Nitrolinoleate inhibits platelet activation by attenuating calcium mobilization and inducing phosphorylation of vasodilator-stimulated phosphoprotein through elevation of cAMP.
Reactive species formed from nitric oxide (NO) nitrate unsaturated fatty acids such as linoleate (LA) to nitrated derivatives including nitrolinoleate (LNO(2)). The effect of LNO(2) on human platelets was examined to define how nitrated lipids might behave in vivo. LNO(2), but not LA or 3-nitrotyrosine, dose dependently (0.5-10 microm) inhibited thrombin-mediated aggregation of washed human platelets, with concomitant attenuation of P-selectin expression and selective phosphorylation of VASP at the cAMP-dependent protein kinase selective site, serine 157. LNO(2) caused slight mobilization of calcium (Ca(2+)) from intracellular stores but significantly inhibited subsequent thrombin-stimulated Ca(2+) elevations. LNO(2) did not elevate platelet cGMP, and its effects were not blocked with inhibitors of NO signaling (oxyhemoglobin, 1H-[1,2,4]oxadiazole[4,3-a]quinoxalin-1-one. 2-fold elevations in cAMP were found following LNO(2) treatment of platelets, and the adenylyl cyclase inhibitors 2',5'-dideoxyadenosine and SQ22536 partially restored thrombin-stimulated aggregation. Finally, LNO(2) significantly inhibited cAMP hydrolysis to AMP by platelet lysates. These data implicate cAMP in the anti-aggregatory action of LNO(2). The platelet inhibitory actions of LNO(2) indicate that nitration reactions that occur following NO generation in an oxidizing environment can alter the activity of lipids and lend insight into mechanisms by which NO-derived species may modulate the progression of vascular injury.
Coles
Barbara
B
Department of Medical Biochemistry, University of Wales College of Medicine, Heath Park, Cardiff CF4 4XN, United Kingdom.
Bloodsworth
Allison
A
Eiserich
Jason P
JP
Coffey
Marcus J
MJ
McLoughlin
Rachel M
RM
Giddings
John C
JC
Lewis
Malcolm J
MJ
Haslam
Richard J
RJ
Freeman
Bruce A
BA
O'Donnell
Valerie B
VB
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D001792:Blood Platelets / Q000187:drug effects / Q000502:physiology; D002118:Calcium / Q000097:blood; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D000242:Cyclic AMP / Q000378:metabolism*; D006152:Cyclic GMP / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D019787:Linoleic Acid / Q000494:pharmacology; D008041:Linoleic Acids / Q000138:chemical synthesis / Q000494:pharmacology*; D009574:Nitro Compounds / Q000138:chemical synthesis / Q000494:pharmacology*; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D015539:Platelet Activation / Q000187:drug effects / Q000502:physiology*; D010974:Platelet Aggregation / Q000187:drug effects / Q000502:physiology; D013917:Thrombin / Q000494:pharmacology; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D014665:Vasodilator Agents / Q000494:pharmacology*
D008041:Linoleic Acids; D009574:Nitro Compounds; D010750:Phosphoproteins; D014665:Vasodilator Agents; C002744:3-nitrotyrosine; D014443:Tyrosine; D019787:Linoleic Acid; D000242:Cyclic AMP; D013917:Thrombin; D006152:Cyclic GMP; D002118:Calcium; D015056:1-Methyl-3-isobutylxanthine