Circulating progenitor cells and their interaction with platelets in patients with an acute coronary syndrome.

CD34+ cells expressing KDR (CD34+/KDR+) represent a small proportion of circulating progenitor cells that have the capacity to interact with platelets and to differentiate into mature endothelial cells, thus contributing to vascular homeostasis and regeneration as well as to re-endothelialization. We investigated the levels of CD34+ and CD34+/KDR+ progenitor cells as well as their interaction with platelets in acute coronary syndrome (ACS) patients before the initiation (baseline) of their treatment with a P2Y12 receptor antagonist, and at 5-days post-treatment (follow-up). Sixty-seven consecutive ACS patients and thirty healthy subjects (controls) participated in the study. On admission, all patients received 325 mg aspirin, followed by 100 mg/day and then were loaded either with 600 mg clopidogrel or 180 mg ticagrelor, followed by 75 mg/day (n = 36) or 90 mg × 2/day (n = 31), respectively. The levels of circulating CD34+ and CD34+/KDR+ progenitor cells, as well as their interaction with platelets, were determined by flow cytometry, before and after activation with ADP, in vitro. The circulating levels of CD34+ and CD34+/KDR+ cells in both patient groups at baseline were lower compared with controls while they were significantly increased at 5-days of follow-up in both groups, this increase being more pronounced in the ticagrelor group. The platelet/CD34+ (CD61+/CD34+) conjugates were higher at baseline and reduced at follow-up while the platelet/KDR+ (CD61+/KDR+) conjugates were lower at baseline and increased at follow-up, both changes being more pronounced in the ticagrelor group. ADP activation of control samples significantly increased the KDR expression by CD34+ cells and the CD61+/KDR+ conjugates, these parameters being unaffected in patients at baseline but increased at follow-up. Short-term dual antiplatelet therapy in ACS patients restores the low platelet/KDR+ conjugates and CD34+ cell levels and improves the low membrane expression levels of KDR in these cells, an effect being more pronounced in ticagrelor-treated patients. This may represent a pleiotropic effect of antiplatelet therapy towards vascular endothelial regeneration.

Comparative Anti-Platelet Profiling Reveals a Potent Anti-Aggregatory Effect of CD34+ Progenitor Cell-Derived Late-Outgrowth Endothelial Cells in vitro.

BACKGROUND/AIMS: Platelets affect endothelial progenitor cell (EPC) functionality, inducing their differentiation into mature endothelial cells. However, it remains to be established whether EPCs can influence platelet functionality. METHODS: Mononuclear proangiogenic cells (MPCs) and early outgrowth cells (EOCs) were prepared from peripheral blood mononuclear cells, whereas late-outgrowth endothelial cells (OECs) were prepared from cord blood CD34+ cells. The effect of the above cells and their supernatants on washed platelet aggregation was studied. The effect of OECs and their supernatant on the adenosine diphosphate (ADP)-induced magnitude of platelet integrin receptor αIIbß3 activation and on P-selectin membrane expression was investigated. The levels of nitric oxide (NO) and prostacyclin (PGI2) that were secreted from EOCs, OECs, and human umbilical vein endothelial cells (HUVECs) were also assessed. RESULTS: Among all progenitors, OECs and their supernatant exhibited the most potent inhibitory activity towards platelet aggregation. Furthermore, OECs and their supernatant, but not CD34+ cells, reduced αIIbß3 activation and P-selectin membrane expression. Finally, OECs secreted higher NO and PGI2 levels than EOCs did. CONCLUSION: The anti-platelet effect of EPCs depends highly on the degree of their endothelial phenotype, with OECs expressing the highest potency. Therefore, the induction of OEC generation and the enhancement of their functionality in vivo could be a new approach for the treatment of patients at a high thrombotic risk.

The Effect of Platelet-Rich Plasma on Endothelial Progenitor Cell Functionality.

Platelets mediate circulating endothelial progenitor cell (EPC) recruitment and maturation, participating in vascular repair, however the underlying mechanism(s) remain unclear. We investigated the effect of platelet-rich plasma (PRP) on the functionality of CD34+-derived late-outgrowth endothelial cells (OECs) in culture. Confluent OECs were coincubated with PRP under platelet aggregation (with adenosine diphosphate; ADP) and nonaggregation conditions, in the presence/absence of the reversible P2Y12 platelet receptor antagonist ticagrelor. Outgrowth endothelial cell activation was evaluated by determining prostacyclin (PGI2) and monocyte chemoattractant protein-1 (MCP-1) release and intercellular adhesion molecule-1 (ICAM-1) membrane expression. Similar experiments were performed using human umbilical vein endothelial cells (HUVECs). Platelet-rich plasma increased ICAM-1 expression and PGI2 and MCP-1 secretion compared with autologous platelet-poor plasma, whereas ADP-aggregated platelets in PRP did not exhibit any effect. Platelet-rich plasma pretreated with ticagrelor prior to activation with ADP increased all markers to a similar extent as PRP. Similar results were obtained using HUVECs. In conclusion, PRP induces OEC activation, a phenomenon not observed when platelets are aggregated with ADP. Platelet inhibition with ticagrelor restores the PRP capability to activate OECs. Since EPC activation is important for endothelial regeneration and angiogenesis, we suggest that agents inhibiting platelet aggregation, such as ticagrelor, may promote platelet-EPC interaction and EPC function.

Comparison of Triflusal with Aspirin in the Secondary Prevention of Atherothrombotic Events; Α Randomised Clinical Trial.

BACKGROUND: Triflusal has demonstrated an efficacy similar to aspirin in the prevention of vascular events in patients with acute myocardial infarction (ΜΙ) and ischaemic stroke but with less bleeding events. OBJECTIVE: We performed a randomised, multicentre, phase 4 clinical trial to compare the clinical efficacy and safety of triflusal versus aspirin, administered for 12 months in patients eligible to receive a cyclooxygenase-1 (COX-1) inhibitor. METHODS: Patients with stable coronary artery disease or with a history of non-cardioembolic ischaemic stroke were randomly assigned to receive either triflusal 300 mg twice or 600 mg once daily or aspirin 100 mg once daily for 12 months. The primary efficacy endpoint was the composite of: (a) ΜΙ, (b) stroke (ischaemic or haemorrhagic), or, (c) death from vascular causes for the entire follow-up period. The primary safety endpoints were the rate of bleeding events as defined by Bleeding Academic Research Consortium (BARC) criteria. RESULTS: At 12-month follow-up, an equivalent result was revealed between the triflusal (n=559) and aspirin (n=560) in primary efficacy endpoint. Specifically, the combined efficacy outcome rate (i.e. MI, stroke or death from vascular causes) difference was equal to -1.3% (95% confidence interval -1.1 to 3.5) and lied within the a-priori defined equivalence interval (p<0.001). Regarding the primary safety endpoints, patients on triflusal treatment were 50% less likely to develop bleeding events according to the BARC criteria, and especially any clinically overt sign of haemorrhage that requires diagnostic studies, hospitalisation or special treatment (BARC type 2). CONCLUSION: The efficacy of triflusal in the secondary prevention of vascular events is similar to aspirin when administered for 12 months. Importantly, triflusal significantly reduced the incidence of ΜΙ and showed a better safety profile compared with aspirin. (ASpirin versus Triflusal for Event Reduction In Atherothrombosis Secondary prevention, ASTERIAS trial; Clinical Trials.gov Identifier: NCT02616497).

Plasma VEGF and IL-8 Levels in Patients with Mixed Dyslipidaemia. Effect of Rosuvastatin Monotherapy or its Combination at a Lower Dose with Omega-3 Fatty Acids: A Pilot Study.

OBJECTIVES: Hypercholesterolaemia is associated with increased plasma levels of vascular endothelial growth factor (VEGF) and interleukin-8 (IL-8). We studied the effect of rosuvastatin monotherapy or its combination at a lower dose with omega-3 polyunsaturated fatty acids (ω-3 PUFAs) in the VEGF and IL-8 plasma levels in patients with mixed dyslipidaemia. METHODS: Fifty patients with mixed dyslipidaemia were recruited. Fifty-five normolipidaemic, apparently healthy, ageand sex- matched subjects acted as controls. Patients were randomized to 40 mg/day rosuvastatin (R group, n=26) or 10 mg/day rosuvastatin plus 2 g/day of ω-3 PUFAs (R+O group, n=24). The levels of VEGF and IL-8 in plasma, were assessed at baseline and 3 months post-treatment. RESULTS: At baseline, both plasma VEGF and IL-8 levels were significantly higher in the R and R+O groups compared with controls (p<0.04, p<0.03 and p<0.02, p<0.03, respectively). Post-treatment levels of VEGF were decreased in the R group while a significant increase was observed in the R+O group, compared with baseline levels (p<0.02 and p<0.03, respectively). Post-treatment IL-8 levels were decreased in both R and R+O groups (p<0.03 and p<0.04, respectively). CONCLUSIONS: We show for the first time that either rosuvastatin monotherapy or its combination at a lower dose with ω-3 PUFAs reduces IL-8 levels in mixed dyslipidaemic patients. High-dose rosuvastatin monotherapy reduces VEGF values, whereas a significant increase is observed in patients receiving lower dose rosuvastatin with ω-3 PUFAs. The clinical significance of the above findings regarding cardiovascular risk remains to be established.

Effect of rosuvastatin or its combination with omega-3 fatty acids on circulating CD34(+) progenitor cells and on endothelial colony formation in patients with mixed dyslipidaemia.

BACKGROUND AND AIMS: Hypercholesterolaemia is associated with a reduced number of circulating progenitor cells, a defect that is restored by statin therapy. We studied the effect of rosuvastatin monotherapy or its combination with omega-3 polyunsaturated fatty acids (ω-3 PUFAs) on progenitor cell number and function in patients with mixed dyslipidaemia. METHODS: Fifty patients with mixed dyslipidaemia and fifty-five normolipidaemic, apparently healthy, age- and sex-matched volunteers participated in the study. Patients were randomized to receive a daily dose of either 40 mg rosuvastatin (R group, n = 26) or 10 mg rosuvastatin plus 2 g of ω-3 PUFAs (R + O group, n = 24). The number of circulating CD34(+) and CD34(+)/KDR(+) progenitor cells as well as the number of colony-forming units-endothelial cells (CFU-ECs) at 10 days of culture were assessed at baseline and 3 months post-treatment. RESULTS: The number of CD34(+) and CD34(+)/KDR(+) cells per 10,000 leukocytes as well as the number of CFU-ECs were significantly lower in both patient groups compared with healthy volunteers (all p = 0.03). A 3-month treatment with either R or R + O significantly increased circulating CD34(+) and CD34(+)/KDR(+) cells as well as the number of CFU-ECs compared with baseline (all p = 0.03). Importantly, the increase in the above parameters was inversely correlated with therapy-induced reduction in lipid parameters in both patient groups. CONCLUSIONS: Patients with mixed dyslipidaemia exhibit low numbers of circulating CD34(+) and CD34(+)/KDR(+) cells as well as CFU-ECs in culture, a defect restored by 3-month treatment with either high-rosuvastatin dose or a combination of low-rosuvastatin dose with ω-3 PUFAs. The pathophysiological meaning of our results regarding the increased cardiovascular risk in these patients remains to be established.