Background The ectoenzymes CD39 and CD73 are expressed by a broad range of immune cells and promote the extracellular degradation of nucleotides to anti-inflammatory adenosine. This study explored the abundance of CD73 and CD39 on circulating and resident cardiac leukocytes and coronary endothelial cells under control conditions and in response to inflammation following myocardial ischemia and reperfusion (I/R). Methods and Results A method was elaborated to permit FACS analysis of non-myocardial cells (resident leukocytes, coronary endothelium and CD31？ CD45？ cells) of the unstressed heart. Under control conditions the murine heart contained 2.3×103 resident leukocytes/mg tissue, the most prominent fraction being antigen-presenting mononuclear cells (CD11b+ CD11c+ F4/80+ MHCII+) followed by B-cells, monocytes and T-cells. CD73 was highly expressed on circulating and resident cardiac lymphoid cells with little expression on myeloid cells, while the opposite was true for CD39. Cardiomyocytes and erythrocytes do not measurably express CD39/CD73 and CD39 dominates on coronary endothelium. Three days after I/R, CD73 was significantly upregulated on invading granulocytes (2.8-fold) and T-cells (1.5-fold). Compared with coronary endothelial cells, CD73 associated with leukocytes comprised 2/3 of the total cardiac CD73. Conclusion Our study suggests that extracellular ATP formed during I/R is preferentially degraded by CD39 present on myeloid cells, while the formation of immunosuppressive adenosine is mainly catalysed by CD73 present on granulocytes and lymphoid cells. Upregulated CD73 on granulocytes and T-cells infiltrating the injured heart is consistent with the existence of an autocrine adenosinergic loop which may promote the healing process.
References
[1]
Rock KL, Latz E, Ontiveros F, Kono H (2010) The sterile inflammatory response. Annu Rev Immunol 28: 321–342.
[2]
Frangogiannis NG (2008) The immune system and cardiac repair. Pharmacol Res 58: 88–111.
[3]
Junger WG (2011) Immune cell regulation by autocrine purinergic signalling. Nat Rev Immunol 11: 201–212.
[4]
Nahrendorf M, Pittet MJ, Swirski FK (2010) Monocytes: protagonists of infarct inflammation and repair after myocardial infarction. Circulation 121: 2437–2445.
[5]
van den Borne SW, Diez J, Blankesteijn WM, Verjans J, Hofstra L, et al. (2010) Myocardial remodeling after infarction: the role of myofibroblasts. Nat Rev Cardiol 7: 30–37.
[6]
Galkina E, Kadl A, Sanders J, Varughese D, Sarembock IJ, et al. (2006) Lymphocyte recruitment into the aortic wall before and during development of atherosclerosis is partially L-selectin dependent. J Exp Med 203: 1273–1282.
[7]
Bulloch K, Miller MM, Gal-Toth J, Milner TA, Gottfried-Blackmore A, et al. (2008) CD11c/EYFP transgene illuminates a discrete network of dendritic cells within the embryonic, neonatal, adult, and injured mouse brain. J Comp Neurol 508: 687–710.
[8]
Ochoa MT, Loncaric A, Krutzik SR, Becker TC, Modlin RL (2008) “Dermal dendritic cells" comprise two distinct populations: CD1+ dendritic cells and CD209+ macrophages. J Invest Dermatol 128: 2225–2231.
[9]
Devey L, Ferenbach D, Mohr E, Sangster K, Bellamy CO, et al. (2009) Tissue-resident macrophages protect the liver from ischemia reperfusion injury via a heme oxygenase-1-dependent mechanism. Mol Ther 17: 65–72.
Koszalka P, Ozuyaman B, Huo Y, Zernecke A, Flogel U, et al. (2004) Targeted disruption of cd73/ecto-5′-nucleotidase alters thromboregulation and augments vascular inflammatory response. Circ Res 95: 814–821.
[12]
Eltzschig HK, Thompson LF, Karhausen J, Cotta RJ, Ibla JC, et al. (2004) Endogenous adenosine produced during hypoxia attenuates neutrophil accumulation: coordination by extracellular nucleotide metabolism. Blood 104: 3986–3992.
[13]
Yegutkin GG (2008) Nucleotide- and nucleoside-converting ectoenzymes: Important modulators of purinergic signalling cascade. Biochim Biophys Acta 1783: 673–694.
[14]
Eltzschig HK, Eckle T, Mager A, Kuper N, Karcher C, et al. (2006) ATP release from activated neutrophils occurs via connexin 43 and modulates adenosine-dependent endothelial cell function. Circ Res 99: 1100–1108.
[15]
Schenk U, Westendorf AM, Radaelli E, Casati A, Ferro M, et al. (2008) Purinergic control of T cell activation by ATP released through pannexin-1 hemichannels. Sci Signal 1: ra6.
[16]
Praetorius HA, Leipziger J (2009) ATP release from non-excitable cells. Purinergic Signal 5: 433–446.
[17]
Pellegatti P, Falzoni S, Pinton P, Rizzuto R, Di VF (2005) A novel recombinant plasma membrane-targeted luciferase reveals a new pathway for ATP secretion. Mol Biol Cell 16: 3659–3665.
[18]
Hasko G, Linden J, Cronstein B, Pacher P (2008) Adenosine receptors: therapeutic aspects for inflammatory and immune diseases. Nat Rev Drug Discov 7: 759–770.
[19]
Fredholm BB, Chern Y, Franco R, Sitkovsky M (2007) Aspects of the general biology of adenosine A2A signaling. Prog Neurobiol 83: 263–276.
[20]
Bours MJ, Swennen EL, Di VF, Cronstein BN, Dagnelie PC (2006) Adenosine 5′-triphosphate and adenosine as endogenous signaling molecules in immunity and inflammation. Pharmacol Ther 112: 358–404.
[21]
Nahrendorf M, Swirski FK, Aikawa E, Stangenberg L, Wurdinger T, et al. (2007) The healing myocardium sequentially mobilizes two monocyte subsets with divergent and complementary functions. J Exp Med 204: 3037–3047.
[22]
Fiset C, Clark RB, Larsen TS, Giles WR (1997) A rapidly activating sustained K+ current modulates repolarization and excitation-contraction coupling in adult mouse ventricle. J Physiol 504(Pt 3): 557–563.
[23]
Zhou YY, Wang SQ, Zhu WZ, Chruscinski A, Kobilka BK, et al. (2000) Culture and adenoviral infection of adult mouse cardiac myocytes: methods for cellular genetic physiology. Am J Physiol Heart Circ Physiol 279: H429–H436.
[24]
Gonzalez-Juarrero M, Shim TS, Kipnis A, Junqueira-Kipnis AP, Orme IM (2003) Dynamics of macrophage cell populations during murine pulmonary tuberculosis. J Immunol 171: 3128–3135.
[25]
Geissmann F, Gordon S, Hume DA, Mowat AM, Randolph GJ (2010) Unravelling mononuclear phagocyte heterogeneity. Nat Rev Immunol 10: 453–460.
[26]
Novobrantseva TI, Majeau GR, Amatucci A, Kogan S, Brenner I, et al. (2005) Attenuated liver fibrosis in the absence of B cells. J Clin Invest 115: 3072–3082.
[27]
Soos TJ, Sims TN, Barisoni L, Lin K, Littman DR, et al. (2006) CX3CR1+ interstitial dendritic cells form a contiguous network throughout the entire kidney. Kidney Int 70: 591–596.
[28]
Gachet C (2006) Regulation of platelet functions by P2 receptors. Annu Rev Pharmacol Toxicol 46: 277–300.
[29]
Arciero JC, Carlson BE, Secomb TW (2008) Theoretical model of metabolic blood flow regulation: roles of ATP release by red blood cells and conducted responses. Am J Physiol Heart Circ Physiol 295: H1562–H1571.
[30]
Wong CW, Christen T, Roth I, Chadjichristos CE, Derouette JP, et al. (2006) Connexin37 protects against atherosclerosis by regulating monocyte adhesion. Nat Med 12: 950–954.
[31]
Idzko M, Hammad H, van Nimwegen M, Kool M, Willart MAM, et al. (2007) Extracellular ATP triggers and maintains asthmatic airway inflammation by activating dendritic cells. Nature Medicine 13: 913–919.
[32]
Mizumoto N, Kumamoto T, Robson SC, Sevigny J, Matsue H, et al. (2002) CD39 is the dominant Langerhans cell associated ecto-NTPDase: Modulatory roles in inflammation and immune responsiveness. Nature Medicine 8: 358–365.
[33]
Elliott MR, Chekeni FB, Trampont PC, Lazarowski ER, Kadl A, et al. (2009) Nucleotides released by apoptotic cells act as a find-me signal to promote phagocytic clearance. Nature 461: 282–U165.
[34]
Robson SC, Sevigny J, Zimmermann H (2006) The E-NTPDase family of ectonucleotidases: Structure function relationships and pathophysiological significance. Purinergic Signal 2: 409–430.
[35]
Levesque SA, Kukulski F, Enjyoji K, Robson SC, Sevigny J (2010) NTPDase1 governs P2X(7)-dependent functions in murine macrophages. European Journal of Immunology 40: 1473–1485.
[36]
Serra S, Horenstein AL, Vaisitti T, Brusa D, Rossi D, et al. (2011) CD73-generated extracellular adenosine in chronic lymphocytic leukemia creates local conditions counteracting drug-induced cell death. Blood 118: 6141–6152.
[37]
Henttinen T, Jalkanen S, Yegutkin GG (2003) Adherent leukocytes prevent adenosine formation and impair endothelial barrier function by Ecto-5′-nucleotidase/CD73-dependent mechanism. J Biol Chem 278: 24888–24895.
[38]
Romio M, Reinbeck B, Bongardt S, Huls S, Burghoff S, et al. (2011) Extracellular purine metabolism and signaling of CD73-derived adenosine in murine Treg and Teff cells. Am J Physiol Cell Physiol 301: C530–C539.
[39]
Salmon JE, Cronstein BN (1990) Fc gamma receptor-mediated functions in neutrophils are modulated by adenosine receptor occupancy. A1 receptors are stimulatory and A2 receptors are inhibitory. J Immunol 145: 2235–2240.
[40]
Save S, Mohlin C, Vumma R, Persson K (2011) Activation of adenosine A2A receptors inhibits neutrophil transuroepithelial migration. Infect Immun 79: 3431–3437.
[41]
Robson SC, Kaczmarek E, Siegel JB, Candinas D, Koziak K, et al. (1997) Loss of ATP diphosphohydrolase activity with endothelial cell activation. Journal of Experimental Medicine 185: 153–163.
[42]
Candinas D, Koyamada N, Miyatake T, Siegel J, Hancock WW, et al. (1996) Loss of rat glomerular ATP diphosphohydrolase activity during reperfusion injury is associated with oxidative stress reactions. Thrombosis and Haemostasis 76: 807–812.
[43]
Eltzschig HK, Ibla JC, Furuta GT, Leonard MO, Jacobson KA, et al. (2003) Coordinated adenine nucleotide phosphohydrolysis and nucleoside signaling in posthypoxic endothelium: role of ectonucleotidases and adenosine A2B receptors. J Exp Med 198: 783–796.
[44]
Enjyoji K, Sevigny J, Lin Y, Frenette PS, Christie PD, et al. (1999) Targeted disruption of cd39/ATP diphosphohydrolase results in disordered hemostasis and thromboregulation. Nat Med 5: 1010–1017.
