title: LDL oxidation by platelets propagates platelet activation via an oxidative stress-mediated mechanism.
abstract: OBJECTIVE
  Platelets generate oxidized LDL (ox-LDL) via NOX2-derived oxidative stress. We investigated if once generated by activated platelets ox-LDL can propagate platelet activation.
  
  
  METHODS
  Experiments were performed in platelets from healthy subjects (HS), hyper-cholesterolemic patients and patients with NOX2 hereditary deficiency.
  
  
  RESULTS
  Agonist-stimulated platelets from HS added with LDL were associated with a dose-dependent increase of reactive oxidant species and ox-LDL. Agonist-stimulated platelets from HS added with a fixed dose of LDL (57.14 μmol/L) or added with homogenized human atherosclerotic plaque showed enhanced ox-LDL formation (approximately +50% and +30% respectively), which was lowered by a NOX2 inhibitor (approximately -35% and -25% respectively). Compared to HS, ox-LDL production was more pronounced in agonist-stimulated platelet rich plasma (PRP) from hyper-cholesterolemic patients but was almost absent in PRP from NOX2-deficient patients. Platelet aggregation and 8-iso-PGF2α-ΙΙΙ formation increased in LDL-treated washed platelets (+42% and +53% respectively) and PRP (+31% and +53% respectively). Also, LDL enhanced platelet-dependent thrombosis at arterial shear rate (+33%) but did not affect platelet activation in NOX2-deficient patients. Platelet activation by LDL was significantly inhibited by CD36 or LOX1 blocking peptides, two ox-LDL receptor antagonists, or by a NOX2 inhibitor. LDL-added platelets showed increased p38MAPK (+59%) and PKC (+51%) phosphorylation, p47(phox) translocation to platelet membrane (+34%) and NOX2 activation (+30%), which were inhibited by ox-LDL receptor antagonists.
  
  
  CONCLUSIONS
  Platelets oxidize LDL, which in turn amplify platelet activation via specific ox-LDL receptors; both effects are mediated by NOX2 activation.
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy.
authors.forename: Roberto
authors.initials: R
authors.lastname: Carnevale
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy.
authors.forename: Simona
authors.initials: S
authors.lastname: Bartimoccia
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy.
authors.forename: Cristina
authors.initials: C
authors.lastname: Nocella
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy.
authors.forename: Serena
authors.initials: S
authors.lastname: Di Santo
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy.
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Loffredo
authors.affiliation: The "Francesco Durante" Department of Surgery, "La Sapienza" University, Rome, Italy.
authors.forename: Giulio
authors.initials: G
authors.lastname: Illuminati
authors.affiliation: Vascular Biology and Stem Cells Unit, Centro di Riferimento Oncologico, Aviano, Italy.
authors.forename: Elisabetta
authors.initials: E
authors.lastname: Lombardi
authors.affiliation: Vascular Biology and Stem Cells Unit, Centro di Riferimento Oncologico, Aviano, Italy.
authors.forename: Valentina
authors.initials: V
authors.lastname: Boz
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy.
authors.forename: Maria
authors.initials: M
authors.lastname: Del Ben
authors.affiliation: Vascular Biology and Stem Cells Unit, Centro di Riferimento Oncologico, Aviano, Italy.
authors.forename: Luigi
authors.initials: L
authors.lastname: De Marco
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy.
authors.forename: Pasquale
authors.initials: P
authors.lastname: Pignatelli
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy. Electronic address: francesco.violi@uniroma1.it.
authors.forename: Francesco
authors.initials: F
authors.lastname: Violi
mesh_terms: D000328:Adult; D050197:Atherosclerosis / Q000378:metabolism; D001792:Blood Platelets / Q000378:metabolism*; D016022:Case-Control Studies; D003094:Collagen / Q000737:chemistry; D015237:Dinoprost / Q000031:analogs & derivatives / Q000378:metabolism; D005260:Female; D006801:Humans; D006937:Hypercholesterolemia / Q000097:blood; D008077:Lipoproteins, LDL / Q000737:chemistry*; D008297:Male; D008562:Membrane Glycoproteins / Q000172:deficiency / Q000235:genetics*; D008875:Middle Aged; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D018384:Oxidative Stress; D010477:Perfusion; D015539:Platelet Activation; D013927:Thrombosis / Q000473:pathology
chemical_list: D008077:Lipoproteins, LDL; D008562:Membrane Glycoproteins; C099252:oxidized low density lipoprotein; C075750:8-epi-prostaglandin F2alpha; D003094:Collagen; D015237:Dinoprost; C056558:CYBB protein, human; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases
keywords: Atherosclerosis; NADPH oxidase; Oxidative stress; ox-LDL