Adenosine protected against pulmonary edema through transporter- and receptor A₂-mediated endothelial barrier enhancement

Abstract

We have previously demonstrated that adenosine plus homocysteine enhanced endothelial basal barrier function and protected against agonist-induced barrier dysfunction in vitro through attenuation of RhoA activation by inhibition of isoprenylcysteine-O-carboxyl methyltransferase. In the current study, we tested the effect of elevated adenosine on pulmonary endothelial barrier function in vitro and in vivo. We noted that adenosine alone dose dependently enhanced endothelial barrier function. While adenosine receptor A(1) or A(3) antagonists were ineffective, an adenosine transporter inhibitor, NBTI, or a combination of DPMX and MRS1754, antagonists for adenosine receptors A(2A) and A(2B), respectively, partially attenuated the barrier-enhancing effect of adenosine. Similarly, inhibition of both A(2A) and A(2B) receptors with siRNA also blunted the effect of adenosine on barrier function. Interestingly, inhibition of both transporters and A(2A)/A(2B) receptors completely abolished adenosine-induced endothelial barrier enhancement. The adenosine receptor A(2A) and A(2B) agonist, NECA, also significantly enhanced endothelial barrier function. These data suggest that both adenosine transporters and A(2A) and A(2B) receptors are necessary for exerting maximal effect of adenosine on barrier enhancement. We also found that adenosine enhanced Rac1 GTPase activity and overexpression of dominant negative Rac1 attenuated adenosine-induced increases in focal adhesion complexes. We further demonstrated that elevation of cellular adenosine by inhibition of adenosine deaminase with Pentostatin significantly enhanced endothelial basal barrier function, an effect that was also associated with enhanced Rac1 GTPase activity and with increased focal adhesion complexes and adherens junctions. Finally, using a non-inflammatory acute lung injury (ALI) model induced by alpha-naphthylthiourea, we found that administration of Pentostatin, which elevated lung adenosine level by 10-fold, not only attenuated the development of edema before ALI but also partially reversed edema after ALI. The data suggest that adenosine deaminase inhibition may be useful in treatment of pulmonary edema in settings of ALI.

Metabolic consequences of sepsis-induced acute lung injury revealed by plasma 1H-nuclear magnetic resonance quantitative metabolomics and computational analysis

“…There are adverse consequences of prolonged elevated levels of adenosine including the propagation of lung inflammation and the activation of apoptosis signaling pathways (17,32,49). Conversely, adenosine may enhance barrier function and be an important and necessary adaptive response to lung injury (14,16,23). One possible direction for future work is to see if the expression of this or any other genes encoding key enzymes regulating the metabolism of the four metabolites that we identified are altered in sepsis-induced ALI patients.…”

Cigarette smoke causes lung vascular barrier dysfunction via oxidative stress-mediated inhibition of RhoA and focal adhesion kinase

“…The lungs were then dried for 48 h in an oven at 90°C and the dry weights were recorded. Data are presented as the ratio of wet weights to dry weights, as we previously described (33).…”

Selective induction of endothelial P2Y6 nucleotide receptor promotes vascular inflammation

“…32,33 As such, adenosine triphosphate (ATP) or adenosine diphosphate (ADP) can be rapidly hydrolyzed to adenosine in a 2-step enzymatic process that involves the ectonucleoside triphosphate diphosphohydrolase 1 (CD39; conversion of ATP/ADP to adenosine monophosphate [AMP]) 11,18,21,22,34 and the ecto-5Ј-nucleotidase (CD73; conversion of AMP to adenosine). [34][35][36][37][38][39] Several studies have implicated extracellular adenosine signaling in attenuating acute inflammatory responses during conditions such as hypoxia-induced vascular leakage and pulmonary edema, [10][11]13,40 sepsis, 41,42 or acute lung injury. 15,16,42,43 As such, experimental approaches that will enhance extracellular nucleotide breakdown and their subsequent conversion to adenosine will simultaneously dampen activation of nucleotide receptors (such as the P2Y 6 ) and increase extracellular adenosine signaling.…”