Exogenous diacylglycerols synergize with PAF with human platelets, but inhibit PAF-induced responses of rabbit platelets.
To investigate whether diacylglycerol (DAG) has a role in reversible platelet aggregation induced by low concentrations of platelet-activating factor (PAF), we attempted to use the DAG kinase inhibitor, R59022, to prevent rapid conversion of DAG to phosphatidic acid. However, we found that R59022 inhibited the binding of [3H]PAF to human and rabbit platelets and to rabbit platelet membranes. We then investigated whether exogenous, cell-penetrating DAGs (1,2-dihexanoyl-sn-glycerol (DHG) and 1-oleoyl-2-acetyl-sn-glycerol (OAG)) act synergistically with low concentrations of PAF that alone induce only reversible aggregation. Platelets were isolated and labeled with [14C]serotonin. DHG (25-75 microM) caused slow, weak aggregation and some release of [14C]serotonin with human, but not rabbit, platelets. OAG (25-75 microM) did not aggregate either species' platelets. Phosphorylation of pleckstrin by DHG was more transient in rabbit platelets than previously observed with human platelets. Both DHG and OAG synergistically potentiated PAF-induced aggregation of human platelets, but, paradoxically, concurrently inhibited the PAF-induced increase in intracellular Ca2+ ([Ca2+]i): potentiation decreased upon incubation with DAGs before PAF addition. In contrast, DHG strongly inhibited PAF-induced aggregation of rabbit platelets; inhibition decreased upon preincubation. OAG, added with PAF, slightly potentiated aggregation of rabbit platelets: upon preincubation, OAG progressively inhibited. Effects of DHG and OAG on PAF-induced increases in [Ca2+]i in rabbit platelets followed a similar pattern; thus, with rabbit platelets, inhibition of the [Ca2+]i increase may at least partially account for inhibition of PAF-induced aggregation by exogenous DAGs. Results with human platelets are consistent with stimulation of protein kinase C by DAGs, and then metabolism of DAGs and/or negative feedback by DAGs, but results with rabbit platelets indicate both an unexpected species difference and a difference between the effects of DHG and OAG on PAF-induced platelet aggregation.
Packham
M A
MA
Department of Biochemistry, University of Toronto, Ontario, Canada. m.packham@utoronto.ca
Ruben
D H
DH
Rand
M L
ML
D000818:Animals; D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000187:drug effects; D019852:Diacylglycerol Kinase; D004075:Diglycerides / Q000097:blood / Q000493:pharmacokinetics / Q000494:pharmacology*; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors; D010972:Platelet Activating Factor / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000494:pharmacology*; D010974:Platelet Aggregation / Q000187:drug effects; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D011744:Pyrimidinones / Q000494:pharmacology; D011817:Rabbits; D013844:Thiazoles / Q000494:pharmacology
D004075:Diglycerides; D004791:Enzyme Inhibitors; D010972:Platelet Activating Factor; D010975:Platelet Aggregation Inhibitors; D011744:Pyrimidinones; D013844:Thiazoles; C044296:1,2-dihexanoylglycerol; C038237:1-oleoyl-2-acetylglycerol; C047505:R 59022; D017853:Phosphotransferases (Alcohol Group Acceptor); D019852:Diacylglycerol Kinase