forename: Alessandro
authors.initials: A
authors.lastname: Barbarulo
authors.forename: Bruna
authors.initials: B
authors.lastname: Pucci
authors.forename: Paola
authors.initials: P
authors.lastname: Gazzaniga
authors.forename: Carlo
authors.initials: C
authors.lastname: Gaudio
authors.forename: Luigi
authors.initials: L
authors.lastname: Frati
authors.forename: Fabio M
authors.initials: FM
authors.lastname: Pulcinelli
mesh_terms: D000328:Adult; D001241:Aspirin / Q000493:pharmacokinetics* / Q000494:pharmacology; D001692:Biological Transport / Q000187:drug effects; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D002478:Cells, Cultured; D001026:Coronary Artery Bypass; D000242:Cyclic AMP / Q000494:pharmacology; D051545:Cyclooxygenase 1 / Q000378:metabolism; D015232:Dinoprostone / Q000378:metabolism; D004347:Drug Interactions; D004351:Drug Resistance; D005260:Female; D005343:Fibrinolytic Agents / Q000493:pharmacokinetics* / Q000494:pharmacology; D006801:Humans; D008297:Male; D008875:Middle Aged; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000378:metabolism / Q000494:pharmacology*; D010974:Platelet Aggregation; D010975:Platelet Aggregation Inhibitors / Q000493:pharmacokinetics* / Q000494:pharmacology; D011422:Propionates / Q000494:pharmacology; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism; D011804:Quinolines / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000378:metabolism; D020156:Salicylic Acid / Q000493:pharmacokinetics; D013929:Thromboxane B2 / Q000378:metabolism
chemical_list: D001241:Aspirin; D000242:Cyclic AMP; D051545:Cyclooxygenase 1; D015232:Dinoprostone; D005343:Fibrinolytic Agents; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D010975:Platelet Aggregation Inhibitors; D011422:Propionates; D011451:Prostaglandin-Endoperoxide Synthases; D011804:Quinolines; D034741:RNA, Small Interfering; D020156:Salicylic Acid; D013929:Thromboxane B2; C073492:ABCC4 protein, human; C059141:verlukast