Restoration of the cAMP second messenger pathway enhances cardiac preservation for transplantation in a heterotopic rat model.

Abstract

Current organ preservation strategies subject graft vasculature to severe hypoxia (Po2 -20 Torr), potentially compromising vascular function and limiting successful transplantation. Previous work has shown that cAMP modulates endothelial cell (EC) antithrombogenicity, barrier function, and leukocyte /EC interactions, and that hypoxia suppresses EC cAMP levels. To explore the possible benefits of cAMP analogs/agonists in organ preservation, we used a rat heterotopic cardiac transplant model; dibutyryl cAMP added to preservation solutions was associated with a time-and dose-dependent increase in the duration of cold storage associated with successful graft function.Preservation was also enhanced by 8-bromo-cAMP, the SP isomer of adenosine 3',5'monophosphorothioate, and types III (indolidan) and IV (rolipram) phosphodiesterase inhibitors. Neither butyrate alone nor 8-bromoadenosine were effective, and the cAMP-dependent protein kinase antagonist Rp isomer of adenosine 3',5'monophosphorothioate prevented preservation enhancement induced by 8-bromo-cAMP. Grafts stored with dibutyryl cAMP demonstrated a 5.5-fold increase in blood flow and a 3.2-fold decreased neutrophil infiltration after transplantation. To explore the role of cAMP in another cell type critical for vascular homeostasis, vascular smooth muscle cells were subjected to hypoxia, causing a time-dependent decline in cAMP levels. Although adenylate cyclase activity was unchanged, diminished oxygen tensions were associated with enhanced phosphodiesterase activity (59 and 30% increase in soluble types III and IV activity, respectively). These data suggest that hypoxia or graft ischemia disrupt vascular homeostasis, at least in part, by perturbing the cAMP second messenger pathway. Supplementation of this pathway provides a new approach for enhancing cardiac preservation, promoting myocardial function, and maintaining vascular homeostatic properties. (J.

cAMP/CREB-mediated Transcriptional Regulation of Ectonucleoside Triphosphate Diphosphohydrolase 1 (CD39) Expression

“…Both adenine nucleotide phosphohydrolysis data and platelet aggregation data, demonstrating that cAMP upregulates CD39 enzyme activity and platelet-inhibitory function, lend credence to this hypothesis. Supporting this, in a rat heterotopic cardiac transplant model, grafts stored with the membrane-permeable cAMP analogues dibutyryl-cAMP or 8-Br-cAMP demonstrated a 5.5-fold increase in blood flow and a 3.2-fold decrease in neutrophil infiltration after transplantation (56). Provision of cAMP also enhances lung preservation in an orthotopic rat left lung transplant model.…”

“…Hypoxia and reoxygenation increase endothelial cell permeability, induce procoagulant activity, and alter endothelial cell/ leukocyte interactions, with a parallel reduction in endothelial cAMP and nitric oxide levels. Under conditions of oxygen deprivation, cAMP promotes endothelial cell antithrombogenicity, maintenance of barrier function, and inhibition of leukocyte/ endothelial cell interactions, although the exact mechanisms remain unclear (56). Our data lead us to hypothesize that enhancement by cAMP of vascular anti-thrombotic properties could be in large measure due to the cAMP-stimulated induction of CD39.…”

Hypoxia-induced exocytosis of endothelial cell Weibel-Palade bodies. A mechanism for rapid neutrophil recruitment after cardiac preservation.

“…When ECs achieved confluence, experiments were performed by placing cultures in an environmental chamber (Coy Laboratory Products, Ann Arbor, MI), which provided a controlled temperature (37°C) and atmosphere with the indicated amount of oxygen and carbon dioxide (5%) and the balance made up of nitrogen. Use of this chamber for cell culture experiments has been described previously (15,18). During exposure of ECs to hypoxia (for a maximum of 16 h), the oxygen tension in the culture medium was 14-18 torr, and there was no change in the medium pH.…”

“…Our experiments were designed to establish whether hypoxia per se (or hypothermic cardiac preservation, as occurs during cardiac surgery, in which the pO 2 within the coronary bed declines to pO 2 Ͻ 20 torr) (15) would result in WP body exocytosis. Furthermore, we wished to determine the role of P-selectin-dependent PMN adhesion in the cardiac graft failure that characteristically follows a period of prolonged hypothermic preservation.…”

Adenosine A2 receptor stimulation protects the predamaged liver from cold preservation through activation of cyclic adenosine monophosphate-protein kinase a pathway

“…13 Enhancement of cellular cAMP in turn has recently been proposed to be protective in ischemic preservation of lung, heart, gut, and liver. 11,[14][15][16] In the present study, we investigated whether adenosine A2-receptor stimulation might protect suboptimal donor livers from cold preservation, emphasizing its role in the modulation of the cAMP-protein kinase A (PKA) pathway.…”