Role of a novel soluble nucleotide phospho-hydrolase from sheep plasma in inhibition of platelet reactivity: hemostasis, thrombosis, and vascular biology.
Ecto- and exoenzymes that metabolize extracellular adenosine diphosphate (ADP), the major promoter of platelet activation and recruitment, are of potential clinical importance because they can metabolically prevent excessive thrombus growth. An ecto-ADPase (CD39, NTPDase1) has been identified on endothelial cells. We demonstrate that ADP and adenosine triphosphate (ATP) are rapidly metabolized to adenosine monophosphate (AMP) in sheep plasma at pH 7.4. This hydrolysis is sensitive to P(1), P(5)-di-(adenosine-5') pentaphosphate (Ap(5)A), and ethylene glycol bis (beta-aminoethyl ether) - N, N, N(-), N(-) tetra-acetate (EGTA) but insensitive to tetramisole (an alkaline phosphatase inhibitor). A specific phosphodiesterase substrate, p -nitrophenol-5'-thymidine monophosphate (TMP) (p -Nph-5'-TMP), was readily hydrolyzed in sheep plasma at a rate of approximately 0.25 nmol/min/mg protein, and this hydrolysis was inhibited by ADP, ATP, and Ap(5)A. Furthermore, 200-fold purified p -Nph-5'-TMP-hydrolyzing activity also hydrolyzed ATP and ADP directly to AMP. When ADP was preincubated in plasma, its ability to induce platelet aggregation was inhibited in a time-dependent manner. This effect was abolished by Ap(5)A. The inhibitory effects on platelet aggregation correlated with hydrolysis of the ADP in plasma. These data suggest that the endogenous soluble plasma phosphohydrolase metabolizes ATP and ADP by means of cleavage of the alpha-beta-phosphodiester bond of nucleoside 5'-phosphate derivatives. This novel biochemical activity inhibits platelet reactivity through hydrolysis of extracellular nucleotides released by activated platelets during (patho)physiological processes, serving a homeostatic and antithrombotic function in vivo.
Birk
Alex V
AV
Division of Hematology and Medical Oncology, Department of Medicine, Weill Medical College of Cornell University, New York, New York 10021, USA.
Bubman
Darya
D
Broekman
M Johan
MJ
Robertson
Hugh D
HD
Drosopoulos
Joan H F
JH
Marcus
Aaron J
AJ
Szeto
Hazel H
HH
D000244:Adenosine Diphosphate / Q000378:metabolism / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001792:Blood Platelets / Q000502:physiology*; D015226:Dinucleoside Phosphates / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005343:Fibrinolytic Agents; D006487:Hemostasis; D006868:Hydrolysis; D007700:Kinetics; D009596:Nitrophenols / Q000378:metabolism; D009708:Nucleotidases / Q000037:antagonists & inhibitors / Q000097:blood*; D010974:Platelet Aggregation / Q000187:drug effects; D010975:Platelet Aggregation Inhibitors; D012756:Sheep; D013927:Thrombosis; D013938:Thymidine Monophosphate / Q000378:metabolism
D015226:Dinucleoside Phosphates; D004791:Enzyme Inhibitors; D005343:Fibrinolytic Agents; D009596:Nitrophenols; D010975:Platelet Aggregation Inhibitors; D013938:Thymidine Monophosphate; C012275:P(1),P(5)-di(adenosine-5'-)pentaphosphate; D004533:Egtazic Acid; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D009708:Nucleotidases; C030287:nucleotidase; C024836:4-nitrophenol