CC BY 4.0 · Thromb Haemost 2020; 120(02): 253-261
DOI: 10.1055/s-0039-3400305
Cellular Haemostasis and Platelets
Georg Thieme Verlag KG Stuttgart · New York

Hypoxia Modulates Platelet Purinergic Signalling Pathways

Gordon G. Paterson
1   APEX (Altitude Physiology Expeditions), Edinburgh, United Kingdom
2   Edinburgh Medical School, University of Edinburgh, Edinburgh, United Kingdom
,
Jason M. Young
1   APEX (Altitude Physiology Expeditions), Edinburgh, United Kingdom
2   Edinburgh Medical School, University of Edinburgh, Edinburgh, United Kingdom
,
Joseph A. Willson
3   University of Edinburgh Centre for Inflammation Research, The Queen's Medical Research Institute, University of Edinburgh, Edinburgh, United Kingdom
,
1   APEX (Altitude Physiology Expeditions), Edinburgh, United Kingdom
2   Edinburgh Medical School, University of Edinburgh, Edinburgh, United Kingdom
,
Rebecca C. Dru
1   APEX (Altitude Physiology Expeditions), Edinburgh, United Kingdom
2   Edinburgh Medical School, University of Edinburgh, Edinburgh, United Kingdom
,
Eleanor W. Lee
1   APEX (Altitude Physiology Expeditions), Edinburgh, United Kingdom
2   Edinburgh Medical School, University of Edinburgh, Edinburgh, United Kingdom
,
Greig S. Torpey
1   APEX (Altitude Physiology Expeditions), Edinburgh, United Kingdom
2   Edinburgh Medical School, University of Edinburgh, Edinburgh, United Kingdom
,
Sarah R. Walmsley
3   University of Edinburgh Centre for Inflammation Research, The Queen's Medical Research Institute, University of Edinburgh, Edinburgh, United Kingdom
,
Melissa V. Chan
4   Centre for Immunobiology, Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London, United Kingdom
,
4   Centre for Immunobiology, Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London, United Kingdom
,
John Kenneth Baillie
1   APEX (Altitude Physiology Expeditions), Edinburgh, United Kingdom
5   Division of Genetics and Genomics, The Roslin Institute, University of Edinburgh, Edinburgh, United Kingdom
6   Department of Anaesthesia, Critical Care and Pain Medicine, Royal Infirmary of Edinburgh, NHS Lothian, Edinburgh, United Kingdom
,
1   APEX (Altitude Physiology Expeditions), Edinburgh, United Kingdom
7   Department of Infection, Immunity and Cardiovascular Disease, University of Sheffield, Sheffield, United Kingdom
› Institutsangaben
Funding This study received funding from the Wilderness Medical Society Charles S. Houston Grant, supported by the Academy of Wilderness Medicine. It also received funding from British Heart Foundation, (Grant/Award Number: ‘FS/18/13/3328’,‘PG/15/47/31591’), Wilderness Medicine Society (Grant/Award Number: ‘Charles S. Houston grant’) and Wellcome Trust, (Grant/Award Number: ‘103258/Z/13/Z,A’).
Weitere Informationen

Publikationsverlauf

24. April 2019

29. September 2019

Publikationsdatum:
13. Dezember 2019 (online)

Abstract

Background Hypoxia resulting from ascent to high-altitude or pathological states at sea level is known to increase platelet reactivity. Previous work from our group has suggested that this may be adenosine diphosphate (ADP)-specific. Given the clinical importance of drugs targeting ADP pathways, research into the impact of hypoxia on platelet ADP pathways is highly important.

Methods Optimul aggregometry was performed on plasma from 29 lowland residents ascending to 4,700 m, allowing systematic assessment of platelet reactivity in response to several platelet agonists. Aggregometry was also performed in response to ADP in the presence of inhibitors of the two main ADP receptors, P2Y1 and P2Y12 (MRS2500 and cangrelor, respectively). Phosphorylation of vasodilator-stimulated phosphoprotein (VASP), a key determinant of platelet aggregation, was analysed using the VASPFix assay.

Results Hypobaric hypoxia significantly reduced the ability of a fixed concentration of cangrelor to inhibit ADP-induced aggregation and increased basal VASP phosphorylation. However, in the absence of P2Y receptor inhibitors, we did not find evidence of increased platelet sensitivity to any of the agonists tested and found reduced sensitivity to thrombin receptor-activating peptide-6 amide.

Conclusion Our results provide evidence of increased P2Y1 receptor activity at high altitude and suggest down-regulation of the P2Y12 pathway through increased VASP phosphorylation. These changes in ADP pathway activity are of potential therapeutic significance to high-altitude sojourners and hypoxic sea level patients prescribed platelet inhibitors and warrant further investigation.

Authors' Contributions

G.G.P., M.V.C., T.D.W. and A.A.R.T. conceived the study and designed the experiments; G.G.P., J.M.Y., J.A.W., C.J.G., R.C.D., E.W.L., G.S.T., M.V.C. and S.R.W. performed experiments and assisted with logistics; G.G.P., M.V.C., T.D.W., J.K.B. and A.A.R.T. analysed and interpreted the data; G.G.P. and A.A.R.T. drafted the manuscript with contributions from all authors.


Supplementary Material

 
  • References

  • 1 Bärtsch P. How thrombogenic is hypoxia?. JAMA 2006; 295 (19) 2297-2299
  • 2 Anand AC, Jha SK, Saha A, Sharma V, Adya CM. Thrombosis as a complication of extended stay at high altitude. Natl Med J India 2001; 14 (04) 197-201
  • 3 Jha SK, Anand AC, Sharma V, Kumar N, Adya CM. Stroke at high altitude: Indian experience. High Alt Med Biol 2002; 3 (01) 21-27
  • 4 Schaber M, Leichtfried V, Fries D. , et al. Influence of acute normobaric hypoxia on hemostasis in volunteers with and without acute mountain sickness. BioMed Res Int 2015; 2015: 593938
  • 5 Toff WD, Jones CI, Ford I. , et al. Effect of hypobaric hypoxia, simulating conditions during long-haul air travel, on coagulation, fibrinolysis, platelet function, and endothelial activation. JAMA 2006; 295 (19) 2251-2261
  • 6 Rocke AS, Paterson GG, Barber MT. , et al. Thromboelastometry and platelet function during acclimatization to high altitude. Thromb Haemost 2018; 118 (01) 63-71
  • 7 Pichler Hefti J, Risch L, Hefti U. , et al. Changes of coagulation parameters during high altitude expedition. Swiss Med Wkly 2010; 140 (7-8): 111-117
  • 8 Ninivaggi M, de Laat M, Lancé MMD. , et al. Hypoxia induces a prothrombotic state independently of the physical activity. PLoS One 2015; 10: e0141797
  • 9 Nieswandt B, Pleines I, Bender M. Platelet adhesion and activation mechanisms in arterial thrombosis and ischaemic stroke. J Thromb Haemost 2011; 9 (Suppl. 01) 92-104
  • 10 Jennings LK. Mechanisms of platelet activation: need for new strategies to protect against platelet-mediated atherothrombosis. Thromb Haemost 2009; 102 (02) 248-257
  • 11 Lehmann T, Mairbäurl H, Pleisch B, Maggiorini M, Bärtsch P, Reinhart WH. Platelet count and function at high altitude and in high-altitude pulmonary edema. J Appl Physiol (1985) 2006; 100 (02) 690-694
  • 12 Tyagi T, Ahmad S, Gupta N. , et al. Altered expression of platelet proteins and calpain activity mediate hypoxia-induced prothrombotic phenotype. Blood 2014; 123 (08) 1250-1260
  • 13 Fox JE, Taylor RG, Taffarel M, Boyles JK, Goll DE. Evidence that activation of platelet calpain is induced as a consequence of binding of adhesive ligand to the integrin, glycoprotein IIb-IIIa. J Cell Biol 1993; 120 (06) 1501-1507
  • 14 Jacobson KA, Deflorian F, Mishra S, Costanzi S. Pharmacochemistry of the platelet purinergic receptors. Purinergic Signal 2011; 7 (03) 305-324
  • 15 Keyes LE, Mather L, Duke C. , et al. Older age, chronic medical conditions and polypharmacy in Himalayan trekkers in Nepal: an epidemiologic survey and case series. J Travel Med 2016; 23 (06) 1-6
  • 16 Chan MV, Warner TD. Standardised optical multichannel (optimul) platelet aggregometry using high-speed shaking and fixed time point readings. Platelets 2012; 23 (05) 404-408
  • 17 Butt E, Abel K, Krieger M. , et al. cAMP- and cGMP-dependent protein kinase phosphorylation sites of the focal adhesion vasodilator-stimulated phosphoprotein (VASP) in vitro and in intact human platelets. J Biol Chem 1994; 269 (20) 14509-14517
  • 18 Hollopeter G, Jantzen H-M, Vincent D. , et al. Identification of the platelet ADP receptor targeted by antithrombotic drugs. Nature 2001; 409 (6817): 202-207
  • 19 Schwarz UR, Geiger J, Walter U, Eigenthaler M. Flow cytometry analysis of intracellular VASP phosphorylation for the assessment of activating and inhibitory signal transduction pathways in human platelets--definition and detection of ticlopidine/clopidogrel effects. Thromb Haemost 1999; 82 (03) 1145-1152
  • 20 Glenn JR, Dovlatova N, White AE, Dhillon K, Heptinstall S, Fox SC. ‘VASPFix’ for measurement of VASP phosphorylation in platelets and for monitoring effects of P2Y12 antagonists. Thromb Haemost 2014; 111 (03) 539-548
  • 21 Belsley DA, Kuh E, Welsch RE. Detecting Influential Observations and Outliers. In: Regression Diagnostics: Identifying Influential Data and Sources of Collinearity. Hoboken: John Wiley & Sons, Inc; 1980: 6-84
  • 22 Jones E, Oliphant T, Peterson P. , et al. SciPy: open source scientific tools for Python [Internet]. 2001 . Available at: http://www.scipy.org/ . Accessed October 16, 2019
  • 23 R Core Team. R: a language and environment for statistical computing. R Foundation for Statistical Computing; Vienna, Austria: [Internet]. 2019. . Available at: http://www.r-project.org/ . Accessed October 16, 2019
  • 24 Würtz M, Hvas A-M, Kristensen SD, Grove EL. Platelet aggregation is dependent on platelet count in patients with coronary artery disease. Thromb Res 2012; 129 (01) 56-61
  • 25 Chan MV, Armstrong PCJ, Papalia F, Kirkby NS, Warner TD. Optical multichannel (optimul) platelet aggregometry in 96-well plates as an additional method of platelet reactivity testing. Platelets 2011; 22 (07) 485-494
  • 26 Würtz M, Hvas A-M, Christensen KH, Rubak P, Kristensen SD, Grove EL. Rapid evaluation of platelet function using the Multiplate® Analyzer. Platelets 2014; 25 (08) 628-633
  • 27 Beck F, Geiger J, Gambaryan S. , et al. Temporal quantitative phosphoproteomics of ADP stimulation reveals novel central nodes in platelet activation and inhibition. Blood 2017; 129 (02) e1-e12
  • 28 Nisar S, Kelly E, Cullen PJ, Mundell SJ. Regulation of P2Y1 receptor traffic by sorting Nexin 1 is retromer independent. Traffic 2010; 11 (04) 508-519
  • 29 Magalhaes AC, Dunn H, Ferguson SS. Regulation of GPCR activity, trafficking and localization by GPCR-interacting proteins. Br J Pharmacol 2012; 165 (06) 1717-1736
  • 30 Jin J, Daniel JL, Kunapuli SP. Molecular basis for ADP-induced platelet activation. II. The P2Y1 receptor mediates ADP-induced intracellular calcium mobilization and shape change in platelets. J Biol Chem 1998; 273 (04) 2030-2034
  • 31 Grocott M, Montgomery H, Vercueil A. High-altitude physiology and pathophysiology: implications and relevance for intensive care medicine. Crit Care 2007; 11 (01) 203
  • 32 Feary JR, Rodrigues LC, Smith CJ, Hubbard RB, Gibson JE. Prevalence of major comorbidities in subjects with COPD and incidence of myocardial infarction and stroke: a comprehensive analysis using data from primary care. Thorax 2010; 65 (11) 956-962
  • 33 Marin JM, Carrizo SJ, Vicente E, Agusti AG. Long-term cardiovascular outcomes in men with obstructive sleep apnoea-hypopnoea with or without treatment with continuous positive airway pressure: an observational study. Lancet 2005; 365 (9464): 1046-1053
  • 34 Sanner BM, Konermann M, Tepel M, Groetz J, Mummenhoff C, Zidek W. Platelet function in patients with obstructive sleep apnoea syndrome. Eur Respir J 2000; 16 (04) 648-652
  • 35 Maclay JD, McAllister DA, Johnston S. , et al. Increased platelet activation in patients with stable and acute exacerbation of COPD. Thorax 2011; 66 (09) 769-774
  • 36 Bokinsky G, Miller M, Ault K, Husband P, Mitchell J. Spontaneous platelet activation and aggregation during obstructive sleep apnea and its response to therapy with nasal continuous positive airway pressure. A preliminary investigation. Chest 1995; 108 (03) 625-630
  • 37 Qureshi Z, Hobson AR. Clopidogrel “resistance”: where are we now?. Cardiovasc Ther 2013; 31 (01) 3-11
  • 38 Topçuoglu MA, Arsava EM, Ay H. Antiplatelet resistance in stroke. Expert Rev Neurother 2011; 11 (02) 251-263
  • 39 Jha PK, Sahu A, Prabhakar A. , et al. Genome-wide expression analysis suggests hypoxia-triggered hyper-coagulation leading to venous thrombosis at high altitude. Thromb Haemost 2018; 118 (07) 1279-1295
  • 40 Duplain H, Sartori C, Lepori M. , et al. Exhaled nitric oxide in high-altitude pulmonary edema: role in the regulation of pulmonary vascular tone and evidence for a role against inflammation. Am J Respir Crit Care Med 2000; 162 (01) 221-224
  • 41 Levett DZ, Fernandez BO, Riley HL. , et al; Caudwell Extreme Everest Research Group. The role of nitrogen oxides in human adaptation to hypoxia. Sci Rep 2011; 1: 109
  • 42 Massberg S, Grüner S, Konrad I. , et al. Enhanced in vivo platelet adhesion in vasodilator-stimulated phosphoprotein (VASP)-deficient mice. Blood 2004; 103 (01) 136-142
  • 43 Srihirun S, Piknova B, Sibmooh N. , et al. Phosphorylated vasodilator-stimulated phosphoprotein (P-VASPSer239) in platelets is increased by nitrite and partially deoxygenated erythrocytes. PLoS One 2018; 13: e0193747
  • 44 Webb AJ, Milsom AB, Rathod KS. , et al. Mechanisms underlying erythrocyte and endothelial nitrite reduction to nitric oxide in hypoxia: role for xanthine oxidoreductase and endothelial nitric oxide synthase. Circ Res 2008; 103 (09) 957-964
  • 45 Versteeg HH, Heemskerk JWM, Levi M, Reitsma PH. New fundamentals in hemostasis. Physiol Rev 2013; 93 (01) 327-358
  • 46 Kiouptsi K, Gambaryan S, Walter E, Walter U, Jurk K, Reinhardt C. Hypoxia impairs agonist-induced integrin αIIbβ3 activation and platelet aggregation. Sci Rep 2017; 7 (01) 7621
  • 47 Kauffenstein G, Bergmeier W, Eckly A. , et al. The P2Y(12) receptor induces platelet aggregation through weak activation of the α(IIb)β(3) integrin--a phosphoinositide 3-kinase-dependent mechanism. FEBS Lett 2001; 505 (02) 281-290
  • 48 Kamae T, Shiraga M, Kashiwagi H. , et al. Critical role of ADP interaction with P2Y12 receptor in the maintenance of alpha(IIb)beta3 activation: association with Rap1B activation. J Thromb Haemost 2006; 4 (06) 1379-1387
  • 49 Horstrup K, Jablonka B, Hönig-Liedl P, Just M, Kochsiek K, Walter U. Phosphorylation of focal adhesion vasodilator-stimulated phosphoprotein at Ser157 in intact human platelets correlates with fibrinogen receptor inhibition. Eur J Biochem 1994; 225 (01) 21-27
  • 50 Caradec J, Sirab N, Keumeugni C. , et al. ‘Desperate house genes’: the dramatic example of hypoxia. Br J Cancer 2010; 102 (06) 1037-1043
  • 51 Becker DM, Segal J, Vaidya D. , et al. Sex differences in platelet reactivity and response to low-dose aspirin therapy. JAMA 2006; 295 (12) 1420-1427