Phospholipase D in human platelets: presence of isoenzymes and participation of autocrine stimulation during thrombin activation

Platelets. 2008 May;19(3):211-24. doi: 10.1080/09537100701777329.

Abstract

Phospholipase D (PLD), which hydrolyzes phosphatidylcholine to phosphatidic acid (PA) and choline, is present in human platelets. Thrombin and other agonists have been shown to activate PLD but the precise mechanisms of activation and PLDs role in platelet activation remains unclear. We measured thrombin-stimulated PLD activity in platelets as formation of phosphatidylethanol. Since no specific PLD inhibitors exist, we investigated possible roles for PLD in platelets by correlating PLD activity with platelet responses such as thrombin-mediated secretion and F-actin formation (part of platelet shape change). Extracellular Ca2+ potentiated thrombin-stimulated PLD, but did not stimulate PLD in the absence of thrombin. Thrombin-induced PLD activity was enhanced by secreted ADP and binding of fibrinogen to its receptors. In contrast to others, we also found a basal PLD activity. Comparison of time courses and dose responses of platelets with PLD showed many points of correlation between PLD activation and lysosomal secretion and F-actin formation. The finding of different PLD activities suggested that different PLD isoenzymes exist in platelets as reported for other cells. Here we present evidence for the presence of both PLD1 and PLD2 in platelets by use of specific antibodies with immunoblotting and immunohistochemistry. Both isoforms were randomly localized in resting platelets, but became rapidly translocated to the proximity of the plasma membrane upon thrombin stimulation, thus indicating a role for PLD in platelet activation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actins / metabolism
  • Adenosine Diphosphate / metabolism
  • Antibodies, Monoclonal / metabolism
  • Autocrine Communication / drug effects
  • Blood Platelets / enzymology*
  • Blood Platelets / ultrastructure
  • Bridged Bicyclo Compounds, Heterocyclic
  • Calcium / metabolism
  • Cell Membrane
  • Fatty Acids, Unsaturated
  • Glycerophospholipids / metabolism
  • Humans
  • Hydrazines / pharmacology
  • Isoenzymes / metabolism
  • Lysosomes / metabolism
  • Oligopeptides / pharmacology
  • Phospholipase D / metabolism*
  • Platelet Activation / physiology*
  • Platelet Aggregation Inhibitors / pharmacology
  • Thrombin / metabolism*
  • Thrombin / pharmacology

Substances

  • Actins
  • Antibodies, Monoclonal
  • Bridged Bicyclo Compounds, Heterocyclic
  • Fatty Acids, Unsaturated
  • Glycerophospholipids
  • Hydrazines
  • Isoenzymes
  • Oligopeptides
  • Platelet Aggregation Inhibitors
  • phosphatidylethanol
  • Adenosine Diphosphate
  • SQ 29548
  • arginyl-glycyl-aspartyl-serine
  • phospholipase D2
  • Phospholipase D
  • phospholipase D1
  • Thrombin
  • Calcium