Results Study group patient characteristics. ACB assay. Cardiac markers. Statistical analysis

Transcription

1 Reduced albumin-cobalt binding with transient myocardial ischemia after elective percutaneous transluminal coronary angioplasty: A preliminary comparison to creatine kinase-mb, myoglobin, and troponin I David Bar-Or, MD, FACEP, a,b,e James V. Winkler, MD, FACEP, c,e Karyl VanBenthuysen, MD, a,c,d Lisbeth Harris, MSPH, a,b Edward Lau, PhD, b,e and Fred W. Hetzel, PhD f Englewood and Denver, Colo Background Previous reports suggest that ischemic conditions rapidly reduce the capacity of human albumin to bind exogenous cobalt. A new assay based on human albumin-cobalt binding (ACB) may help detect early myocardial ischemia. We investigated altered ACB during the first 24 hours after transient ischemia induced during elective percutaneous transluminal coronary angioplasty (PTCA). We then compared ACB assay results with creatine kinase isoenzyme (CK-MB), myoglobin, and cardiac troponin I (ctn-i) values after PTCA. Methods and Results In 41 patients undergoing elective PTCA, plasma samples were tested for the ACB assay, CK-MB, myoglobin, and ctn-i before, immediately after, and 6 and 24 hours after PTCA. Thirteen additional patients served as a control group with albumin-cobalt assays performed before and after diagnostic coronary catheterization without angioplasty. ACB assay results demonstrated a significant mean percent difference (10.1%) immediately after PTCA compared with baseline (P < ) and returned to baseline by 6 hours after PTCA. ACB assay differences immediately after PTCA were significantly greater than in the control group (10.1% vs 0.9%, P <.001). Mean CK-MB, myoglobin, and ctn-i values were not elevated above baseline immediately after PTCA but were significantly elevated above baseline 6 and 24 hours after PTCA. Conclusions These preliminary results suggest that human albumin undergoes a significant reduction in its capacity to bind exogenous cobalt soon after transient coronary occlusion during human PTCA and before significant elevations of CK- MB, myoglobin, or ctn-i. Further confirmatory investigations are warranted to determine if the ACB assay is a useful diagnostic test for early myocardial ischemia. (Am Heart J 2001;141: ) Cardiac markers of cellular necrosis, such as creatine kinase isoenzymes (CK-MB), myoglobin, or troponin, are unreliable markers of transient myocardial ischemia, particularly when measured in the first 2 to 6 hours after an ischemic event. 1-4 Patients who are examined soon after the onset of ischemic symptoms typically require prolonged observation to rule out myocardial infarction or myocardial ischemia. 5-8 We From a Swedish Medical Center; b Trauma Research, Swedish Medical Center; c Centura Health Porter Adventist Hospital; d South Denver Cardiology; e DMI Bio- Sciences, Inc; and f HealthOne Research. Supported by DMI BioSciences, Inc, Englewood, Colo, and Ischemia Technologies, Inc, Denver, Colo. David Bar-Or, Edward Lau, and James V. Winkler are employees of DMI Bio- Sciences, Inc, and own stock in the company. Submitted August 18, 2000; accepted February 2, Reprint requests: David Bar-Or, MD, DMI BioSciences, Inc, 3601 S Clarkson, #335, Englewood, CO dbaror@dmibio.com Copyright 2001 by Mosby, Inc /2001/$ /1/ doi: /mhj previously reported a novel blood assay method to measure reduced exogenous cobalt binding to human albumin in patients with myocardial ischemia. 9 The albumin-cobalt binding (ACB) assay measures the binding capacity of exogenous cobalt to the amino terminus (N-terminus) of human albumin. Under normal conditions, transition metals, including cobalt, are tightly bound to the exposed N-terminus of albumin. 10 The ACB assay is based on observations that ischemic conditions may alter the N-terminus of albumin and rapidly reduce its binding capacity for transition metals. 11,12 Ischemia-induced alterations to albumin would be predicted to occur minutes or hours before abnormal levels of CK-MB, myoglobin, or troponin could be detected. Previous reports have demonstrated that percutaneous transluminal coronary angioplasty (PTCA) can be a useful in vivo model of mild transient myocardial ischemia in human beings The purpose of this preliminary study was to evaluate the percent differences from baseline in ACB assay values drawn immediately after

2 986 Bar-Or et al June 2001 elective PTCA and 6 hours and 24 hours after PTCA and to compare the assay differences with CK-MB, myoglobin, and cardiac troponin I (ctn-i) differences. Methods In this prospective study, 42 patients without evidence for acute myocardial infarction scheduled for elective PTCA and 13 control patients scheduled for elective diagnostic coronary angiography were enrolled at two hospitals: Swedish Medical Center, Englewood, Colo, and Centura Health Porter Adventist Hospital, Denver, Colo. Institutional review boards of both hospitals approved the study protocol, and all patients gave informed consent. Patients were excluded if they had any of the following: signs or symptoms of other acute or chronic ischemic conditions (including but not limited to stroke, transient ischemic attack, claudication, diabetes mellitus with signs of peripheral vascular disease, kidney failure, or shock), evidence of collateral coronary circulation around the target angioplasty lesion, 18,19 use of a continuous-perfusion balloon catheter or intra-aortic balloon pump during PTCA, >72 hours of known occlusion of the coronary artery targeted for angioplasty, cardiac arrest or another coronary revascularization procedure during or in the 24 hours after PTCA, or previous enrollment in this study. A clinical cardiologist blinded to all ACB assay results retrospectively reviewed each patient s complete medical record to exclude any patient who demonstrated objective evidence of acute myocardial infarction <72 hours before PTCA (unequivocal electrocardiographic abnormalities, serial CK-MB or troponin elevations, or angiographic evidence of acute myocardial infarction), angiographic evidence of >72 hours of known occlusion of the coronary artery lesion targeted for PTCA, or significant collateral coronary circulation at the angioplasty site documented during PTCA. One patient was excluded because of lost baseline blood samples, leaving a final study group of 41 patients with PTCA. A control group for preprocedure and postprocedure ACB assay values included 13 patients who received diagnostic coronary angiography without PTCA. Interventional cardiologists blinded to all study assay results performed PTCA and diagnostic coronary angiography according to standard hospital protocols. All patients received standardized medical care before and after the procedure. Decisions regarding the number of balloon inflations, whether or not to use intraluminal stents, and when to terminate the procedure were based entirely on the clinical judgment of the interventional cardiologists. All patients had baseline blood samples collected from the distal aorta catheter port into tubes with sodium heparin before receiving any contrast material. Blood samples were drawn in a similar fashion immediately after the last balloon inflation in the PTCA group and at the end of diagnostic coronary angiography in the control group. For follow-up samples, patients in the study group had additional venous blood samples collected in heparinized tubes 6 hours and 24 hours after PTCA. Plasma aliquots were prepared for CK-MB, myoglobin, and ctn-i analysis, and additional aliquots were stored at 4 C for ACB assay batched analysis <48 hours after the last sample collection. ACB assay Colorimetric ACB assays were performed with the use of a Beckman DU-62 model spectrophotometer (470 nm) at the Trauma Research Laboratory, Swedish Medical Center, by one technician blinded to sample time and collection order. Triplicate values were averaged for all ACB assay calculations for each patient sample. The ACB assay laboratory method has been described in detail in a previous report. 9 No ACB assay normal values have yet been established for patients with known preexisting coronary disease. Thus ACB assay values for patients with PTCA and control patients were recorded as a percent change from baseline to lessen any effect of silent ischemia or ischemic preconditioning on baseline values. 20,21 A mean of all replicate assay measurements was determined before calculation of percent change. Cardiac markers All CK-MB, myoglobin, and ctn-i assays were performed in a blinded fashion at the clinical laboratory at Swedish Medical Center. CK-MB mass, myoglobin, and ctn-i were all determined by means of a 2-site sandwich monoclonal antibody immunoassay with the Dade Behring Stratus CS STAT Fluorometric Analyzer (Dade Behring Inc, Newark, Del). Upper reference limits for the CK-MB, myoglobin, and ctn-i assays were 4.5 ng/ml, 110 ng/ml, and 1.4 ng/ml, respectively. As with the ACB assay, CK-MB, myoglobin, and ctn-i values were reported as a percent change from baseline. Statistical analysis Study group and control group data are expressed as mean value ± SD and as median and 25th and 75th percentiles. Percent change was calculated as follows: (Assay level after PTCA or angiography alone Baseline assay level before PTCA or angiography alone) 100/Baseline assay level before PTCA or angiography alone. Data analysis was performed with Microsoft Excel 2000 (Redmond, Wash). Intraoperator variability for the ACB assay was reported as percent coefficient of variation. A Student t test was used to analyze the difference between means. The mean difference between preprocedure and postprocedure was compared by means of a 2- tailed, 2-sample t test. A value of P.05 was considered significant. Results Study group patient characteristics Study group clinical characteristics are represented in Table I. The PTCA study group of 41 patients included 26 (63%) men and 15 (37%) women; the mean age was 62.8 years (range, 41 to 83), with 30 (73%) of 41 having 2 risk factors for coronary artery disease, 10 (24%) of 41 having unstable angina, and 20 (49%) of 41 having had a prior revascularization procedure (Table II). A high percentage of study group patients (33 [81%] of 41) had at least one of the following signs of myocardial ischemia during the procedure: significant chest pain, ST-segment changes >1 mm, or ventricular rhythm disturbances. Stents were used with angioplasty for 37

3 Volume 141, Number 6 Bar-Or et al 987 Table I. Clinical characteristics Control group PTCA group (n = 13) (n = 41) Age (y) 50 ± 8 63 ± 12 Women/men 4/9 15/26 Unstable angina 0 10 (24%) Cardiac risk factors Smoking 6 (46%) 22 (54%) Diabetes mellitus 0 5 (12%) High cholesterol 3 (23%) 24 (59%) Hypertension 9 (69%) 22 (54%) Family history 6 (46%) 26 (63%) Medications with the procedure Aspirin 11 (85%) 35 (85%) Calcium blockers 0 12 (29%) Heparin 11 (85%) 41 (100%) Intravenous nitrites 0 36 (88%) Abciximab 0 7 (17%) (90%) of 41 study group patients, and 5 (12%) of 41 study group patients had side branch occlusions (SBO). No PTCA study group patients received β-blockers before or during PTCA; 12 (29%) of 41 received calcium channel blockers; 7 (17%) of 41 received abciximab; 35 (85%) of 41 received aspirin; and 41 (100%) of 41 received intravenous heparin. Angiographic characteristics for the 41 PTCA study group patients are listed in Table II. Control group patient characteristics Control group clinical characteristics are represented in Table I. The control group of 13 patients undergoing diagnostic coronary angiography without PTCA included 9 (69%) men and 4 (31%) women; the mean age was 49.9 years (range, 33 to 63), with 10 (77%) of 13 having 2 risk factors for coronary artery disease. Four (31%) of the 13 control patients had angiographic coronary lesions that did not require angioplasty, and 2 of the patients with coronary lesions had multivessel disease. No control group patient had significant chest pain, ST-segment changes >1 mm, or ventricular rhythm disturbances during the angiographic procedure. None of the control group patients received abciximab with the procedure, but 11 (85%) of 13 received aspirin, and 11 (85%) of 13 received intravenous heparin (Table II). Albumin-cobalt assay results in the study group ACB assay intraoperator variability for the PTCA study group replicate values was considered acceptable, with a mean coefficient of variation of 3.4% (range, 0.2% to 13.7%). Of the 41 patients in the PTCA group, 34 (83%) showed an increase in ACB assay values after PTCA compared with baseline. Mean ACB assay values were Table II. Angiographic characteristics Control PTCA group* group (n = 13) (n = 41) Single-vessel disease 2 (15%) 19 (46%) Multivessel disease 2 (15%) 22 (54%) Coronary artery lesions (>50%) Left main 0 1 (2%) Left anterior descending 3 (23%) 21 (51%) Left circumflex 0 17 (42%) Right 3 (23%) 25 (61%) Diagonal 1 (8%) 9 (22%) Vein graft 0 1 (2%) Average No. of dilated lesions 1.6 ± 0.7 Chest pain 0 19 (46%) ST-segment deviation 0 24 (59%) Dysrhythmias 0 9 (22%) SBO 5 (12%) Intracoronary stent 37 (90%) Total balloon occlusion time per patient (s) ± 64.7 Mean balloon occlusion time (s) 69.7 ± 32.7 Mean No. of balloon occlusions 5.9 ± 3.0 *Angiography only; no angioplasty ± ABSU (absorbance units) at baseline and ± ABSU immediately after PTCA. Mean percent difference in ACB values after PTCA compared with baseline was 10.1% ± 11.1% (P < ) (95% confidence interval, 6.7% to 13.5%). The mean percent difference for the study group was significantly greater compared with the 13 patients in the control group (10.1% ± 11.1% vs 0.9% ± 8.2%, P <.001). Of the 41 PTCA patients enrolled in the study group, 2 patients (2 [5%] of 41) refused further blood collection at 6 hours, and 15 patients (15 [37%] of 41) either refused further blood collection or were discharged from the hospital and could not be located for the 24-hour blood sample. None of the study group or control group patients died or had adverse reactions during the 24 hours after the procedure. In the final PTCA study group, ACB assay values were recorded in 41 of 41 patients at baseline and immediately after PTCA, in 39 of 41 patients at baseline and 6 hours after PTCA, and in 26 of 41 patients at baseline and 24 hours after PTCA. Data from 25 PTCA study group patients with all sample data available for ACB assay and all cardiac markers demonstrated that the results were essentially the same as for the entire 41-patient cohort (data not shown). The 39 patients sampled at 6 hours after PTCA and 26 patients sampled at 24 hours after PTCA had ACB values essentially unchanged from baseline, with mean percent differences of 2.7% and 1.8 %, respectively (Table III). Median and 25th and 75th percentile ACB assay values are shown in Table IV. Within the PTCA study group, 7 (17%) patients had no increase in ACB assay levels compared with baseline at any time sampled after PTCA.

4 988 Bar-Or et al June 2001 Table III. Albumin-cobalt assay and cardiac markers mean percent difference for PTCA study group at baseline, after PTCA, and at 6 hours and 24 hours Percent difference from baseline Assay After PTCA (n = 41) P value 6 h (n = 39) P value 24 h (n = 26) P value Albumin-cobalt 10.1 ± 11.1 < ± ± CK-MB ± ± ± Myoglobin 3.3 ± ± ± ctn-i 4.9 ± ± ± Values are mean percent difference ± SD. Table IV. ACB assay and cardiac markers median and 25th and 75th percentile values for PTCA study group at baseline, after PTCA, and at 6 and 24 hours Median and 25th and 75th percentile assay values Assay Baseline (n = 41) After PTCA (n = 41) 6 h (n = 39) 24 h (n = 26) ACB assay (ABSU) (0.321, 0.373) (0.360, 0.413) (0.331, 0.392) (0.307, 0.377) CK-MB (ng/ml) 0.30 (0.30, 1.05) 0.30 (0.30, 2.00) 0.80 (0.30, 3.35) 1.60 (0.30, 7.10) Myoglobin (ng/ml) 39.0 (30.0, 49.0) 39.0 (26.0, 46.0) 51.0 (38.0, 69.0) 44.0 (34.0, 63.0) ctn-i (ng/ml) 0.40 (0.40, 0.40) 0.40 (0.40, 0.40) 0.40 (0.40, 0.40) 0.40 (0.40, 1.33) Values are median (25th and 75th percentiles). Results of ACB assay mean percent differences for the 19 (46%) of 41 PTCA patients who had chest pain and the 24 (59%) of 41 patients with ST-segment deviations during balloon inflation were not significantly different from the entire 41-patient PTCA cohort. The 5 patients with SBO during PTCA had somewhat higher than average ACB assay mean percent differences immediately after PTCA (19.7% vs 10.1%) and 6 hours later (6.9% vs 2.7%), but the number of patients was too small to be statistically meaningful. Cardiac marker results in the study group ACB and cardiac marker assay mean differences from baseline are listed in Table III, and median and 25th and 75th percentiles are listed in Table IV. CK-MB, myoglobin, and ctn-i mean percent differences from baseline were not significantly elevated immediately after PTCA for the 41 study group patients. CK-MB, myoglobin, and ctn-i mean percent differences from baseline were significantly elevated at 6 hours and at 24 hours after PTCA (Table III), but none of the median cardiac marker assay values were elevated above upper limits of normal at any time point after PTCA (Table IV). In this preliminary study, we found no significant correlation between patients with post-ptca elevated ACB assay values and later elevation of cardiac markers. Albumin-cobalt assay results in the control group ACB assay intraoperator variability for all control patient replicate values was considered acceptable, with a mean coefficient of variation of 3.8% (range, 0.0% to 19.4%). The percent difference in ACB values for the control group was 0.9% ± 8.2% (95% confidence interval, 5.4% to 3.6%). The control group demonstrated no change in ACB assay values between baseline and samples drawn immediately after angiography. Discussion This preliminary study is the first to report significantly reduced exogenous cobalt binding to the N- terminus of human albumin after transient arterial occlusion and reperfusion with PTCA. These data suggest that alterations of ACB, compared with baseline, occur within minutes after PTCA-induced ischemic changes and can be detected by elevated values of a new colorimetric ACB assay. 9 Additionally, the results indicate that elevated ACB assay levels return to baseline within 6 hours after PTCA. Cardiac markers of necrosis such as CK-MB, myoglobin, and troponin have been reported to be unreliable as diagnostic tests for the presence of mild, reversible myocardial ischemia; when used to help identify acute myocardial infarction they are often not elevated above normal during the first 3 to 6 hours after the onset of symptoms. 1-4,7,8 CK-MB, myoglobin, and ctn-i results in this study provide additional evidence that these cardiac markers do not appear to be significantly elevated immediately after elective PTCA.

5 Volume 141, Number 6 Bar-Or et al 989 Alterations of cobalt-binding capacity of human albumin In vitro studies of human albumin have demonstrated that the transition metals cobalt, copper, and nickel bind tightly at the N-terminus binding site. 10 Conditions such as acidosis, 22 reduced oxygen tension, membrane energy-dependent sodium, and calcium pump disruptions, which occur during myocardial ischemia, have been shown to structurally alter the N-terminus of albumin and thereby reduce the transition metal-binding capacity of the protein. 11,23 Reperfusion after transient ischemic events includes free radical and iron and copper exposure, which may cause even more alterations to albumin than ischemia itself. 12,23-25 Albumin may act as a sacrificial antioxidant to reduce localized injury during reperfusion. 26 ACB assay The ACB assay reported here has been previously described as a measurement of reduced cobalt-binding capacity in albumin in patients with acute myocardial ischemia. 9 In the current assay format, a predetermined amount of exogenous cobalt is added to plasma samples and indirectly detects ACB by measuring the remaining unbound cobalt. Elevated ACB assay results in this format indicate reduced cobalt-binding capacity of albumin. Other reported assay methods for albumincobalt binding involve liquid chromatography, mass spectrometry, and nuclear magnetic resonance and are not well suited for routine clinical analysis. 10,11,22 The ACB assay used in this study is a simple biochemical procedure that can be adapted for hospital laboratories. Albumin-cobalt assay kinetics after PTCA The results of this study suggest that mean plasma levels of the ACB assay are significantly elevated a few minutes after transient PTCA-induced coronary artery occlusion and reperfusion. The data correlate chronologically with the previously reported time period most likely to encompass intravascular and endothelial ischemic conditions that could alter the N-terminus of albumin in the coronary circulation. 12,24,25 This suggests that a significant reduction of the transition metalbinding capacity of human albumin occurs within minutes after transient mild myocardial ischemia. Patients in the control group without angioplasty did not demonstrate a mean elevation in ACB assay levels, which suggests that the study group differences were caused by transient balloon inflations rather than by coronary catheterization and contrast angiography alone. Mean ACB assay levels were not significantly different from baseline at 6 hours and 24 hours after PTCA. These results suggest that reduced ACB occurs in the first minutes after brief coronary occlusion and reperfusion and is best detected in blood samples drawn soon after the onset of a suspected ischemic event. The availability of an assay that quickly indicates the presence of myocardial ischemia could help fill a diagnostic need for those occasions when patients with suspected heart disease are examined soon after the onset of symptoms. Additionally, any blood assay that consistently detects a brief ischemic event soon after its occurrence and returns to baseline within several hours may be clinically useful to identify repeated ischemic events occurring after the initial episode. Comparison with cardiac markers In this preliminary study, CK-MB, myoglobin, and ctn-i did not increase significantly immediately after PTCA when compared with baseline levels, but mean percent differences were significantly elevated at 6 and 24 hours after PTCA. However, in this study of patients with no evidence of a prior acute myocardial infarction, median values of these traditional cardiac markers were not elevated above established upper limits of normal at any time in the first 24 hours after elective PTCA. These results are similar to previous reports showing that CK-MB, myoglobin, and troponin I are not routinely elevated in the 2 to 6 hours after ischemic injury and that only a relatively small percentage of patients have abnormal elevations of these cardiac markers after PTCA. 14,17 No correlation between the ACB assay and cardiac markers was demonstrated in this study. Study limitations This preliminary study has several limitations. We acknowledge that the PTCA cohort was small and that the control group was younger and had less extensive coronary artery disease than did the PTCA group. Also, missing several blood samples at 6 hours and 24 hours reduced the sample size for those time points. Blood samples drawn before and after angioplasty were drawn from arterial sites, whereas later samples were venous, and it is possible that comparing arterial and venous samples could have introduced a degree of error. The study design included only 3 sample collection time points after PTCA for comparison to baseline values. Having no blood samples drawn between the end of PTCA and 6 hours later may have missed the time of maximum ACB assay, myoglobin, and CK-MB elevation and may have missed a more precise time when ACB assay values return to baseline. Additionally, this preliminary study was not designed to distinguish transient myocardial ischemia from small areas of myocardial necrosis; it lacked a clearly defined gold standard for the diagnosis of transient myocardial ischemia and did not have clearly established normal values for the ACB assay. Chest pain, rhythm disturbances, ST changes, and ventricular wall motion abnormalities have been previously recorded with balloon

6 990 Bar-Or et al June 2001 inflation during angioplasty, but no symptom or electrocardiographic abnormality appears to occur with enough consistency to be a valid standard. 13,15,16,27 Other variables such as preconditioning during PTCA, 20,21,28 undocumented small collateral coronary arteries at the site of angioplasty, 18,19 and the use of intraluminal stents, heparin, abciximab, or other medications may have either prevented PTCA from being an optimal model of myocardial ischemia or interfered with ACB during this study. That may help to explain why some patients did not have elevated ACB assay values after PTCA. It is beyond the scope of this preliminary study to fully explore potential false-positive or false-negative ACB assay values, but several factors, such as severe hypoalbuminemia, the presence of metal chelators, and the sample collection time in relation to the onset of ischemia, could affect ABC assay results. In its current format, the ACB assay method does not directly measure ischemia-altered albumin but rather measures free unbound cobalt. Future formats may be developed that can directly measure human albumin that has been altered during ischemia. Clinical implications and conclusions Early myocardial ischemia without definitive electrocardiographic abnormalities is difficult to diagnose soon after the onset of clinical symptoms, particularly because of the absence of reliable diagnostic blood tests. 1,4,7,8,27 Current treatment protocols for the presence of suspected early myocardial ischemia often require prolonged hospital observation, serial cardiac marker results to rule out myocardial infarction, and stress tests and cardiac imaging tests. 1,4-7 This preliminary study suggests that a blood assay of reduced ACB of the N-terminus of human albumin is significantly elevated almost immediately after transient arterial occlusion during PTCA, and these results are detected long before significant elevations of CK-MB, myoglobin, or ctn-i. Future investigations are warranted to establish whether the ACB assay is a clinically useful diagnostic test for very early myocardial ischemia. We thank Andrea van Woudenberg, JoBeth Northrop, RN, and Deborah L. Morris, MS, for their assistance in the preparation of the manuscript. References 1. Kontos MC, Jesse RL. Evaluation of the emergency department chest pain patient. Am J Cardiol 2000;85:32B-9B. 2. Ishikawa Y, Saffitz JE, Mealman TL, et al. Reversible myocardial ischemic injury is not associated with increased creatine kinase activity in plasma. Clin Chem 1997;43: Brogan GX, Hollander JE, McCuskey CF, et al. Evaluation of a new assay for cardiac troponin I vs creatine kinase-mb for the diagnosis of acute myocardial infarction. Acad Emerg Med 1997;4: Hedges JR, Swanson JR, Heeter C, et al. Prospective assessment of presenting serum markers for cardiac risk stratification. Acad Emerg Med 1996;3: Gomez MA, Anderson JL, Karagounis LA, et al. An emergency department-based protocol for rapidly ruling out myocardial ischemia reduces hospital time and expense: results of a randomized study (ROMIO). 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Characterization of the Co 2+ and Ni 2+ binding amino-acid residues of the N-terminus of human albumin: an insight into the mechanism of a new assay for myocardial ischemia. Eur J of Biochem 2001;268: Berenshtein E, Mayer B, Goldberg C, et al. Patterns of mobilization of copper and iron following myocardial ischemia: possible predictive criteria for tissue injury. J Mol Cell Cardiol 1997;29: Bush HS, Ferguson JJ, Angelini P, et al. Twelve-lead electrocardiographic evaluation of ischemia during percutaneous transluminal coronary angioplasty and its correlation with acute reocclusion. Am Heart J 1991;121: Bertinchant JP, Polge A, Ledermann B, et al. Relation of minor cardiac troponin I elevation to late cardiac events after uncomplicated elective successful percutaneous transluminal coronary angioplasty for angina pectoris. Am J Cardiol 1999;84: Dellborg M, Emanuelsson H, Swedberg K. Silent myocardial ischemia during coronary angioplasty. 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7 Volume 141, Number 6 Bar-Or et al 991 auto degradation of human serum albumin. Eur J Biochem 1995; 227: McCord JM. Oxygen-derived free radicals in postischemic tissue injury. N Engl J Med 1985;312: Cobbe SM, Poole-Wilson PA. The time of onset and severity of acidosis in myocardial ischaemia. J Mol Cell Cardiol 1980;12: Reimer KA, Lowe JE, Rasmussen MM, et al. The wavefront phenomenon of ischemic cell death, I: myocardial infarct size vs duration of coronary occlusion in dogs. Circulation 1977;56: Halliwell B. Albumin: an important extracellular antioxidant? Biochem Pharmacol 1988;37: Singer AJ, Brogan GX, Valentine SM, et al. Effect of duration from symptom onset on the negative predictive value of a normal ECG for exclusion of acute myocardial infarction. Ann Emerg Med 1997; 29: Vaitkus PT, Miller JM, Buxton AE, et al. Ischemia-induced changes in human endocardial electrograms during percutaneous transluminal coronary angioplasty. Am Heart J 1994;127: Receive tables of contents by To receive the tables of contents by , sign up through our web site at mosby.com/ahj Choose notification Simply type your address in the box and click on the subscribe button Alternatively, you may send an message to majordomo@mosby.com Leave the subject line blank, and type the following as the body of your message: subscribe ahj_toc You will receive an to confirm that you have been added to the mailing list. Note that TOC s will be sent out when a new issue is posted to the Web site.