Adenosinergic System Involvement in Ischemic Stroke Patients’ Lymphocytes
Abstract
:1. Introduction
2. Materials and Methods
2.1. Ischemic Stroke Patients and Healthy Subjects
2.2. Sample Collection and Human Lymphocyte Preparation
2.3. Real-Time Quantitative Polymerase Chain Reaction Assays
2.4. Saturation Binding Assays to A1, A2A, A2B, and A3ARs
2.5. Flow Cytometry Analysis
2.6. Statistical Analysis
3. Results
3.1. A2AAR mRNA Expression is Up-Regulated in Lymphocytes of Ischemic Stroke Patients
3.2. A2AAR Increase in Density and Affinity in Lymphocytes of Ischemic Stroke Patients
3.3. CD4 and CD8 Expression on Lymphocytes of Ischemic Stroke Patients
3.4. CD73 and CD39 Expression on Lymphocytes of Ischemic Stroke Patients
4. Discussion
5. Conclusions
Author Contributions
Funding
Conflicts of Interest
References
- Borea, P.A.; Varani, K.; Vincenzi, F.; Baraldi, P.G.; Tabrizi, M.A.; Merighi, S.; Gessi, S. The A3 adenosine receptor: History and perspectives. Pharmacol. Rev. 2015, 67, 74–102. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Borea, P.A.; Gessi, S.; Merighi, S.; Varani, K. Adenosine as a Multi-Signalling Guardian Angel in Human Diseases: When, Where and How Does it Exert its Protective Effects? Trends Pharmacol. Sci. 2016, 37, 419–434. [Google Scholar] [CrossRef] [PubMed]
- Borea, P.A.; Gessi, S.; Merighi, S.; Vincenzi, F.; Varani, K. Pathological overproduction: The bad side of adenosine. Br. J. Pharmacol. 2017, 174, 1945–1960. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Borea, P.A.; Gessi, S.; Merighi, S.; Vincenzi, F.; Varani, K. Pharmacology of Adenosine Receptors: The State of the Art. Physiol. Rev. 2018, 98, 1591–1625. [Google Scholar] [CrossRef] [PubMed]
- Yang, Q.; Huang, Q.; Hu, Z.; Tang, X. Potential Neuroprotective Treatment of Stroke: Targeting Excitotoxicity, Oxidative Stress, and Inflammation. Front. Neurosci. 2019, 13, 1036. [Google Scholar] [CrossRef]
- Khanna, S.; Briggs, Z.; Rink, C. Inducible Glutamate Oxaloacetate Transaminase as a Therapeutic Target Against Ischemic Stroke. Antioxid Redox Signal. 2015, 22, 175–186. [Google Scholar] [CrossRef] [Green Version]
- Pedata, F.; Dettori, I.; Coppi, E.; Melani, A.; Fusco, I.; Corradetti, R.; Pugliese, A.M. Purinergic signalling in brain ischemia. Neuropharmacology 2016, 104, 105–130. [Google Scholar] [CrossRef]
- Melani, A.; Corti, F.; Stephan, H.; Müller, C.E.; Donati, C.; Bruni, P.; Vannucchi, M.G.; Pedata, F. Ecto-ATPase inhibition: ATP and adenosine release under physiological and ischemic in vivo conditions in the rat striatum. Exp. Neurol. 2012, 233, 193–204. [Google Scholar] [CrossRef]
- Burnstock, G. An introduction to the roles of purinergic signalling in neurodegeneration, neuroprotection and neuroregeneration. Neuropharmacology 2016, 104, 4–17. [Google Scholar] [CrossRef]
- Burnstock, G. Purinergic Signalling and Neurological Diseases: An Update. CNS Neurol. Disord. Drug Targets 2017, 16, 257–265. [Google Scholar] [CrossRef]
- Varani, K.; Abbracchio, M.P.; Cannella, M.; Cislaghi, G.; Giallonardo, P.; Mariotti, C.; Cattabriga, E.; Cattabeni, F.; Borea, P.A.; Squitieri, F.; et al. Aberrant A2A receptor function in peripheral blood cells in Huntington’s disease. FASEB J. 2003, 17, 2148–2150. [Google Scholar] [CrossRef] [PubMed]
- Varani, K.; Vincenzi, F.; Tosi, A.; Gessi, S.; Casetta, I.; Granieri, G.; Fazio, P.; Leung, E.; MacLennan, S.; Granieri, E.; et al. A2A adenosine receptor overexpression and functionality, as well as TNF-alpha levels, correlate with motor symptoms in Parkinson’s disease. FASEB J. 2010, 24, 587–598. [Google Scholar] [CrossRef] [PubMed]
- Yamaguchi, D.; Terayama, R.; Omura, S.; Tsuchiya, H.; Sato, T.; Ichikawa, H.; Sugimoto, T. Effect of adenosine A1 receptor agonist on the enhanced excitability of spinal dorsal horn neurons after peripheral nerve injury. Int. J. Neurosci. 2014, 124, 213–222. [Google Scholar] [CrossRef] [PubMed]
- Melani, A.; Pugliese, A.M.; Pedata, F. Chapter Thirteen-Adenosine Receptors in Cerebral Ischemia. In International Review of Neurobiology; Mori, A., Ed.; Adenosine Receptors in Neurology and Psychiatry; Academic Press: Cambridge, MA, USA, 2014; Volume 119, pp. 309–348. [Google Scholar]
- Pedata, F.; Pugliese, A.M.; Coppi, E.; Dettori, I.; Maraula, G.; Cellai, L.; Melani, A. Adenosine A2A receptors modulate acute injury and neuroinflammation in brain ischemia. Mediat. Inflamm. 2014, 2014, 805198. [Google Scholar] [CrossRef] [Green Version]
- Melani, A.; Corti, F.; Cellai, L.; Vannucchi, M.G.; Pedata, F. Low doses of the selective adenosine A2A receptor agonist CGS21680 are protective in a rat model of transient cerebral ischemia. Brain Res. 2014, 1551, 59–72. [Google Scholar] [CrossRef]
- Antonioli, L.; Csóka, B.; Fornai, M.; Colucci, R.; Kókai, E.; Blandizzi, C.; Haskó, G. Adenosine and inflammation: what’s new on the horizon? Drug Discov. Today 2014, 19, 1051–1068. [Google Scholar] [CrossRef]
- Trincavelli, M.L.; Melani, A.; Guidi, S.; Cuboni, S.; Cipriani, S.; Pedata, F.; Martini, C. Regulation of A(2A) adenosine receptor expression and functioning following permanent focal ischemia in rat brain. J. Neurochem. 2008, 104, 479–490. [Google Scholar]
- Gessi, S.; Varani, K.; Merighi, S.; Ongini, E.; Borea, P.A. A2A adenosine receptors in human peripheral blood cells. Br. J. Pharmacol. 2000, 129, 2–11. [Google Scholar] [CrossRef] [Green Version]
- Varani, K.; Portaluppi, F.; Gessi, S.; Merighi, S.; Vincenzi, F.; Cattabriga, E.; Dalpiaz, A.; Bortolotti, F.; Belardinelli, L.; Borea, P.A. Caffeine intake induces an alteration in human neutrophil A2A adenosine receptors. Cell. Mol. Life Sci. 2005, 62, 2350–2358. [Google Scholar] [CrossRef]
- Liu, Y.-J.; Chen, J.; Li, X.; Zhou, X.; Hu, Y.-M.; Chu, S.-F.; Peng, Y.; Chen, N.-H. Research progress on adenosine in central nervous system diseases. CNS Neurosci. Ther. 2019, 25, 899–910. [Google Scholar] [CrossRef]
- Ham, J.; Rees, D.A. The adenosine a2b receptor: Its role in inflammation. Endocr. Metab. Immune. Disord. Drug Targets 2008, 8, 244–254. [Google Scholar] [CrossRef] [PubMed]
- Borea, P.A.; Gessi, S.; Bar-Yehuda, S.; Fishman, P. A3 adenosine receptor: Pharmacology and role in disease. Handb. Exp. Pharmacol. 2009, 297–327. [Google Scholar]
- Petrovic-Djergovic, D.; Hyman, M.C.; Ray, J.J.; Bouis, D.; Visovatti, S.H.; Hayasaki, T.; Pinsky, D.J. Tissue-Resident Ecto-5′ Nucleotidase (CD73) Regulates Leukocyte Trafficking in the Ischemic Brain. J. Immunol. 2012, 188, 2387–2398. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Antonioli, L.; Pacher, P.; Vizi, E.S.; Haskó, G. CD39 and CD73 in immunity and inflammation. Trends Mol. Med. 2013, 19, 355–367. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Synnestvedt, K.; Furuta, G.T.; Comerford, K.M.; Louis, N.; Karhausen, J.; Eltzschig, H.K.; Hansen, K.R.; Thompson, L.F.; Colgan, S.P. Ecto-5′-nucleotidase (CD73) regulation by hypoxia-inducible factor-1 mediates permeability changes in intestinal epithelia. J. Clin. Invest. 2002, 110, 993–1002. [Google Scholar] [CrossRef]
- Kiss, J.; Yegutkin, G.G.; Koskinen, K.; Savunen, T.; Jalkanen, S.; Salmi, M. IFN-beta protects from vascular leakage via up-regulation of CD73. Eur. J. Immunol. 2007, 37, 3334–3338. [Google Scholar] [CrossRef]
- Regateiro, F.S.; Howie, D.; Nolan, K.F.; Agorogiannis, E.I.; Greaves, D.R.; Cobbold, S.P.; Waldmann, H. Generation of anti-inflammatory adenosine by leukocytes is regulated by TGF-β. Eur. J. Immunol. 2011, 41, 2955–2965. [Google Scholar] [CrossRef]
- Narravula, S.; Lennon, P.F.; Mueller, B.U.; Colgan, S.P. Regulation of endothelial CD73 by adenosine: Paracrine pathway for enhanced endothelial barrier function. J. Immunol. 2000, 165, 5262–5268. [Google Scholar] [CrossRef] [Green Version]
- Liang, D.; Zuo, A.; Zhao, R.; Shao, H.; Born, W.K.; O’Brien, R.L.; Kaplan, H.J.; Sun, D. CD73 Expressed on γδ T Cells Shapes Their Regulatory Effect in Experimental Autoimmune Uveitis. PLoS ONE 2016, 11, e0150078. [Google Scholar] [CrossRef] [Green Version]
- Blume, C.; Felix, A.; Shushakova, N.; Gueler, F.; Falk, C.S.; Haller, H.; Schrader, J. Autoimmunity in CD73/Ecto-5′-nucleotidase deficient mice induces renal injury. PLoS ONE 2012, 7, e37100. [Google Scholar] [CrossRef] [Green Version]
- Sturm, J.W.; Donnan, G.A.; Dewey, H.M.; Macdonell, R.A.L.; Gilligan, A.K.; Srikanth, V.; Thrift, A.G. Quality of life after stroke: The North East Melbourne Stroke Incidence Study (NEMESIS). Stroke 2004, 35, 2340–2345. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Varani, K.; Bachoud-Lévi, A.-C.; Mariotti, C.; Tarditi, A.; Abbracchio, M.P.; Gasperi, V.; Borea, P.A.; Dolbeau, G.; Gellera, C.; Solari, A.; et al. Biological abnormalities of peripheral A(2A) receptors in a large representation of polyglutamine disorders and Huntington’s disease stages. Neurobiol. Dis. 2007, 27, 36–43. [Google Scholar] [CrossRef] [PubMed]
- Mizuno, Y.; Hasegawa, K.; Kondo, T.; Kuno, S.; Yamamoto, M.; Japanese Istradefylline Study Group. Clinical efficacy of istradefylline (KW-6002) in Parkinson’s disease: A randomized, controlled study. Mov. Disord. 2010, 25, 1437–1443. [Google Scholar] [CrossRef] [PubMed]
- Vincenzi, F.; Corciulo, C.; Targa, M.; Casetta, I.; Gentile, M.; Granieri, E.; Borea, P.A.; Popoli, P.; Varani, K. A2A adenosine receptors are up-regulated in lymphocytes from amyotrophic lateral sclerosis patients. Amyotroph. Lateral Scler. Frontotemporal Degener. 2013, 14, 406–413. [Google Scholar] [CrossRef]
- Gomes, C.; Ferreira, R.; George, J.; Sanches, R.; Rodrigues, D.I.; Gonçalves, N.; Cunha, R.A. Activation of microglial cells triggers a release of brain-derived neurotrophic factor (BDNF) inducing their proliferation in an adenosine A2A receptor-dependent manner: A2A receptor blockade prevents BDNF release and proliferation of microglia. J. Neuroinflamm. 2013, 10, 16. [Google Scholar] [CrossRef] [Green Version]
- Varani, K.; Padovan, M.; Vincenzi, F.; Targa, M.; Trotta, F.; Govoni, M.; Borea, P.A. A2A and A3 adenosine receptor expression in rheumatoid arthritis: Upregulation, inverse correlation with disease activity score and suppression of inflammatory cytokine and metalloproteinase release. Arthritis Res. Ther. 2011, 13, R197. [Google Scholar] [CrossRef] [Green Version]
- Vincenzi, F.; Corciulo, C.; Targa, M.; Merighi, S.; Gessi, S.; Casetta, I.; Gentile, M.; Granieri, E.; Borea, P.A.; Varani, K. Multiple sclerosis lymphocytes upregulate A2A adenosine receptors that are antiinflammatory when stimulated. Eur. J. Immunol. 2013, 43, 2206–2216. [Google Scholar] [CrossRef]
- Bortoluzzi, A.; Vincenzi, F.; Govoni, M.; Padovan, M.; Ravani, A.; Borea, P.A.; Varani, K. A2A adenosine receptor upregulation correlates with disease activity in patients with systemic lupus erythematosus. Arthritis Res. Ther. 2016, 18, 192. [Google Scholar] [CrossRef] [Green Version]
- Ravani, A.; Vincenzi, F.; Bortoluzzi, A.; Padovan, M.; Pasquini, S.; Gessi, S.; Merighi, S.; Borea, P.A.; Govoni, M.; Varani, K. Role and Function of A2A and A₃ Adenosine Receptors in Patients with Ankylosing Spondylitis, Psoriatic Arthritis and Rheumatoid Arthritis. Int. J. Mol. Sci. 2017, 18. [Google Scholar] [CrossRef]
- Linden, J. Molecular Approach to Adenosine Receptors: Receptor-Mediated Mechanisms of Tissue Protection. Annu. Rev. Pharmacol. Toxicol. 2001, 41, 775–787. [Google Scholar] [CrossRef]
- Thompson, L.F.; Eltzschig, H.K.; Ibla, J.C.; Van De Wiele, C.J.; Resta, R.; Morote-Garcia, J.C.; Colgan, S.P. Crucial role for ecto-5′-nucleotidase (CD73) in vascular leakage during hypoxia. J. Exp. Med. 2004, 200, 1395–1405. [Google Scholar] [CrossRef] [PubMed]
- Eltzschig, H.K.; Thompson, L.F.; Karhausen, J.; Cotta, R.J.; Ibla, J.C.; Robson, S.C.; Colgan, S.P. Endogenous adenosine produced during hypoxia attenuates neutrophil accumulation: Coordination by extracellular nucleotide metabolism. Blood 2004, 104, 3986–3992. [Google Scholar] [CrossRef]
- Deaglio, S.; Dwyer, K.M.; Gao, W.; Friedman, D.; Usheva, A.; Erat, A.; Chen, J.-F.; Enjyoji, K.; Linden, J.; Oukka, M.; et al. Adenosine generation catalyzed by CD39 and CD73 expressed on regulatory T cells mediates immune suppression. J. Exp. Med. 2007, 204, 1257–1265. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Eltzschig, H.K.; Ibla, J.C.; Furuta, G.T.; Leonard, M.O.; Jacobson, K.A.; Enjyoji, K.; Robson, S.C.; Colgan, S.P. Coordinated adenine nucleotide phosphohydrolysis and nucleoside signaling in posthypoxic endothelium: Role of ectonucleotidases and adenosine A2B receptors. J. Exp. Med. 2003, 198, 783–796. [Google Scholar] [CrossRef] [PubMed] [Green Version]
- Sitkovsky, M.V. Use of the A(2A) adenosine receptor as a physiological immunosuppressor and to engineer inflammation in vivo. Biochem. Pharmacol. 2003, 65, 493–501. [Google Scholar] [CrossRef]
- Botta Gordon-Smith, S.; Ursu, S.; Eaton, S.; Moncrieffe, H.; Wedderburn, L.R. Correlation of low CD73 expression on synovial lymphocytes with reduced adenosine generation and higher disease severity in juvenile idiopathic arthritis. Arthritis Rheumatol. 2015, 67, 545–554. [Google Scholar] [CrossRef] [PubMed]
- Laghi Pasini, F.; Guideri, F.; Picano, E.; Parenti, G.; Petersen, C.; Varga, A.; Di Perri, T. Increase in plasma adenosine during brain ischemia in man: A study during transient ischemic attacks, and stroke. Brain Res. Bull. 2000, 51, 327–330. [Google Scholar] [CrossRef]
Ischemic Stroke Patients | n = 71 |
Demographic features | |
N° female/male | 35/36 |
Age | 78.31 ± 13.13 |
Hypertension | 48 |
Diabetes | 12 |
Smoke | 10 |
Atrial fibrillation | 26 |
OCSP patients classification | |
TACI | 19 |
PACI | 36 |
POCI | 8 |
LACI | 8 |
NIHSS | |
Slight impairment (1–5) | 26 |
Moderate impairment (6–14) | 26 |
Severe impairment (15–25) | 19 |
Very severe impairment (>25) | 0 |
TOAST Classification | |
LAA | 20 |
CE | 34 |
SVA | 7 |
Other | 1 |
Undetermined | 9 |
Healthy Subjects | n = 70 |
Demographic features | |
N° female/male | 31/39 |
Age | 61.24 ± 12.17 |
A1ARs | A2AARs | A2BARs | A3ARs | |
---|---|---|---|---|
Healthy subjects (n = 50) | ||||
KD (nM) | 1.74 ± 0.11 | 1.48 ± 0.10 | 2.11 ± 0.20 | 1.87 ± 0.14 |
Bmax (fmol/mg protein) | 52 ± 4 | 63 ± 5 | 53 ± 5 | 76 ± 8 |
Ischemic stroke patients (n = 50) | ||||
KD (nM) | 1.58 ± 0.12 | 0.97 ± 0.04** | 2.34 ± 0.16 | 1.91 ± 0.11 |
Bmax (fmol/mg protein) | 45 ± 3 | 168 ± 14** | 58 ± 6 | 87 ± 9 |
CD39 (%) | CD73 (%) | |||
---|---|---|---|---|
Healthy Subjects | Ischemic Stroke Patients | Healthy Subjects | Ischemic Stroke Patients | |
All lymphocytes | 14.82 ± 0.66 | 16.22 ± 1.16 | 10.04 ± 1.10 | 6.98 ± 0.77* |
CD4+ lymphocytes | 8.56 ± 0.74 | 11.35 ± 1.29 | 2.96 ± 0.45 | 1.47 ± 1.10* |
CD8+ lymphocytes | 4.09 ± 0.53 | 6.50 ± 0.90 | 21.04 ± 2.54 | 10.54 ± 1.48** |
CD39 (MFI) | CD73 (MFI) | |||
---|---|---|---|---|
Healthy Subjects | Ischemic Stroke Patients | Healthy Subjects | Ischemic Stroke Patients | |
All lymphocytes | 5389 ± 515 | 4502 ± 416 | 10084 ± 588 | 11436 ± 986 |
CD4+ lymphocytes | 4783 ± 258 | 4715 ± 380 | 6983 ± 234 | 7058 ± 396 |
CD8+ lymphocytes | 2640 ± 208 | 2357 ± 212 | 5197 ± 246 | 4239 ± 184** |
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Pasquini, S.; Vincenzi, F.; Casetta, I.; Laudisi, M.; Merighi, S.; Gessi, S.; Borea, P.A.; Varani, K. Adenosinergic System Involvement in Ischemic Stroke Patients’ Lymphocytes. Cells 2020, 9, 1072. https://doi.org/10.3390/cells9051072
Pasquini S, Vincenzi F, Casetta I, Laudisi M, Merighi S, Gessi S, Borea PA, Varani K. Adenosinergic System Involvement in Ischemic Stroke Patients’ Lymphocytes. Cells. 2020; 9(5):1072. https://doi.org/10.3390/cells9051072
Chicago/Turabian StylePasquini, Silvia, Fabrizio Vincenzi, Ilaria Casetta, Michele Laudisi, Stefania Merighi, Stefania Gessi, Pier Andrea Borea, and Katia Varani. 2020. "Adenosinergic System Involvement in Ischemic Stroke Patients’ Lymphocytes" Cells 9, no. 5: 1072. https://doi.org/10.3390/cells9051072
APA StylePasquini, S., Vincenzi, F., Casetta, I., Laudisi, M., Merighi, S., Gessi, S., Borea, P. A., & Varani, K. (2020). Adenosinergic System Involvement in Ischemic Stroke Patients’ Lymphocytes. Cells, 9(5), 1072. https://doi.org/10.3390/cells9051072