A2 adenosine receptor contributes to the development of cow’s milk protein allergy via regulating regulatory T cells

Document Type : Original Article


Department of Gastroenterology, Anhui Provincial Children’s Hospital (Children’s Hospital of Anhui Medical University), Wangjiang East Road No.39, Hefei, 230051, China


Objective(s): A2 adenosine receptor (A2AR) is a novel promising target for the treatment of inflammatory and allergic diseases. However, its role in the development of cow’s milk protein allergy (CMPA) has not been elucidated. The present study was designed to investigate the function of A2AR in CMPA development.
Materials and Methods: BALB/c mice were sensitized and challenged with ovalbumin (OVA) to induce allergic responses. The model was assessed by detecting allergic responses and plasma-specific IgE levels. The levels of A2AR were measured by PCR and flow cytometry. The subpopulation of Treg cells was analysed.
Results: The mice sensitized and challenged with OVA showed classic allergic symptoms, such as acute allergic skin responses, increased anaphylactic shock symptom scores, and higher levels of total IgE, OVA-specific IgE, IgG1 and IgG2a. OVA-sensitized mice and CMPA patients showed decreased levels of A2AR and Treg cells. Interestingly, we observed a positive correlation between A2AR expression and Treg levels in CMPA patients. Further study showed that the A2AR agonist CGS21680 blocked OVA-induced allergic reactions, and the A2AR antagonist KW-6002 amplified allergic responses. Interestingly, CGS21680 not only activated the A2AR-mediated signalling pathway but also caused an increase in the population of Treg cells. In contrast, KW-6002 therapy decreased the levels of Tregs in allergic mice.
Conclusion: A2AR expression is downregulated in CMPA. The A2AR-mediated pathway negatively regulates the development of CMPA, at least in part, by amplifying the differentiation of Tregs.


1. Kostadinova AI, Willemsen LE, Knippels LM, Garssen J. Immunotherapy-risk/benefit in food allergy. Pediatr Allergy Immunol 2013; 24:633-644.
2. Vandenplas Y. Prevention and management of cow’s milk allergy in non-exclusively breastfed infants. Nutrients 2017; 9:731.
3. Hochwallner H, Schulmeister U, Swoboda I, Spitzauer S, Valenta R. Cow’s milk allergy: From allergens to new forms of diagnosis, therapy and prevention. Methods 2014; 66:22-33.
4. Taniuchi S, Takahashi M, Soejima K, Hatano Y, Minami H. Immunotherapy for cow’s milk allergy. Hum Vaccin Immunother 2017; 13:2443-2451.
5. Borea PA, Gessi S, Merighi S, Vincenzi F, Varani K. Pharmacology of adenosine Receptors: The state of the art. Physiol Rev 2018;98:1591-1625.
6. Chen JF, Eltzschig HK, Fredholm BB. Adenosine receptors as drug targets--what are the challenges? Nat Rev Drug Discov 2013; 12:265-286.
7. Al-Attraqchi OHA, Attimarad M, Venugopala KN, Nair A, Al-Attraqchi NHA. Adenosine A2A receptor as a potential drug target - current status and future perspectives. Curr Pharm Des 2019; 25:2716-2740.
8. Nadeem A, Ponnoth DS, Ansari HR, Batchelor TP, Dey RD, Ledent C, et al. A2A adenosine receptor deficiency leads to impaired tracheal relaxation via NADPH oxidase pathway in allergic mice. J Pharmacol Exp Ther 2009; 330:99-108.
9. Caruso M, Varani K, Tringali G, Polosa R. Adenosine and adenosine receptors: their contribution to airway inflammation and therapeutic potential in asthma. Curr Med Chem 2009; 16:3875-3885.
10. Luyt D, Ball H, Makwana N, Green MR, Bravin K, Nasser SM, et al. Standards of care committee (socc) of the british society for allergy and clinical immunology (BSACI). BSACI guideline for the diagnosis and management of cow’s milk allergy. Clin Exp Allergy 2014; 44:642-672.
11. Lebetwa N, Suzuki Y, Tanaka S, Nakamura S, Katayama S. Enhanced anti-allergic activity of milk casein phosphopeptide by additional phosphorylation in ovalbumin-sensitized mice. Molecules 2019; 24:738-752.
12. van Sadelhoff JHJ, Hogenkamp A, Wiertsema SP, Harthoorn LF, Loonstra R, Hartog A, et al. A free amino acid-based diet partially prevents symptoms of cow’s milk allergy in mice after oral sensitization with whey. Immun Inflamm Dis 2020; 8:93-105.
13. Vonk MM, Blokhuis BRJ, Diks MAP, Wagenaar L, Smit JJ, Pieters RHH, et al. Butyrate enhances desensitization induced by oral immunotherapy in cow’s milk allergic mice. mediators inflamm 2019; 2019:9062537.
14. Song J, Xi JY, Yu WB, Yan C, Luo SS, Zhou L, et al. Inhibition of ROCK activity regulates the balance of Th1, Th17 and Treg cells in myasthenia gravis. Clin Immunol 2019; 203:142-153.
15. Masjedi A, Hassannia H, Atyabi F, Rastegari A, Hojjat-Farsangi M, Namdar A, et al. Downregulation of A2AR by siRNA loaded PEG-chitosan-lactate nanoparticles restores the T cell mediated anti-tumor responses through blockage of PKA/CREB signaling pathway. Int J Biol Macromol 2019; 133:436-445.
16.Kanagaratham C, Sallis BF, Fiebiger E. Experimental models for studying food allergy. Cell Mol Gastroenterol Hepatol 2018; 6:356-369.
17. Larsen JM, Bøgh KL. Animal models of allergen-specific immunotherapy in food allergy: Overview and opportunities. Clin Exp Allergy 2018; 48:1255-1274.
18. Cronstein BN, Sitkovsky M. Adenosine and adenosine receptors in the pathogenesis and treatment of rheumatic diseases. Nat Rev Rheumatol 2017; 13:41-51.
19. Huang S, Apasov S, Koshiba M, Sitkovsky M. Role of A2a extracellular adenosine receptor- mediated signaling in adenosine-mediated inhibition of T-cell activation and expansion. Blood 1997; 90:1600-1610.
20. Koshiba M, Kojima H, Huang S, Apasov S, Sitkovsky MV. Memory of extracellular adenosine A2A purinergic receptor-mediated signaling in murine T cells. J Biol Chem 1997; 272:25881-25889.
21. Lappas CM, Rieger JM, Linden J. A2A adenosine receptor induction inhibits IFN-gamma production in murine CD4+ T cells. J Immunol 2005; 174:1073-1080.
22. Versluis M, van den Berge M, Timens W, Luijk B, Rutgers B, Lammers JW, et al. Allergen inhalation decreases adenosine receptor expression in sputum and blood of asthma patients. Allergy 2008; 63:1186-1194.
23. Pei H, Linden J. Adenosine influences myeloid cells to inhibit aeroallergen sensitization. Am J Physiol Lung Cell Mol Physiol 2016;310: L985-992.
24. Palomares O, Yaman G, Azkur AK, Akkoc T, Akdis M, Akdis CA. Role of Treg in immune regulation of allergic diseases. Eur J Immunol 2010; 40:1232-1240.
25. Kerperien J, Veening-Griffioen D, Wehkamp T, van Esch BCAM, Hofman GA, Cornelissen P, et al. IL-10 receptor or TGF-β neutralization abrogates the protective effect of a specific nondigestible oligosaccharide mixture in cow-milk-allergic mice. J Nutr 2018; 148:1372-1379.
26. Hong JY, Kim M, Sol IS, Kim KW, Lee CM, Elias JA, et al. Chitotriosidase inhibits allergic asthmatic airways via regulation of TGF-β expression and Foxp3+ Treg cells. Allergy 2018; 73: 1686-1699.
27. Ren J, Zhao Y, Huang S, Lv D, Yang F, Lou L, et al. Immunomodulatory effect of bifidobacterium breve on experimental allergic rhinitis in BALB/c mice. Exp Ther Med 2018; 16:3996-4004.
28. Adel-Patient K, Wavrin S, Bernard H, Meziti N, Ah-Leung S, Wal JM. Oral tolerance and Treg cells are induced in BALB/c mice after gavage with bovine β-lactoglobulin. Allergy 2011; 66:1312-1321.
29. Schouten B, van Esch BC, Hofman GA, Boon L, Knippels LM, Willemsen LE, et al. Oligosaccharide-induced whey-specific CD25(+) regulatory T-cells are involved in the suppression of cow milk allergy in mice. J Nutr 2010; 140:835-841.
30. Wang L, Wan H, Tang W, Ni Y, Hou X, Pan L, et al. Critical roles of adenosine A2A receptor in regulating the balance of Treg/Th17 cells in allergic asthma. Clin Respir J 2018; 12:149-157.
31. Ohta A, Kini R, Ohta A, Subramanian M, Madasu M, Sitkovsky M. The development and immunosuppressive functions of CD4(+) CD25(+) FoxP3(+) regulatory T cells are under influence of the adenosine-A2A adenosine receptor pathway. Front Immunol 2012; 3:190-201.