TNF-α upregulates the A2B adenosine receptor gene: The role of NAD(P)H oxidase 4

https://doi.org/10.1016/j.bbrc.2008.07.059Get rights and content

Abstract

Proliferation of vascular smooth muscle cells (VSMC), oxidative stress, and elevated inflammatory cytokines are some of the components that contribute to plaque formation in the vasculature. The cytokine tumor necrosis factor-alpha (TNF-α) is released during vascular injury, and contributes to lesion formation also by affecting VSMC proliferation. Recently, an A2B adenosine receptor (A2BAR) knockout mouse illustrated that this receptor is a tissue protector, in that it inhibits VSMC proliferation and attenuates the inflammatory response following injury, including the release of TNF-α. Here, we show a regulatory loop by which TNF-α upregulates the A2BAR in VSMC in vitro and in vivo. The effect of this cytokine is mimicked by its known downstream target, NAD(P)H oxidase 4 (Nox4). Nox4 upregulates the A2BAR, and Nox inhibitors dampen the effect of TNF-α. Hence, our study is the first to show that signaling associated with Nox4 is also able to upregulate the tissue protecting A2BAR.

Section snippets

Materials and methods

Vascular smooth muscle cell culture conditions. Aortic VSMC were isolated from neonatal Sprague–Dawley rats, as described previously [26], [27], and were seeded at a density of 1.5 × 105 cells per well of a 6-well plate; all experiments were performed with cells in their first passage. VSMC were cultured in VSMC growth medium containing DMEM, 10% bovine calf serum (BCS), 1 mM sodium pyruvate, 1 mM nonessential amino acids, 100 U/mL penicillin, and 100 μg/mL streptomycin (all from Invitrogen,

Effect of an inflammation-inducing agent and TNF-α on A2BAR mRNA and promoter activity in vitro and in vivo

In addition to our findings describing the inducibility of the A2BAR gene in proliferating VSMC by the transcription factor B-Myb [30], a recent paper by Kong et al. identified HIF-1α as the responsible transcription factor for the induction of A2BAR expression in endothelial cells under hypoxic conditions [31]. In the context of atherosclerosis, proliferation and hypoxia are stress responses, and A2BAR activation has been shown to be protective in these instances [14], [31]. Additionally,

Acknowledgments

We thank Dr. Barbara Schreiber and the cell core for preparation of smooth muscle cells and for the insight, as well as Dr. Xu Yong for initial assistance with qPCR. This work was supported by National Institutes of Health (NIH) Public Health Services Grant HL13262. K.R. is an Established Investigator with the American Heart Association.

References (41)

  • J. Mattana et al.

    Leukocyte-polytetrafluoroethylene interaction enhances proliferation of vascular smooth muscle cells via tumor necrosis factor-alpha secretion

    Kidney Int.

    (1997)
  • P. Libby

    Inflammation in atherosclerosis

    Nature

    (2002)
  • G. Hasko et al.

    Adenosine inhibits IL-12 and TNF-[alpha] production via adenosine A2a receptor-dependent and independent mechanisms

    FASEB J.

    (2000)
  • G. Hasko et al.

    Adenosine receptor agonists differentially regulate IL-10, TNF-alpha, and nitric oxide production in RAW 264.7 macrophages and in endotoxemic mice

    J. Immunol.

    (1996)
  • J. Linden

    Molecular approach to adenosine receptors: receptor-mediated mechanisms of tissue protection

    Annu. Rev. Pharmacol. Toxicol.

    (2001)
  • L.M. Kreckler et al.

    Adenosine inhibits tumor necrosis factor-alpha release from mouse peritoneal macrophages via A2A and A2B but not the A3 adenosine receptor

    J. Pharmacol. Exp. Ther.

    (2006)
  • V. Kolachala et al.

    TNF-alpha upregulates adenosine 2b (A2b) receptor expression and signaling in intestinal epithelial cells: a basis for A2bR overexpression in colitis

    Cell Mol. Life Sci.

    (2005)
  • M.L. Trincavelli et al.

    Regulation of A2B adenosine receptor functioning by tumour necrosis factor a in human astroglial cells

    J. Neurochem.

    (2004)
  • V. Kolachala et al.

    TNF-a upregulates adenosine 2b (A2b) receptor expression and signaling in intestinal epithelial cells: a basis for A2bR overexpression in colitis

    Cell. Mol. Life Sci.

    (2005)
  • R.K. Dubey et al.

    Adenosine inhibits growth of rat aortic smooth muscle cells. Possible role of A2b receptor

    Hypertension

    (1996)
  • Cited by (45)

    • The adenosine A<inf>2B</inf> G protein-coupled receptor: Recent advances and therapeutic implications

      2019, Pharmacology and Therapeutics
      Citation Excerpt :

      There is also a positive feedback mechanism whereby the A2BAR increases HIF-1α through stabilisation of the circadian rhythm protein period 2 (Per2) (Eckle et al., 2012). Factors present in an inflammatory environment such as the pro-inflammatory cytokines; tumour necrosis factor (TNF)-α (Kolachala et al., 2005), interferon (IFN)-γ (Xaus et al., 1999), interleukin (IL)-1β (Khoa, Montesinos, Williams, Kelly, & Cronstein, 2003), and the reactive oxygen species-generating enzyme Nox4 (St Hilaire, Koupenova, Carroll, Smith, & Ravid, 2008) also upregulate A2BAR expression at the mRNA and protein level. In addition, the endogenous agonist adenosine has been demonstrated to recruit intracellular pools of A2BAR to the plasma membrane on intestinal epithelial cells (Sitaraman et al., 2002; Lixin Wang et al., 2004) and may represent a more general mechanism of regulating A2BAR expression at the cell-surface.

    • Adenosine receptors and caffeine in retinopathy of prematurity

      2017, Molecular Aspects of Medicine
      Citation Excerpt :

      Similar to A2ARs, the A2BRs are adenylyl-cyclase activating receptors, known to be expressed in the eye, and more specifically in the vasculature, microglia and macrophages (Saura et al., 2005; Brambilla et al., 2003; Boison et al., 1071; van Calker and Biber, 2005). Their expression is induced by oxidative stress (St Hilaire et al., 2008), and they control the expression of VEGF in various cells, including macrophages (Granata et al.,). In the cultured human retinal endothelial cells, pharmacological profiles are consistent with A2BR-mediated (but not with A1R- or A2AR-mediated) effects on growth factor expression and cell proliferation (Grant et al., 1999, 2001).

    View all citing articles on Scopus
    1

    Present address: Laboratory for Comparative Carcinogenesis, National Cancer Institute-Frederick, Building 538, Room 144, Frederick, MD 21702, USA.

    View full text