Skip Navigation


European Heart Journal Advance Access originally published online on December 24, 2008
European Heart Journal 2009 30(5):584-593; doi:10.1093/eurheartj/ehn566
This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow All Versions of this Article:
30/5/584    most recent
ehn566v1
Right arrow E-letters: Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when E-letters are posted
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Stellos, K.
Right arrow Articles by Gawaz, M.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Stellos, K.
Right arrow Articles by Gawaz, M.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2008. For permissions please email: journals.permissions@oxfordjournals.org

Expression of stromal-cell-derived factor-1 on circulating platelets is increased in patients with acute coronary syndrome and correlates with the number of CD34+ progenitor cells

Konstantinos Stellos1,*, Boris Bigalke1, Harald Langer1, Tobias Geisler1, Annika Schad1, Andreas Kögel3, Florian Pfaff1, Dimitrios Stakos2, Peter Seizer1, Iris Müller1, Patrick Htun1, Stephan Lindemann1 and Meinrad Gawaz1,*

1 Medizinische Klinik III, Kardiologie und Kreislauferkrankungen, Eberhard Karls-Universität Tübingen, Otfried-Müller Str. 10, 72076 Tübingen, Germany
2 University Cardiology Clinic, Democritus University of Thrace, Alexandroupolis, Greece
3 Institut Schreier, Statistical Support, Tübingen, Germany

Received 2 January 2008; revised 4 November 2008; accepted 27 November 2008; online publish-ahead-of-print 24 December 2008.

* Corresponding author. Tel: +49 70712983688, Fax: +49 7071295040, Email: konstantinos.stellos{at}med.uni-tuebingen.de (K.S.) or meinrad.gawaz{at}med.uni-tuebingen.de (M.G.)


   Abstract
Top
 Abstract
Introduction
 Subjects and methods
 Enzyme-linked immunosorbent...
 Results
 Discussion
 Funding
 References
 
Aims: Previous experimental studies have suggested that platelet stromal-cell-derived factor-1 (SDF-1) regulates mobilization and recruitment of haematopoietic progenitor cells supporting revascularization in mice. However, there are no clinical data available regarding platelet-bound SDF-1 in patients with acute coronary syndrome (ACS). The objective of this study was to evaluate the platelet-surface expression of SDF-1 in patients with ACS.

Methods and results: Patients with ACS (n = 418) showed a significantly enhanced SDF-1 expression on admission compared with those with stable angina pectoris (SAP, n = 486) [SAP (mean fluorescence intensity (MFI) ± SD): 13.48 ± 5.27; ACS: 18.45 ± 12.85; P < 0.001) independent of cardiovascular risk factors and medication. Enhanced platelet-bound SDF-1 expression was found in patients with reduced left ventricular ejection fraction (LVEF <55%) in comparison to patients with normal LVEF (P = 0.005). Platelet-bound SDF-1 expression positively correlated with the degree of platelet activation [CD62P: r = 0.325; glycoprotein VI (GPVI): r = 0.277; PAC-1: r = 0.501; P < 0.001 for all] and showed a significant, but slight association with plasma levels of SDF-1 (r = 0.084; P = 0.045). In a subgroup of patients with coronary artery disease, platelet-bound SDF-1, but not other platelet activation markers, significantly correlated with the number of circulating CD34+ progenitor cells (r = 0.252; P = 0.002) or CD34+/CD133+ endothelial progenitor cells (r = 0.352; P = 0.008).

Conclusion: Platelet-bound SDF-1 may play an important role in peripheral homing of circulating progenitor cells thus in tissue regeneration.

Key Words: SDF-1 • Platelets • Progenitor cells • Acute coronary syndrome


   Introduction
Top
 Abstract
 Introduction
Subjects and methods
 Enzyme-linked immunosorbent...
 Results
 Discussion
 Funding
 References
 
Circulating endothelial progenitor cells (EPCs) are mobilized endogenously in response to tissue ischaemia and thereby augment neovascularization of ischaemic tissues.1,2 In patients with acute myocardial infarction (AMI), EPCs, usually defined as CD34+ progenitor cells, are increased in peripheral blood and are rapidly recruited to myocardium mediating a protective effect of ischaemic preconditioning.3–5 Myocardial regeneration via stem-cell mobilization at the time of myocardial infarction is known to occur, although the mechanisms for stem-cell mobilization and subsequent domiciliation to infarcted tissue are poorly understood.

Stromal-cell-derived factor-1 (SDF-1, also known as CXCL12) is a powerful chemoattractant for stem and progenitor cells and is essential for both homing (migration, retention, and development) and trafficking (mobilization) of CD34+ progenitor cells to bone marrow or peripheral blood, respectively.6–10 Human CD34+ stem-cell transendothelial migration, engraftment, and repopulation of murine bone marrow11,12 and ischaemia-induced recruitment and differentiation to EPCs13–15 are found to be regulated by the chemokine SDF-1 binding to its receptor CXC chemokine receptor 4 (CXCR4), as thoroughly described in different in vitro and in vivo models. Overexpression of CXCR4 on human CD34+ progenitors increases their proliferation, migration, and bone marrow repopulation.16 Locally injected SDF-1 into ischaemic hindlimb muscle of immuno-deficient mice combined with human EPC transplantation results in an augmentation of EPC-induced vasculogenesis. This contribution to ischaemic neovascularization in vivo was mediated by increasing EPC recruitment into the ischaemic tissues.7 The SDF-1/CXCR4 axis seems particularly important in stem/muscle progenitor cell homing, chemotaxis, engraftment, and retention in ischaemic myocardium in vivo. SDF-1 is sufficient to induce therapeutic stem cells homing to injured myocardium and suggests a strategy for directed stem-cell engraftment into injured tissues.17 SDF-1/CXCR4 interactions play a crucial role in the recruitment of bone-marrow-derived cells to the heart after MI and can further increase homing in the presence, but not in the absence of injury in vivo.18 Overexpression of SDF-1 enhances endothelium-supported transmigration, maintenance, and proliferation of haematopoietic progenitor cells.19 Furthermore, hypoxia-inducible factor-1 (HIF-1)-induced SDF-1 expression increases the adhesion, migration, and homing of circulating CXCR4+ progenitor cells to ischaemic tissue.20 Blockade of SDF-1 in ischaemic tissue or inhibition of CXCR4 on circulating cells prevents progenitor cell recruitment to sites of injury, indicating that the recruitment of CXCR4+ progenitor cells to regenerating tissues is mediated by hypoxic gradients via HIF-1-induced expression of SDF-1.20 However, the origin, regulation, and the impact of SDF-1 in circumstances where tissue regeneration is needed are incompletely understood. Nevertheless, the pathophysiological role of SDF-1 in myocardial ischaemia has been mainly studied in animal models. These results cannot be uncritically transferred into human situation. Therefore, clinical studies are needed to elucidate the role of SDF-1 in humans.

Platelets are the first circulating blood cells that interact with the injured vessel wall playing a role not only in atherogenesis, but also in vascular and tissue regeneration.21,22 Recently, we reported that platelets recruit bone-marrow-derived progenitor cells to arterial thrombi in vivo involving platelet P-selectin and GPIIb/IIIa integrin and that activated platelets secrete SDF-1 supporting the migration of murine embryonic EPCs and the accumulation of bone-marrow-derived progenitor cells into the platelet-rich thrombus in vivo.14,23 Moreover, haematopoietic cytokines, through graded deployment of SDF-1 from platelets, support mobilization and recruitment of CXCR4+ vascular endothelial growth factor receptor-1 (VEGFR1)-positive haemangiocytes in mice.24 Furthermore, we recently showed that platelet-derived SDF-1 regulates vascular injury-dependent adhesion and promotes differentiation of human CD34+ cells to an endothelial phenotype.15 However, no study has addressed the role of platelet-bound SDF-1 in humans, specifically in patients with myocardial ischaemia or infarction.

The aim this study was to evaluate the platelet-bound SDF-1 expression and its correlation with platelet activation, plasma levels of SDF-1, and number of CD34+ progenitor cells in patients with coronary artery disease and/or reduced left ventricular (LV) function.


   Subjects and methods
Top
 Abstract
 Introduction
 Subjects and methods
Enzyme-linked immunosorbent...
 Results
 Discussion
 Funding
 References
 
Patients
Nine hundred and forty-three patients were initially assessed for inclusion into the study. Thirty-nine patients refused to give their consent and therefore were not included in our analysis. A total of 904 consecutive patients were recruited in a pre-planned time period. Four hundred and eighty-six patients with suspected or known coronary artery disease with typical symptoms for stable angina pectoris (SAP) were referred to our hospital for coronary angiography according to the ACC/AHA guidelines for coronary angiography.25 Patients with SAP had either typical angina on exertion and/or a pathological exercise test and were negative for markers of myocardial ischaemia (troponin, creatinine kinase). Four hundred and eighteen patients presented in our emergency room with an acute coronary syndrome (ACS), as previously defined,26,27 were immediately proceeded to percutaneous coronary intervention (PCI). In the means of a subgroup analysis, patients with an ACS were further classified as patients with a non-ST-segment elevation myocardial infarction (NSTEMI), a ST-segment myocardial infarction (STEMI), or an unstable angina with co-existence of positive troponin (Tn+ACS). Coronary interventions were performed within 6 h after admission to the hospital in patients with ACS. Patients with SAP were investigated on schedule. In a subgroup of 101 consecutive patients, we determined the expression of platelet-bound SDF-1 in relation to the time. The study was approved by the institutional ethical committee and all subjects gave their written informed consent.

Sample collection
Arterial blood was drawn from the sheath that was introduced into the femoral artery at the beginning of coronary intervention and after administration of 2500 U of unfractionated heparin. Arterial blood was filled into 5 mL vials containing citrate phosphate dextrose adenine (CPDA) and analysed by flow cytometry according to standard methods.28 Cardiac markers were determined at time of hospital admission and 4–6 and 12–24 h after cardiac catheterization. In the subgroup of 101 consecutive patients, we determined the expression of platelet-bound SDF-1 in relation to the time and therefore blood probes were collected every day after the coronary intervention till the fourth day.

Left ventricular function determination
Left ventricular function was determined by standard LV angiography (single-panel method) for evaluation of ejection fraction (EF). Based on a previous study describing the association of LVEF with mortality in stable outpatients with heart failure,29 we analysed the association between LVEF as a continuous variable with platelet-bound SDF-1.

Whole blood flow cytometry
Platelets obtained from 904 consecutive patients were studied for surface expression of SDF-1, platelet collagen receptor GPVI (4C9), and GPIb (CD42b) by flow cytometric analysis as previously described.30 Surface expression of P-selectin (CD62P) and activated fibrinogen receptor (PAC-1) was also analysed in subgroups of the above cohort. Conjugated monoclonal antibodies were used to measure platelet SDF-1 surface expression [R&D Systems, Minneapolis, MN, USA; clone 79014; fluorescein isothiocyanate (FITC)], P-selectin (CD62P, Immunotec, Marseille, France; clone CLB-Thromb/6; FITC), GPIb [CD42b, Immunotec, Marseille, France; clone SZ2; phycoerythrin (PE)], and activated form of GPIIb/IIIa (PAC-1, Becton Dickinson, USA; clone SP-2) with a two-colour flow cytometry in patients' whole blood as previously described. The anti-GPVI mAb 4C9 was generated and characterized as previously described.31,32 In brief, 10 µl of CPDA-blood was resuspended 50:1 with phosphate-buffered saline (PBS; Invitrogen Corporation, Paisley, Scotland, UK) and was incubated for 30 min with the relevant conjugated antibodies in the dark at room temperature. After staining, the cells were fixed with 0.5% paraformaldehyde and stored at 4°C until fluorescence-activated cell sorting (FACS) was performed with a FACS-Calibur Flow Cytometer (Becton-Dickinson, Heidelberg, Germany). CD42b-PE served as a control antibody to identify the platelet population in the whole blood. Specific monoclonal antibody binding was expressed as mean fluorescence intensity (MFI) and was used as a quantitative measurement of platelet proteins' surface expression.


   Enzyme-linked immunosorbent assay
Top
 Abstract
 Introduction
 Subjects and methods
 Enzyme-linked immunosorbent...
Results
 Discussion
 Funding
 References
 
Plasma levels of SDF-1 were determined in a subgroup of 566 consecutive patients with symptomatic coronary artery disease on time of platelet determination using a commercially available enzyme-linked immunosorbent assay kit according to the manufacturer's guidelines (R&D Systems, Minneapolis, MN, USA). Ethylenediamine tetraacetic acid plasma probes were centrifuged for 15 min at 10 000 g within 30 min of collection. Probes were aliquotted and stored at –20°C before analysis. The lower detection limit of this assay is 18 pg/mL. The mean centred coefficient of variation for soluble SDF-1 was 3.2%, thus allowing a relatively good reproducibility of our measurements.

Peripheral blood mononuclear cell isolation and flow cytometry
Arterial blood was drawn from the femoral artery before cardiac catheterization and buffered with 20 mL of sodium citrate. Mononuclear cells were isolated using a Ficoll density gradient (Biocoll, Biochrom, Berlin, Germany) according to standard protocols, as previously described.33 For FACS analysis, mononuclear cells were resuspended in 100 µL of PBS. Immunofluorescence cell staining was performed in duplicate with the use of the fluorescent conjugated antibody CD34–FITC (10 µL; Becton Dickinson, San Jose, USA; clone 8G12). IgG1–FITC antibody (BD Biosciences Pharmingen, USA; clone MOPC-21) served as a negative isotype control. Cell fluorescence was measured immediately after staining, and data were analysed with the use of CellQuest software (FACSCalibur, Becton Dickinson, Heidelberg, Germany). Units of all measured components are absolute cell counts obtained after the measurement of 250 000 events in the lymphocyte gate.

Data presentation and statistical analysis
Data are presented as mean ± standard deviation (SD). Continuous variables were tested for normal distribution with the Kolmogorov–Smirnov test. The Mann–Whitney U-test and the Kruskal–Wallis test were used to assess differences between two or three groups, respectively. The paired-sample t-test was applied to test the difference of platelet SDF-1 MFI in patients with ACS between day 1 and days 2–4 and the difference of the number of CD34+ cells between time point 0 and 24 h. Comparison of categorical variables was generated by the Pearson {chi}2 test. Correlations were assessed with the Pearson correlation coefficient test after logarithmic transformation of the data. A univariate analysis of variance was used to estimate the impact of different factors on SDF-1 expression or number of CD34+ cells. All tests were two-tailed, and statistical significance was considered for P-values <0.05. All statistical analyses were performed using SPSS version 13 for windows (Chicago, IL, USA).


   Results
Top
 Abstract
 Introduction
 Subjects and methods
 Enzyme-linked immunosorbent...
 Results
Discussion
 Funding
 References
 
Platelet-surface expression of stromal-cell-derived factor-1 is increased in acute coronary syndrome
We first analysed the surface expression of SDF-1 in a consecutive cohort of 904 patients with symptomatic coronary artery disease including ACS (n = 418) and stable coronary artery disease (SAP; n = 486). In all patients, the diagnosis was verified and the severity of the disease was assessed by coronary angiography. The demographic details are given in Table 1. We found that platelet SDF-1 surface expression was significantly enhanced in ACS compared with SAP (ACS (MFI ± SD): 18.45 ± 12.85; SAP: 13.48 ± 5.27; P < 0.001; Figure 1A and B). Patients with ACS were further subdivided into patients with STEMI, NSTEMI, or troponin I-positive ACS (Tn+ACS). We did not observe any statistical significant difference between these subgroups. However, each of these subgroups showed a significant increase in platelet SDF-1 expression levels when compared with patients with SAP (Figure 1C).


View this table:
[in this window]
[in a new window]

  		Table 1 Baseline patients’ characteristics

 

Figure 1
View larger version (26K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Figure 1 Platelet-surface expression of stromal-cell-derived factor-1 is elevated in patients with acute coronary syndrome (ACS) compared with patients with stable angina pectoris (SAP). (A) Representative immunofluorescence histogram showing an overlay of platelet-surface expression of stromal-cell-derived factor-1 of a patient with stable angina pectoris (grey line) or acute coronary syndrome (black line). (B) Increased platelet-bound stromal-cell-derived factor-1 expression was observed in patients with acute coronary syndrome in comparison to those with stable angina pectoris. (C) Expression of platelet stromal-cell-derived factor-1 in patients with troponin I-positive acute coronary syndrome (Tn+ACS), ST-segment myocardial infarction, or non-ST-segment elevation myocardial infarction. The double-line indicates the mean stromal-cell-derived factor-1 value in patients with stable angina pectoris. *P ≤ 0.05 when compared with stable angina pectoris. (D) Patients with stable angina pectoris or acute coronary syndrome were followed-up for up to 1 or 4 days, respectively, in order to examine the expression of platelet stromal-cell-derived factor-1 in relation to time. *P ≤ 0.05 when compared with respective stable angina pectoris. #P ≤ 0.05 when compared days 2–4 with day 1.

 
SDF-1 expression neither correlated nor was influenced, respectively, by cardiovascular risk factors (arterial hypertension, hyperlipidaemia, diabetes mellitus, family history of coronary artery disease, and smoking), the extent of the disease (1/2/3-vessel disease), initial creatinine kinase, troponin, gender, or the received baseline medications (Table 2). C-reactive protein (CRP) slightly, but significantly, influences the levels of SDF-1 (point estimate equal to 0.52). On the other hand, no significant correlation was observed between platelet-bound SDF-1 and CRP (data not shown).


View this table:
[in this window]
[in a new window]

  		Table 2 Univariate analysis of variance for platelet-bound stromal-cell-derived factor-1 and possible confounders for acute coronary syndrome

 
In order to determine the expression of SDF-1 over the time, 101 consecutive patients with SAP (n = 73) or ACS (n = 28) were followed-up for 24 h. The patients with ACS were further followed-up for up to 4 days measuring their SDF-1 expression every 24 h. We found that platelet-surface expression of SDF-1 remained significantly increased for 24 h in patients with ACS being followed by a significant decrease in the following days after the acute event (Figure 1D).

Stromal-cell-derived factor-1 is increased in patients with reduced left ventricular function
Enhanced platelet-bound SDF-1 expression was found in patients with reduced left ventricular ejection fraction (LVEF <55%; n = 434) in comparison to patients with normal LVEF (>55%; n = 470; P = 0.005; Figure 2A). In patients with SAP, platelet-bound SDF-1 expression was increased in patients with reduced LVEF (n = 209) when compared with patients with normal LVEF (n = 277; MFI ± SD: normal vs. reduced LVEF: 13.35 ± 7.08 vs. 15.03 ± 8.59; P = 0.019; Figure 2B). In a similar manner, in patients with ACS, platelet-bound SDF-1 expression was increased in patients with reduced LVEF (n = 225) when compared with patients with normal LVEF (n = 193; MFI ± SD: normal vs. reduced LVEF: 15.88 ± 10.47 vs. 19.33 ± 12.23; P = 0.009; Figure 2B). Assessing LVEF and SDF-1 expression as continuous variables, we observed a poor, but significant correlation between LVEF (%) and SDF-1 MFI (r =–0.087, P = 0.014). In a similar manner like in patients with ACS, platelet-bound SDF-1 is increased in patients with reduced LVEF independent of other factors including baseline medication and cardiovascular risk factors (data not shown). Furthermore, we assessed potential correlations among LVEF and initial or maximal myocardial necrosis markers (initial Tn: r =–0.11, P = 0.003; initial CK: r =–0.13, P = 0.724; maximal Tn: r =–0.142, P < 0.001; maximal CK: r =–0.089, P = 0.011).


Figure 2
View larger version (14K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Figure 2 Platelet-bound stromal-cell-derived factor-1 is increased in patients with reduced left ventricular function. (A) Patients with reduced left ventricular ejection fraction (LVEF <55%) presented with significantly increased platelet-bound stromal-cell-derived factor-1 expression. (B) Enhanced stromal-cell-derived factor-1 expression was observed in patients with stable angina pectoris or acute coronary syndrome and reduced left ventricular ejection fraction. *P ≤ 0.05 when compared with normal left ventricular ejection fraction.

 
Similar to platelet-bound SDF-1 expression, plasma levels of SDF-1 were increased in patients with reduced LV function (LVEF <55%; n = 275) compared with patients with normal LVEF (>55%; n = 291; P = 0.007). In patients with SAP, plasma levels of SDF-1 did not show a significant difference between normal and reduced LV function (P = 0.393). In patients with ACS, plasma levels of SDF-1 were significantly increased in patients with reduced LV function (n = 137) when compared with patients with normal LV function (n = 117; mean ± SD: normal vs. reduced: 1940 ± 390 pg/mL vs. 2122.4 ± 512 pg/mL; P = 0.004).

Platelet-bound stromal-cell-derived factor-1 and plasma levels of stromal-cell-derived factor-1
In a subgroup of 566 consecutive patients, we determined both platelet SDF-1 expression and plasma levels of SDF-1. We observed that platelet-bound SDF-1 poorly correlates with plasma SDF-1 (n = 566, r = 0.084, P = 0.045; see Figure 3). In order to further investigate the association of platelet-bound SDF-1 with plasma SDF-1, we further analysed the correlation between these parameters. Checking interaction effects via a univariate ANOVA model showed no significant effect of either ACS/SAP group or the interaction between ACS/SAP group and platelet-bound SDF-1 on the levels of plasma SDF-1. The only significant effect was the main effect of platelet-bound SDF-1 (P = 0.041, point estimate: 5.917; lower–upper 95% CI: 0.844–10.991).


Figure 3
View larger version (26K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Figure 3 Platelet-bound stromal-cell-derived factor-1 correlates with plasma stromal-cell-derived factor-1 in patients with acute coronary syndrome. The correlation of platelet-surface expression with plasma levels of stromal-cell-derived factor-1 in patients with coronary artery disease is shown (n = 566).

 
Platelet-bound stromal-cell-derived factor-1 correlates with platelet activation markers
It was previously reported that platelet-bound P-selectin, GPVI, and fibrinogen receptor GPIIb/IIIa are increased in patients with ACS.28,30 We found in this study that surface expression of SDF-1 on circulating platelets of patients with coronary artery disease correlated with the degree of P-selectin expression, thus systemic platelet activation (r = 0.325, P < 0.001; Figure 4), GPVI (r = 0.277, P < 0.001; Figure 4), and PAC-1 (r = 0.501, P < 0.001).


Figure 4
View larger version (34K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Figure 4 Platelet-bound stromal-cell-derived factor-1 correlates with known platelet activation biomarkers. Platelet-surface expression of stromal-cell-derived factor-1 correlates with platelet-bound P-selectin (Plt-CD62P; upper panel) and glycoprotein VI (Plt-GPVI; lower panel) at the time of hospital admission.

 
Platelet-bound stromal-cell-derived factor-1 correlates with the number of CD34+ progenitor cells
As SDF-1 is a potent chemokine for CD34+ progenitor cells, we asked whether there was a correlation between the expression of platelet SDF-1 and the number of circulating CD34+ progenitor cells. In 144 consecutive patients with coronary artery disease, i.e. SAP (n = 68) and ACS (n = 76), we measured the number of CD34+ progenitor cells using a standard protocol.33 We found that increased progenitor cells circulate in the blood of patients with ACS in comparison to patients with SAP (absolute number of CD34+ cells: ACS vs. SAP: mean ± SD: 243.2 ± 148 vs. 152.3 ± 96.3, P = 0.001; Figure 5). The number of CD34+ progenitor cells in our population is increased in patients with ACS independent of cardiovascular risk factors, age, gender, and extension of coronary artery disease, documented as number of coronary arteries affected (Table 3). A not significant tendency (P = 0.055) was observed for left ventricular function. On the other hand, intake of aspirin, but not other medication, seems to influence the number of CD34+ cells in patients with ACS (P = 0.017).


Figure 5
View larger version (27K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		Figure 5 Platelet-bound stromal-cell-derived factor-1 correlates with the number of CD34+ progenitor cells in patients with coronary artery disease. (A) The left panel shows a representative forward scatter and side scatter histogram of peripheral blood mononuclear cells. CD34+ progenitor cells were counted using the lymphocyte gate (R1); the middle panel or the right panel demonstrates a representative forward scatter and CD34-FITC immunofluorescence histogram in a patient with stable angina pectoris or acute coronary syndrome, respectively. (B) Increased number of CD34+ progenitor cells was observed in patients with acute coronary syndrome compared with patients with stable angina pectoris. (C) Platelet-bound stromal-cell-derived factor-1 significantly correlated with the absolute number of CD34+ progenitor cells (left) and CD34+/CD133+ endothelial progenitor cells (right).

 

View this table:
[in this window]
[in a new window]

  		Table 3 Univariate analysis of variance for number of CD34+ cells and possible confounders for acute coronary syndrome

 
In a subgroup of 12 patients with ACS and 4 patients with SAP, we measured the number of CD34+ cells at time point 0 (admission to hospital) and 24 h after the first measurement. The number of CD34+ progenitor cells is significantly increased on the time course in patients with ACS (time point 0 vs. time point 24 h: mean ± SD: 166.17 ± 65.9 vs. 205.92 ± 93.19; P = 0.031), whereas there was no significant difference in patients with SAP (time point 0 vs. time point 24 h: mean ± SD: 100.25 ± 41.8 vs. 121.00 ± 31.5; P = 0.5).

Moreover, the number of CD34+ progenitor cells correlated with the expression of platelet-bound SDF-1 (n = 144, r = 0.253, P = 0.002; Figure 5C), but not with the expression of P-selectin (r = 0.086, P = 0.304) or PAC-1 (r =– 0.70, P = 0.448). Similar correlation was observed in a subpopulation of our study (n = 56) between EPCs, defined as CD34+/CD133+ cells, and platelet-bound SDF-1 expression (r = 0.352, P = 0.008; Figure 5C).


   Discussion
Top
 Abstract
 Introduction
 Subjects and methods
 Enzyme-linked immunosorbent...
 Results
 Discussion
Funding
 References
 
The major findings of the present study are: (1) platelet-surface expression of SDF-1 is elevated in patients with ACS compared with patients with SAP; (2) it is increased in patients with reduced LV function compared with patients with normal LVEF; (3) it is slightly associated with plasma SDF-1 on admission day and correlates well with the degree of platelet activation; and (4) in a subgroup of patients, platelet-bound SDF-1 correlates with the number of circulating CD34+ progenitor cells.

CD34+ cells have been described to be recruited to the ischaemic myocardium, differentiating into cardiac and vascular cells, and restoring cardiac function.34 Recent studies have identified specific molecular signals, such as SDF-1/CXCR4, required for the interaction of bone-marrow-derived stem cells and damaged host tissues. SDF-1, the ligand for CXCR4, plays a crucial role in trafficking of CXCR4+ bone-marrow-derived cells into peripheral blood and their recruitment to damaged organs in vivo. SDF-1 secretion is increased during tissue damage such as myocardial infarction34 and hind-limb ischaemia.7,20,35 Local delivery of SDF-1 can enhance progenitor cell recruitment and neovascularization.17 Combined delivery of SDF-1 and EPCs into sites of limb ischaemia promotes local EPC-mediated vasculogenesis.17 SDF-1 has also been shown to promote bone marrow cell proliferation and angiogenesis.17 Platelet-derived SDF-1 has been recently described to play a role in chemotaxis, adhesion, and differentiation of both murine and human progenitor cells mediating vascular regeneration in vivo.14,15,24 Although SDF-1 effects have been relatively extensively studied in vitro and animal models, the role of SDF-1 in humans and especially in patients with myocardial ischaemia has been poorly described so far. The present study describes the expression of platelet-bound SDF-1 in circulating platelets in patients with ACS and/or reduced LV function, indicating that in cases where regeneration is needed platelet-bound SDF-1 is found to be increased.

Endothelial progenitor cells are mobilized in response to vascular injury as a physiologic response.36 Cardiovascular risk factors contribute to atherogenesis not only by inducing endothelial cell injury and dysfunction, but also by influencing the number and the function of EPCs, the cells responsible for vascular homeostasis and regeneration.37 There are numerous physiological and pathological stimuli which influence the number of circulating EPCs such as regular physical activity, age, medications (statins, estrogens, PPAR-gamma agonists), as well as numerous inflammatory and haematopoietic cytokines including VEGF, SDF-1, granulocyte- colony-stimulating factor, angiopoietin, and erythropoietin.38 Endothelial injury in the absence of sufficient circulating progenitor cells affects the progression of cardiovascular disease in subjects with various cardiovascular risk factors.37 Taking into consideration the known role of SDF-1 in mobilization and function of progenitor cells, the present study reports for the first time correlative findings between platelet-bound SDF-1 and number of progenitor cells in patients with coronary artery disease. On the other hand, ischaemia alone or any of its confounders could influence the expression of platelet-bound SDF-1, the LV function, and the number of circulating EPCs in patients with ACS, and therefore further studies are needed to elucidate this pathophysiological mechanism.

The present findings imply that increased surface expression of platelet-bound SDF-1 takes place in cases of acute or chronic tissue ischaemia, such as ACS or reduced systolic ventricular function, and in diseases or pathophysiological phenomena characterized by enhanced platelet activation such as stroke, sepsis, and inflammation. Therefore, in diseases where tissue regeneration is demanded, platelet-bound SDF-1 may be increased. In addition, platelet-surface-bound SDF-1 slightly correlates with plasma SDF-1 and with the number of circulating CD34+ progenitor cells in patients with ACS, a phenomenon which could potentially and/or partially explain the reported mobilization of EPCs in those patients.

Domiciliation of progenitor cells in peripheral tissue is a multistep cascade including the initial adhesion to activated endothelium or exposed matrix, transmigration through the endothelium, and invasion of the target tissue.34 Platelets are the first circulating blood cells that adhere to vascular lesions and that accumulate in the microcirculation within ischaemic tissue.31 Recently, we described that adherent platelets express substantial amounts of SDF-1, recruit EPCs in vitro and in vivo, and induce their subsequent differentiation towards an endothelial phenotype.14,39 Moreover, we have observed that platelet-bound SDF-1 regulates recruitment of CD34+ progenitor cells in vitro and in vivo, and differentiation of the latter into endothelial cells.15 Therefore, although the exact clinical significance of platelet-bound SDF-1 in patients with ACS remains to be elucidated, it is tempting to speculate that platelet-bound SDF-1 may play a role in vascular and myocardial remodelling or regeneration in patients with AMI. Understanding the role of platelets and platelet-derived SDF-1 in progenitor cell function and tissue regeneration may help to develop novel therapeutic strategies in treatment of patients with myocardial damage.


   Funding
Top
 Abstract
 Introduction
 Subjects and methods
 Enzyme-linked immunosorbent...
 Results
 Discussion
 Funding
References
 
The study was supported by the Deutsche Forschungsgemeinschaft (DFG) Sonderforschungsbereich/Transregio 19 ‘Inflammatorische Kardiomyopathie- Molekulare Pathogenese und Therapie’ to K.S., S.L., and M.G. and by the DFG 849/3/1 to S.L.

Conflict of interest: The authors declare that none of them pertain a relationship with pharmaceutical companies, biomedical device manufacturers, or other corporations whose products or services are related to the subject matter of the article.


   References
Top
 Abstract
 Introduction
 Subjects and methods
 Enzyme-linked immunosorbent...
 Results
 Discussion
 Funding
 References
 

  1. Takahashi T, Kalka C, Masuda H, Chen D, Silver M, Kearney M, Magner M, Isner JM, Asahara T. Ischemia- and cytokine-induced mobilization of bone-marrow-derived endothelial progenitor cells for neovascularization. Nat Med (1999) 5:434–438.[CrossRef][Web of Science][Medline]
  2. Iwami Y, Masuda H, Asahara T. Endothelial progenitor cells: past, state of the art, and future. J Cell Mol Med (2004) 8:488–497.[Web of Science][Medline]
  3. Feugier P, Li N, Jo DY, Shieh JH, MacKenzie KL, Lesesve JF, Latger-Cannard V, Bensoussan D, Crystal RG, Rafii S, Stoltz JF, Moore MA. Osteopetrotic mouse stroma with thrombopoietin, c-kit ligand, and flk-2 ligand supports long-term mobilized CD34+ hematopoiesis in vitro. Stem Cells Dev (2005) 14:505–516.[CrossRef][Web of Science][Medline]
  4. Shintani S, Murohara T, Ikeda H, Ueno T, Honma T, Katoh A, Sasaki K, Shimada T, Oike Y, Imaizumi T. Mobilization of endothelial progenitor cells in patients with acute myocardial infarction. Circulation (2001) 103:2776–2779.[Abstract/Free Full Text]
  5. Wojakowski W, Tendera M, Michalowska A, Majka M, Kucia M, Maslankiewicz K, Wyderka R, Ochala A, Ratajczak MZ. Mobilization of CD34/CXCR4+, CD34/CD117+, c-met+ stem cells, and mononuclear cells expressing early cardiac, muscle, and endothelial markers into peripheral blood in patients with acute myocardial infarction. Circulation (2004) 110:3213–3220.[Abstract/Free Full Text]
  6. Petit I, Goichberg P, Spiegel A, Peled A, Brodie C, Seger R, Nagler A, Alon R, Lapidot T. Atypical PKC-zeta regulates SDF-1-mediated migration and development of human CD34+ progenitor cells. J Clin Invest (2005) 115:168–176.[CrossRef][Web of Science][Medline]
  7. Yamaguchi J, Kusano KF, Masuo O, Kawamoto A, Silver M, Murasawa S, Bosch-Marce M, Masuda H, Losordo DW, Isner JM, Asahara T. Stromal-cell- derived factor-1 effects on ex vivo expanded endothelial progenitor cell recruitment for ischemic neovascularization. Circulation (2003) 107:1322–1328.[Abstract/Free Full Text]
  8. Aiuti A, Webb IJ, Bleul C, Springer T, Gutierrez-Ramos JC. The chemokine SDF-1 is a chemoattractant for human CD34+ hematopoietic progenitor cells and provides a new mechanism to explain the mobilization of CD34+ progenitors to peripheral blood. J Exp Med (1997) 185:111–120.[Abstract/Free Full Text]
  9. Goichberg P, Kalinkovich A, Borodovsky N, Tesio M, Petit I, Nagler A, Hardan I, Lapidot T. cAMP-induced PKCzeta activation increases functional CXCR4 expression on human CD34+ hematopoietic progenitors. Blood (2006) 107:870–879.[Abstract/Free Full Text]
  10. Peled A, Grabovsky V, Habler L, Sandbank J, Arenzana-Seisdedos F, Petit I, Ben-Hur H, Lapidot T, Alon R. The chemokine SDF-1 stimulates integrin- mediated arrest of CD34(+) cells on vascular endothelium under shear flow. J Clin Invest (1999) 104:1199–1211.[Web of Science][Medline]
  11. Peled A, Petit I, Kollet O, Magid M, Ponomaryov T, Byk T, Nagler A, Ben-Hur H, Many A, Shultz L, Lider O, Alon R, Zipori D, Lapidot T. Dependence of human stem cell engraftment and repopulation of NOD/SCID mice on CXCR4. Science (1999) 283:845–848.[Abstract/Free Full Text]
  12. Peled A, Kollet O, Ponomaryov T, Petit I, Franitza S, Grabovsky V, Slav MM, Nagler A, Lider O, Alon R, Zipori D, Lapidot T. The chemokine SDF-1 activates the integrins LFA-1, VLA-4, and VLA-5 on immature human CD34(+) cells: role in transendothelial/stromal migration and engraftment of NOD/SCID mice. Blood (2000) 95:3289–3296.[Abstract/Free Full Text]
  13. De Falco E, Porcelli D, Torella AR, Straino S, Iachininoto MG, Orlandi A, Truffa S, Biglioli P, Napolitano M, Capogrossi MC, Pesce M. SDF-1 involvement in endothelial phenotype and ischemia-induced recruitment of bone marrow progenitor cells. Blood (2004) 104:3472–3482.[Abstract/Free Full Text]
  14. Massberg S, Konrad I, Schurzinger K, Lorenz M, Schneider S, Zohlnhoefer D, Hoppe K, Schiemann M, Kennerknecht E, Sauer S, Schulz C, Kerstan S, Rudelius M, Seidl S, Sorge F, Langer H, Peluso M, Goyal P, Vestweber D, Emambokus NR, Busch DH, Frampton J, Gawaz M. Platelets secrete stromal-cell-derived factor 1alpha and recruit bone-marrow-derived progenitor cells to arterial thrombi in vivo. J Exp Med (2006) 203:1221–1233.[Abstract/Free Full Text]
  15. Stellos K, Langer H, Daub K, Schoenberger T, Gauss A, Geisler T, Bigalke B, Mueller I, Schumm M, Schaefer I, Seizer P, Kraemer B, Siegel-Axel D, May A, Lindemann S, Gawaz M. Platelet-derived SDF-1 regulates adhesion and promotes differentiation of human CD34+ cells to endothelial progenitor cells. Circulation (2008) 117:206–215.[Abstract/Free Full Text]
  16. Kahn J, Byk T, Jansson-Sjostrand L, Petit I, Shivtiel S, Nagler A, Hardan I, Deutsch V, Gazit Z, Gazit D, Karlsson S, Lapidot T. Overexpression of CXCR4 on human CD34+ progenitors increases their proliferation, migration, and NOD/SCID repopulation. Blood (2004) 103:2942–2949.[Abstract/Free Full Text]
  17. Askari AT, Unzek S, Popovic ZB, Goldman CK, Forudi F, Kiedrowski M, Rovner A, Ellis SG, Thomas JD, DiCorleto PE, Topol EJ, Penn MS. Effect of stromal-cell-derived factor 1 on stem-cell homing and tissue regeneration in ischaemic cardiomyopathy. Lancet (2003) 362:697–703.[CrossRef][Web of Science][Medline]
  18. Abbott JD, Huang Y, Liu D, Hickey R, Krause DS, Giordano FJ. Stromal-cell-derived factor-1alpha plays a critical role in stem cell recruitment to the heart after myocardial infarction but is not sufficient to induce homing in the absence of injury. Circulation (2004) 110:3300–3305.[Abstract/Free Full Text]
  19. Hwang JH, Kim SW, Park SE, Yun HJ, Lee Y, Kim S, Jo DY. Overexpression of stromal-cell-derived factor-1 enhances endothelium-supported transmigration, maintenance, and proliferation of hematopoietic progenitor cells. Stem Cells Dev (2006) 15:260–268.[CrossRef][Web of Science][Medline]
  20. Ceradini DJ, Kulkarni AR, Callaghan MJ, Tepper OM, Bastidas N, Kleinman ME, Capla JM, Galiano RD, Levine JP, Gurtner GC. Progenitor cell trafficking is regulated by hypoxic gradients through HIF-1 induction of SDF-1. Nat Med (2004) 10:858–864.[CrossRef][Web of Science][Medline]
  21. Stellos K, Gawaz M. Platelet interaction with progenitor cells: potential implications for regenerative medicine. Thromb Haemost (2007) 98:922–929.[Web of Science][Medline]
  22. Lindemann S, Kraemer B, Daub K, Stellos K, Gawaz M. Molecular pathways used by platelets to initiate and accelerate atherogenesis. Curr Opin Lipidol (2007) 18:566–573.[Medline]
  23. Stellos K, Gawaz M. Platelets and stromal-cell-derived factor-1 in progenitor cell recruitment. Semin Thromb Hemost (2007) 33:159–164.[CrossRef][Web of Science][Medline]
  24. Jin DK, Shido K, Kopp HG, Petit I, Shmelkov SV, Young LM, Hooper AT, Amano H, Avecilla ST, Heissig B, Hattori K, Zhang F, Hicklin DJ, Wu Y, Zhu Z, Dunn A, Salari H, Werb Z, Hackett NR, Crystal RG, Lyden D, Rafii S. Cytokine-mediated deployment of SDF-1 induces revascularization through recruitment of CXCR4+ hemangiocytes. Nat Med (2006) 12:557–567.[CrossRef][Web of Science][Medline]
  25. Scanlon PJ, Faxon DP, Audet AM, Carabello B, Dehmer GJ, Eagle KA, Legako RD, Leon DF, Murray JA, Nissen SE, Pepine CJ, Watson RM, Ritchie JL, Gibbons RJ, Cheitlin MD, Gardner TJ, Garson A Jr, Russell RO Jr, Ryan TJ, Smith SC Jr. ACC/AHA guidelines for coronary angiography: executive summary and recommendations. A report of the American College of Cardiology/American Heart Association Task Force on Practice Guidelines (Committee on Coronary Angiography) developed in collaboration with the Society for Cardiac Angiography and Interventions. Circulation (1999) 99:2345–2357.[Free Full Text]
  26. Gibler WB, Cannon CP, Blomkalns AL, Char DM, Drew BJ, Hollander JE, Jaffe AS, Jesse RL, Newby LK, Ohman EM, Peterson ED, Pollack CV. Practical implementation of the guidelines for unstable angina/non-ST-segment elevation myocardial infarction in the emergency department: a scientific statement from the American Heart Association Council on Clinical Cardiology (Subcommittee on Acute Cardiac Care), Council on Cardiovascular Nursing, and Quality of Care and Outcomes Research Interdisciplinary Working Group, in Collaboration with the Society of Chest Pain Centers. Circulation (2005) 111:2699–2710.[Abstract/Free Full Text]
  27. Alpert JS, White HD, Jaffe ASAF, Galvani M, Katus HA, Newby LK, Ravkilde J, Chaitman B, Clemmensen PMDM, Hod H, Porela P, Underwood R, Bax JJ, Beller GABR, Van Der Wall EE, Bassand JP, Wijns W, Ferguson TBSP, Uretsky BF, Williams DO, Armstrong PW, Fox KA, Hamm CW, Ohman EM, Simoons ML, Gurfinkel EP, Lopez-Sendon JL, Pais P, Mendis SZJIG, Wallentin LC, Fernández-Avilés F, Fox KM, Parkhomenko ANPS, Tendera M, Voipio-Pulkki LM, Camm AJ, De Caterina R, Dean V, Dickstein K, Filippatos GF-BC, Hellemans I, Kristensen SD, McGregor K, Sechtem USS, Tendera M, Widimsky P, Zamorano JL, Morais J, Brener SHR, Morrow D, Sechtem U, Lim M, Martinez-Rios MA, Steinhubl SLG, Gibler WB, Goff D, Tubaro M, Dudek D, Al-Attar N, Biomarker Group, Task Force Members TK, ECG Group, Imaging Group, Intervention Group, Clinical Investigation Group, Global, Perspective Group P-WP, ESC Committee for, Practice Guidelines VA, Document Reviewers. Universal definition of myocardial infarction: Kristian Thygesen, Joseph S. Alpert and Harvey D. White on behalf of the Joint ESC/ACCF/AHA/WHF Task Force for the Redefinition of Myocardial Infarction. Eur Heart J (2007) 28:2525–2538.[Free Full Text]
  28. Bigalke B, Lindemann S, Ehlers R, Seizer P, Daub K, Langer H, Schonberger T, Kremmer E, Siegel-Axel D, May AE, Gawaz M. Expression of platelet collagen receptor glycoprotein VI is associated with acute coronary syndrome. Eur Heart J (2006) 27:2165–2169.[Abstract/Free Full Text]
  29. Curtis JP, Sokol SI, Wang Y, Rathore SS, Ko DT, Jadbabaie F, Portnay EL, Marshalko SJ, Radford MJ, Krumholz HM. The association of left ventricular ejection fraction, mortality, and cause of death in stable outpatients with heart failure. J Am Coll Cardiol (2003) 42:736–742.[Abstract/Free Full Text]
  30. Gawaz M, Neumann FJ, Schomig A. Evaluation of platelet membrane glycoproteins in coronary artery disease: consequences for diagnosis and therapy. Circulation (1999) 99:E1–E11.[Medline]
  31. Massberg S, Konrad I, Bultmann A, Schulz C, Munch G, Peluso M, Lorenz M, Schneider S, Besta F, Muller I, Hu B, Langer H, Kremmer E, Rudelius M, Heinzmann U, Ungerer M, Gawaz M. Soluble glycoprotein VI dimer inhibits platelet adhesion and aggregation to the injured vessel wall in vivo. FASEB J (2004) 18:397–399.[Abstract/Free Full Text]
  32. Cabeza N, Li Z, Schulz C, Kremmer E, Massberg S, Bultmann A, Gawaz M. Surface expression of collagen receptor Fc receptor-gamma/glycoprotein VI is enhanced on platelets in type 2 diabetes and mediates release of CD40 ligand and activation of endothelial cells. Diabetes (2004) 53:2117–2121.[Abstract/Free Full Text]
  33. Werner N, Kosiol S, Schiegl T, Ahlers P, Walenta K, Link A, Bohm M, Nickenig G. Circulating endothelial progenitor cells and cardiovascular outcomes. N Engl J Med (2005) 353:999–1007.[Abstract/Free Full Text]
  34. Urbich C, Aicher A, Heeschen C, Dernbach E, Hofmann WK, Zeiher AM, Dimmeler S. Soluble factors released by endothelial progenitor cells promote migration of endothelial cells and cardiac resident progenitor cells. J Mol Cell Cardiol (2005) 39:733–742.[CrossRef][Web of Science][Medline]
  35. Korbling M, Estrov Z. Adult stem cells for tissue repair—a new therapeutic concept? N Engl J Med (2003) 349:570–582.[Free Full Text]
  36. Banerjee S, Brilakis E, Zhang S, Roesle M, Lindsey J, Philips B, Blewett C, Terada L. Endothelial progenitor cell mobilization after percutaneous coronary intervention. Atherosclerosis (2006) 189:70–75.[CrossRef][Web of Science][Medline]
  37. Hill J, Zalos G, Halcox J, Schenke W, Waclawiw M, Quyyumi A, Finkel T. Circulating endothelial progenitor cells, vascular function, and cardiovascular risk. N Engl J Med (2003) 348:593–600.[Abstract/Free Full Text]
  38. Wojakowski W, Tendera M. Mobilization of bone-marrow-derived progenitor cells in acute coronary syndromes. Folia Histochem Cytobiol (2005) 43:229–232.[Medline]
  39. Daub K, Langer H, Seizer P, Stellos K, May A, Goyal P, Bigalke B, Schönberger T, Geisler T, Siegel-Axel D, Oostendorp R, Lindemann SMG. Platelets induce differentiation of human CD34+-progenitor cells into foam cells and endothelial cells. FASEB J (2006) 20:2559–2561.[Abstract/Free Full Text]

Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?


This article has been cited by other articles:


Home page
 J Am Coll CardiolHome page
J. J. Bax, B. Casadei, C. Di Mario, R. Fagard, G. Filippatos, K. A.A. Fox, M. Metra, P. Nihoyannopoulos, J. Perk, F. Rademakers, et al.
Highlights of the 2009 scientific sessions of the European society of cardiology.
J. Am. Coll. Cardiol., December 15, 2009; 54(25): 2447 - 2458.
[Full Text] [PDF]

Home page
 J Am Coll CardiolHome page
R. P. Giugliano and E. Braunwald
The Year in Non-ST-Segment Elevation Acute Coronary Syndrome.
J. Am. Coll. Cardiol., October 13, 2009; 54(16): 1544 - 1555.
[Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow All Versions of this Article:
30/5/584    most recent
ehn566v1
Right arrow E-letters: Submit a response
Right arrow Alert me when this article is cited
Right arrow Alert me when E-letters are posted
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Stellos, K.
Right arrow Articles by Gawaz, M.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Stellos, K.
Right arrow Articles by Gawaz, M.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?