Skip to main content
SpringerLink
Log in

P2X1 receptor blockers reduce the number of circulating thrombocytes and the overall survival of urosepsis with haemolysin-producing Escherichia coli

  • Original Article
  • Published:
Purinergic Signalling Aims and scope Submit manuscript

Abstract

Urosepsis is a severe condition often caused by Escherichia coli that spontaneously have ascended the urinary tract to the kidneys causing pyelonephritis and potentially bacteraemia. The number of sepsis cases has been steadily increasing over the last decades, and there are still no specific, molecular supportive therapies for sepsis to supplement antibiotic treatment. P2X1 receptors are expressed by a number of immune cells including thrombocytes, which presently have been established as an important player in the acute immune response to bacterial infections. P2X1 receptor-deficient mice have been shown to be relatively protected against urosepsis, with markedly reduced levels of circulating proinflammatory cytokines and intravascular coagulation. However, here we show that continuous intravenous infusion with P2X1 receptor antagonist markedly accelerates development of a septic response to induced bacteraemia with uropathogenic E. coli. Mice exposed to the P2X1 receptor antagonists die very early with haematuria, substantially elevated plasma levels of proinflammatory cytokines, massive intravascular coagulation and a concomitant reduction in circulating thrombocytes. Interestingly, infusion of P2X1 receptor antagonists causes a marked acute reduction in circulating thrombocytes and a higher number of bacteria in the blood. These data support the notion that the number of functional thrombocytes is important for the acute defence against bacteria in the circulation and that the P2X1 receptor potentially could be essential for this response.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  1. Foxman B (2003) Epidemiology of urinary tract infections: incidence, morbidity, and economic costs. Dis Mon 49(2):53–70

    Article  PubMed  Google Scholar 

  2. Ragnarsdottir B, Svanborg C (2012) Susceptibility to acute pyelonephritis or asymptomatic bacteriuria: host-pathogen interaction in urinary tract infections. Pediatr Nephrol 27(11):2017–2029

    Article  PubMed  Google Scholar 

  3. Foxman B (2002) Epidemiology of urinary tract infections: incidence, morbidity, and economic costs. Am J Med 113(Suppl 1A):5S–13S

    Article  PubMed  Google Scholar 

  4. Johnson JR (1991) Virulence factors in Escherichia coli urinary tract infection. Clin Microbiol Rev 4(1):80–128

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  5. Connell I, Agace W, Klemm P, Schembri M, Marild S, Svanborg C (1996) Type 1 fimbrial expression enhances Escherichia coli virulence for the urinary tract. Proc Natl Acad Sci U S A 93(18):9827–9832

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  6. Svanborg C (2013) Urinary tract infections in children: microbial virulence versus host susceptibility. Adv Exp Med Biol 764:205–210

    Article  CAS  PubMed  Google Scholar 

  7. Wullt B, Bergsten G, Connell H, Rollano P, Gebretsadik N, Hull R, Svanborg C (2000) P fimbriae enhance the early establishment of Escherichia coli in the human urinary tract. Mol Microbiol 38(3):456–464

    Article  CAS  PubMed  Google Scholar 

  8. Johnson JR, Stell AL (2000) Extended virulence genotypes of Escherichia coli strains from patients with urosepsis in relation to phylogeny and host compromise. J Infect Dis 181(1):261–272

    Article  CAS  PubMed  Google Scholar 

  9. Cavalieri SJ, Bohach GA, Snyder IS (1984) Escherichia coli α-hemolysin: characteristics and probable role in pathogenicity. Microbiol Rev 48(4):326–343

    PubMed Central  CAS  PubMed  Google Scholar 

  10. Skals M, Bjaelde RG, Reinholdt J, Poulsen K, Vad BS, Otzen DE, Leipziger J, Praetorius HA (2014) Bacterial RTX toxins allow acute ATP release from human erythrocytes directly through the toxin pore. J Biol Chem 289:19098–19109

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  11. Skals MG, Jorgensen NR, Leipziger J, Praetorius HA (2009) α -hemolysin from Escherichia coli uses endogenous amplification through P2X receptor activation to induce hemolysis. Proc Natl Acad Sci U S A 106:4030–4035

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  12. Larsen CK, Skals M, Wang T, Cheema MU, Leipziger J, Praetorius HA (2011) Python erythrocytes are resistant to α-Hemolysin from Escherichia coli. J Membr Biol 244(3):131–140

    Article  CAS  PubMed  Google Scholar 

  13. Fagerberg SK, Jakobsen MR, Skals M, Praetorius HA (2016) Inhibition of P2X receptors protects human monocytes against damage by leukotoxin from Aggregatibacter actinomycetemcomitans and ⍺-hemolysin from Escherichia coli. Infect Immun 84(11):3114–3130

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  14. Christensen MG, Fagerberg SK, de Bruijn PI, Bjaelde RG, Jakobsen H, Leipziger J, Skals M, Praetorius HA (2015) [Ca2+]i oscillations and IL-6 release induced by alpha-hemolysin from Escherichia coli require P2 receptor activation in renal epithelia. J Biol Chem 290(23):14776–14784

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  15. Greve AS, Skals M, Fagerberg SK, Tonnus W, Ellermann-Eriksen S, Evans RJ, Linkermann A, Praetorius HA (2017) P2X1, P2X4, and P2X7 receptor knock out mice expose differential outcome of sepsis induced by ⍺-haemolysin producing Escherichia coli. Front Cell Infect Microbiol 7:113

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  16. Fagerberg SK, Patel P, Andersen LW, Lui X, Donnino MW, Praetorius HA (2018) Erythrocyte P2X1 receptor expression is correlated with change in haematocrit in patients admitted to the ICU with blood pathogen-positive sepsis. Crit Care 22(1):181

    Article  PubMed Central  PubMed  Google Scholar 

  17. Maitre B, Magnenat S, Heim V, Ravanat C, Evans RJ, de la Salle H, Gachet C, Hechler B (2015) The P2X1 receptor is required for neutrophil extravasation during lipopolysaccharide-induced lethal endotoxemia in mice. J Immunol 194(2):739–749

    Article  CAS  PubMed  Google Scholar 

  18. Adriouch S, Dox C, Welge V, Seman M, Koch-Nolte F, Haag F (2002) Cutting edge: a natural P451L mutation in the cytoplasmic domain impairs the function of the mouse P2X7 receptor. J Immunol 169(8):4108–4112

    Article  CAS  PubMed  Google Scholar 

  19. Boyden ED, Dietrich WF (2006) Nalp1b controls mouse macrophage susceptibility to anthrax lethal toxin. Nat Genet 38(2):240–244

    Article  CAS  PubMed  Google Scholar 

  20. Lecut C, Faccinetto C, Delierneux C, van Oerle R, Spronk HM, Evans RJ, El Benna J, Bours V, Oury C (2012) ATP-gated P2X1 ion channels protect against endotoxemia by dampening neutrophil activation. J Thromb Haemost 10(3):453–465

    Article  CAS  PubMed  Google Scholar 

  21. Johnsen N, Hamilton ADM, Greve AS, Christensen MG, Therkildsen JR, Wehmoller J, Skals M, Praetorius HA (2019) Alpha-haemolysin production, as a single factor, causes fulminant sepsis in a model of Escherichia coli-induced bacteraemia. Cell Microbiol e13017. https://www.ncbi.nlm.nih.gov/pubmed/30761726

  22. Hechler B, Magnenat S, Zighetti ML, Kassack MU, Ullmann H, Cazenave JP, Evans R, Cattaneo M, Gachet C (2005) Inhibition of platelet functions and thrombosis through selective or nonselective inhibition of the platelet P2 receptors with increasing doses of NF449 [4,4′,4″,4″'-(carbonylbis(imino-5,1,3-benzenetriylbis-(carbonylimino)))tetrakis -benzene-1,3-disulfonic acid octasodium salt]. J Pharmacol Exp Ther 314(1):232–243

    Article  CAS  PubMed  Google Scholar 

  23. Martin GS, Mannino DM, Eaton S, Moss M (2003) The epidemiology of sepsis in the United States from 1979 through 2000. N Engl J Med 348(16):1546–1554. https://doi.org/10.1056/NEJMoa022139

    Article  PubMed  Google Scholar 

  24. Gotts JE, Matthay MA (2016) Sepsis: pathophysiology and clinical management. BMJ 353:i1585

    Article  PubMed  Google Scholar 

  25. Skals M, Praetorius HA (2013) Mechanisms of cytolysin-induced cell damage - a role for auto- and paracrine signalling. Acta Physiol 209:95–113

    Article  CAS  Google Scholar 

  26. Land WG, Agostinis P, Gasser S, Garg AD, Linkermann A (2016) DAMP - induced allograft and tumor rejection: the circle is closing. Am J Transplant 16:3322–3337

    Article  CAS  PubMed  Google Scholar 

  27. Jin J, Daniel JL, Kunapuli SP (1998) 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 273(4):2030–2034

    Article  CAS  PubMed  Google Scholar 

  28. Daniel JL, Dangelmaier C, Jin J, Ashby B, Smith JB, Kunapuli SP (1998) Molecular basis for ADP-induced platelet activation. I Evidence for three distinct ADP receptors on human platelets. J Biol Chem 273(4):2024–2029

    Article  CAS  PubMed  Google Scholar 

  29. Jarvis GE, Humphries RG, Robertson MJ, Leff P (2000) ADP can induce aggregation of human platelets via both P2Y(1) and P(2T) receptors. Br J Pharmacol 129(2):275–282

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  30. Darbousset R, Delierneux C, Mezouar S, Hego A, Lecut C, Guillaumat I, Riederer MA, Evans RJ, Dignat-George F, Panicot-Dubois L, Oury C, Dubois C (2014) P2X1 expressed on polymorphonuclear neutrophils and platelets is required for thrombosis in mice. Blood 124(16):2575–2585

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  31. Oury C, Kuijpers MJ, Toth-Zsamboki E, Bonnefoy A, Danloy S, Vreys I, Feijge MA, De VR, Vermylen J, Heemskerk JW, Hoylaerts MF (2003) Overexpression of the platelet P2X1 ion channel in transgenic mice generates a novel prothrombotic phenotype. Blood 101(10):3969–3976

    Article  CAS  PubMed  Google Scholar 

  32. Oury C, Toth-Zsamboki E, Thys C, Tytgat J, Vermylen J, Hoylaerts MF (2001) The ATP-gated P2X1 ion channel acts as a positive regulator of platelet responses to collagen. Thromb Haemost 86(5):1264–1271

    CAS  PubMed  Google Scholar 

  33. Kazama I, Baba A, Endo Y, Toyama H, Ejima Y, Matsubara M, Tachi M (2015) Salicylate inhibits thrombopoiesis in rat megakaryocytes by changing the membrane micro-architecture. Cell Physiol Biochem 35(6):2371–2382

    Article  CAS  PubMed  Google Scholar 

  34. Siao WZ, Chuang WY, Su CH, Huang SF, Tu WK, Chan KC (2017) A rare case of ticagrelor-induced profound isolated thrombocytopenia. Acta Cardiol Sin 33(5):556–558

    PubMed Central  PubMed  Google Scholar 

  35. Rubano JA, Chen K, Sullivan B, Vosswinkel JA, Jawa RS (2015) Clopidogrel-associated thrombotic thrombocytopenic purpura following endovascular treatment of spontaneous carotid artery dissection. J Neurol Surg Rep 76(2):e287–e290

    Article  PubMed Central  PubMed  Google Scholar 

  36. Guo YL, Li JJ, Yuan JQ, Qin XW, Zheng X, Mu CW, Hua YH (2010) Profound thrombocytopenia induced by clopidogrel with a prior history of long-term safe administration. World J Cardiol 2(6):160–162

    Article  PubMed Central  PubMed  Google Scholar 

  37. Vanderschueren S, De Weerdt A, Malbrain M, Vankersschaever D, Frans E, Wilmer A, Bobbaers H (2000) Thrombocytopenia and prognosis in intensive care. Crit Care Med 28(6):1871–1876

    Article  CAS  PubMed  Google Scholar 

  38. Akca S, Haji-Michael P, de Mendonca A, Suter P, Levi M, Vincent JL (2002) Time course of platelet counts in critically ill patients. Crit Care Med 30(4):753–756

    Article  PubMed  Google Scholar 

  39. Crowther MA, Cook DJ, Meade MO, Griffith LE, Guyatt GH, Arnold DM, Rabbat CG, Geerts WH, Warkentin TE (2005) Thrombocytopenia in medical-surgical critically ill patients: prevalence, incidence, and risk factors. J Crit Care 20(4):348–353

    Article  PubMed  Google Scholar 

  40. Venkata C, Kashyap R, Farmer JC, Afessa B (2013) Thrombocytopenia in adult patients with sepsis: incidence, risk factors, and its association with clinical outcome. J Intensive Care 1(1):9

    Article  PubMed Central  PubMed  Google Scholar 

  41. Zhou H, Deng M, Liu Y, Yang C, Hoffman R, Zhou J, Loughran PA, Scott MJ, Neal MD, Billiar TR (2018) Platelet HMGB1 is required for efficient bacterial clearance in intra-abdominal bacterial sepsis in mice. Blood Adv 2(6):638–648

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  42. Yeaman MR (2014) Platelets: at the nexus of antimicrobial defence. Nat Rev Microbiol 12(6):426–437

    Article  CAS  PubMed  Google Scholar 

  43. Hamzeh-Cognasse H, Damien P, Chabert A, Pozzetto B, Cognasse F, Garraud O (2015) Platelets and infections - complex interactions with bacteria. Front Immunol 6:82

    Article  PubMed Central  PubMed  CAS  Google Scholar 

  44. Gaertner F, Ahmad Z, Rosenberger G, Fan S, Nicolai L, Busch B, Yavuz G, Luckner M, Ishikawa-Ankerhold H, Hennel R, Benechet A, Lorenz M, Chandraratne S, Schubert I, Helmer S, Striednig B, Stark K, Janko M, Bottcher RT, Verschoor A, Leon C, Gachet C, Gudermann T, Mederos YSM, Pincus Z, Iannacone M, Haas R, Wanner G, Lauber K, Sixt M, Massberg S (2017) Migrating platelets are mechano-scavengers that collect and bundle bacteria. Cell 171(6):1368–1382 e1323

    Article  CAS  PubMed  Google Scholar 

  45. Nauseef WM, Borregaard N (2014) Neutrophils at work. Nat Immunol 15(7):602–611

    Article  CAS  PubMed  Google Scholar 

  46. Minasyan H (2018) Phagocytosis and oxycytosis: two arms of human innate immunity. Immunol Res 66(2):271–280

    Article  CAS  PubMed  Google Scholar 

  47. Clifford EE, Parker K, Humphreys BD, Kertesy SB, Dubyak GR (1998) The P2X1 receptor, an adenosine triphosphate-gated cation channel, is expressed in human platelets but not in human blood leukocytes. Blood 91(9):3172–3181

    Article  CAS  PubMed  Google Scholar 

  48. Lecut C, Frederix K, Johnson DM, Deroanne C, Thiry M, Faccinetto C, Maree R, Evans RJ, Volders PG, Bours V, Oury C (2009) P2X1 ion channels promote neutrophil chemotaxis through rho kinase activation. J Immunol 183(4):2801–2809

    Article  CAS  PubMed  Google Scholar 

  49. Wang X, Qin W, Xu X, Xiong Y, Zhang Y, Zhang H, Sun B (2017) Endotoxin-induced autocrine ATP signaling inhibits neutrophil chemotaxis through enhancing myosin light chain phosphorylation. Proc Natl Acad Sci U S A 114(17):4483–4488

    Article  PubMed Central  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We would like to cordially thank Richard J. Evans for supplying the P2X1 breeding pairs and Helle Jakobsen for skilled technical support and Julia Wehmöller for help with correction of the manuscript.

Funding

The study was funded by the Independent Research Fund, Denmark (Danmarks Frie Forskningsfond): DFF-1331-00203A and the Lundbeck Foundation: R-192-2015-1362.

Author information

Author notes

  1. Marianne Skals and Anne-Sofie Greve contributed equally to this work.

Authors and Affiliations

  1. Department of Biomedicine, Aarhus University, Ole Worms Alle 4, build 1160, 8000, Aarhus C, Denmark

    Marianne Skals, Anne-Sofie Greve, Steen K. Fagerberg, Nanna Johnsen, Mette G. Christensen & Helle A. Praetorius

Authors
  1. Marianne Skals
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Anne-Sofie Greve
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Steen K. Fagerberg
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Nanna Johnsen
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Mette G. Christensen
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Helle A. Praetorius
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Marianne Skals.

Ethics declarations

Conflict of interest

Marianne Skals declares that she has no conflict of interest.

Anne-Sofie Greve declares that he has no conflict of interest.

Nanna Johnsen declares that she has no conflict of interest.

Mette G. Christensen declares that she has no conflict of interest.

Helle A. Praetorius declares that she has no conflict of interest.

Ethical approval

The experiments performed in this study were approved by the Danish ethic committee for animal research “Dyreforsøgstilsynet” (2014-15-0201-00316).

Additional information

Publisher’s note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Fig. S1

(PDF 45 kb)

Fig. S2

(PDF 40 kb)

Fig. S3

(PDF 45 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Skals, M., Greve, AS., Fagerberg, S.K. et al. P2X1 receptor blockers reduce the number of circulating thrombocytes and the overall survival of urosepsis with haemolysin-producing Escherichia coli. Purinergic Signalling 15, 265–276 (2019). https://doi.org/10.1007/s11302-019-09658-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11302-019-09658-1

Keywords

Search

Navigation