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Abstract
Background Increased serum osteoprotegerin has been shown to be associated with increased mortality and heart failure development in patients with acute coronary syndromes. The aim of the present study was to elucidate a possible association between serum osteoprotegerin measured acutely in patients with ST-elevation myocardial infarction (STEMI) and final infarct size.
Methods Serum osteoprotegerin was measured in fasting blood samples from 199 patients with acute STEMI, sampled at a median time of 16 h after primary percutaneous coronary intervention (PCI). After 3 months, final infarct size (in percentage of left ventricular mass; LVM) was assessed by single-photon emission CT. The outcome variable final infarct size was dichotomised using the 75th percentile as the cutoff value (large infarct size ≥29.0%). A multivariable analysis was performed adjusting for multiple clinical and biochemical covariates.
Results Median (IQR) osteoprotegerin concentration was 1.4 (1.0, 2.1) ng ml−1 and patients with high osteoprotegerin level (> median) at baseline had larger infarct size at 3 months compared with patients with low osteoprotegerin levels (< median) (25 (8, 40) vs 6 (0, 19)% of LVM, respectively, p<0.0001). A high osteoprotegerin level was also associated with an approximately sevenfold increase in the odds of developing a large myocardial infarct (OR 7.0; 3.2, 15.5, p<0.001). After adjustment for potential confounders including peak troponin T, the adjusted OR was 5.2 (2.0, 13.1) p<0.001.
Conclusion High levels of circulating osteoprotegerin measured the first morning after a PCI-treated acute STEMI were strongly associated with final infarct size.
- Final infarct size
- inflammation
- myocardial infarction
- osteoprotegerin
- STEMI
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Serum levels of osteoprotegerin, a member of the tumour necrosis factor receptor superfamily acting as a soluble decoy receptor of the receptor activator of nuclear factor kappa B ligand (RANKL), have been shown to be associated with the presence and severity of coronary and peripheral artery disease, coronary artery calcium, incident cardiovascular disease and vascular mortality.1–3 Serum osteoprotegerin has been reported to be higher in patients with acute myocardial infarction (MI) compared with both healthy controls and patients with stable coronary artery disease.4 5 Furthermore, elevated circulating osteoprotegerin was associated with increased mortality and heart failure development in patients with acute coronary syndrome (ACS) including MI, and high levels were related to N-terminal proB-type natriuretic peptide (proBNP) and markers of myocardial necrosis such as troponin I.5 6 However, the relationship between osteoprotegerin levels and final infarct size has, to our knowledge, not previously been investigated.
The aim of the present study was to elucidate a possible association between serum osteoprotegerin levels measured within the first 24 h of admission in patients with ST-elevation myocardial infarction (STEMI) and final infarct size measured after 3 months as assessed by single-photon emission CT (SPECT) imaging.
Methods
Study population
This was an observational cohort study prospectively enrolling 224 primary percutaneous coronary intervention (PCI)-treated STEMI patients admitted to the coronary care unit at Ullevål University Hospital, Oslo, Norway, between November 2005 and May 2007. Patients without known diabetes were included in order to study the prevalence of undiagnosed abnormal glucose regulation classified by an oral glucose tolerance test.7 A total of 199 patients completed a 3-month follow-up and were investigated for a possible association between circulating levels of osteoprotegerin and final infarct size. At enrolment, patients were clinically stable, without persistent glyseroltrinitrate infusion, chest pain, nausea or symptoms of heart failure. Patients with known diabetes, serum creatinine concentration of 200 umol/l or greater, and age over 85 years were excluded. STEMI was defined according to the universal definition of MI8 as typical rise and fall of the cardiac biomarker troponin T (TnT) with at least one value above the 99th percentile of the upper reference limit in patients presenting with symptoms of ischaemia together with new ST elevation at the J-point in two contiguous leads with the cutoff points: 0.2 mV in men or 0.15 mV in women in leads V2–V3 and/or 0.1 mV in other leads or new left bundle-branch block.
The regional ethics committee approved the study and all patients provided written and oral informed consent.
Biochemical analyses
Blood samples were drawn after an overnight fast (median 16.5 h after acute PCI), centrifuged within 1 h at 2500g for 10 min and serum isolated for routine analyses by use of conventional methods or aliquoted and stored at −80°C. Serum levels of osteoprotegerin were quantified by enzyme immunoassay using commercially available matched antibodies (R&D Systems, Minneapolis, Minnesota, USA) as previously described and validated.9 The mean recovery of samples spiked with recombinant osteoprotegerin was 93%. The intra-assay and inter-assay coefficients of variation (CV) were 3.6% and 10.6%, respectively. The sensitivity was calculated to be 15 pg/ml. C-reactive protein (CRP) was determined by enzyme-linked immunosorbent assays (DRG Instruments, Marburg/Lahn, Germany). The inter-assay CV was less than 5%. Serum cardiac-specific troponin T (TnT) was measured by electrochemiluminescence technology for quantitative measurement (Elecsys 2010, Roche, Mannheim, Germany). The inter-assay CV was 7%.
Levels of N-terminal proBNP were determined with Elecsys proBNP sandwich immunoassay on Elecsys 2010 (Roche Diagnotics, Indianapolis, Indiana, USA). The inter-assay CV was 7%. The TnT maximum level (TnTmax) was defined as the maximum (peak) value measured in each patient during the acute STEMI. Peak TnT levels have previously been shown to correlate well with infarct size measured by SPECT.10 Inter-assay and intra-assay CV% were less than 10% for all other assays.
SPECT imaging
Left ventricular function and infarct size were assessed at rest at the 3-month follow-up. Data on left ventricular function during the index hospitalisation were not available. Perfusion imaging was performed according to ECG-gated SPECT after the injection of 99m-tetrofosmin (Myoview, Amersham Health, UK). An Exeleris processing station (GE Medical Systems, Buckinghamshire, UK) with 4D-MSPECT software (University of Michigan), was used for the processing of all recordings and assessment of left ventricular volumes, left ventricular ejection fraction (LVEF) and infarct size (proportion perfusion defect) expressed as a percentage of left ventricular mass (LVM).11
Clinical follow-up
Of the 224 patients originally included in this cohort, 201 patients attended the 3-month follow-up, which included a clinical examination, SPECT imaging and blood sampling for biochemical analyses. One (of the 224) patient died during the first 3 months, 22 patients were unwilling to attend the follow-up and 199 patients were included in the present study because blood samples were missing from two of the 201 patients.
Statistical analyses
As a result of skewness in most of the measured variables, non-parametric statistics were used throughout. Categorical variables are presented as proportions and continuous variables as median values with interquartile ranges. Final infarct size was dichotomised into large or small categories using the 75th percentile as a cut-off value (large infarct size ≥29.0% of LVM).12 Osteoprotegerin was analysed both as a continuous variable and a dichotomised variable based on linear trend analysis of the association between the quartiles of osteoprotegerin and infarct size. Correlation was measured as Spearman rank correlation.
A multivariable logistic regression analysis was performed and the following covariates were included in the model based on an association with either high osteoprotegerin levels or large MI with a p value less than 0.20: Age, gender, body mass index (BMI), smoking, treated hypertension, previous MI, history of heart failure, multivessel disease, infarct localisation (anterior MI or left-bundle branch block vs inferior or posterior MI), proBNP, serum cholesterol, CRP, creatinine, fasting plasma glucose, previous use of diuretics, heart rate, systolic blood pressure and Killip class at admission and time from symptoms to PCI. Infarct localisation, previous MI and TnTmax were included in the final step of the model building in addition to high levels of osteoprotegerin. The analysis was performed with and without TnTmax due to its well-known association with final infarct size. A two-sided p value of p<0.05 was considered statistically significant. All analyses were performed by SPSS software version 15.0.
Results
Median age was 58 years (range 51–68) and 199 patients were included. A majority of the patients were men (18% women), only 7% of the patients had a history of previous MI, while 27% were treated for hypertension. All patients were treated by primary PCI. Final infarct size measured by SPECT 3 months after the acute STEMI was 14% (0–29) of LVM. Patients who developed a large infarct size (>75th percentile) were older and had higher TnTmax, proBNP, fasting plasma glucose, creatinine and osteoprotegerin levels at baseline (table 1). In addition, patients who developed large MI had a higher frequency of either anterior MI or left-bundle branch block and more patients had symptoms of heart failure at admission (Killip class >I).
Characteristics of patients with large final infarct size (>75th percentile) after acute STEMI (n=199)
The median (IQR) value of osteoprotegerin measured the first day was 1.4 ng ml−1 (1.0, 2.1) (n=199). The osteoprotegerin values were significantly higher in patients with large infarct size (figure 1, table 1). Characteristics of the study population according to osteoprotegerin concentrations at baseline are given in detail in table 2. Patients with a high osteoprotegerin level (> median value) at baseline were characterised by higher age, longer time from symptoms to treatment, lower BMI and waist circumference, higher fasting plasma glucose the first morning after admission and higher proBNP and TnTmax levels. In addition, patients with high osteoprotegerin levels had significantly lower LVEF measured after 3 months, although the difference was small (table 2).
Osteoprotegerin (OPG) levels in relation to infarct size in patients with ST-elevation myocardial infarction (MI) (n=199). Serum concentrations of osteoprotegerin were measured at baseline (median time 16.5 h after acute percutaneous coronary intervention) and infarct size 3 months later by single-photon emission CT. (A) Box plot showing osteoprotegerin levels according to quartiles of infarct size (in % of left ventricular mass). The bar represents the median value of osteoprotegerin, the top and bottom of the boxes represent the 75th and the 25th percentile, respectively. The ends of the whiskers represent minimum and maximum values. *p<0.05. (B) Dot plot showing osteoprotegerin values in patients with large (>75th percentile, n=51) compared with small (or medium size) MI (<75th percentile, n=148). The lines represent median values with IQR. ANOVA, analysis of variance.
Characteristics of the study population (n=199) related to osteoprotegerin levels (above or below median value) measured in patients with acute STEMI
There was significant correlation between osteoprotegerin concentrations (analysed as a continuous variable) at baseline and TnTmax (r=0.29, p<0.001), proBNP (r=0.45, p<0.001), infarct size at 3 months (r=0.34, p<0.001) and LVEF at 3 months (0.18 (p=0.012), respectively (Spearman ρ). There was, however, no significant correlation between osteoprotegerin and CRP (r=0.04, p=0.58).
Osteoprotegerin concentrations at baseline were strongly indicative of the presence of large infarct size defined as infarct size above the 75th percentile. The crude OR per 1 SD increase in osteoprotegerin level of developing a large infarct size at 3 months was 2.4 (1.6, 3.5, p<0.0001). The ability of serum concentrations of osteoprotegerin at baseline to predict large MI was also assessed by the area under the receiver operating characteristics curve (figure 2). Area under the curve (AUC) for osteoprotegerin and large MI (AUC 0.73; 95% CI 0.65 to 0.80, p<0.001) was comparable to TnTmax (AUC 0.81; 95% CI 0.74 to 0.88, p<0.001) as well as AUC for a single TnT value (not peak TnT) measured at the same time point as osteoprotegerin (AUC 0.66; 95% CI 0.57 to 0.75, p=0.001).
Receiver operating characteristic curve for differentiating between large versus small myocardial infarction (MI) by baseline osteoprotegerin in patients with acute ST-elevation MI (n=199). Area under the curve (AUC) (95% CI) for osteoprotegerin and large MI.
Linear trend analysis of the association between the quartiles of osteoprotegerin and the OR of large infarct size (p for trend <0.001) identified median levels as a cut-off value for dichotomising osteoprotegerin into high and low levels (figure 3A). Patients with a high osteoprotegerin level (> median) at baseline had a larger infarct size at 3 months compared with patients with a low osteoprotegerin level (< median) (25% (8, 40) vs 6% (0, 19) of LVM, respectively, p<0.001; figure 3B). In univariate analyses, a high osteoprotegerin level (> median value) was associated with an approximately sevenfold increase in the odds of developing a large MI (crude OR 7.04; 3.2, 15.5, p<0.001). After adjustment for all available clinical and biochemical covariates (see Methods section) in a multivariable model, osteoprotegerin remained a strong, independent predictor of developing a large MI with an adjusted OR 8.2 (3.6, 19.2, p<0.001). If TnTmax was included in the model, the adjusted OR of developing a large MI in patients with a high osteoprotegerin level was 5.2 (2.0, 13.2, p=0.001; table 3). The results were not influenced significantly by adding proBNP to the model.
Associations between osteoprotegerin (OPG) levels at baseline and final infarct size measured by single-photon emission CT 3 months after acute ST-elevation myocardial infarction (MI) (n=199). (A) OR of having large infarct size (>75th percentile) by quartiles of osteoprotegerin (ng ml−1) are shown (p for trend <0.001). The median value (arrow) was used as a cut-off for dichotomising osteoprotegerin into high and low levels. (B) Box plot showing final infarct size (in percentage of left ventricular (LV) mass) in patients with high osteoprotegerin level (> median) at baseline compared with patients with low osteoprotegerin level (< median). The bar represents the median value of osteoprotegerin, the top and bottom of the boxes represent the 75th and the 25th percentile, respectively. The ends of the whiskers represent minimum and maximum values.
Crude and multivariable adjusted OR for having large infarct size measured by SPECT 3 months after acute STEMI when high (above median) levels of osteoprotegerin are present at baseline
Potential confounders included in the analysis: age, gender, BMI, treated hypertension, previous MI, history of heart failure, multivessel disease, infarct localisation (anterior MI or left-bundle branch block vs inferior or posterior MI), proBNP, TnTmax, serum cholesterol, CRP, creatinine, fasting plasma glucose, previous use of diuretics, heart rate, systolic blood pressure and Killip class at admission, and time from symptoms to PCI.
Discussion
Our main finding was that circulating osteoprotegerin levels during acute STEMI were strongly and independently associated with final infarct size. High levels of osteoprotegerin were associated with an increased risk of developing large MI as assessed by SPECT imaging after 3 months.
Circulating osteoprotegerin has been demonstrated to be an independent prognostic marker of adverse outcomes in patients with ACS, including STEMI.6 Whether the association between increased osteoprotegerin and cardiovascular events in patients with ACS is explained by a relation with atherosclerotic plaque rupture or whether elevated serum levels are just part of a systemic or local inflammatory response is still unclear. RANKL circulates in low concentrations and is cumbersome to determine accurately while serum osteoprotegerin has been regarded as a reliable and stable marker of the overall activity in the osteoprotegerin/RANKL/RANK axis, including the ability of this system to promote inflammation and matrix degradation. Circulating osteoprotegerin may thus mirror two interacting pathogenic processes in atherogenesis and plaque destabilisation. In addition, while osteoprotegerin is thought to neutralise RANKL activity, at least at high concentrations, osteoprotegerin may enhance the matrix metalloproteinase-inducing effect of RANKL,13 suggesting an active role of osteoprotegerin in the promotion of plaque instability and ACS.
The strong association between osteoprotegerin and infarct size in STEMI patients could also indicate the ability of osteoprotegerin to reflect myocardial necrosis. We have previously reported increased myocardial expression of osteoprotegerin/RANKL/RANK in an experimental model of postinfarction heart failure with increased gene expression in both the ischaemic and non-ischemic part of the left ventricle.14 Moreover, osteoprotegerin is produced together with RANKL and its receptor RANK in a wide range of cells with relation to the myocardium such as vascular endothelial and smooth muscle cells.15 16 Also, we have previously shown a strong expression of osteoprotegerin within the failing myocardium at least partly localised to cardiomyocytes.14 The strong association between serum levels of osteoprotegerin and infarct size in STEMI patients may thus reflect the contribution of the damaged myocardium itself to circulating osteoprotegerin levels and partly explain the association between osteoprotegerin and long-term mortality and heart failure development in ACS patients.6 The association between osteoprotegerin and infarct size was also present after adjustment for peak levels of TnT. It has previously been shown that TnT levels peak within the first 24 h and that peak TnT correlated significantly, and almost as good as AUC TnT (0.63 and 0.65, respectively), with final infarct size.10
The present study has certain limitations such as a relatively low number of patients, the exclusion of patients with high age, kidney failure, heart failure or known diabetes, lack of information about ST segment deviation at baseline and lack of clinical outcome data. In addition, we did not perform serial measurements of osteoprotegerin and may have missed the peak osteoprotegerin value in some of the patients. However, a possible clinical implication of our study could be measurements of osteoprotegerin in patients with acute STEMI in order to identify patients at increased risk of developing a large MI, heart failure and, as a consequence, sudden cardiac death. The clinical relevance may be limited by the moderate AUC of osteoprotegerin to predict large MI, but this should be investigated further in forthcoming studies including a larger number of patients, longitudinal sampling and long-time follow-up with clinical events as primary endpoints.
Conclusion
Our findings suggest that high levels of circulating osteoprotegerin, measured the first morning after a PCI-treated acute STEMI, are strongly associated with final infarct size. These data further support a role for osteoprotegerin as a biomarker in cardiovascular disorders.
References
Footnotes
Funding The south-eastern Norway regional health authority supported this work.
Competing interests None.
Patient consent Obtained.
Ethics approval This study was conducted with the approval of the national committees for research ethics, Norway.
Provenance and peer review Not commissioned; externally peer reviewed.