Arjit Nigam, Voddarahally N Manjuprasanna, Meghna U Naik, Ulhas P Naik
{"title":"Platelet spreading and clot retraction are regulated by 2 distinct α<sub>IIb</sub>β<sub>3</sub> outside-in signaling pathways.","authors":"Arjit Nigam, Voddarahally N Manjuprasanna, Meghna U Naik, Ulhas P Naik","doi":"10.1124/jpet.124.002149","DOIUrl":null,"url":null,"abstract":"<p><p>Bidirectional signaling through platelet integrin α<sub>IIb</sub>β<sub>3</sub> is essential in hemostasis and thrombosis. In quiescent platelets, α<sub>IIb</sub>β<sub>3</sub> is in a low-affinity ligand binding state. However, on platelet activation by agonists through inside-out signaling, a rapid switch in the conformation of the integrin results in a high affinity ligand binding state capable of binding soluble fibrinogen. Ligand binding to the α<sub>IIb</sub>β<sub>3</sub> induces a signaling termed outside-in signaling that regulate platelet spreading and clot retraction. These events are often interchangeably used to represent outside-in signaling pathway. Using pharmacological inhibitors of known signaling molecules that have been implicated to regulate outside-in signaling, we assessed human platelet spreading and clot retraction. We found that inhibition of phosphoinositide-3-kinase, phospholipase C, protein kinase C, and focal adhesion kinase strongly attenuated both platelet spreading and clot retraction suggesting that they are essential for both clot retraction and platelet spreading, whereas inhibition of Rac1, rho-associated, coiled-coil containing protein kinase, p38, and MEK did not affect platelet spreading but significantly delayed clot retraction suggesting that these signaling molecules do not participate in platelet spreading. Interestingly, Src family kinases are required for platelet spreading and FAK activation but suppress clot retraction because their inhibition causes faster clot retraction. Thus, it becomes evident that platelet spreading, and clot retraction are differently regulated through α<sub>IIb</sub>β<sub>3</sub> outside-in signaling and should not be used interchangeably as readout for α<sub>IIb</sub>β<sub>3</sub> outside-in signaling assessment. SIGNIFICANCE STATEMENT: Current antiplatelet drugs have increased risk of bleeding and low efficacy. There is an increased effort to identify novel antiplatelet agents that have improved efficacy with reduced risk of bleeding. It is increasingly felt that inhibition of α<sub>IIb</sub>β<sub>3-</sub>induced outside-in signaling may inhibit thrombosis without compromising hemostasis. However, the signaling entities regulating outside-in signaling are poorly understood. The work included in this article delineates the distinct signaling pathways involved in outside-in signaling and identify potential novel targets for intervention of thrombosis.</p>","PeriodicalId":16798,"journal":{"name":"Journal of Pharmacology and Experimental Therapeutics","volume":"392 1","pages":"100012"},"PeriodicalIF":3.1000,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Pharmacology and Experimental Therapeutics","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1124/jpet.124.002149","RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2024/11/22 0:00:00","PubModel":"Epub","JCR":"Q2","JCRName":"PHARMACOLOGY & PHARMACY","Score":null,"Total":0}
引用次数: 0
Abstract
Bidirectional signaling through platelet integrin αIIbβ3 is essential in hemostasis and thrombosis. In quiescent platelets, αIIbβ3 is in a low-affinity ligand binding state. However, on platelet activation by agonists through inside-out signaling, a rapid switch in the conformation of the integrin results in a high affinity ligand binding state capable of binding soluble fibrinogen. Ligand binding to the αIIbβ3 induces a signaling termed outside-in signaling that regulate platelet spreading and clot retraction. These events are often interchangeably used to represent outside-in signaling pathway. Using pharmacological inhibitors of known signaling molecules that have been implicated to regulate outside-in signaling, we assessed human platelet spreading and clot retraction. We found that inhibition of phosphoinositide-3-kinase, phospholipase C, protein kinase C, and focal adhesion kinase strongly attenuated both platelet spreading and clot retraction suggesting that they are essential for both clot retraction and platelet spreading, whereas inhibition of Rac1, rho-associated, coiled-coil containing protein kinase, p38, and MEK did not affect platelet spreading but significantly delayed clot retraction suggesting that these signaling molecules do not participate in platelet spreading. Interestingly, Src family kinases are required for platelet spreading and FAK activation but suppress clot retraction because their inhibition causes faster clot retraction. Thus, it becomes evident that platelet spreading, and clot retraction are differently regulated through αIIbβ3 outside-in signaling and should not be used interchangeably as readout for αIIbβ3 outside-in signaling assessment. SIGNIFICANCE STATEMENT: Current antiplatelet drugs have increased risk of bleeding and low efficacy. There is an increased effort to identify novel antiplatelet agents that have improved efficacy with reduced risk of bleeding. It is increasingly felt that inhibition of αIIbβ3-induced outside-in signaling may inhibit thrombosis without compromising hemostasis. However, the signaling entities regulating outside-in signaling are poorly understood. The work included in this article delineates the distinct signaling pathways involved in outside-in signaling and identify potential novel targets for intervention of thrombosis.
期刊介绍:
A leading research journal in the field of pharmacology published since 1909, JPET provides broad coverage of all aspects of the interactions of chemicals with biological systems, including autonomic, behavioral, cardiovascular, cellular, clinical, developmental, gastrointestinal, immuno-, neuro-, pulmonary, and renal pharmacology, as well as analgesics, drug abuse, metabolism and disposition, chemotherapy, and toxicology.