Joanne C Clark1,2, Deirdre M Kavanagh1,2, Stephanie Watson1, Jeremy A Pike1,2, Robert K Andrews3, Elizabeth E Gardiner4, Natalie S Poulter1,2, Stephen J Hill2,5, Steve P Watson1,2. 1. Institute of Cardiovascular Sciences, Level 1 IBR, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham, United Kingdom. 2. Centre of Membrane Proteins and Receptors (COMPARE), The Universities of Birmingham and Nottingham, The Midlands, United Kingdom. 3. Australian Centre for Blood Diseases, Monash University, Melbourne, Australia. 4. Department of Cancer Biology and Therapeutics, John Curtin School of Medical Research, Australian National University, Canberra, Australia. 5. Division of Physiology, Pharmacology and Neuroscience, School of Life Sciences, University of Nottingham Medical School, Queen's Medical Centre, Nottingham, United Kingdom.
Abstract
BACKGROUND: The G protein-coupled receptor, adenosine A2A, signals through the stimulatory G protein, Gs, in platelets leading to activation of adenylyl cyclase and elevation of cyclic adenosine monophosphate (cAMP) and inhibition of platelet activation. OBJECTIVE: This article investigates the effect of A2A receptor activation on signalling by the collagen receptor glycoprotein (GP) VI in platelets. METHODS: Washed human platelets were stimulated by collagen or the GPVI-specific agonist collagen-related peptide (CRP) in the presence of the adenosine receptor agonist, 5'-N-ethylcarboxamidoadenosine (NECA) or the adenylyl cyclase activator, forskolin and analysed for aggregation, adenosine triphosphate secretion, protein phosphorylation, spreading, Ca2+ mobilisation, GPVI receptor clustering, cAMP, thromboxane B2 (TxB2) and P-selectin exposure. RESULTS: NECA, a bioactive adenosine analogue, partially inhibits aggregation and secretion to collagen or CRP in the absence or presence of the P2Y12 receptor antagonist, cangrelor and the cyclooxygenase inhibitor, indomethacin. The inhibitory effect in the presence of the three inhibitors is largely overcome at higher concentrations of collagen but not CRP. Neither NECA nor forskolin altered clustering of GPVI, elevation of Ca2+ or spreading of platelets on a collagen surface. Further, neither NECA nor forskolin, altered collagen-induced tyrosine phosphorylation of Syk, LAT nor PLCγ2. However, NECA and forskolin inhibited platelet activation by the TxA2 mimetic, U46619, but not the combination of adenosine diphosphate and collagen. CONCLUSION: NECA and forskolin have no effect on the proximal signalling events by collagen. They inhibit platelet activation in a response-specific manner in part through inhibition of the feedback action of TxA2. Georg Thieme Verlag KG Stuttgart · New York.
BACKGROUND: The G protein-coupled receptor, adenosine A2A, signals through the stimulatory G protein, Gs, in platelets leading to activation of adenylyl cyclase and elevation of cyclic adenosine monophosphate (cAMP) and inhibition of platelet activation. OBJECTIVE: This article investigates the effect of A2A receptor activation on signalling by the collagen receptor glycoprotein (GP) VI in platelets. METHODS: Washed human platelets were stimulated by collagen or the GPVI-specific agonist collagen-related peptide (CRP) in the presence of the adenosine receptor agonist, 5'-N-ethylcarboxamidoadenosine (NECA) or the adenylyl cyclase activator, forskolin and analysed for aggregation, adenosine triphosphate secretion, protein phosphorylation, spreading, Ca2+ mobilisation, GPVI receptor clustering, cAMP, thromboxane B2 (TxB2) and P-selectin exposure. RESULTS: NECA, a bioactive adenosine analogue, partially inhibits aggregation and secretion to collagen or CRP in the absence or presence of the P2Y12 receptor antagonist, cangrelor and the cyclooxygenase inhibitor, indomethacin. The inhibitory effect in the presence of the three inhibitors is largely overcome at higher concentrations of collagen but not CRP. Neither NECA nor forskolin altered clustering of GPVI, elevation of Ca2+ or spreading of platelets on a collagen surface. Further, neither NECA nor forskolin, altered collagen-induced tyrosine phosphorylation of Syk, LAT nor PLCγ2. However, NECA and forskolin inhibited platelet activation by the TxA2 mimetic, U46619, but not the combination of adenosine diphosphate and collagen. CONCLUSION: NECA and forskolin have no effect on the proximal signalling events by collagen. They inhibit platelet activation in a response-specific manner in part through inhibition of the feedback action of TxA2. Georg Thieme Verlag KG Stuttgart · New York.
Authors: Raluca A I Neagoe; Elizabeth E Gardiner; David Stegner; Bernhard Nieswandt; Steve P Watson; Natalie S Poulter Journal: Int J Mol Sci Date: 2022-03-29 Impact factor: 5.923