The Pathophysiological Role of Platelet-Derived Extracellular Vesicles.

Platelets are very abundant in the blood, where they play a role in hemostasis, inflammation, and immunity. When activated, platelets undergo a conformational change that allows the release of numerous effector molecules as well as the production of extracellular vesicles, which are circulating submicron vesicles (10 to 1,000 nm in diameter) released into the extracellular space. Extracellular vesicles are formed by the budding of platelet and they carry some of its contents, including nucleic acids, surface proteins, and organelles. While platelets cannot cross tissue barriers, platelet-derived extracellular vesicles can enter the lymph, bone marrow, and synovial fluid. This allows the transfer of diverse contents carried by these platelet-derived vesicles to cell recipients and organs inaccessible to platelets where they can perform many functions. This review highlights the importance of these platelet-derived extracellular vesicles under different physiological and pathophysiological conditions.

Dissecting Platelet's Role in Viral Infection: A Double-Edged Effector of the Immune System.

Platelets play a major role in the processes of primary hemostasis and pathological inflammation-induced thrombosis. In the mid-2000s, several studies expanded the role of these particular cells, placing them in the "immune continuum" and thus changing the understanding of their function in both innate and adaptive immune responses. Among the many receptors they express on their surface, platelets express Toll-Like Receptors (TLRs), key receptors in the inflammatory cell-cell reaction and in the interaction between innate and adaptive immunity. In response to an infectious stimulus, platelets will become differentially activated. Platelet activation is variable depending on whether platelets are activated by a hemostatic or pathogen stimulus. This review highlights the role that platelets play in platelet modulation count and adaptative immune response during viral infection.

University of Montreal's Clinician-Investigator Program: A 10-Year Descriptive Evaluation.

PURPOSE: Clinician-investigators have an important role in the development and implantation of new therapies and treatment modalities; however, there have been several reports highlighting a pending shortage in the clinician-investigators' workforce. In Canada, the Royal College has promoted the development of clinician-investigators programs (CIP) to facilitate the training of these individuals. There is currently a paucity of data regarding the outcomes of such programs. This study aims to identify the strengths and areas of improvement of the Montreal University CIP.  Methods: An internet-based 51-question survey was distributed to all the alumni from the University of Montreal CIP. Participation was voluntary and no incentives were provided. The response rate was 64%.  Results: Among respondents, 50% (n=16) had completed their clinical residency and all CIP requirements. The majority of these individuals (63%) had become independent investigators and had secured provincial and national funding. Satisfaction of the respondents was high regarding the overall program (85%), the research skills developed during the CIP (84%) and the financial support obtained during the program (72%). The satisfaction rate regarding career planning was lower (63%).  Conclusion: This survey demonstrates that, while indicators are favorable, some areas still require improvement. Several steps to improve the CIP have been identified; notably, the transition from the CIP to early independent career has been identified as critical in the development of clinician-investigators and steps have been taken to improve this progression.

The Role of the Proteasome in Platelet Function.

Platelets are megakaryocyte-derived acellular fragments prepped to maintain primary hemostasis and thrombosis by preserving vascular integrity. Although they lack nuclei, platelets harbor functional genomic mediators that bolster platelet activity in a signal-specific manner by performing limited de novo protein synthesis. Furthermore, despite their limited protein synthesis, platelets are equipped with multiple protein degradation mechanisms, such as the proteasome. In nucleated cells, the functions of the proteasome are well established and primarily include proteostasis among a myriad of other signaling processes. However, the role of proteasome-mediated protein degradation in platelets remains elusive. In this review article, we recapitulate the developing literature on the functions of the proteasome in platelets, discussing its emerging regulatory role in platelet viability and function and highlighting how its functional coupling with the transcription factor NF-κB constitutes a novel potential therapeutic target in atherothrombotic diseases.

Role of NF-κB in Platelet Function.

Platelets are megakaryocyte-derived fragments lacking nuclei and prepped to maintain primary hemostasis by initiating blood clots on injured vascular endothelia. Pathologically, platelets undergo the same physiological processes of activation, secretion, and aggregation yet with such pronouncedness that they orchestrate and make headway the progression of atherothrombotic diseases not only through clot formation but also via forcing a pro-inflammatory state. Indeed, nuclear factor-κB (NF-κB) is largely implicated in atherosclerosis and its pathological complication in atherothrombotic diseases due to its transcriptional role in maintaining pro-survival and pro-inflammatory states in vascular and blood cells. On the other hand, we know little on the functions of platelet NF-κB, which seems to function in other non-genomic ways to modulate atherothrombosis. Therein, this review will resemble a rich portfolio for NF-κB in platelets, specifically showing its implications at the levels of platelet survival and function. We will also share the knowledge thus far on the effects of active ingredients on NF-κB in general, as an extrapolative method to highlight the potential therapeutic targeting of NF-κB in coronary diseases. Finally, we will unzip a new horizon on a possible extra-platelet role of platelet NF-κB, which will better expand our knowledge on the etiology and pathophysiology of atherothrombosis.

Interaction of CD154 with different receptors and its role in bidirectional signals.

In addition to its classical receptor, CD40, it is now well established that CD154 also binds αIIbß3, α5ß1, and αMß2 integrins. Although these integrins are all members of the same family, they bind CD154 differently. The current investigation aims to analyze the interaction of CD154 with α5ß1 and αMß2 and investigate its role in bidirectional signals in various human cell lines. Results obtained herein indicate that the CD154 residues involved in the interaction with α5ß1 are N151 and Q166, whereas those involved in αMß2 binding are common to residues required for CD40, namely Y145 and R203. Soluble CD40/CD154 or αMß2/CD154 complexes do not interfere with the binding of CD154 to α5ß1-positive cells, but inhibit the binding of CD154 to CD40- or αMß2-positive cells, respectively. Ligation of CD154 on CD154-positive cells with soluble CD40, αIIbß3, α5ß1, or αMß2 stimulates intracellular signaling, including MAPK phosphorylation. Given that CD154 exists as a trimer, our data strongly suggest that CD154 may bind concomitantly to two receptors of the same or different family, and biologically activate cells expressing both receptors. The characterization of CD154/receptor interactions helps the identification of new therapeutic targets for the prevention and/or treatment of CD154-associated autoimmune and inflammatory diseases.

Endothelial progenitor cells inhibit platelet function in a P-selectin-dependent manner.

BACKGROUND: The role of endothelial progenitor cells (EPCs) in vascular repair is related to their recruitment at the sites of injury and their interaction with different components of the circulatory system. We have previously shown that EPCs bind and inhibit platelet function and impair thrombus formation via prostacyclin secretion, but the role of EPC binding to platelet P-selectin in this process has not been fully characterized. In the present study, we assessed the impact of EPCs on thrombus formation and we addressed the implication of P-selectin in this process. METHODS: EPCs were generated from human peripheral blood mononuclear cells cultured on fibronectin in conditioned media. The impact of EPCs on platelet aggregation and thrombus formation was investigated in P-selectin deficient (P-sel(-/-)) mice and their wild-type (WT) counterparts. RESULTS: EPCs significantly and dose-dependently impaired collagen-induced whole blood platelet aggregation in WT mice, whereas no effects were observed in P-sel(-/-) mice. Moreover, in a ferric chloride-induced arterial thrombosis model, infusion of EPCs significantly reduced thrombus formation in WT, but not in P-sel(-/-) mice. Furthermore, the relative mass of thrombi generated in EPC-treated P-sel(-/-) mice were significantly larger than those in EPC-treated WT mice, and the number of EPCs recruited within the thrombi and along the arterial wall was reduced in P-sel(-/-) mice as compared to WT mice. CONCLUSION: This study shows that EPCs impair platelet aggregation and reduce thrombus formation via a cellular mechanism involving binding to platelet P-selectin. These findings add new insights into the role of EPC-platelet interactions in the regulation of thrombotic events during vascular repair.

Early outgrowth cells versus endothelial colony forming cells functions in platelet aggregation.

BACKGROUND: Endothelial progenitor cells (EPCs) have been implicated in neoangiogenesis, endothelial repair and cell-based therapies for cardiovascular diseases. We have previously shown that the recruitment of EPCs to sites of vascular lesions is facilitated by platelets where EPCs, in turn, modulate platelet function and thrombosis. However, EPCs encompass a heterogeneous population of progenitor cells that may exert different effects on platelet function. Recent evidence suggests the existence of two EPC subtypes: early outgrowth cells (EOCs) and endothelial colony-forming cells (ECFCs). We aimed at characterizing these two EPC subtypes and at identifying their role in platelet aggregation. METHODS: EOCs and ECFCs were generated from human peripheral blood mononuclear cells (PBMCs) seeded in conditioned media on fibronectin and collagen, respectively. The morphological, phenotypical and functional characteristics of EOCs and ECFCs were assessed by optical and confocal laser scanning microscopes, cell surface markers expression, and Matrigel tube formation. The impact of EOCs and ECFCs on platelet aggregation was monitored in collagen-induced optical aggregometry and compared with PBMCs and human umbilical vein endothelial cells (HUVECs). The levels of the anti-platelet agents' nitric oxide (NO) and prostacyclin (PGI2) released from cultured cells as well as the expression of their respective producing enzymes NO synthases (NOS) and cyclooxygenases (COX) were also assessed. RESULTS: We showed that EOCs display a monocytic-like phenotype whereas ECFCs have an endothelial-like phenotype. We demonstrated that both EOCs and ECFCs and their supernatants inhibited platelet aggregation; however ECFCs were more efficient than EOCs. This could be related to the release of significantly higher amounts of NO and PGI2 from ECFCs, in comparison to EOCs. Indeed, ECFCs, like HUVECs, constitutively express the endothelial (eNOS)-and inducible (iNOS)-NOS isoforms, and COX-1 and weakly express COX-2, whereas EOCs do not constitutively express these NO and PGI2 producing enzymes. CONCLUSION: The different morphological, phenotypic and more importantly the release of the anti-aggregating agents PGI2 and NO in each EPC subtype are implicated in their respective roles in platelet function and thus, may be linked to the increased efficiency of ECFCs in inhibiting platelet aggregation as compared to EOCs.

Development of a Polyester Coating Combining Antithrombogenic and Cell Adhesive Properties: Influence of Sequence and Surface Density of Adhesion Peptides.

Biofunctionalization strategies have been developed to improve small-diameter vascular grafts. However, a fully successful coating featuring antithrombogenic properties while allowing for endothelialization has not been achieved yet. In this report, we explored the combination of low-fouling polyethylene glycol (PEG) and adhesion peptides, namely, RGD, YIGSR, and REDV, grafted on top of polyvinylamine (PVAm)-coated polyester. The peptides were grafted over a wide range of density (ca. 20-2000 pmol/cm(2)) on top of a dense PEG underlayer. The coating performances were assessed through HUVEC adhesion, platelet attachment, and protein adsorption, which were all drastically diminished on PEG-coated samples. RGD exhibited the expected high adhesive properties, toward both HUVEC and platelets. REDV had no effect neither on platelet attachment, as expected, nor on HUVEC adhesion, in contrast with previous reports. YIGSR was the most promising sequence even though its combination with other agents should be explored to further decrease thrombogenicity for vascular graft applications.

Chondroitin sulfate coatings display low platelet but high endothelial cell adhesive properties favorable for vascular implants.

This study highlights the advantages of chondroitin sulfate (CS) as a sublayer combining selective low-fouling properties, low-platelet adhesion and pro-adhesive properties on endothelial cells, making CS promising for vascular graft applications. These properties were evaluated by comparing CS with well-known low-fouling coatings such as poly(ethylene glycol) (PEG) and carboxymethylated dextran (CMD), which were covalently grafted on primary amine-rich plasma polymerized (LP) films. Protein adsorption studies by quartz crystal microbalance with dissipation monitoring (QCM-D) and fluorescence measurements showed that CS is as effective as PEG in reducing fibrinogen adsorption (~90% reduction). CS also largely reduced adsorption of bovine serum albumin (BSA) as well as fetal bovine serum (FBS) but to a lower extent than PEG and CMD surfaces (72% vs 85% for BSA and 66% vs 89% for FBS). Whole blood perfusion assays indicated that, while LP surfaces were highly reactive with platelets, PEG, CMD, and CS grafted surfaces drastically decreased platelet adhesion and activation to levels significantly lower than polyethylene terephthalate (PET) surfaces. Finally, while human umbilical vein endothelial cell (HUVEC) adhesion and growth were found to be very limited on PEG and CMD, they were significantly increased on CS compared to that on bare PET and reached similar values as those for tissue culture polystyrene positive controls. Interestingly, HUVEC retention during perfusion with blood was found to be excellent on CS but poor on PET. Overall, our results suggest that the CS surface has the advantage of promoting HUVEC growth and resistance to flow-induced shear stress while preventing fibrinogen and platelet attachment. Such a nonthrombogenic but endothelial-cell adhesive surface is thus promising to limit vascular graft occlusion.

Functional interaction of CD154 protein with α5ß1 integrin is totally independent from its binding to αIIbß3 integrin and CD40 molecules.

In addition to its classical CD40 receptor, CD154 also binds to αIIbß3, α5ß1, and αMß2 integrins. Binding of CD154 to these receptors seems to play a key role in the pathogenic processes of chronic inflammation. This investigation was aimed at analyzing the functional interaction of CD154 with CD40, αIIbß3, and α5ß1 receptors. We found that the binding affinity of CD154 for αIIbß3 is ∼4-fold higher than for α5ß1. We also describe the generation of sCD154 mutants that lost their ability to bind CD40 or αIIbß3 and show that CD154 residues involved in its binding to CD40 or αIIbß3 are distinct from those implicated in its interaction to α5ß1, suggesting that sCD154 may bind simultaneously to different receptors. Indeed, sCD154 can bind simultaneously to CD40 and α5ß1 and biologically activate human monocytic U937 cells expressing both receptors. The simultaneous engagement of CD40 and α5ß1 activates the mitogen-activated protein kinases, p38, and extracellular signal-related kinases 1/2 and synergizes in the release of inflammatory mediators MMP-2 and -9, suggesting a cross-talk between these receptors.

ARC15105 is a potent antagonist of von Willebrand factor mediated platelet activation and adhesion.

OBJECTIVE: We investigated the stability, pharmacokinetic, and pharmacodynamic profile of the 2(nd) generation anti-von Willeband factor aptamer ARC15105. METHODS AND RESULTS: Platelet plug formation was measured by collagen/adenosine diphosphate-induced closure time with the platelet function analyzer-100 and platelet aggregation by multiple electrode aggregometry. Platelet adhesion was measured on denuded porcine aortas and in a flow chamber. Aptamer stability was assessed by incubation in nuclease rich human, monkey, and rat serum for up to 72 hours. Pharmacokinetic and pharmacodynamic profiles were tested in cynomolgus monkeys after IV and SC administration. The median IC(100) and IC(50) to prolong collagen/adenosine diphosphate-induced closure timewere 27 nmol/L and 12 nmol/L, respectively. ARC15105 (1.3 µmol/L) completely inhibited ristocetin-induced platelet aggregation in whole blood (P<0.001), but also diminished collagen, ADP, arachidonic acid, and thrombin receptor activating peptide-induced platelet aggregation to some extent (P<0.05). ARC15105 (40 nmol/L) decreased platelet adhesion by >90% on denuded porcine aortas (P<0.001), which was comparable to the degree of inhibition obtained with abciximab. ARC15105 (100 nmol/L) also inhibited platelet adhesion to collagen under arterial shear in a flow chamber by >90% (P<0.001). The IV and SC terminal half-lives in cynomolgus monkeys were 67 and 65 hours, respectively, and the SC bioavailability was ≈98%. Allometric scaling estimates the human T(1/2) would be ≈217 hours. Pharmacodynamic analysis confirms that ARC15105 inhibits von Willeband factor activity >90% in blood samples taken 300 hours after a 20 mg/kg IV or SC dose in monkeys. CONCLUSIONS: The potency, pharmacokinetic profile, and SC bioavailability of ARC15105 support its clinical investigation for chronic inhibition of von Willeband factor -mediated platelet activation.

Involvement of nuclear factor κB in platelet CD40 signaling.

CD40 ligand (CD40L) is a thrombo-inflammatory molecule that predicts cardiovascular events. Platelets constitute the major source of soluble CD40L (sCD40L), which has been shown to potentiate platelet activation and aggregation, in a CD40-dependent manner, via p38 mitogen activated protein kinase (MAPK) and Rac1 signaling. In many cells, the CD40L/CD40 dyad also induces activation of nuclear factor kappa B (NF-κB). Given that platelets contain NF-κB, we hypothesized that it may be involved in platelet CD40 signaling and function. In human platelets, sCD40L induces association of CD40 with its adaptor protein the tumor necrosis factor receptor associated factor 2 and triggers phosphorylation of IκBα, which are abolished by CD40L blockade. Inhibition of IκBα phosphorylation reverses sCD40L-induced IκBα phosphorylation without affecting p38 MAPK phosphorylation. On the other hand, inhibition of p38 MAPK phosphorylation has no effect on IκBα phosphorylation, indicating a divergence in the signaling pathway originating from CD40 upon its ligation. In functional studies, inhibition of IκBα phosphorylation reverses sCD40L-induced platelet activation and potentiation of platelet aggregation in response to a sub-threshold concentration of collagen. This study demonstrates that the sCD40L/CD40 axis triggers NF-κB activation in platelets. This signaling pathway plays a critical role in platelet activation and aggregation upon sCD40L stimulation and may represent an important target against thrombo-inflammatory disorders.

Implication of Platelets in Immuno-Thrombosis and Thrombo-Inflammation.

In addition to their well-described hemostatic function, platelets are active participants in innate and adaptive immunity. Inflammation and immunity are closely related to changes in platelet reactions and enhanced platelet function in thrombo-inflammation, as well as in microbial and virus infections. A platelet's immune function is incompletely understood, but an important balance exists between its protective and pathogenic responses and its thrombotic and inflammatory functions. As the mediator of vascular homeostasis, platelets interact with neutrophils, bacteria and virus by expressing specific receptors and releasing granules, transferring RNA, and secreting mitochondria, which controls hemostasis and thrombosis, infection, and innate and adaptive immunity. This review focuses on the involvement of platelets during immuno-thrombosis and thrombo-inflammation.

Characterization of FGF receptor expression in human neutrophils and their contribution to chemotaxis.

Several members of the fibroblast growth factor (FGF) family are potent endothelial cell (EC) mitogens and angiogenic factors, and their activities can be mediated by four tyrosine kinase receptors (FGFR1-4). In addition, FGFs can induce the release of inflammatory mediators by ECs and the expression of adhesion molecules at their surface, thereby favoring the recruitment and transvascular migration of inflammatory cells such as neutrophils. Neither the expression nor the biological activities that could be mediated by FGFRs have been investigated in human neutrophils. By biochemical and cytological analyses, we observed that purified circulating human neutrophils from healthy individuals expressed varying levels of FGFRs in their cytosol and at their cytoplasmic membrane. FGFR-2 was identified as the sole cell surface receptor, with FGFR-1 and -4 localizing in the cytosol and FGFR-3 being undetectable. We assessed the capacity of FGF-1 and FGF-2 to induce neutrophil chemotaxis in a modified Boyden microchamber and observed that they increase neutrophil transmigration at 10(-10) and 10(-9) M and by 1.77- and 2.34-fold, respectively, as compared with PBS-treated cells. Treatment with a selective anti-FGFR-2 antibody reduced FGF-1-mediated chemotaxis by 75% and abrogated the effect of FGF-2, while the blockade of FGFR-1 and -4 partially inhibited (15-40%) FGF-chemotactic activities. In summary, our data are the first to report the expression of FGF receptors in human neutrophils, with FGF-1 and FGF-2 promoting neutrophil chemotaxis mainly through FGFR-2 activation.

Investigation of layer-by-layer assembly of polyelectrolytes on fully functional human red blood cells in suspension for attenuated immune response.

The encapsulation of live cells with polymeric coat-ings is a versatile approach to modulate or control the response cells to their environment. The layer-by-layer (LbL) self-assembly of nonimmunogenic polyelectrolytes is employed here to attenuate or suppress the binding of antibodies to live red blood cells (RBCs) and, consequently, decrease their inherent immunogenicity toward foreign RBCs. The optimized shell was composed of four bilayers of alginate (AL) and chitosan-graft-phosphorylcholine (CH-PC) surrounded by two bilayers of AL and poly-l-lysine-graft-polyethylene glycol (PLL-PEG). Experimental parameters, including the polyelectrolytes and RBCs concentrations and the cell handling and purification protocols, were optimized to achieve effective encapsulation of live and functional RBCs in suspension. The viability and functionality of coated RBCs were confirmed by a hemolysis assay and by their ability to take up oxygen. The successful immunocamouflage of RBCs was confirmed by observing that the recognition of the ABO/D (Rh) blood group antigens present on the surface of RBCs by their respective antibodies was muted in the case of coated RBCs. The results of this studies mark an important step toward the production of universal RBCs.

Enhanced levels of soluble CD40 ligand exacerbate platelet aggregation and thrombus formation through a CD40-dependent tumor necrosis factor receptor-associated factor-2/Rac1/p38 mitogen-activated protein kinase signaling pathway.

OBJECTIVE: CD40 ligand is a thromboinflammatory molecule that predicts cardiovascular events. Platelets constitute the major source of soluble CD40 ligand (sCD40L), which has been shown to influence platelet activation, although its exact functional impact on platelets and the underlying mechanisms remain undefined. We aimed to determine the impact and the signaling mechanisms of sCD40L on platelets. METHODS AND RESULTS: sCD40L strongly enhances platelet activation and aggregation. Human platelets treated with a mutated form of sCD40L that does not bind CD40, and CD40(-/-) mouse platelets failed to elicit such responses. Furthermore, sCD40L stimulation induces the association of the tumor necrosis factor receptor-associated factor-2 with platelet CD40. Notably, sCD40L primes platelets through activation of the small GTPase Rac1 and its downstream target p38 mitogen-activated protein kinase, which leads to platelet shape change and actin polymerization. Moreover, sCD40L exacerbates thrombus formation and leukocyte infiltration in wild-type mice but not in CD40(-/-) mice. CONCLUSIONS: sCD40L enhances agonist-induced platelet activation and aggregation through a CD40-dependent tumor necrosis factor receptor-associated factor-2/Rac1/p38 mitogen-activated protein kinase signaling pathway. Thus, sCD40L is an important platelet primer predisposing platelets to enhanced thrombus formation in response to vascular injury. This may explain the link between circulating levels of sCD40L and cardiovascular diseases.

Detailed analysis of a series of explanted Talent AAA stent-grafts: biofunctionality assessment.

Six Talent stent-grafts were harvested at reoperations (N=5) and autopsy (N=1). The explants were observed nondestructively, including gross morphology, X-rays, CT scans and closed pressure system analysis. The Nitinol frames in three devices harvested at reoperations and another harvested at autopsy were intact. One had a stent fracture of the proximal bare stent, and one had a wire fracture of a thin proximal external supporting stent as well as a hole in the fabric just above the bifurcation. For the three devices structurally intact, reoperations were performed for a type 1A endoleak (one patient) and aorto-enteric fistulas (two patients). The healing characteristics were poor or absent. The fabric in the main body of the grafts harvested after aorto-enteric fistula was devoid of biological deposits. Two of the grafts harvested at reoperation demonstrated fabric holes of up to 4 mm 2. The device obtained at autopsy showed an almost continuous internal capsule with variable thickness. The luminal surface was smooth, but the capsule detached easily. The devices explanted at reoperations showed various levels of impaired biofunctionality associated with adverse outcomes. The stent-graft retrieved from autopsy was intact.

Endothelial progenitor cells bind and inhibit platelet function and thrombus formation.

BACKGROUND: Interactions of endothelial progenitor cells (EPCs) with vascular and blood cells contribute to vascular homeostasis. Although platelets promote the homing of EPCs to sites of vascular injury and their differentiation into endothelial cells, the functional consequences of such interactions on platelets remain unknown. Herein, we addressed the interactions between EPCs and platelets and their impact on platelet function and thrombus formation. METHODS AND RESULTS: Cultured on fibronectin in conditioned media, human peripheral blood mononuclear cells differentiated, within 10 days of culture, into EPCs, which uptake acetylated low-density lipoprotein, bind ulex-lectin, lack monocyte/leukocyte markers (CD14, P-selectin glycoprotein ligand-1, L-selectin), express progenitor/endothelial markers (CD34, vascular endothelial growth factor receptor-2, von Willebrand factor, and vascular endothelial cadherin), and proliferate in culture. These EPCs bound activated platelets via CD62P and inhibited its translocation, glycoprotein IIb/IIIa activation, aggregation, and adhesion to collagen, mainly via prostacyclin secretion. Indeed, this was associated with upregulation of cyclooxygenase-2 and inducible nitric oxide synthase. However, the effects on platelets in vitro were reversed by cyclooxygenase and cyclooxygenase-2 inhibition but not by nitric oxide or inducible nitric oxide synthase inhibition. Moreover, in a ferric chloride-induced murine arterial thrombosis model, injection of EPCs led to their incorporation into sites of injury and impaired thrombus formation, leading to an incomplete occlusion with 50% residual flow. CONCLUSIONS: Peripheral blood mononuclear cell-derived EPCs bind platelets via CD62P and inhibit platelet activation, aggregation, adhesion to collagen, and thrombus formation, predominantly via upregulation of cyclooxygenase-2 and secretion of prostacyclin. These findings add new insights into the biology of EPCs and define their potential roles in regulating platelet function and thrombosis.