In mid-March of this year, I was fortunate to attend a pivotal scientific meeting on climate chan... more In mid-March of this year, I was fortunate to attend a pivotal scientific meeting on climate change-the 2009 International Alliance of Research Universities Climate Congress in Copenhagen.'In an act of symbolic foreshadowing, the summit was hosted in the same ...
Extracellular adenosine triphosphate (ATP) regulates platelet reactivity by way of direct action ... more Extracellular adenosine triphosphate (ATP) regulates platelet reactivity by way of direct action on platelet purinergic receptors or by hydrolysis to adenosine diphosphate (ADP). Subsequent metabolism of ATP and ADP to adenosine monophosphate (AMP) and adenosine inhibits platelet aggregation. Endothelial cell membrane-bound ecto-ATP/ADPase (CD39, E-NTP-Dase1) is thought to be the main regulator of platelet responsiveness. However, the findings in studies of CD39-knockout mice imply that nucleotidase(s) in plasma regulates circulating adenine nucleotides levels. Understanding extracellular ATP metabolism by CD39 and plasma nucleotidases is therefore important. In this study, ␣-phosphorus 32-and ␥-phosphorus 32labeled ATP were rapidly metabolized directly to AMP and pyrophosphate in human plasma at pH 7.4, suggesting the presence of pyrophosphatase/phosphodiesterase-like activity. A specific phosphodiesterase substrate, p-nitrophenol-5-TMP (p-Nph-5-TMP), was readily hydrolyzed in human plasma. The antiaggregatory action of ,␥-methylene-ATP (AMPPCP) (5 mol/L) was blocked by DMPX, an adenosine-receptor antagonist, suggesting that in plasma, AMPPCP was metabolized to AMP and adenosine. Recombinant soluble CD39 (solCD39) was used to assess the role of CD39 in ATP metabolism. As little as 0.25 g/mL of solCD39 inhibited ADP-induced platelet aggregation. However, in the presence of ADP-free ATP (10 mol/L), solCD39 induced platelet aggregation in a dose-dependent manner. Because AMPPCP could not substitute for ATP in solCD39-stimulated platelet aggregation, it is likely that ADP formation from ATP was required. Endogenous CD39 may thus have a hemostatic function by promoting ADP formation from released ATP, in addition to its antiaggregatory properties. A plasma nucleotidase hydrolyzes ATP directly to AMP. This prevents ADP accumulation and generates adenosine, a potent, locally acting inhibitor of platelet reactivity. The presence of both endothelial CD39 and plasma nucleotidase appears to be important in the maintenance of normal hemostasis and prevention of excessive platelet responsiveness. (J Lab Clin Med 2002;140: 166-75) Abbreviations: ADP ϭ adenosine diphosphate; AMP ϭ adenosine monophosphate; AMPPCP ϭ ,␥-methylene-adenosine triphosphate; ATP ϭ adenosine triphosphate; AUC ϭ area under the curve; DEAE ϭ diethylaminoethyl [cellulose]; DMPX ϭ 3,7-dimethyl-1-propargylxanthine; EDTA ϭ ethylenediaminetetraacetate; solCD39 ϭ soluble CD39; p-Nph-5Ј-TMP ϭ p-nitrophenol-5Ј-thymidine monophosphate; p-Nph-PO 4 ϭ p-Nph-phosphate; PPP ϭ platelet-poor plasma; PRP ϭ platelet-rich plasma; SEM ϭ standard error of the mean From the
In mid-March of this year, I was fortunate to attend a pivotal scientific meeting on climate chan... more In mid-March of this year, I was fortunate to attend a pivotal scientific meeting on climate change-the 2009 International Alliance of Research Universities Climate Congress in Copenhagen.'In an act of symbolic foreshadowing, the summit was hosted in the same ...
Extracellular adenosine triphosphate (ATP) regulates platelet reactivity by way of direct action ... more Extracellular adenosine triphosphate (ATP) regulates platelet reactivity by way of direct action on platelet purinergic receptors or by hydrolysis to adenosine diphosphate (ADP). Subsequent metabolism of ATP and ADP to adenosine monophosphate (AMP) and adenosine inhibits platelet aggregation. Endothelial cell membrane-bound ecto-ATP/ADPase (CD39, E-NTP-Dase1) is thought to be the main regulator of platelet responsiveness. However, the findings in studies of CD39-knockout mice imply that nucleotidase(s) in plasma regulates circulating adenine nucleotides levels. Understanding extracellular ATP metabolism by CD39 and plasma nucleotidases is therefore important. In this study, ␣-phosphorus 32-and ␥-phosphorus 32labeled ATP were rapidly metabolized directly to AMP and pyrophosphate in human plasma at pH 7.4, suggesting the presence of pyrophosphatase/phosphodiesterase-like activity. A specific phosphodiesterase substrate, p-nitrophenol-5-TMP (p-Nph-5-TMP), was readily hydrolyzed in human plasma. The antiaggregatory action of ,␥-methylene-ATP (AMPPCP) (5 mol/L) was blocked by DMPX, an adenosine-receptor antagonist, suggesting that in plasma, AMPPCP was metabolized to AMP and adenosine. Recombinant soluble CD39 (solCD39) was used to assess the role of CD39 in ATP metabolism. As little as 0.25 g/mL of solCD39 inhibited ADP-induced platelet aggregation. However, in the presence of ADP-free ATP (10 mol/L), solCD39 induced platelet aggregation in a dose-dependent manner. Because AMPPCP could not substitute for ATP in solCD39-stimulated platelet aggregation, it is likely that ADP formation from ATP was required. Endogenous CD39 may thus have a hemostatic function by promoting ADP formation from released ATP, in addition to its antiaggregatory properties. A plasma nucleotidase hydrolyzes ATP directly to AMP. This prevents ADP accumulation and generates adenosine, a potent, locally acting inhibitor of platelet reactivity. The presence of both endothelial CD39 and plasma nucleotidase appears to be important in the maintenance of normal hemostasis and prevention of excessive platelet responsiveness. (J Lab Clin Med 2002;140: 166-75) Abbreviations: ADP ϭ adenosine diphosphate; AMP ϭ adenosine monophosphate; AMPPCP ϭ ,␥-methylene-adenosine triphosphate; ATP ϭ adenosine triphosphate; AUC ϭ area under the curve; DEAE ϭ diethylaminoethyl [cellulose]; DMPX ϭ 3,7-dimethyl-1-propargylxanthine; EDTA ϭ ethylenediaminetetraacetate; solCD39 ϭ soluble CD39; p-Nph-5Ј-TMP ϭ p-nitrophenol-5Ј-thymidine monophosphate; p-Nph-PO 4 ϭ p-Nph-phosphate; PPP ϭ platelet-poor plasma; PRP ϭ platelet-rich plasma; SEM ϭ standard error of the mean From the
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