TY - JOUR
T1 - Activation of human platelets by 2-arachidonoylglycerol
T2 - Role of PKC in NO/cGMP pathway modulation
AU - Signorello, Maria Grazia
AU - Giacobbe, Enrica
AU - Segantin, Alessia
AU - Avigliano, Luciana
AU - Sinigaglia, Fabiola
AU - Maccarrone, Mauro
AU - Leoncini, Giuliana
PY - 2011/8
Y1 - 2011/8
N2 - We demonstrated that the endocannabinoid 2-arachidonoylglycerol (2-AG) activated dose-dependently washed human platelets and increased intracellular calcium levels. Moreover 2-AG activated protein kinase C measured as p47pleckstrin phosphorylation. These parameters were prevented by the tromboxane A
2 receptor antagonist SQ29548, by phospholipase C pathway (U73122) and protein kinase C (GF109203X) inhibitors. No effect on 2-AG-induced platelet activation and calcium elevation in the presence of inhibitors of fatty acid amide hydrolase or monoacylglycerol lipase was observed. In addition we have shown that 2-AG dose-dependently increased NO and cGMP levels. These effects were abolished by U73122, GF109203X, EGTA and the intracellular calcium chelator BAPTA/AM. Moreover, 2-AG enhanced eNOS activity through the phosphorylation of its positive regulatory residue ser1177 and by dephosphorylation of the negative one thr495. The eNOS ser1177 phosphorylation was inhibited by U73122 and GF109203X but it was unaffected by the PI3K/AKT pathway inhibitors LY294002 and MK2206. The dephosphorylation of thr495 was reversed by low concentrations of calyculin A. Taken together these data suggest that 2-AG behaves as a true platelet agonist stimulating PKC activation and calcium elevation. Likely 2-AG can modulate platelet activation by increasing NO levels through eNOS activation.
AB - We demonstrated that the endocannabinoid 2-arachidonoylglycerol (2-AG) activated dose-dependently washed human platelets and increased intracellular calcium levels. Moreover 2-AG activated protein kinase C measured as p47pleckstrin phosphorylation. These parameters were prevented by the tromboxane A
2 receptor antagonist SQ29548, by phospholipase C pathway (U73122) and protein kinase C (GF109203X) inhibitors. No effect on 2-AG-induced platelet activation and calcium elevation in the presence of inhibitors of fatty acid amide hydrolase or monoacylglycerol lipase was observed. In addition we have shown that 2-AG dose-dependently increased NO and cGMP levels. These effects were abolished by U73122, GF109203X, EGTA and the intracellular calcium chelator BAPTA/AM. Moreover, 2-AG enhanced eNOS activity through the phosphorylation of its positive regulatory residue ser1177 and by dephosphorylation of the negative one thr495. The eNOS ser1177 phosphorylation was inhibited by U73122 and GF109203X but it was unaffected by the PI3K/AKT pathway inhibitors LY294002 and MK2206. The dephosphorylation of thr495 was reversed by low concentrations of calyculin A. Taken together these data suggest that 2-AG behaves as a true platelet agonist stimulating PKC activation and calcium elevation. Likely 2-AG can modulate platelet activation by increasing NO levels through eNOS activation.
KW - 2-Arachidonoylglycerol
KW - Enos phosphorylation/dephosphorylation
KW - Human platelets
KW - Nitric oxide
KW - PKC
KW - PP1
KW - PP2A
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U2 - 10.2174/156720211796558041
DO - 10.2174/156720211796558041
M3 - Article
C2 - 21675954
AN - SCOPUS:80051533854
VL - 8
SP - 200
EP - 209
JO - Current Neurovascular Research
JF - Current Neurovascular Research
SN - 1567-2026
IS - 3
ER -