Although both inflammation and apoptosis occur in acute coronary syndromes (ACSs), previous studies have not tested the diagnostic and prognostic utility of an approach that measures circulating markers of these pathways. The aim of the present study was to assess whether measuring soluble Fas (sFas) and high-sensitivity C-reactive protein (hs-CRP), as markers of apoptosis and inflammation, improve ACS diagnostic and prognostic accuracy. In a prospective cohort of consecutive subjects admitted to the hospital for suspicion of ACS, we measured sFas, hs-CRP, and troponin T in those who had a final noncardiac chest pain diagnosis (n = 100), those who had an ACS diagnosis and experienced (n = 218) or did not experience (n = 170) recurrent cardiac events during 1 year of follow-up. sFas was strongly and independently associated with a discharge diagnosis of an ACS versus noncardiac chest pain during the index hospitalization (odds ratio 16.16 for the second vs first tertile, 95% confidence interval [CI] 7.07 to 36.91; and odds ratio 25.40 for the third vs first tertile, 95% CI 9.38 to 68.75). However, hs-CRP was not. sFas significantly improved the diagnostic accuracy for ACSs (C statistic increased from 0.85 to 0.93, difference +0.08, 95% CI for the difference 0.05 to 0.11). The sFas levels were high and did not vary with time in the subjects having early versus late measurements (β 0.00 ln pg/ml/hour, 95% CI −0.01 to 0.01). In contrast, troponin increased with time since the beginning of the symptoms (β 0.07 ln μg/L/hour, 95% CI 0.04 to 0.10). Baseline sFas and hs-CRP did not predict recurrent cardiac events. In conclusion, our results suggest that in suspected ACS cases, sFas, but not hs-CRP, helps to improve the diagnostic accuracy and timeliness over and above standard diagnostic criteria.
The diagnosis of acute coronary syndromes (ACSs) remains challenging. Chest pain can be atypical, and myocardial necrosis marker levels are often normal at admission and can remain negative in patients with unstable angina pectoris (UAP). Identifying the subjects who are at greatest risk of recurrence is also important for clinical management. A strategy that takes into account multiple relevant pathophysiologic pathways might help in improving both ACS diagnostic and prognostic accuracy. Hence, the aim of the present study was to assess whether a multimarker approach that includes soluble Fas (sFas), high-sensitivity C-reactive protein (hs-CRP), and troponin as markers of apoptosis, inflammation, and necrosis can improve the diagnostic accuracy and timeliness in patients with suspected ACS. We also wished to determine whether this approach could predict recurrent cardiac events in those who experienced an initial ACS.
Methods
This was a case-control study, which was nested within a prospective cohort, the Récurrence et inflammation dans les syndromes coronariens aigus (RISCA) study. A nested case-control approach was chosen for efficiency motives and yielded unbiased estimates of the hazard ratio from a cohort study, provided that controls can become future cases. The recruitment phase of the RISCA study began in 2000 and ended in early 2002. In all, 1,210 patients participated in the present study. To fully reflect the spectrum of practice patterns, 4 community and 4 tertiary participating centers were included. All consecutive patients hospitalized with an admitting diagnosis of an ACS (UAP or acute myocardial infarction [AMI]) were eligible for inclusion if they were approached within 24 hours of the end of ischemic symptoms. Clinical management was left to the discretion of the treating physicians. For inclusion in this cohort, AMI was defined as characteristic discomfort or pain with an elevation of creatine kinase-MB to ≥1.5 times the upper limit of normal and/or troponin I or T in the positive range according to each center’s cutoff value diagnostic of AMI. The diagnosis of UAP required either one episode of typical chest pain at rest or with minimal exertion lasting ≥10 minutes or ≥2 episodes, each lasting ≥5 minutes. This could be either new-onset angina or an abrupt and significant change in the pattern of established angina, with creatine kinase-MB and/or cardiac troponin I or T levels remaining below the threshold for the diagnosis of AMI. No specific electrocardiographic criteria were necessary to be included in the RISCA study with an initial diagnosis of ACS.
From this cohort, we selected all patients who had experienced our primary outcome (recurrent AMI, UAP, or cardiac death within 1 year after their initial hospitalization) and randomly selected controls, matched for age (±5 years), gender, and follow-up duration (±0 days) in a 1:1 ratio. In a nested case-control approach, controls can serve more than once for different cases, the reason the control group could be smaller than the case group, even when matched in a 1:1 ratio. Although the RISCA cohort was specifically selected to have a high probability of ACS (only hospitalized patients were approached), a small group of discharged patients had a final noncardiac chest pain diagnosis. Biomarkers were not measured in the whole cohort but only in patients with recurrence, their matched controls, and the noncardiac chest pain group. The institutional review committee of each of the participating hospitals approved the study, and all subjects gave informed consent.
Experimental blood samples were taken within 24 hours of the end of chest pain. After centrifugation, the plasma samples were distributed in aliquots of 2 ml each, stored locally at −70°C, and sent on dry ice to the central repository where they were stored at −80°C. Clinical data collection was performed prospectively by trained research nurses at each study site and systematically validated initially by the local principal investigator and finally by on-site chart review by the central investigators. The patients were followed up at 1 month after discharge (on-site visit) and 1 year after discharge (telephone interview), obtaining, as necessary, confirmatory hospital files.
Citrated plasma levels of sFas were measured at baseline using a commercially available enzyme-linked immunosorbent assay kit (Bender MedSystems, Vienna, Austria). The coefficient of variability among plaques was 6.33%. hs-CRP was measured using the N High-Sensitivity CRP mono assay using the BN ProsPec Nephelometer (Dade Behring, Deerfield, Illinois). In addition to each center measuring cardiac troponin T or I for clinical purposes, cardiac troponin T was measured centrally using a commercial assay (Roche, Mannheim, Germany) with a detection limit for myocardial injury of 0.1 μg/L.
Detailed clinical variables, as well as the baseline laboratory values for maximal creatine kinase, creatine kinase-MB, and creatinine (to estimate the glomerular filtration rate using the Modification of Diet in Renal Disease abbreviated equation ) were recorded prospectively. The admission electrocardiogram was centrally adjudicated for all patients. ST-segment elevation or depression ≥1 mm (not necessarily new or transient) in ≥2 contiguous leads was recorded. The baseline left ventricular ejection fraction was measured using echocardiography, scintigraphy, or angiography.
The initial discharge diagnoses of UAP, AMI, or noncardiac chest pain were made by the treating physicians and recorded from the medical chart. The principal investigator of the RISCA study (PB) reviewed all charts to ensure that these discharge diagnoses were consistent with the local cardiac enzyme and troponin definition limits, as well as the central laboratory troponin T measurements. When this was not the case, the diagnosis was adjudicated to reach conformity with these criteria. The diagnostic criteria for recurrent UAP and AMI were the same as those used for the initial diagnoses. All recurrent end points, with their relevant electrocardiograms, were centrally adjudicated independently by 2 investigators. In the case of disagreement, the final decision was reached by a third investigator. All diagnoses were made and fixed before the sFas and hs-CRP measurements were performed. The loss to follow-up at 1 year was minimal (0.3%).
The primary objectives were to determine: (1) the independent relation among sFas, hs-CRP, and an initial hospitalization discharge diagnosis of ACS (vs noncardiac chest pain); and (2) the independent relation among sFas, hs-CRP, and recurrent cardiac events and death during 1 year of follow-up. Normally distributed variables are presented as the mean ± SD, and non-normally distributed variables as the median with the interquartile range (twenty-fifth and seventy-fifth percentiles). Categorical variables are summarized using proportions. We performed initial bivariate analyses to identify potential confounders. Spearman correlation coefficients (continuous covariates) were also used to assess the relation among sFas, hs-CRP, and the covariates, including the necrosis markers. The association between the biomarker levels and the discharge diagnosis of ACS was assessed using standard logistic regression analysis, in univariate and multivariate models. Linearity on the logit scale was checked by graphing methods for each continuous variable. Because they were nonlinear, sFas and hs-CRP were categorized into tertiles and analyzed as categorical variables. The incremental value of the sFas measurement for the diagnosis of ACS was determined by comparing the area under the receiver operating curve for models based on standard diagnostic criteria (electrocardiographic changes, elevated necrosis markers) with and without the addition of sFas and hs-CRP. We examined temporal trends in sFas and troponin T levels in patients with ACS using simple linear regression models in which the log transformed values of these markers were included as dependent variables and the interval since symptom onset was included as the independent variable. The normality assumption for residuals was verified visually and met. Associations among sFas, hs-CRP, and recurrent cardiac events were assessed using conditional logistic regression analysis in univariate and multivariate models. All statistical analyses were performed using Statistical Analysis Systems, version 9.1 (SAS Institute, Cary, North Carolina).
Results
To address the importance of apoptosis, inflammation, and necrosis biomarker levels in the diagnosis and prognosis of ACS, we measured sFas, hs-CRP, and troponin T in all patients with recurrent ACS (n = 218), their matched control (n = 170), and all subjects with a final noncardiac chest pain diagnosis (n = 100). The patients with a final discharge diagnosis of noncardiac chest pain were more likely to be younger and female and to have better cardiac and renal function and were less likely to have insulin-dependent diabetes ( Table 1 ). By definition, their troponin levels were not elevated. Three subjects in this group had 1-mm ST-segment depression in ≥2 contiguous leads on their baseline electrocardiogram. However, these abnormalities were already present on previous electrocardiograms and judged to be nonevolutive. Among those who were discharged with a diagnosis of ACS, the subjects who experienced recurrent cardiac events were more likely to have a history of heart failure, diabetes, or coronary or peripheral vascular disease. Their mean body mass index was slightly greater, and they had experienced more heart failure episodes at baseline. They underwent revascularization slightly less often than subjects without recurrence. sFas and hs-CRP were weakly associated with each other, as well as with troponin, maximal creatine kinase, and a lower glomerular filtration rate.
