apyrase and Myocardial-Ischemia

Extracellular nucleotides play a critical role in vascular thrombosis and inflammation. Alterations in purinergic extracellular nucleotide concentrations activate pathways that result in platelet degranulation and aggregation, and endothelial and leukocyte activation and recruitment. CD39, the dominant vascular nucleotidase, hydrolyzes ATP and ADP to provide the substrate for generation of the anti-inflammatory and antithrombotic mediator adenosine. The purinergic signaling system, with CD39 at its center, plays an important role in modulating vascular homeostasis and the response to vascular injury, as seen in clinically relevant diseases such as stroke, ischemia-reperfusion injury, and pulmonary hypertension. A growing body of knowledge of the purinergic signaling pathway implicates CD39 as a critical modulator of vascular thrombosis and inflammation. Therapeutic strategies targeting CD39 offer promising opportunities in the management of vascular thromboinflammatory diseases.

Topics: Antigens, CD; Apyrase; Atherosclerosis; Endothelium, Vascular; Humans; Hypertension, Pulmonary; Inflammation; Myocardial Ischemia; Myocardial Reperfusion Injury; Signal Transduction; Stroke; Thrombosis; Vasculitis

Thrombus formation at sites of disrupted atherosclerotic plaques is a leading cause of death and disability worldwide. Although the platelet is now recognized to be a central regulator of thrombus formation, development of antiplatelet reagents that selectively target thrombosis over hemostasis represents a challenge. Existing prophylactic antiplatelet therapies are centered on the use of aspirin, an irreversible cyclooxygenase inhibitor, and a thienopyridine such as clopidogrel, which inactivates the adenosine diphosphate-stimulated P2Y(12) receptor. Although these compounds are widely used and have beneficial effects for patients, their antithrombotic benefit is complicated by an elevated bleeding risk and substantial or partial "resistance." Moreover, combination therapy with these two drugs increases the hemorrhagic risk even further. This review explores the possibility of inhibiting the platelet-surface ionotropic P2X(1) receptor and/or elevating CD39/NTPDase1 activity as new therapeutic approaches to reduce overall platelet reactivity and recruitment of surrounding platelets at prothrombotic locations. Because both proteins affect platelet activation at an early stage in the events leading to thrombosis but are less crucial in hemostasis, they provide new strategies to widen the cardiovascular therapeutic window without compromising safety.

Topics: Adenosine Triphosphate; Antigens, CD; Apyrase; Aspirin; Biomedical Research; Cardiovascular Diseases; Cerebral Hemorrhage; Clinical Trials as Topic; Clopidogrel; Cyclooxygenase Inhibitors; Drug Therapy, Combination; Endothelium, Vascular; Humans; Myocardial Ischemia; Platelet Aggregation Inhibitors; Receptors, Purinergic P2; Receptors, Purinergic P2X; Receptors, Purinergic P2Y12; Ticlopidine; Treatment Outcome

Blood levels of extracellular nucleotides (e.g. ATP) are greatly increased during heart ischaemia, but, despite the presence of their specific receptors on cardiomyocytes (both P2X and P2Y subtypes), their effects on the subsequent myocardial damage are still unknown. In this study, we aimed at investigating the role of ATP and specific P2 receptors in the appearance of cell injury in a cardiac model of ischaemic/hypoxic stress. Cells were maintained in a modular incubator chamber in a controlled humidified atmosphere of 95% N(2) for 16 hrs in a glucose-free medium. In this condition, we detected an early increase in the release of ATP in the culture medium, which was followed by a massive increase in the release of cytoplasmic histone-associated-DNA-fragments, a marker of apoptosis. Addition of either apyrase, which degrades extracellular ATP, or various inhibitors of ATP release via connexin hemichannels fully abolished ischaemic/hypoxic stress-associated apoptosis. To dissect the role of specific P2 receptor subtypes, we used a combined approach: (i) non-selective and, when available, subtype-selective P2 antagonists, were added to cardiomyocytes before ischaemic/hypoxic stress; (ii) selected P2 receptors genes were silenced via specific small interfering RNAs. Both approaches indicated that the P2Y(2) and P2χ(7) receptor subtypes are directly involved in the induction of cell death during ischaemic/hypoxic stress, whereas the P2Y(4) receptor has a protective effect. Overall, these findings indicate a role for ATP and its receptors in modulating cardiomyocyte damage during ischaemic/hypoxic stress.

Topics: Adenosine Triphosphate; Animals; Apoptosis; Apyrase; Cell Line; Cells, Cultured; DNA Fragmentation; Gene Silencing; Histones; Mice; Myocardial Ischemia; Myocytes, Cardiac; Purinergic P2 Receptor Antagonists; Receptors, Purinergic P2

A clopidogrel loading dose administered during stenting attenuates inflammation marker release. However, less is known of the anti-inflammatory effect of clopidogrel maintenance therapy. Platelet reactivity to adenosine diphosphate and inflammation markers were measured in 110 consecutive patients (69 clopidogrel-naive patients and 41 patients receiving long-term clopidogrel therapy for >6 months) before nonemergent stenting by turbidimetric aggregometry and flow cytometry and multianalyte profiling, respectively. All patients were treated with aspirin. Prestenting adenosine diphosphate-induced platelet aggregation, P-selectin, and activated glycoprotein IIb/IIIa expression were lower in patients receiving long-term clopidogrel therapy compared with the clopidogrel-naive group (p <0.001), accompanied by lower levels of selected inflammation markers (p < or = 0.05). Additionally, there were strong correlations between platelet aggregation and flow cytometric measurements (p < or = 0.04) and between specific inflammation markers (p < or = 0.02). In conclusion, in addition to markedly lowering platelet reactivity to adenosine diphosphate, long-term clopidogrel therapy is associated with an anti-inflammatory effect.

Topics: Aged; Angioplasty, Balloon, Coronary; Apyrase; Blood Platelets; Clopidogrel; Female; Flow Cytometry; Humans; Inflammation; Male; Middle Aged; Myocardial Ischemia; P-Selectin; Pilot Projects; Platelet Aggregation; Platelet Aggregation Inhibitors; Platelet Glycoprotein GPIIb-IIIa Complex; Stents; Ticlopidine

Focal mechanical stimulation of single neonatal mouse cardiac myocytes in culture induced intercellular Ca(2+) waves that propagated with mean velocities of approximately 14 micrometer/s, reaching approximately 80% of the cells in the field. Deletion of connexin43 (Cx43), the main cardiac gap junction channel protein, did not prevent communication of mechanically induced Ca(2+) waves, although the velocity and number of cells communicated by the Ca(2+) signal were significantly reduced. Similar effects were observed in wild-type cardiac myocytes treated with heptanol, a gap junction channel blocker. Fewer cells were involved in intercellular Ca(2+) signaling in both wild-type and Cx43-null cultures in the presence of suramin, a P(2)-receptor blocker; blockage was more effective in Cx43-null than in wild-type cells. Thus gap junction channels provide the main pathway for communication of slow intercellular Ca(2+) signals in wild-type neonatal mouse cardiac myocytes. Activation of P(2)-receptors induced by ATP release contributes a secondary, extracellular pathway for transmission of Ca(2+) signals. The importance of such ATP-mediated Ca(2+) signaling would be expected to be enhanced under ischemic conditions, when release of ATP is increased and gap junction channels conductance is significantly reduced.

Topics: Adenosine Triphosphate; Animals; Animals, Newborn; Antineoplastic Agents; Apyrase; Calcium Signaling; Cell Communication; Cells, Cultured; Connexin 43; Extracellular Space; Gap Junctions; Image Processing, Computer-Assisted; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle Fibers, Skeletal; Myocardial Ischemia; Myocardium; Paracrine Communication; Receptors, Purinergic; Stress, Mechanical; Suramin