Adenosine binding to adenylyl cyclase stimulatory (A2-type) adenosine receptors (ARs) induces coronary vasodilatation, inhibits human aortic vascular smooth muscle cell proliferation, and affects arterial wall matrix production, all of which are important components of atherosclerosis/restenosis. The role of the low affinity A2bARs in regulating vascular pathology in vivo had not been examined prior to our study. To further address this, we generated the first A2bAR-knockout (KO)/reporter gene-knock-in mouse model and showed reported gene expression primarily in VSMC and macrophages, in accordance with the endogenous profile in control mice. Augmentation of proinflammatory cytokines, such as tumor necrosis factor-? (TNF-?), is the underlying mechanism for an observed upregulation of leukocyte adhesion in the vasculature of these A2bAR KO mice, as compared with age-, sex-, and strain-matched control mice. On the other hand, the A2bAR gene receptor, itself, is induced by TNF-?, suggesting a regulatory loop. A2bAR KO mice display greater lesion formation after guidewire-induced femoral artery injury. This occurs in association with the upregulation of CXCR4, a protein known to promote mobilization of progenitors and inflammatory cells to the vessel by interaction with its ligand stromal cell-derived factor-1 (SDF-1). Furthermore, bone marrow (BM) transplantation experiments indicate that the inflammatory response and vascular lesion formation are significantly regulated by BM-derived A2bARs. Building upon these novel findings, we hypothesize that A2bAR-mediated signaling regulates the expression of CXRC4 and, hence, plays a vital role in vascular lesion formation. It is our contention that macrophage A2bARs, via their ability to control the level of inflammatory cytokines, are capable of significantly protecting against vascular pathology on their own, consistent with the BM transplantation experiments. Finally, identifying mechanisms of A2bAR gene regulation by TNF-? should lead the way for the development of strategies for controlling vascular dysfunction during inflammation.
Three specific aims of research are proposed:
Aim 1. To examine the direct participation of BM cells and/or their signals in mediating effects of the A2bAR on lesion formation during vascular injury and atherosclerosis, and to study related mechanisms, with a focus on the contribution of macrophage A2bARs.
Aim 2. To elucidate the mechanism of control of the CXCR4/SDF-1 axis by the A2bAR.
Aim 3. To study the mechanism of A2bAR gene activation by TNF-? in primary cultures and in vivo. Taken together, our proposed investigations should shed new light on the role of A2bARs in the pathogenesis of vascular dysfunction, and could focus attention on A2bAR activation as a therapeutic target.

Public Health Relevance

Atherosclerosis is a leading cause of vascular disease in the Western world. Our research proposal builds upon novel findings, including the identification of the A2b adenosine receptor as protective against atherosclerosis/restenosis. The mechanisms leading to this protection and the control of expression of this receptor gene are the focus of research in this proposal.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL093149-04
Application #
8235840
Study Section
Atherosclerosis and Inflammation of the Cardiovascular System Study Section (AICS)
Program Officer
Olive, Michelle
Project Start
2009-04-01
Project End
2014-03-31
Budget Start
2012-04-01
Budget End
2014-03-31
Support Year
4
Fiscal Year
2012
Total Cost
$402,188
Indirect Cost
$154,688
Name
Boston University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
604483045
City
Boston
State
MA
Country
United States
Zip Code
02118
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Eisenstein, Anna; Ravid, Katya (2014) G protein-coupled receptors and adipogenesis: a focus on adenosine receptors. J Cell Physiol 229:414-21
Johnston-Cox, Hillary; Eisenstein, Anna S; Koupenova, Milka et al. (2014) The macrophage A2B adenosine receptor regulates tissue insulin sensitivity. PLoS One 9:e98775
Koupenova, Milka; Ravid, Katya (2013) Adenosine, adenosine receptors and their role in glucose homeostasis and lipid metabolism. J Cell Physiol :
Carroll, Shannon H; Wigner, Nathan A; Kulkarni, Nitin et al. (2012) A2B adenosine receptor promotes mesenchymal stem cell differentiation to osteoblasts and bone formation in vivo. J Biol Chem 287:15718-27
Kataoka, Shinji; Baquero, Arian; Yang, Dan et al. (2012) A2BR adenosine receptor modulates sweet taste in circumvallate taste buds. PLoS One 7:e30032
Johnston-Cox, Hillary A; Koupenova, Milka; Ravid, Katya (2012) A2 adenosine receptors and vascular pathologies. Arterioscler Thromb Vasc Biol 32:870-8
Koupenova, Milka; Johnston-Cox, Hillary; Vezeridis, Alexander et al. (2012) A2b adenosine receptor regulates hyperlipidemia and atherosclerosis. Circulation 125:354-63
Chen, Hongjie; Koupenova, Milka; Yang, Dan et al. (2011) Regulation of MMP-9 expression by the A2b adenosine receptor and its dependency on TNF-ýý signaling. Exp Hematol 39:525-30
van der Hoeven, Dharini; Wan, Tina C; Gizewski, Elizabeth T et al. (2011) A role for the low-affinity A2B adenosine receptor in regulating superoxide generation by murine neutrophils. J Pharmacol Exp Ther 338:1004-12

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