A2B adenosine receptor dampens hypoxia-induced vascular leak. Blood

Departments of Anesthesiology and Intensive Care , Tübingen University Hospital, Tübingen, Germany.
Blood (Impact Factor: 10.43). 03/2008; 111(4):2024-35. DOI: 10.1182/blood-2007-10-117044
Source: PubMed

ABSTRACT Extracellular adenosine has been implicated in adaptation to hypoxia and previous studies demonstrated a central role in vascular responses. Here, we examined the contribution of individual adenosine receptors (ARs: A1AR/A2AAR/A2BAR/A3AR) to vascular leak induced by hypoxia. Initial profiling studies revealed that siRNA-mediated repression of the A2BAR selectively increased endothelial leak in response to hypoxia in vitro. In parallel, vascular permeability was significantly increased in vascular organs of A2BAR(-/-)-mice subjected to ambient hypoxia (8% oxygen, 4 hours; eg, lung: 2.1 +/- 0.12-fold increase). By contrast, hypoxia-induced vascular leak was not accentuated in A1AR(-/-)-, A2AAR(-/-)-, or A3AR(-/-)-deficient mice, suggesting a degree of specificity for the A2BAR. Further studies in wild type mice revealed that the selective A2BAR antagonist PSB1115 resulted in profound increases in hypoxia-associated vascular leakage while A2BAR agonist (BAY60-6583 [2-[6-amino-3,5-dicyano-4-[4-(cyclopropylmethoxy)-. phenyl]pyridin-2-ylsulfanyl]acetamide]) treatment was associated with almost complete reversal of hypoxia-induced vascular leakage (eg, lung: 2.0 +/- 0.21-fold reduction). Studies in bone marrow chimeric A2BAR mice suggested a predominant role of vascular A2BARs in this response, while hypoxia-associated increases in tissue neutrophils were, at least in part, mediated by A2BAR expressing hematopoietic cells. Taken together, these studies provide pharmacologic and genetic evidence for vascular A2BAR signaling as central control point of hypoxia-associated vascular leak.

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Available from: Linda F Thompson, Aug 04, 2014
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    • "After blocking for 2 h at room temperature with 5% milk in Trisbuffered saline, pH 7.6, containing 0.1% Tween 20 (TBS-T), the membranes were incubated overnight at 4 °C with a rabbit anti- A 2B R antibody (1:250; sc-28996 from Santa Cruz Biotechnology, Santa Cruz, CA, USA). This particular antibody was selected as its selectivity has been previously been validated by eliminating its immunoreactivity upon neutralising A 2B R with a siRNA (Eckle et al., 2008). After four washing periods for 10 min with TBS-T containing 0.5% milk, the membranes were incubated with the alkaline phosphatase-conjugated anti-rabbit secondary antibody (1:2000; GE Healthcare) in TBS-T containing 1% milk for 90 min at room temperature. "
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    ABSTRACT: Adenosine is a neuromodulator mostly acting through A1 (inhibitory) and A2A (excitatory) receptors in the brain. A2B receptors (A2 B R) are Gs/q -protein-coupled receptors with low expression in the brain. As A2 B R function is largely unknown, we have now explored their role in the mouse hippocampus. We performed electrophysiological extracellular recordings in mouse hippocampal slices, and immunological analysis of nerve terminals and glutamate release in hippocampal slices and synaptosomes. Additionally, A2 B R-knockout (A2 B R-KO) and C57/BL6 mice were submitted to a behavioural test battery (open field, elevated plus-maze, Y-maze). The A2 B R agonist BAY60-6583 (300 nm) decreased the paired-pulse stimulation ratio, an effect prevented by the A2 B R antagonist MRS 1754 (200 nM) and abrogated in A2 B R-KO mice. Accordingly, A2 B R immunoreactivity was present in 73 ± 5% of glutamatergic nerve terminals, i.e. those immunopositive for vesicular glutamate transporters. Furthermore, BAY 60-6583 attenuated the A1 R control of synaptic transmission, both the A1 R inhibition caused by 2-chloroadenosine (0.1-1 μm) and the disinhibition caused by the A1 R antagonist DPCPX (100 nm), both effects prevented by MRS 1754 and abrogated in A2 B R-KO mice. BAY 60-6583 decreased glutamate release in slices and also attenuated the A1 R inhibition (CPA 100 nm). A2 B R-KO mice displayed a modified exploratory behaviour with an increased time in the central areas of the open field, elevated plus-maze and the Y-maze and no alteration of locomotion, anxiety or working memory. We conclude that A2 B R are present in hippocampal glutamatergic terminals where they counteract the predominant A1 R-mediated inhibition of synaptic transmission, impacting on exploratory behaviour. © 2015 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.
    European Journal of Neuroscience 02/2015; 41(7). DOI:10.1111/ejn.12851 · 3.67 Impact Factor
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    • "This in turn leads to recruitment of immune cells to the lung and loss of pulmonary barrier function, producing the inflammation and pulmonary edema that are characteristics of hyperoxic lung injury. Adenosine is a nucleoside signaling molecule that has been recognized to play an important role in the regulation of inflammation following acute lung injury (Eckle et al. 2008a,b; Schingnitz et al. 2010; Karmouty-Quintana et al. 2013). The concentration of adenosine is normally low in the extracellular compartment, but following acute lung injury, extracellular adenosine concentrations rapidly increase (Volmer et al. 2006; Eckle et al. 2007). "
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    ABSTRACT: Hyperoxic lung injury is characterized by cellular damage from high oxygen concentrations that lead to an inflammatory response in the lung with cellular infiltration and pulmonary edema. Adenosine is a signaling molecule that is generated extracellularly by CD73 in response to injury. Extracellular adenosine signals through cell surface receptors and has been found to be elevated and plays a protective role in acute injury situations. In particular, ADORA2B activation is protective in acute lung injury. However, little is known about the role of adenosine signaling in hyperoxic lung injury. We hypothesized that hyperoxia-induced lung injury leads to CD73-mediated increases in extracellular adenosine, which is protective through ADORA2B signaling pathways. To test this hypothesis, we exposed C57BL6, CD73−/−, and Adora2B−/− mice to 95% oxygen or room air and examined markers of pulmonary inflammation, edema, and monitored lung histology. Hyperoxic exposure caused pulmonary inflammation and edema in association with elevations in lung adenosine levels. Loss of CD73-mediated extracellular adenosine production exacerbated pulmonary edema without affecting inflammatory cell counts. Furthermore, loss of the ADORA2B had similar results with worsening of pulmonary edema following hyperoxia exposure without affecting inflammatory cell infiltration. This loss of barrier function correlated with a decrease in occludin in pulmonary vasculature in CD73−/− and Adora2B−/− mice following hyperoxia exposure. These results demonstrate that exposure to a hyperoxic environment causes lung injury associated with an increase in adenosine concentration, and elevated adenosine levels protect vascular barrier function in hyperoxic lung injury through the ADORA2B-dependent regulation of occludin.
    09/2014; 2(9). DOI:10.14814/phy2.12155
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    • "Thus, it is likely adenosine release at inflamed sites contributes to the erythema (rubor) and resulting heat loss (calor) associated with inflammation. Interestingly, diminished production of adenosine leads to dramatic vascular leakage resulting from diminished activation of adenosine A 2B receptors on the vascular endothelium (Thompson et al., 2004; Eckle et al., 2008) suggesting that the adenosine released at inflamed sites diminishes the swelling (tumor) that is so prominent at inflamed sites. "
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    ABSTRACT: Adenosine, a purine nucleoside generated by the dephosphorylation of adenine nucleotides, is a potent endogenous physiologic and pharmacologic regulator of many functions. Adenosine was first reported to inhibit the inflammatory actions of neutrophils nearly 30 years ago and since then the role of adenosine and its receptors as feedback regulators of inflammation has been well established. Here we review the effects of adenosine, acting at its receptors, on neutrophil and monocyte/macrophage function in inflammation. Moreover, we review the role of adenosine in mediating the anti-inflammatory effects of methotrexate, the anchor drug in the treatment of Rheumatoid Arthritis and other inflammatory disorders.
    Frontiers in Immunology 04/2013; 4:85. DOI:10.3389/fimmu.2013.00085
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