@phdthesis{Thielmann2014,
  author    = {Thielmann, Ina},
  title     = {Function and regulation of phospholipase D in blood platelets: in vitro and in vivo studies in mice},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-99179},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2014},
  abstract  = {Summary Platelet activation and aggregation are crucial for primary hemostasis but can also result in occlusive thrombus formation. Agonist induced platelet activation involves different signaling pathways leading to the activation of phospholipases (PL) which produce second messengers. While the role of PLCs in platelet activation is well established, less is known about the relevance of PLDs. In the current study, the function and regulation of PLD in platelets was investigated using genetic and pharmacological approaches. In the first part of this thesis, adhesion, activation and aggregation of platelets from mice lacking PLD2 or both PLD1 and PLD2 were analyzed in vitro and in vivo. While the absence of PLD2 resulted in slightly reduced PLD activity in platelets, it had no detectable effect on the platelet function in vitro and in vivo. However, the combined deficiency of both PLD isoforms resulted in defective alpha-granule release and protection in a model of ferric chloride induced arteriolar thrombosis, effects that were not observed in mice lacking only one PLD isoform. These results revealed, for the first time, redundant roles of PLD1 and PLD2 in platelet alpha-granule secretion and indicate that this may be relevant for pathological thrombus formation. Thus, PLD might represent a promising target for antithrombotic therapy. Thus, this hypothesis was tested more directly in the second part of this thesis. The effects of pharmacological inhibition of PLD activity on hemostasis, thrombosis and thrombo-inflammatory brain infarction in mice were assessed. Treatment of platelets with the reversible, small molecule PLD inhibitor 5-Fluoro-2-indolyl des-chlorohalopemide (FIPI) led to a specific blockade of PLD activity that was associated with reduced -granule release and integrin activation. Mice that received FIPI at a dose of 3 mg/kg displayed reduced occlusive thrombus formation upon chemical injury of carotid arteries or mesenterial arterioles. Similarly, FIPI-treated mice had smaller infarct sizes and significantly better motor and neurological function 24 hours after transient middle cerebral artery occlusion. This protective effect was not associated with major intracerebral hemorrhage or prolonged tail bleeding times. Thus, pharmacological PLD inhibition might represent a safe therapeutic strategy to prevent arterial thrombosis or ischemic stroke. After revealing a central role for PLD in thrombo-inflammation, the regulation of PLD activity in platelets was analyzed in the last part of the thesis. Up to date, most studies made use of inhibitors potentially exerting off-target effects and consequently PLD regulation is discussed controversially. Therefore, PLD activity in mice genetically lacking potential modulators of PLD activity was determined to address these controversies. These studies revealed that PLD is tightly regulated during initial platelet activation. While integrin outside-in signaling and Gi signaling was dispensable for PLD activation, it was found that PLC dependent pathways were relevant for the regulation of PLD enzyme activity.},
  subject      = {Phospholipase D},
  language  = {en}
}