Human Recombinant Apyrase Therapy Protects Against Myocardial Ischemia/Reperfusion Injury and Preserves Left Ventricular Systolic Function in Rats, as Evaluated by 7T Cardiovascular Magnetic Resonance Imaging.

OBJECTIVE: The occurrence of intramyocardial hemorrhage (IMH) and microvascular obstruction (MVO) in myocardial infarction (MI), known as severe ischemia/reperfusion injury (IRI), has been associated with adverse remodeling. APT102, a soluble human recombinant ecto-nucleoside triphosphate diphosphohydrolase-1, can hydrolyze extracellular nucleotides to attenuate their prothrombotic and proinflammatory effects. The purpose of this study was to temporally evaluate the therapeutic effect of APT102 on IRI in rats and to elucidate the evolution of IRI in the acute stage using cardiovascular magnetic resonance imaging (CMRI). MATERIALS AND METHODS: Fifty-four rats with MI, induced by ligation of the origin of the left anterior descending coronary artery for 60 minutes, were randomly divided into the APT102 (n = 27) or control (n = 27) group. Intravenous infusion of APT102 (0.3 mg/kg) or placebo was administered 15 minutes before reperfusion, and then 24 hours, 48 hours, 72 hours, and on day 4 after reperfusion. CMRI was performed at 24 hours, 48 hours, 72 hours, and on day 5 post-reperfusion using a 7T system and the hearts were collected for histopathological examination. Cardiac function was quantified using cine imaging and IMH/edema using T2 mapping, and infarct/MVO using late gadolinium enhancement. RESULTS: The extent of infarction (p < 0.001), edema (p < 0.001), IMH (p = 0.013), and MVO (p = 0.049) was less severe in the APT102 group than in the control group. IMH size at 48 hours was significantly greater than that at 24 hours, 72 hours, and 5 days after reperfusion (all p < 0.001). The left ventricular ejection fraction (LVEF) was significantly greater in the APT102 group than in the control group (p = 0.006). There was a negative correlation between LVEF and IMH (r = -0.294, p = 0.010) and a positive correlation between IMH and MVO (r = 0.392, p < 0.001). CONCLUSION: APT102 can significantly alleviate damage to the ischemic myocardium and microvasculature. IMH size peaked at 48 hours post reperfusion and IMH is a downstream consequence of MVO. IMH may be a potential therapeutic target to prevent adverse remodeling in MI.

Kinetics of MSC-based enzyme therapy for immunoregulation.

BACKGROUND: Mesenchymal stromal cells (MSC) demonstrate innate and regulatory immunologic functions and have been widely explored for cell therapy applications. Mechanisms by which MSCs achieve therapeutic effects are theorized, though appropriate dosing and duration of these mechanisms in vivo warrant deeper investigation. One, rapid immunosuppressive function of MSCs is through ectoenzyme expression of CD73 and CD39 which cooperatively hydrolyze inflammatory, extracellular adenosine triphosphate (ATP) to anti-inflammatory adenosine. Extracellular ATP has a key role in autoimmune and inflammatory diseases, which administered MSCs have the potential to modulate in a timescale that is befitting of shorter acting therapeutic function. METHODS: In vitro experiments were performed to determine the hydrolysis rates of ATP by MSCs. Through kinetic modeling from experimental results, the rate at which a single cell can metabolize ATP was determined. Based on these rates, the ability of MSCs to downregulate inflammatory signaling pathways was prospectively validated using model system parameters with respect to two different mechanisms: extracellular ATP stimulates lymphocytes to suppress proliferation and induce apoptosis and with co-stimulation, it stimulates monocytes to release pro-inflammatory IL-1ß. MSCs were co-cultured with immune cells using transwell inserts and compared to immune cell only groups. RESULTS: Hydrolysis of ATP was efficiently modeled by first-order enzyme kinetics. For in vitro culture, the rate at which a single cell can hydrolyize ATP is 8.9 nmol/min. In the presence of extracellular ATP, cocultures of MSCs reduced cytotoxicity and allows for proliferation of lymphocytes while limiting IL-1ß secretion from monocytes. CONCLUSIONS: Such use of these models may allow for better dosing predictions for MSCs to be used therapeutically for chronic inflammatory diseases such as rheumatoid arthritis, diabetes, pancreatitis, and other systemic inflammatory response syndromes. For the first time, the effect of MSCs on ATP hydrolysis in immune cell response is quantitatively analyzed on a cell-molecule basis by modeling the hydrolysis as an enzyme-substrate reaction. The results also give insight into MSCs' dynamic response mechanisms to ameliorate effects of extracellular ATP whether it be through positive or negative regulation.

Apyrase Elicits Host Antimicrobial Responses and Resolves Infection in Burns.

The authors previously reported that adenosine triphosphate (ATP) stimulates biofilm formation and removal of the ATP could reduce biofilm formation. The main objective of this study was to evaluate the effects of the ATP-hydrolyzing enzyme, apyrase, on control of Acinetabacter baumannii infection in the burn wound as well as to assess host skin antimicrobial responses. The authors found that apyrase stimulated nitric oxide formation at the wound site and reduced CD55 expression, thereby inducing the assembly of membrane attack complexes. Apyrase treatment nearly eradicated multidrug-resistant A. baumannii from burn wounds in the absence of antibiotics. Apyrase may be an effective therapy against antibiotic-resistant bacterial infections in burns.

Modulation of radiochemoimmunotherapy-induced B16 melanoma cell death by the pan-caspase inhibitor zVAD-fmk induces anti-tumor immunity in a HMGB1-, nucleotide- and T-cell-dependent manner.

One prerequisite that radiotherapy (RT) and chemotherapy (CT) result in anti-tumor immune responses is triggering of immunogenic cell death forms such as necroptosis. The latter is inducible by inhibition of apoptosis with the pan-caspase inhibitor zVAD-fmk. The design of multimodal therapies that overcome melanoma's resistance to apoptosis is a big challenge of oncoimmunology. As hints exist that immune stimulation by hyperthermia (HT) augments the efficacy of melanoma therapies and that tumors can be sensitized for RT with zVAD-fmk, we asked whether combinations of RT with dacarbazine (DTIC) and/or HT induce immunogenic melanoma cell death and how this is especially influenced by zVAD-fmk. Necroptosis was inducible in poorly immunogenic B16-F10 melanoma cells and zVAD-fmk generally increased melanoma cell necrosis concomitantly with the release of HMGB1. Supernatants (SNs) of melanoma cells whose cell death was modulated with zVAD-fmk induced an upregulation of the activation markers CD86 and MHCII on macrophages. The same was seen on dendritic cells (DCs), but only when zVAD-fmk was added to multimodal tumor treatments including DTIC. DCs of MyD88 KO mice and DCs incubated with SNs containing apyrase did not increase the expression of these activation markers on their surface. The in vivo experiments revealed that zVAD-fmk decreases the tumor growth significantly and results in a significantly reduced tumor infiltration of Tregs when added to multimodal treatment of the tumor with RT, DTIC and HT. Further, a significantly increased DC and CD8+ T-cell infiltration into the tumor and in the draining lymph nodes was induced, as well as an increased expression of IFNγ by CD8+ T cells. However, zVAD-fmk did not further reduce tumor growth in MyD88 KO mice, mice treated with apyrase or RAG KO mice. We conclude that HMGB1, nucleotides and CD8+ T cells mediate zVAD-fmk induced anti-melanoma immune reactions in multimodal therapy settings.

Human recombinant apyrase therapy protects against canine pulmonary ischemia-reperfusion injury.

BACKGROUND: There is accumulating evidence that extracellular adenosine triphosphate (eATP) promotes many of the underlying mechanisms that exacerbate acute lung injury. However, much of these data are from inbred rodent models, indicating the need for further investigation in higher vertebrates to better establish clinical relevance. To this end we evaluated a human recombinant apyrase therapy in a canine warm pulmonary ischemia-reperfusion injury (IRI) model and measured eATP levels in human lung recipients with or without primary lung graft dysfunction (PGD). METHODS: Warm ischemia was induced for 90 minutes in the left lung of 14 mongrel dogs. Seven minutes after reperfusion, the apyrase APT102 (1 mg/kg, n = 7) or saline vehicle (n = 7) was injected into the pulmonary artery. Arterial blood gases were obtained every 30 minutes up to 180 minutes after reperfusion. Bronchioalveolar lavage fluid (BALF) was analyzed for eATP concentration, cellularity, and inflammatory mediator accumulation. Thirty bilateral human lung transplant recipients were graded for immediate early PGD and assessed for BALF eATP levels. RESULTS: APT102-treated dogs had progressively better lung function and less pulmonary edema during the 3-hour reperfusion period compared with vehicle-treated controls. Protection from IRI was observed, with lower BALF eATP levels, fewer airway leukocytes, and blunted inflammatory mediator expression. Human lung recipients with moderate to severe PGD had significantly higher eATP levels compared with recipients without this injury. CONCLUSIONS: Extracellular ATP accumulates in acutely injured canine and human lungs. Strategies that target eATP reduction may help protect lung recipients from IRI.

Optimizing human apyrase to treat arterial thrombosis and limit reperfusion injury without increasing bleeding risk.

In patients with acute myocardial infarction undergoing reperfusion therapy to restore blood flow through blocked arteries, simultaneous inhibition of platelet P2Y12 receptors with the current standard of care neither completely prevents recurrent thrombosis nor provides satisfactory protection against reperfusion injury. Additionally, these antiplatelet drugs increase the risk of bleeding. To devise a different strategy, we engineered and optimized the apyrase activity of human nucleoside triphosphate diphosphohydrolase-3 (CD39L3) to enhance scavenging of extracellular adenosine diphosphate, a predominant ligand of P2Y12 receptors. The resulting recombinant protein, APT102, exhibited greater than four times higher adenosine diphosphatase activity and a 50 times longer plasma half-life than did native apyrase. Treatment with APT102 before coronary fibrinolysis with intravenous recombinant human tissue-type plasminogen activator in conscious dogs completely prevented thrombotic reocclusion and significantly decreased infarction size by 81% without increasing bleeding time. In contrast, clopidogrel did not prevent coronary reocclusion and increased bleeding time. In a murine model of myocardial reperfusion injury caused by transient coronary artery occlusion, APT102 also decreased infarct size by 51%, whereas clopidogrel was not effective. These preclinical data suggest that APT102 should be tested for its ability to safely and effectively maximize the benefits of myocardial reperfusion therapy in patients with arterial thrombosis.

Delayed targeting of CD39 to activated platelet GPIIb/IIIa via a single-chain antibody: breaking the link between antithrombotic potency and bleeding?

The ecto-nucleoside triphosphate diphosphohydrolase CD39 represents a promising antithrombotic therapeutic. It degrades adenosine 5'-diphosphate (ADP), a main platelet activating/recruiting agent. We hypothesized that delayed enrichment of CD39 on developing thrombi will allow for a low and safe systemic concentration and thus avoid bleeding. We use a single-chain antibody (scFv, specific for activated GPIIb/IIIa) for targeting CD39. This should allow delayed enrichment on growing thrombi but not on the initial sealing layer of platelets, which do not yet express activated GPIIb/IIIa. CD39 was recombinantly fused to an activated GPIIb/IIIa-specific scFv (targ-CD39) and a nonfunctional scFv (non-targ-CD39). Targ-CD39 was more effective at preventing ADP-induced platelet activation than non-targ-CD39. In a mouse carotid artery thrombosis model, non-targ-CD39, although protective against vessel occlusion, was associated with significant bleeding on tail transection. In contrast, targ-CD39 concentrated at the thrombus site; hence, a dose ∼10 times less of CD39 prevented vessel occlusion to a similar extent as high-dose non-targ-CD39, without prolonged bleeding time. An equimolar dose of non-targ-CD39 at this low concentration was ineffective at preventing vessel occlusion. Thus, delayed targeting of CD39 via scFv to activated platelets provides strong antithrombotic potency and yet prevents bleeding and thereby promotes CD39 toward clinical use.

Targeting stenosis with nucleotide-hydrolyzing enzymes.

Well-established evidence links extracellular nucleotides to numerous vascular pathologies, including restenosis associated with angioplasty, atherosclerosis and transplant arteriosclerosis. Through activation of purinergic P2 receptors, extracellular nucleotides contribute to the pathogenesis of occlusive vascular diseases by mediating thrombosis, and vascular smooth muscle proliferation and migration. Therefore, there is a growing interest in the enzymes that hydrolyze nucleotides for their capability to modulate nucleotide-triggered pathologies. In this review, we present the current data addressing the therapeutic potential of nucleoside triphosphate diphosphohydrolases (NTPDases) to prevent intimal hyperplasia and treat vascular intimal disease. In addition, we discuss the mechanisms by which NTPDases exert protective effects in vascular function.

Apyrase treatment of myocardial infarction according to a clinically applicable protocol fails to reduce myocardial injury in a porcine model.

BACKGROUND: Ectonucleotidase dependent adenosine generation has been implicated in preconditioning related cardioprotection against ischemia-reperfusion injury, and treatment with a soluble ectonucleotidase has been shown to reduce myocardial infarct size (IS) when applied prior to induction of ischemia. However, ectonucleotidase treatment according to a clinically applicable protocol, with administration only after induction of ischemia, has not previously been evaluated. We therefore investigated if treatment with the ectonucleotidase apyrase, according to a clinically applicable protocol, would reduce IS and microvascular obstruction (MO) in a large animal model. METHODS: A percutaneous coronary intervention balloon was inflated in the left anterior descending artery for 40 min, in 16 anesthetized pigs (40-50 kg). The pigs were randomized to 40 min of 1 ml/min intracoronary infusion of apyrase (10 U/ml, n = 8) or saline (0.9 mg/ml, n = 8), twenty minutes after balloon inflation. Area at risk (AAR) was evaluated by ex vivo SPECT. IS and MO were evaluated by ex vivo MRI. RESULTS: No differences were observed between the apyrase group and saline group with respect to IS/AAR (75.7 +/- 4.2% vs 69.4 +/- 5.0%, p = NS) or MO (10.7 +/- 4.8% vs 11.4 +/- 4.8%, p = NS), but apyrase prolonged the post-ischemic reactive hyperemia. CONCLUSION: Apyrase treatment according to a clinically applicable protocol, with administration of apyrase after induction of ischemia, does not reduce myocardial infarct size or microvascular obstruction.

Human solCD39 inhibits injury-induced development of neointimal hyperplasia.

Blood platelets provide the initial response to vascular endothelial injury, becoming activated as they adhere to the injured site. Activated platelets recruit leukocytes, and initiate proliferation and migration of vascular smooth muscle cells (SMC) within the injured vessel wall, leading to development of neointimal hyperplasia. Endothelial CD39/NTPDase1 and recombinant solCD39 rapidly metabolise nucleotides, including stimulatory ADP released from activated platelets, thereby suppressing additional platelet reactivity. Using a murine model of vascular endothelial injury, we investigated whether circulating human solCD39 could reduce platelet activation and accumulation, thus abating leukocyte infiltration and neointimal formation following vascular damage. Intraperitoneally-administered solCD39 ADPase activity in plasma peaked 1 hour (h) post-injection, with an elimination half-life of 43 h. Accordingly, mice were administered solCD39 or saline 1 h prior to vessel injury, then either sacrificed 24 h post-injury or treated with solCD39 or saline (three times weekly) for an additional 18 days. Twenty-four hours post-injury, solCD39-treated mice displayed a reduction in platelet activation and recruitment, P-selectin expression, and leukocyte accumulation in the arterial lumen. Furthermore, repeated administration of solCD39 modulated the late stage of vascular injury by suppressing leukocyte deposition, macrophage infiltration and smooth muscle cell (SMC) proliferation/migration, resulting in abrogation of neointimal thickening. In contrast, injured femoral arteries of saline-injected mice exhibited massive platelet thrombus formation, marked P-selectin expression, and leukocyte infiltration. Pronounced neointimal growth with macrophage and SMC accretion was also observed (intimal-to-medial area ratio 1.56 +/- 0.34 at 19 days). Thus, systemic administration of solCD39 profoundly affects injury-induced cellular responses, minimising platelet deposition and leukocyte recruitment, and suppressing neointimal hyperplasia.

APT102, a novel adpase, cooperates with aspirin to disrupt bone metastasis in mice.

Platelets contribute to the development of metastasis, the most common cause of mortality in cancer patients, but the precise role that anti-platelet drugs play in cancer treatment is not defined. Metastatic tumor cells can produce platelet alphaIIb beta3 activators, such as ADP and thromboxane A(2) (TXA(2)). Inhibitors of platelet beta3 integrins decrease bone metastases in mice but are associated with significant bleeding. We examined the role of a novel soluble apyrase/ADPase, APT102, and an inhibitor of TXA(2) synthesis, acetylsalicylic acid (aspirin or ASA), in mouse models of experimental bone metastases. We found that treatment with ASA and APT102 in combination (ASA + APT102), but not either drug alone, significantly decreased breast cancer and melanoma bone metastases in mice with fewer bleeding complications than observed with alphaIIb beta3 inhibition. ASA + APT102 diminished tumor cell induced platelet aggregation but did not directly alter tumor cell viability. Notably, APT102 + ASA treatment did not affect initial tumor cell distribution and similar results were observed in beta3-/- mice. These results show that treatment with ASA + APT102 decreases bone metastases without significant bleeding complications. Anti-platelet drugs such as ASA + APT102 could be valuable experimental tools for studying the role of platelet activation in metastasis as well as a therapeutic option for the prevention of bone metastases.

In vivo glioblastoma growth is reduced by apyrase activity in a rat glioma model.

BACKGROUND: ATP is an important signalling molecule in the peripheral and central nervous system. Both glioma growth and tumor resection induces cell death, thus liberating nucleotides to the extracellular medium. Nucleotides are hydrolyzed very slowly by gliomas when compared with astrocytes and induce neuronal cell death and glioma proliferation. The objective of the present study was to test the involvement of extracellular ATP in glioblastoma growth in a rat glioma model. METHODS: To deplete the extracellular ATP, the enzyme apyrase was tested on the treatment of gliomas implanted in the rats CNS. One million glioma C6 cells in 3 microliters of DMEM/FCS were injected in the right striata of male Wistar rats, 250-270 g. After 20 days, the rats were decapitated and the brain sectioning and stained with hematoxylin and eosine. We performed immunohistochemical experiments with Ki67, CD31 and VEGF. Total RNA was isolated from cultured glioma C6 cells and the cDNA was analyzed by Real Time-PCR with primers for the NTPDase family. RESULTS: C6 glioma cells effectively have a low expression of all NTPDases investigated, in comparison with normal astrocytes. The implanted glioma co-injected with apyrase had a significant reduction in the tumor size (p < 0.05) when compared with the rats injected only with gliomas or with gliomas plus inactivated apyrase. According to the pathological analysis, the malignant gliomas induced by C6 injection and co-injected with apyrase presented a significant reduction in the mitotic index and other histological characteristics that indicate a less invasive/proliferative tumor. Reduction of proliferation induced by apyrase co-injection was confirmed by counting the percentage of Ki67 positive glioma cell nuclei. According to counts with CD31, vessel density and neoformation was higher in the C6 group 20 days after implantation. Confirming this observation, rats treated with apyrase presented less VEGF staining in comparison to the control group. CONCLUSION: These results indicate that the participation of extracellular ATP and the ecto-nucleotidases may be associated with the development of this type of brain tumor in an in vivo glioma model.

Ecto-nucleotidases of the CD39/NTPDase family modulate platelet activation and thrombus formation: Potential as therapeutic targets.

Extracellular nucleotide P2-receptor-mediated effects on platelets, leukocytes and endothelium are modulated by ecto-nucleotidases. These ecto-enzymes hydrolyze extracellular nucleotides to the respective nucleosides. The dominant ecto-nucleotidase expressed by the endothelium, by monocytes and vascular smooth muscle cells is CD39/NTPDase1. Ecto-nucleotidase biochemical activity of CD39 is lost at sites of acute vascular injury, such as in ischemia reperfusion and immune graft rejection. CD39L(Like)1/NTPDase2, a related protein, is associated with the basolateral surface of endothelium, the adventitia of vessels and microvascular pericytes. CD39/NTPDase1 hydrolyzes both tri- and diphosphonucleosides and blocks platelet aggregation responses to ADP. In contrast, CD39L1/NTPDase2, a preferential nucleoside triphosphatase, activates platelets by preferentially converting ATP to ADP, the major agonist of platelet P2 receptors. Spatial and temporal expression of NTPDases in the vasculature appears to control platelet activation, thrombus size and stability by regulating phosphohydrolytic activity and consequent P2 receptor signaling. Constitutively circulating microparticles appear to be associated with functional NTPDases, and accumulation of these at sites of vascular injury might influence local thrombus formation and evolution. The phenotype of the cd39-null mouse is in keeping with disordered thromboregulation with heightened susceptibility to inflammatory vasculary reactions, increased permeability and high levels of tissue fibrin. Paradoxically, these mutant mice also exhibit a bleeding phenotype with differential platelet P2Y1 desensitization. Over-expression of CD39 at sites of vascular injury and inflammation by adenoviral vectors, by transgenesis or by the use of pharmacological modalities with soluble derivatives has been shown to have major potential in several animal models tested to date. Future clinical applications will involve the development of new therapeutic strategies to various inflammatory vascular diseases and in transplantation.

Effects of SolCD39, a novel inhibitor of Platelet Aggregation, on Platelet Deposition and Aggregation after PTCA in a Porcine Model.

INTRODUCTION: This study evaluated CD39 in a porcine model of balloon angioplasty and in plasma of patients undergoing percutaneous intervention. CD39 (E-NTPDase1), is the endothelial ecto-ADPase inhibiting platelet function via hydrolysis of released platelet ADP. METHODS AND RESULTS: A recombinant soluble form of CD39 (solCD39) given intravenously to pigs had an elimination half life of 5--7 days, increased the bleeding time to an extent similar to aspirin, and inhibits platelet aggregation by>90%. Platelet counts and clot retraction remained normal following solCD39 administration. In a pig model of acute coronary balloon injury, solCD39 resulted in non-statistically significant decreases in platelet (7.7+/-1.4 versus 11.7+/- 3.4) and fibrin (3.5+/- 0.4 versus 4.2+/- 0.7) deposition ratios. Adding ex vivo to human platelet rich plasma (PRP) solCD39 produced nearly 100% inhibition of ADP-induced platelet aggregation. A dose-response effect of solCD39 on platelet aggregation induced by collagen or a thrombin receptor activating peptide (TRAP(SFLLRN)) was noted in PRP obtained from volunteers and patients receiving aspirin, clopidogrel or ticlopidine. SolCD39 also provided additional and complete inhibition of TRAP-induced platelet aggregation in PRP from patients who had received abciximab, aspirin and clopidogrel. CONCLUSIONS: SolCD39, a novel inhibitor of platelet activation and recruitment with a relatively long half-life appears to be well tolerated and is a potent inhibitor of ADP-, collagen-, or TRAP-induced platelet activation. Its potential use in percutaneous coronary intervention requires further study. ABBREVIATED ABSTRACT: E-NTPDase1/CD39 is the endothelial ecto-ADPase responsible for inhibition of platelet function. A recombinant soluble form (solCD39) had an elimination half life of 5-7 days in pigs, elevated bleeding times similar to aspirin, did not affect clot retraction, and inhibited platelet aggregation by > 90%. When combined with standard heparin therapy in a pig model of acute coronary balloon injury, solCD39 resulted in a trend toward a decrease in platelet and fibrin deposition. SolCD39 added ex vivo to human platelet rich plasma yielded nearly 100% inhibition of ADP-induced platelet aggregation and provided further inhibition when combined with standard therapy.

Ectonucleotidases of CD39 family modulate vascular inflammation and thrombosis in transplantation.

Transplantation results in exposure of the graft vasculature to warm and cold ischemia, followed by perfusion by circulating blood constituents and obligatory oxidant stress. Further graft injury occurs as consequences of acute humoral cellular rejection or chronic transplant vasculopathy, or both. Extracellular nucleotide stimulation of purinergic type 2 (P2) receptors are key components of platelet, endothelial cell (EC), and leukocyte activation resulting in vascular thrombosis and inflammation in vivo. CD39, the prototype nucleoside triphosphate diphosphohydrolase (NTPDase-1) is highly expressed on endothelium; in contrast, CD39L1/NTPDase-2 (a preferential adenosine triphosphatase [ATPase]) is found on vascular adventitial cells. Both ectoenzymes influence thrombogenesis by the regulated hydrolysis of extracellular nucleotides that differentially regulate P2-receptor activity and function in platelets and vascular cells. The intracytoplasmic domains of NTPDase-1 may also independently influence cellular activation and proliferation. NTPDase activity is substantively lost in the vasculature of injured or rejected grafts. A role for NTPDase-1 in thromboregulation has been validated by generation of mutant mice either null for cd39 or overexpressing human CD39. Administration of soluble NTPDase or induction of CD39 by adenoviral vectors, or both, are also of benefit in several models of transplantation. Administration of soluble CD39 or targeted expression may have future therapeutic application in transplantation-associated and other vascular diseases.