Allergic sensitization increases the amount of extracellular ATP hydrolyzed by guinea pig leukocytes

  • Jaime ChávezEmail author
  • Mario H. Vargas
  • Jesús Martínez-Zúñiga
  • Ramcés Falfán-Valencia
  • Enrique Ambrocio-Ortiz
  • Verónica Carbajal
  • Rosa Sandoval-Roldán
Original Article


Increased levels of ATP have been found in the bronchoalveolar lavage of patients with asthma, and subjects with this disease, but not healthy subjects, develop bronchospasm after nebulization with ATP. Because the main mechanism for controlling the noxious effects of extracellular ATP is its enzymatic hydrolysis, we hypothesized that allergic sensitization is accompanied by a decreased functioning of such hydrolysis. In the present study, peripheral blood leukocytes from sensitized and non-sensitized guinea pigs were used for determining the extracellular metabolism (as assessed by inorganic phosphate production) of ATP, ADP, AMP, or adenosine, and for detecting possible changes in the expression (qPCR and Western blot) of major ectonucleotidases (NTPDase1, NTPDase3, and NPP1) and purinoceptors (P2X1, P2X7, P2Y4, and P2Y6). Contrary to our hypothesis, we found that leukocytes from allergic animals produced higher amounts of inorganic phosphate after stimulation with ATP and ADP, as compared with leukocytes from non-sensitized animals. Although at first glance, this result suggested that sensitization caused higher efficiency of ectonucleotidases, their mRNA and protein expressions were unaffected. On the other hand, after sensitization, we found a significant increase in the protein expression of P2X7 and P2Y4, two purinoceptors known to be responsible for ATP release after activation. We concluded that allergic sensitization increased the amount of ATP hydrolyzed by ectonucleotidases, the latter probably not due to the enhanced efficiency of its enzymatic breakdown, but rather due to an increased release of endogenous ATP or other nucleotides, partly mediated by enhanced expression or P2X7 and P2Y4 receptors.


Extracellular ATP ATP hydrolysis Ectonucleotidase Purinoceptors Allergic sensitization Guinea pig asthma model 


Compliance with ethical standards

Conflict of interest

Jaime Chávez declares that he has no conflict of interest.

Mario H. Vargas declares that he has no conflict of interest.

Jesús Martínez-Zúñiga declares that he has no conflict of interest.

Ramcés Falfán-Valencia declares that he has no conflict of interest.

Enrique Ambrocio-Ortiz declares that he has no conflict of interest.

Verónica Carbajal declares that she has no conflict of interest.

Rosa Sandoval-Roldán declares that she has no conflict of interest.

Ethical approval

Animal management was done according with the 2011 Guide for the Care and Use of Laboratory Animals. All procedures performed in guinea pigs were in accordance with the ethical standards of the institution, and the study protocol was approved by our institutional scientific and bioethics committees, with the approval number B23-12.

Supplementary material

11302_2019_9644_MOESM1_ESM.doc (34 kb)
ESM 1 (DOC 33 kb)
11302_2019_9644_MOESM2_ESM.pptt (53 kb)
ESM 2 (PPTT 52 kb)


  1. 1.
    Lipmann F (1941) Metabolic generation and utilization of phosphate bond energy. In: Advances in enzymology and related subjects, vol 1. Interscience Publishers, New York, pp 99–162Google Scholar
  2. 2.
    Abbracchio MP, Burnstock G (1994) Purinoceptors: are there families of P2X and P2Y purinoceptors? Pharmacol Ther 64(3):445–475CrossRefGoogle Scholar
  3. 3.
    Muller CE (2002) P2-pyrimidinergic receptors and their ligands. Curr Pharm Des 8(26):2353–2369CrossRefGoogle Scholar
  4. 4.
    North RA (2002) Molecular physiology of P2X receptors. Physiol Rev 82(4):1013–1067CrossRefGoogle Scholar
  5. 5.
    Atkinson B, Dwyer K, Enjyoji K, Robson SC (2006) Ecto-nucleotidases of the CD39/NTPDase family modulate platelet activation and thrombus formation: potential as therapeutic targets. Blood Cells Mol Dis 36:217–222CrossRefGoogle Scholar
  6. 6.
    Gachet C (2008) P2 receptors, platelet function and pharmacological implications. Thromb Haemost 99:466–472CrossRefGoogle Scholar
  7. 7.
    Chen Z, He L, Li L, Chen L (2018) The P2X7 purinergic receptor: An emerging therapeutic target in cardiovascular diseases. Clin Chim Acta 479:196–207. CrossRefGoogle Scholar
  8. 8.
    Castillo-Leon E, Dellepiane S, Fiorina P (2018) ATP and T-cell-mediated rejection. Curr Opin Organ Transplant 23(1):34–43. CrossRefGoogle Scholar
  9. 9.
    Polosa R (2002) Adenosine-receptor subtypes: their relevance to adenosine-mediated responses in asthma and chronic obstructive pulmonary disease. Eur Respir J 20:488–496CrossRefGoogle Scholar
  10. 10.
    Chávez J, Vargas MH, Rebollar-Ayala DC, Díaz-Hernández V, Cruz-Valderrama JE, Flores-Soto E, Flores-García M, Jiménez-Vargas NN, Barajas-López C, Montaño LM (2013) Inhibition of extracellular nucleotides hydrolysis intensifies the allergic bronchospasm. A novel protective role of ectonucleotidases. Allergy 68(4):462–471. CrossRefGoogle Scholar
  11. 11.
    Pelleg A, Schulman ES, Barnes PJ (2016) Extracellular adenosine 5′-triphosphate in obstructive airway diseases. Chest 150(4):908–915. CrossRefGoogle Scholar
  12. 12.
    Katchanov G, Xu J, Schulman ES, Pelleg A (1998) ATP causes neurogenic bronchoconstriction in the dog. Drug Dev Res 45(3–4):342–349CrossRefGoogle Scholar
  13. 13.
    Schulman ES, Glaum MC, Post T, Wang Y, Raible DG, Mohanty J, Butterfield JH, Pelleg A (1999) ATP modulates anti-IgE-induced release of histamine from human lung mast cells. Am J Respir Cell Mol Biol 20:530–537CrossRefGoogle Scholar
  14. 14.
    Idzko M, Hammad H, van Nimwegen M, Kool M, Willart MA, Muskens F, Hoogsteden HC, Luttmann W, Ferrari D, Di Virgilio F, Virchow JCJ, Lambrecht BN (2007) Extracellular ATP triggers and maintains asthmatic airway inflammation by activating dendritic cells. Nat Med 13:913–919CrossRefGoogle Scholar
  15. 15.
    Pellegrino R, Wilson O, Jenouri G, Rodarte JR (1996) Lung mechanics during induced bronchoconstriction. J Appl Physiol 81:964–975CrossRefGoogle Scholar
  16. 16.
    Basoglu OK, Pelleg A, Kharitonov SA, Barnes PJ (2017) Contrasting effects of ATP and adenosine on capsaicin challenge in asthmatic patients. Pulm Pharmacol Ther 45:13–18. CrossRefGoogle Scholar
  17. 17.
    Zhang F, Su X, Huang G, Xin XF, Cao EH, Shi Y, Song Y (2017) Adenosine triphosphate promotes allergen-induced airway inflammation and Th17 cell polarization in neutrophilic asthma. J Immunol Res 2017:5358647. Google Scholar
  18. 18.
    Zimmermann H (2001) Ectonucleotidases: some recent developments and note on nomenclature. Drug Dev Res 52(1–2):44–56CrossRefGoogle Scholar
  19. 19.
    Robson SC, Sévigny J, Zimmermann H (2006) The E-NTPDase family of ectonucleotidases: structure function relationships and pathophysiological significance. Purinergic Signalling 2:409–430CrossRefGoogle Scholar
  20. 20.
    Montaño LM, Vargas MH, Díaz-Hernández V, De Ita M, Kazakova R, Barajas-López C (2016) Decreased expression of ectonucleotidase E-NPP1 in leukocytes from subjects with severe asthma exacerbation. Allergy 71(1):124–128. CrossRefGoogle Scholar
  21. 21.
    Picher M (2011) Mechanisms regulating airway nucleotides. In: Picher M, Boucher RC (eds) Purinergic regulation of respiratory diseases. Springer, LondonCrossRefGoogle Scholar
  22. 22.
    Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCT method. Methods 25(4):402–408CrossRefGoogle Scholar
  23. 23.
    De Ita M, Vargas MH, Carbajal V, Ortiz-Quintero B, López-López C, Miranda-Morales M, Barajas-López C, Montaño LM (2016) ATP releases ATP or other nucleotides from human peripheral blood leukocytes through purinergic P2 receptors. Life Sci 145:85–92. CrossRefGoogle Scholar
  24. 24.
    Esquerré T, Moisan A, Chiapello H, Arike L, Vilu R, Gaspin C, Cocaign-Bousquet M, Girbal L (2015) Genome-wide investigation of mRNA lifetime determinants in Escherichia coli cells cultured at different growth rates. BMC Genomics 16:275CrossRefGoogle Scholar
  25. 25.
    Greenbaum D, Colangelo C, Williams K, Gerstein M (2003) Comparing protein abundance and mRNA expression levels on a genomic scale. Genome Biol 4:117CrossRefGoogle Scholar
  26. 26.
    Nie L, Wu G, Zhang W (2006) Correlation between mRNA and protein abundance in Desulfovibrio vulgaris: a multiple regression to identify sources of variations. Biochem Biophys Res Commun 339:603–610CrossRefGoogle Scholar
  27. 27.
    Suadicani SO, Brosnan CF, Scemes E (2006) P2X7 receptors mediate ATP release and amplification of astrocytic intercellular Ca2+ signaling. J Neurosci 26(5):1378–1385CrossRefGoogle Scholar
  28. 28.
    Bogdanov YD, Wildman SS, Clements MP, King BF, Burnstock G (1998) Molecular cloning and characterization of rat P2Y4 nucleotide receptor. Br J Pharmacol 124(3):428–430CrossRefGoogle Scholar
  29. 29.
    Manthei DM, Jackson DJ, Evans MD, Gangnon RE, Tisler CJ, Gern JE, Lemanske RFJ, Denlinger LC (2012) Protection from asthma in a high-risk birth cohort by attenuated P2X7 function. J Allergy Clin Immunol 130(2):496–502. CrossRefGoogle Scholar
  30. 30.
    Migita K, Lu L, Zhao Y, Honda K, Iwamoto T, Kita S, Katsuragi T (2005) Adenosine induces ATP release via an inositol 1,4,5-trisphosphate signaling pathway in MDCK cells. Biochem Biophys Res Commun 328(4):1211–1215CrossRefGoogle Scholar
  31. 31.
    Migita K, Zhao Y, Katsuragi T (2007) Mitochondria play an important role in adenosine-induced ATP release from Madin-Darby canine kidney cells. Biochem Pharmacol 73(10):1676–1682CrossRefGoogle Scholar
  32. 32.
    Graven P, Tambalo M, Scapozza L, Perozzo R (2014) Purine metabolite and energy charge analysis of Trypanosoma brucei cells in different growth phases using an optimized ion-pair RP-HPLC/UV for the quantification of adenine and guanine pools. Exp Parasitol 141:28–38. CrossRefGoogle Scholar
  33. 33.
    Yegutkin GG (2008) Nucleotide- and nucleoside-converting ectoenzymes: important modulators of purinergic signalling cascade. Biochim Biophys Acta 1783(5):673–694. CrossRefGoogle Scholar
  34. 34.
    Praetorius HA, Leipziger J (2009) ATP release from non-excitable cells. Purinergic Signal 5(4):433–446. CrossRefGoogle Scholar
  35. 35.
    Pillai P, Chan YC, Wu SY, Ohm-Laursen L, Thomas C, Durham SR, Menzies-Gow A, Rajakulasingam RK, Ying S, Gould HJ, Corrigan CJ (2016) Omalizumab reduces bronchial mucosal IgE and improves lung function in non-atopic asthma. Eur Respir J 48(6):1593–1601. CrossRefGoogle Scholar
  36. 36.
    World Health Organization (2017) Asthma; Fact sheet (updated April 2017)Google Scholar

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  • Jaime Chávez
    • 1
    Email author return OK on get
  • Mario H. Vargas
    • 1
  • Jesús Martínez-Zúñiga
    • 1
  • Ramcés Falfán-Valencia
    • 2
  • Enrique Ambrocio-Ortiz
    • 2
  • Verónica Carbajal
    • 1
  • Rosa Sandoval-Roldán
    • 1
  1. 1.Departamento de Investigación en Hiperreactividad BronquialInstituto Nacional de Enfermedades Respiratorias Ismael Cosío VillegasMexico CityMexico
  2. 2.Laboratorio HLAInstituto Nacional de Enfermedades Respiratorias Ismael Cosío VillegasMexico CityMexico

Personalised recommendations