title: Investigation of cyclooxygenase and signaling pathways involved in human platelet aggregation mediated by synergistic interaction of various agonists.
abstract: In the present study, the mechanism(s) of synergistic interaction of various platelet mediators such as arachidonic acid (AA) when combined with 5-hydroxytryptamine (5-HT) or adenosine diphosphate (ADP) on human platelet aggregation were examined. The results demonstrated that 5-HT had no or negligible effect on aggregation but it did potentiate the aggregation response of AA. Similarly, the combination of subeffective concentrations of ADP and AA exhibited noticeable rise in platelet aggregation. Moreover, the observed synergistic effect of AA with 5-HT on platelets was inhibited by different cyclooxygenase (COX) inhibitors, namely ibuprofen and celecoxib, with half maximal inhibitory effect (IC50) values of 18.0 ± 1.8 and 15.6 ± 3.4 μmol/L, respectively. Interestingly, the synergistic effect observed for AA with 5-HT was, also, blocked by the 5-HT receptor blockers cyproheptadine (IC50=22.0 ± 7 μmol/L), ketanserin (IC50=152 ± 23 μmol/L), phospholipase C (PLC) inhibitor (U73122; IC50=6.1 ± 0.8 μmol/L), and mitogen activated protein kinase (MAPK) inhibitor (PD98059; IC50=3.8 ± 0.5 μmol/L). Likewise, the synergism of AA and ADP was, also, attenuated by COX inhibitors (ibuprofen; IC50=20 ± 4 μmol/L and celecoxib; IC50=24 ± 7 μmol/L), PLC inhibitor (U73122; IC50=3.7 ± 0.3 μmol/L), and MAPK inhibitor (PD98059; IC50=2.8 ± 1.1 μmol/L). Our observed data demonstrate that the combination of subthreshold concentrations of agonists amplifies platelet aggregation and that these synergistic effects largely depend on activation of COX/thromboxane A2, receptor-operated Ca(2+) channels, Gq/PLC, and MAPK signaling pathways. Moreover, our data revealed that inhibition of COX pathways by using both selective and/or non-selective COX inhibitors blocks not only AA metabolism and thromboxane A2 formation, but also its binding to Gq receptors and activation of receptor-operated Ca(2+) channels in platelets. Overall, our results show that PLC and MAPK inhibitors proved to inhibit the synergistic activation of platelets by several/multiple agonists.
authors.affiliation: Dr Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical and Biological Sciences, University of Karachi, Karachi, Pakistan ; Department of Pharmacology and Therapeutics, College of Medicine and Health Sciences, United Arab Emirates University, Al Ain, United Arab Emirates.
authors.forename: Nadia
authors.initials: N
authors.lastname: Khan
authors.affiliation: Dr Panjwani Center for Molecular Medicine and Drug Research, International Center for Chemical and Biological Sciences, University of Karachi, Karachi, Pakistan.
authors.forename: Ahsana Dar
authors.initials: AD
authors.lastname: Farooq
authors.affiliation: Department of Pharmacology and Therapeutics, College of Medicine and Health Sciences, United Arab Emirates University, Al Ain, United Arab Emirates.
authors.forename: Bassem
authors.initials: B
authors.lastname: Sadek
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism*; D000328:Adult; D016718:Arachidonic Acid / Q000378:metabolism*; D016861:Cyclooxygenase Inhibitors / Q000008:administration & dosage / Q000494:pharmacology; D006801:Humans; D020128:Inhibitory Concentration 50; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D010974:Platelet Aggregation / Q000187:drug effects / Q000502:physiology*; D011451:Prostaglandin-Endoperoxide Synthases / Q000187:drug effects / Q000378:metabolism; D012701:Serotonin / Q000378:metabolism*; D012702:Serotonin Antagonists / Q000008:administration & dosage / Q000494:pharmacology; D015398:Signal Transduction; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors / Q000378:metabolism; D055815:Young Adult
chemical_list: D016861:Cyclooxygenase Inhibitors; D012702:Serotonin Antagonists; D016718:Arachidonic Acid; D012701:Serotonin; D000244:Adenosine Diphosphate; D011451:Prostaglandin-Endoperoxide Synthases; D020928:Mitogen-Activated Protein Kinases; D010738:Type C Phospholipases
keywords: 5-hydroxytryptophan; adenosine-5-diphosphate; arachidonic acid; cyclooxygenase; platelet aggregation; signaling pathway; synergism