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Ischemia-modified albumin increases after skeletal muscle ischemia during arthroscopic knee surgery
Clinica Chimica Acta 366 (2006) 264 – 268 www.elsevier.com/locate/clinchim
Ischemia-modified albumin increases after skeletal muscle ischemia during arthroscopic knee surgery Majed A. Refaai a, Rick W. Wright b, Curtis A. Parvin a, Ann M. Gronowski a, Mitchell G. Scott a, Charles S. Eby a,* a
Division of Laboratory Medicine, Department of Pathology and Immunology, Washington University School of Medicine, 660 South Euclid Avenue, Box 8118, Saint Louis, MO 63110, United States b Department of Orthopedic Surgery, Washington University School of Medicine, St. Louis, MO, United States Received 21 July 2005; received in revised form 24 October 2005; accepted 25 October 2005 Available online 7 December 2005
Abstract Background: Ischemia can alter the ability of albumin to bind free metal atoms. Based on these biochemical changes, methods to quantify ischemia modified albumin (IMA) were developed to assist in the evaluation of patients with symptoms of cardiac ischemia. Since ischemia can occur in any vascular bed, the specificity of IMA for cardiac muscle ischemia is unclear and requires further investigation. Methods: We evaluated the specificity of an IMA test in patients with skeletal muscle ischemia during arthroscopic knee surgery. A pressurized thigh cuff was continuously inflated to 300 mm Hg on the operative leg, in order to arrest blood flow during the procedure. Samples were collected before surgery, 15 min after surgery, and prior to discharge. Results: Twenty-three patients were enrolled in the study. Median tourniquet time was 29 min (range 19 – 108). Median pre-operative IMA was 90.2 KU/l (range 77 – 101.6). Statistically significant ( p < 0.05) increases in IMA and myoglobin concentrations, and decreases in albumin concentrations were observed following tourniquet release and before discharge. Conclusions: Post-operative myoglobin elevations indicated that skeletal muscle ischemia was sufficient to produce detectable myocyte necrosis. Post-operative IMA increases are consistent with ischemic modification of albumin during exposure to ischemic conditions in skeletal muscle during and /or immediately after tourniquet application. However, the negative correlations between IMA and albumin results suggest that increases in IMA were in part due to lower post-operative albumin concentrations resulting in decreased cobalt binding. D 2005 Elsevier B.V. All rights reserved. Keywords: Albumin; Cobalt binding test; Skeletal muscle; Ischemia
1. Introduction Ischemia, hypoxia, acidosis, and free radical formation can alter the ability of the first 3 amino acids in the albumin N-terminus, Asp – Ala – His, to bind free metal atoms including cobalt [1 –4]. These modifications are transient, and albumin regains its cobalt binding capacity within 6 h [1,2,4]. Based on these biochemical changes, the albumin cobalt binding test (ACB\) (Inverness Medical Innovations Inc.,
* Corresponding author. Tel.: +1 314 362 1302; fax: +1 314 362 1461. E-mail address: [email protected] (C.S. Eby). 0009-8981/$ - see front matter D 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.cca.2005.10.020
Waltham, MA) was developed to quantify ischemia-modified albumin (IMA) in serum, with the intent of identifying patients with cardiac ischemia. Its use, in conjunction with ECG and cardiac troponin, has been suggested to be useful in evaluating patients presenting with symptoms of cardiac damage. When ACB\ results were used in combination with an ECG and serial troponins, physicians were 70% accurate in ruling out cardiac ischemia. When using only ECG and troponin data, physicians’ diagnostic accuracy decreased to 50% for ruling out ischemia [3,5]. Because ischemia, and the resulting biochemical changes that modify albumin, can occur in any vascular bed, the specificity of IMA for cardiac muscle ischemia is unclear [6 – 9] and requires further investigation. We evaluated the
M.A. Refaai et al. / Clinica Chimica Acta 366 (2006) 264 – 268
The study was approved by the Washington University School of Medicine IRB committee. Informed consent was obtained from patients undergoing elective arthroscopic knee surgery. Potential subjects were excluded from participation in the study if any of the following conditions were noted during pre-operative assessment: pregnancy, renal insufficiency, chronic liver disease, active cancer, acute chest pain, or a history of cardiovascular disease. Patients were instructed to stop eating and drinking at midnight prior to surgery the following morning. Intravenous hydration with lactated ringers was begun before surgery and discontinued at discharge (average 1.3 l). During surgery, a pressurized thigh cuff on the operative leg was continuously inflated to 300 mm Hg, in order to arrest blood flow during the procedure, until shortly before patients were transferred to the recovery room. Venous blood samples were collected via separate venipunctures into tubes prior to insertion of an intravenous catheter and within an hour before application of the pressure cuff, again within approximately 15 min of arrival in recovery, and finally, prior to discharge (range 1.0 – 3.25 h after arrival in recovery). Processing, aliquoting, and freezing of serum samples at 80 -C for later testing were completed within 2 h of sample collection. IMA was measured with the ACB\ test adapted to a Roche/Hitachi MODULAR P instrument (Roche Diagnostics, Basel, Switzerland) according to the manufacturer’s instructions and previous studies [2]. Within-assay (n = 10) and between-assay (n = 2 5 days) imprecision (CV) was determined at IMA concentrations of 57, 78, and 124 KU/ l, and was respectively < 2.0% and < 4.5% for each concentration. Albumin concentrations were measured using the MODULAR P instrument and reagents (Roche). Myoglobin concentrations were measured using the Dade Dimension
Range (median) Myoglobin (ng/ml) Pre-operative Recovery Pre-discharge IMA (KU/l) Pre-operative Recovery Pre-discharge Albuminb (g/dl) Pre-operative Recovery Pre-discharge
P valuea
19 – 86 (37) 24 – 160 (47) 24 – 139 (48)
. . . . . .. <0.001 0.0001
77 – 101.6 (90.2) 82.4 – 102.4 (97.1) 81.8 – 113 (96.7)
. . . . . .. <0.005 <0.01
3.7 – 4.8 (4.45) 3.7 – 4.6 (4.20) 3.4 – 4.6 (4.05)
. . . . . .. <0.05 <0.05
a
P value, determined by sign-rank test, denotes statistical differences compared to pre-operative concentration. b Albumin was measured in 14 patients.
RxL instrument and reagents (Dade Behring Inc, Newark, DE) [10,11]. IMA concentration was modeled using mixed model analysis of variance (ANOVA). The model treated period (pre-operative, recovery, discharge) as a fixed effect, albumin concentration as a continuous covariate, and patients as a random sample (random effect). The model was used to estimate the patient’s change in IMA 150
Myoglobin (ng/ml)
2. Methods and materials
Table 2 Range, median, and P values of IMA, myoglobin, and albumin concentrations of pre-operative, recovery, and pre-discharge time points
100
50
0 120 110
IMA (KU/l)
specificity of the ACB\ test by measuring serum IMA concentrations in patients with skeletal muscle ischemia due to a pressurized tourniquet cuff during arthroscopic knee surgery.
265
100 90 80
Age (median) Sex
Body surface area (m2) Pressurized cuff tourniquet (mm Hg) Tourniquet duration (min) Cuff release to recovery (min) Cuff release to discharge (min)
33 – 79 (50) M=9 F = 10 Mean (SD) 2.10 (0.25) 300 (0) Range (median) 19 – 108 (29) 7 – 32 (12) 65 – 208 (136)
Albumin (g/dl)
5
Table 1 Clinical characteristics of arthroscopic surgery patients (n = 19)
4.5
4
3.5 Pre-Op
Recovery
Discharge
Fig. 1. Ischemic-modified albumin, myoglobin, and albumin concentrations at 3 different time periods; pre-operative, recovery, and discharge. Dotted lines indicate the normal reference interval for each analyte.
266
M.A. Refaai et al. / Clinica Chimica Acta 366 (2006) 264 – 268 Recovery
concentration from pre-operative level that could be attributed to the patient’s change in albumin concentration. Non-parametric sign-rank tests were used to compare changes from pre-operative levels to recovery and discharge levels.
20
10
0
3. Results Twenty-three consecutive patients in excellent general health were enrolled in the study. Three patients had medically controlled hypertension and 1 had diet-controlled diabetes mellitus type II. Four subjects were excluded from analysis due to hemolyzed (2), insufficient quantity (1), and mislabeled (1) samples. Demographic data are summarized in Table 1. Median tourniquet time was 29 min (range 19– 108). Test results are summarized in Table 2. All 19 subjects had IMA results, 18 had myoglobin results, and 14 had albumin results. Based on monitoring heart rate and rhythm, blood pressure, respiratory rate, and lack of any complaints of chest pain or discomfort, no patients developed signs or symptoms of cardiac ischemia peri-operatively or prior to an out-patient follow-up visit 1 week after surgery. Preoperative serum IMA, myoglobin, and albumin concentrations served as the baseline values. Statistically significant ( p < 0.05) increases in IMA and myoglobin concentrations Pre-Op
Delta IMA (KU/l)
-10
-20
Discharge 20
10
0
-10
-20 -1
-.5
0
.5
1
Delta Albumin (g/dL) Fig. 3. IMA concentration changes versus albumin concentration changes from pre-operative concentrations to recovery and discharge. The solid lines are the mixed model estimates of the linear relationship between changes in IMA concentration and changes in albumin concentration.
were observed following tourniquet release (recovery) and remained significantly elevated at discharge (Table 2, Fig. 1). Serum albumin concentrations decreased significantly following tourniquet release and at discharge (Table 2, Fig. 1). Albumin and IMA concentrations are inversely correlated. IMA decreases an average of 20 KU/l for each 1 g/dl increase in albumin (Fig 2). When a patient’s change in IMA concentration is plotted versus his or her
110 100 90 80
Recovery
10 100 90 80
Discharge 110 100 90 80 3.5
4
4.5
5
IMA Change (Adjusted for Albumin) (KU/l)
IMA (KU/l)
110
8 6 4 2 0 -2 -4 -6 -8
Albumin (g/dL) -10
Fig. 2. Relationship between IMA concentrations and albumin concentrations at pre-operative, recovery, and discharge time periods. The solid lines are the mixed model estimates of the linear relationship between IMA concentration and albumin concentration.
Recovery
Discharge
Fig. 4. Changes in IMA concentration from pre-operative to recovery and discharge after adjusting for associated changes in albumin concentration.
M.A. Refaai et al. / Clinica Chimica Acta 366 (2006) 264 – 268
change in albumin concentration it is clear that a significant fraction of the observed change in IMA is explained by the change in albumin (Fig 3). Fig. 4 plots each patient’s change in IMA concentration after adjusting for their change in albumin concentration. The pvalue for the change in IMA concentrations from preoperative levels to recovery was not significant. A p < 0.05 was obtained for the change in IMA concentrations from pre-operative levels to discharge.
4. Conclusions A reliable biomarker that detects reversible cardiac ischemia in the absence of infarction would be a significant advancement in the management of patients presenting with symptoms of acute coronary artery ischemia. IMA is a promising candidate for this role, but several unresolved issues must be addressed before it becomes a routine test, including effect of variation in patient albumin concentration [6,7], lactic acid interference [6], and specificity for cardiac, as opposed to skeletal, muscle ischemia [6– 8]. This study investigated the effect of skeletal muscle ischemia on IMA measurements using a clinically relevant model: prolonged lower extremity ischemia during arthroscopic knee surgery. A thigh tourniquet is commonly used to interrupt arterial flow during this procedure. Neuromuscular injuries attributable to tourniquet use are uncommon [12]. However, complication rates increase with tourniquet time > 60 min and patient age > 40 y [13]. Metabolic affects of prolonged tourniquet ischemia of extremities include elevated creatine kinase [14], and lactate and progressive acidosis [15]. Our study showed a statistically significant increase in serum myoglobin and IMA concentrations shortly after tourniquet release that persisted until discharge. Elevated myoglobin, an early marker of cardiac and skeletal myonecrosis, indicates that the severity of lower extremity muscle ischemia was sufficient to produce detectable myocyte necrosis. We did not obtain additional samples to determine IMA concentration >3 h after tourniquet release. However, previous studies have shown that IMA concentrations return to baseline within 6 – 12 h after cardiac ischemia during cardioversion and percutaneous coronary interventions, respectively [16,17]. Our IMA results are different than those obtained by other investigators using alternative experimental models of skeletal muscle ischemia. Apple et al. reported lower IMA concentrations in 19 runners upon completion of a marathon compared to before and 24 –48 h after the race, despite an 18 fold increase in median post-race myoglobin concentration [7]. A similar pattern of post-exercise decrease in IMA was reported by Roy et al. [8]. Twenty-three patients with symptomatic lower extremity peripheral vascular disease underwent treadmill exercise testing. Mean IMA concentrations were statistically lower immediately after subjects
267
experienced peak exercise-induced claudication pain compared to baseline and one hour recovery time points. Unlike both of these models, which combined skeletal muscle mechanical work and ischemia, our model involved isolated muscle ischemia. Thus, comparisons of results may not be appropriate. Other investigators have identified a likely explanation for immediate post-exercise skeletal muscle ischemia declines in IMA. Suspected temporal lactate interference in the ACB\ test for IMA was confirmed by the addition of increasing concentrations of lactic acid to pooled serum producing a linear decrease in IMA concentrations [6]. In the present study, post-tourniquet release samples could not be drawn until patients arrived in recovery (median of 12 min) for practical reasons. Thus, peak systemic lactic acid concentrations and associated interference with IMA measurements may have been averted. The ABC test for ischemia modified albumin measures unbound cobalt after incubation with patient serum. The amount of unbound cobalt is directly correlated to the concentration of ischemia modified albumin [1– 4] but is also inversely correlated to the concentration of unmodified albumin [6]. Therefore, changes in serum albumin concentration during periods of muscle ischemia, such as decreased intravascular volume during a marathon race [7] or increased intravascular volume due to intravenous fluid infusion during surgery, as in our study, will affect cobalt binding independent of changes in the concentration of ischemia modified albumin. In the present study, noncardiac ischemic increases in IMA could be a result of both skeletal muscle ischemia modification of albumin as well as decreased total albumin due to dilution from IV fluids. While the experimental design does not permit accurate determinations of the contribution from each mechanism to the total change in IMA, our results indicate that postarthroscopic knee surgery increases in IMA are nonspecific, and if an elevated IMA were obtained in an orthopedic patient with signs and or symptoms suggestive of post-operative ischemia, the positive predictive value of an elevated IMA is likely to be very low. The optimum IMA cutoff point for ruling out cardiac ischemia concurrent with, or following, regional skeletal muscle ischemia has not been determined, and additional studies are required to evaluate the ACB\ test performance in surgery and trauma patients. In addition, the literature lacks evaluations of IMA in patients with common medical conditions, such as congestive heart failure, diabetes mellitus, renal failure, and hypertension. Finally, albumin concentration is an important confounding variable when measuring single or serial IMA concentrations in patients suspected of cardiac ischemia. References [1] Bar-Or D, Lau E, Winkler JV. A novel assay for cobalt – albumin binding and its potential as a marker for myocardial ischemia—a preliminary report. J Emerg Med 2000;19:311 – 5.
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[2] Christenson RH, Duh SH, Sanhai WR, et al. Characteristic of an albumin cobalt binding test for assessment of acute coronary syndrome patients: a multicenter study. Clin Chem 2001;47:464 – 70. [3] Bhagavan NV, Lai EM, Rios PA, et al. Evaluation of human serum albumin cobalt binding assay for the assessment of myocardial ischemia and myocardial infarction. Clin Chem 2003;49:581 – 5. [4] Bar-Or D, Curtis G, Rao N, Bampos N, Lau E. Characterization of the Co(2+) and Ni(2+) binding amino-acid residues of the N-terminus of human albumin. Eur J Biochem 2001;268:42 – 8. [5] Morrow DA, de Lemos JA, Sabatine MS, Antman EM. The search for a biomarker of cardiac ischemia. Clin Chem 2003;49:537 – 9. [6] Zapico-Mun˜iz E, Santalo´-Bel M, Merce´-Muntan˜ola J, Montiel JA, Martı´nez-Rubio A, Ordo´n˜ez-Llanos J. Ischemia-modified albumin during skeletal muscle ischemia. Clin Chem 2004;50:1063 – 5. [7] Apple FS, Quist HE, Otto AP, Mathews WE, Murakami MM. Release characteristics of cardiac biomarkers and ischemia-modified albumin as measured by the albumin cobalt-binding test after a marathon race. Clin Chem 2002;48:1097 – 100. [8] Roy D, Quiles J, Sharma R, et al. Ischemia-modified albumin concentrations in patients with peripheral vascular disease and exercise-induced skeletal muscle ischemia. Clin Chem 2004;50: 1656 – 60. [9] Apple FS, Wu AHB, Mair J, et al. Future biomarkers for detection of ischemia and risk stratification in acute coronary syndrome. Clin Chem 2005;51:810 – 24.
[10] Carraro P, Plebani M, Varagnolo MC, Zaninotto M, Rossetti M, Burlina A. A new immunoassay for the measurement of myoglobin in serum. J Clin Lab Anal 1994;8:70 – 5. [11] Tietz NW. Clinical guide to laboratory tests. Philadelphia’ W.B. Saunders Co., 1990. p. 348. [12] Allum R. Complications of arthroscopy of the knee. J Bone Joint Surg Br 2002;84-B:937 – 45. [13] Sherman OH, Fox JM, Snyder SJ, et al. Arthroscopy – ‘‘no problem surgery’’: an analysis of complications in two thousand six hundred and forty cases. J Bone Joint Surg Am 1986;68-A:256 – 65. [14] Thorblad J, Ekstrand J, Hamberg P, Gillquist J. Muscle rehabilitation after arthroscopic meniscectomy with or without tourniquet control. Am J Sports Med 1985;13:133 – 5. [15] De´ry R, Pelletier J, Jacques A, Calvet M, Houde JJ. Metabolic changes induced in the limb during tourniquet. Can Anaesth Soc J 1965;12: 367 – 378. [16] Roy D, Quiles J, Sinha M, et al. Effect of direct-current cardioversion on ischemia-modified albumin levels in patients with atrial fibrillation. Am J Cardiol 2004;93:366 – 8. [17] Sinha MK, Gaze DC, Tippins JR, Collinson PO, Kaski JC. Ischemia modified albumin is a sensitive marker of myocardial ischemia after percutaneous coronary intervention. Circulation 2003;107:2403 – 5.
Ischemia-modified albumin increases after skeletal muscle ischemia during arthroscopic knee surgery Majed A. Refaai a, Rick W. Wright b, Curtis A. Parvin a, Ann M. Gronowski a, Mitchell G. Scott a, Charles S. Eby a,* a
Division of Laboratory Medicine, Department of Pathology and Immunology, Washington University School of Medicine, 660 South Euclid Avenue, Box 8118, Saint Louis, MO 63110, United States b Department of Orthopedic Surgery, Washington University School of Medicine, St. Louis, MO, United States Received 21 July 2005; received in revised form 24 October 2005; accepted 25 October 2005 Available online 7 December 2005
Abstract Background: Ischemia can alter the ability of albumin to bind free metal atoms. Based on these biochemical changes, methods to quantify ischemia modified albumin (IMA) were developed to assist in the evaluation of patients with symptoms of cardiac ischemia. Since ischemia can occur in any vascular bed, the specificity of IMA for cardiac muscle ischemia is unclear and requires further investigation. Methods: We evaluated the specificity of an IMA test in patients with skeletal muscle ischemia during arthroscopic knee surgery. A pressurized thigh cuff was continuously inflated to 300 mm Hg on the operative leg, in order to arrest blood flow during the procedure. Samples were collected before surgery, 15 min after surgery, and prior to discharge. Results: Twenty-three patients were enrolled in the study. Median tourniquet time was 29 min (range 19 – 108). Median pre-operative IMA was 90.2 KU/l (range 77 – 101.6). Statistically significant ( p < 0.05) increases in IMA and myoglobin concentrations, and decreases in albumin concentrations were observed following tourniquet release and before discharge. Conclusions: Post-operative myoglobin elevations indicated that skeletal muscle ischemia was sufficient to produce detectable myocyte necrosis. Post-operative IMA increases are consistent with ischemic modification of albumin during exposure to ischemic conditions in skeletal muscle during and /or immediately after tourniquet application. However, the negative correlations between IMA and albumin results suggest that increases in IMA were in part due to lower post-operative albumin concentrations resulting in decreased cobalt binding. D 2005 Elsevier B.V. All rights reserved. Keywords: Albumin; Cobalt binding test; Skeletal muscle; Ischemia
1. Introduction Ischemia, hypoxia, acidosis, and free radical formation can alter the ability of the first 3 amino acids in the albumin N-terminus, Asp – Ala – His, to bind free metal atoms including cobalt [1 –4]. These modifications are transient, and albumin regains its cobalt binding capacity within 6 h [1,2,4]. Based on these biochemical changes, the albumin cobalt binding test (ACB\) (Inverness Medical Innovations Inc.,
* Corresponding author. Tel.: +1 314 362 1302; fax: +1 314 362 1461. E-mail address: [email protected] (C.S. Eby). 0009-8981/$ - see front matter D 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.cca.2005.10.020
Waltham, MA) was developed to quantify ischemia-modified albumin (IMA) in serum, with the intent of identifying patients with cardiac ischemia. Its use, in conjunction with ECG and cardiac troponin, has been suggested to be useful in evaluating patients presenting with symptoms of cardiac damage. When ACB\ results were used in combination with an ECG and serial troponins, physicians were 70% accurate in ruling out cardiac ischemia. When using only ECG and troponin data, physicians’ diagnostic accuracy decreased to 50% for ruling out ischemia [3,5]. Because ischemia, and the resulting biochemical changes that modify albumin, can occur in any vascular bed, the specificity of IMA for cardiac muscle ischemia is unclear [6 – 9] and requires further investigation. We evaluated the
M.A. Refaai et al. / Clinica Chimica Acta 366 (2006) 264 – 268
The study was approved by the Washington University School of Medicine IRB committee. Informed consent was obtained from patients undergoing elective arthroscopic knee surgery. Potential subjects were excluded from participation in the study if any of the following conditions were noted during pre-operative assessment: pregnancy, renal insufficiency, chronic liver disease, active cancer, acute chest pain, or a history of cardiovascular disease. Patients were instructed to stop eating and drinking at midnight prior to surgery the following morning. Intravenous hydration with lactated ringers was begun before surgery and discontinued at discharge (average 1.3 l). During surgery, a pressurized thigh cuff on the operative leg was continuously inflated to 300 mm Hg, in order to arrest blood flow during the procedure, until shortly before patients were transferred to the recovery room. Venous blood samples were collected via separate venipunctures into tubes prior to insertion of an intravenous catheter and within an hour before application of the pressure cuff, again within approximately 15 min of arrival in recovery, and finally, prior to discharge (range 1.0 – 3.25 h after arrival in recovery). Processing, aliquoting, and freezing of serum samples at 80 -C for later testing were completed within 2 h of sample collection. IMA was measured with the ACB\ test adapted to a Roche/Hitachi MODULAR P instrument (Roche Diagnostics, Basel, Switzerland) according to the manufacturer’s instructions and previous studies [2]. Within-assay (n = 10) and between-assay (n = 2 5 days) imprecision (CV) was determined at IMA concentrations of 57, 78, and 124 KU/ l, and was respectively < 2.0% and < 4.5% for each concentration. Albumin concentrations were measured using the MODULAR P instrument and reagents (Roche). Myoglobin concentrations were measured using the Dade Dimension
Range (median) Myoglobin (ng/ml) Pre-operative Recovery Pre-discharge IMA (KU/l) Pre-operative Recovery Pre-discharge Albuminb (g/dl) Pre-operative Recovery Pre-discharge
P valuea
19 – 86 (37) 24 – 160 (47) 24 – 139 (48)
. . . . . .. <0.001 0.0001
77 – 101.6 (90.2) 82.4 – 102.4 (97.1) 81.8 – 113 (96.7)
. . . . . .. <0.005 <0.01
3.7 – 4.8 (4.45) 3.7 – 4.6 (4.20) 3.4 – 4.6 (4.05)
. . . . . .. <0.05 <0.05
a
P value, determined by sign-rank test, denotes statistical differences compared to pre-operative concentration. b Albumin was measured in 14 patients.
RxL instrument and reagents (Dade Behring Inc, Newark, DE) [10,11]. IMA concentration was modeled using mixed model analysis of variance (ANOVA). The model treated period (pre-operative, recovery, discharge) as a fixed effect, albumin concentration as a continuous covariate, and patients as a random sample (random effect). The model was used to estimate the patient’s change in IMA 150
Myoglobin (ng/ml)
2. Methods and materials
Table 2 Range, median, and P values of IMA, myoglobin, and albumin concentrations of pre-operative, recovery, and pre-discharge time points
100
50
0 120 110
IMA (KU/l)
specificity of the ACB\ test by measuring serum IMA concentrations in patients with skeletal muscle ischemia due to a pressurized tourniquet cuff during arthroscopic knee surgery.
265
100 90 80
Age (median) Sex
Body surface area (m2) Pressurized cuff tourniquet (mm Hg) Tourniquet duration (min) Cuff release to recovery (min) Cuff release to discharge (min)
33 – 79 (50) M=9 F = 10 Mean (SD) 2.10 (0.25) 300 (0) Range (median) 19 – 108 (29) 7 – 32 (12) 65 – 208 (136)
Albumin (g/dl)
5
Table 1 Clinical characteristics of arthroscopic surgery patients (n = 19)
4.5
4
3.5 Pre-Op
Recovery
Discharge
Fig. 1. Ischemic-modified albumin, myoglobin, and albumin concentrations at 3 different time periods; pre-operative, recovery, and discharge. Dotted lines indicate the normal reference interval for each analyte.
266
M.A. Refaai et al. / Clinica Chimica Acta 366 (2006) 264 – 268 Recovery
concentration from pre-operative level that could be attributed to the patient’s change in albumin concentration. Non-parametric sign-rank tests were used to compare changes from pre-operative levels to recovery and discharge levels.
20
10
0
3. Results Twenty-three consecutive patients in excellent general health were enrolled in the study. Three patients had medically controlled hypertension and 1 had diet-controlled diabetes mellitus type II. Four subjects were excluded from analysis due to hemolyzed (2), insufficient quantity (1), and mislabeled (1) samples. Demographic data are summarized in Table 1. Median tourniquet time was 29 min (range 19– 108). Test results are summarized in Table 2. All 19 subjects had IMA results, 18 had myoglobin results, and 14 had albumin results. Based on monitoring heart rate and rhythm, blood pressure, respiratory rate, and lack of any complaints of chest pain or discomfort, no patients developed signs or symptoms of cardiac ischemia peri-operatively or prior to an out-patient follow-up visit 1 week after surgery. Preoperative serum IMA, myoglobin, and albumin concentrations served as the baseline values. Statistically significant ( p < 0.05) increases in IMA and myoglobin concentrations Pre-Op
Delta IMA (KU/l)
-10
-20
Discharge 20
10
0
-10
-20 -1
-.5
0
.5
1
Delta Albumin (g/dL) Fig. 3. IMA concentration changes versus albumin concentration changes from pre-operative concentrations to recovery and discharge. The solid lines are the mixed model estimates of the linear relationship between changes in IMA concentration and changes in albumin concentration.
were observed following tourniquet release (recovery) and remained significantly elevated at discharge (Table 2, Fig. 1). Serum albumin concentrations decreased significantly following tourniquet release and at discharge (Table 2, Fig. 1). Albumin and IMA concentrations are inversely correlated. IMA decreases an average of 20 KU/l for each 1 g/dl increase in albumin (Fig 2). When a patient’s change in IMA concentration is plotted versus his or her
110 100 90 80
Recovery
10 100 90 80
Discharge 110 100 90 80 3.5
4
4.5
5
IMA Change (Adjusted for Albumin) (KU/l)
IMA (KU/l)
110
8 6 4 2 0 -2 -4 -6 -8
Albumin (g/dL) -10
Fig. 2. Relationship between IMA concentrations and albumin concentrations at pre-operative, recovery, and discharge time periods. The solid lines are the mixed model estimates of the linear relationship between IMA concentration and albumin concentration.
Recovery
Discharge
Fig. 4. Changes in IMA concentration from pre-operative to recovery and discharge after adjusting for associated changes in albumin concentration.
M.A. Refaai et al. / Clinica Chimica Acta 366 (2006) 264 – 268
change in albumin concentration it is clear that a significant fraction of the observed change in IMA is explained by the change in albumin (Fig 3). Fig. 4 plots each patient’s change in IMA concentration after adjusting for their change in albumin concentration. The pvalue for the change in IMA concentrations from preoperative levels to recovery was not significant. A p < 0.05 was obtained for the change in IMA concentrations from pre-operative levels to discharge.
4. Conclusions A reliable biomarker that detects reversible cardiac ischemia in the absence of infarction would be a significant advancement in the management of patients presenting with symptoms of acute coronary artery ischemia. IMA is a promising candidate for this role, but several unresolved issues must be addressed before it becomes a routine test, including effect of variation in patient albumin concentration [6,7], lactic acid interference [6], and specificity for cardiac, as opposed to skeletal, muscle ischemia [6– 8]. This study investigated the effect of skeletal muscle ischemia on IMA measurements using a clinically relevant model: prolonged lower extremity ischemia during arthroscopic knee surgery. A thigh tourniquet is commonly used to interrupt arterial flow during this procedure. Neuromuscular injuries attributable to tourniquet use are uncommon [12]. However, complication rates increase with tourniquet time > 60 min and patient age > 40 y [13]. Metabolic affects of prolonged tourniquet ischemia of extremities include elevated creatine kinase [14], and lactate and progressive acidosis [15]. Our study showed a statistically significant increase in serum myoglobin and IMA concentrations shortly after tourniquet release that persisted until discharge. Elevated myoglobin, an early marker of cardiac and skeletal myonecrosis, indicates that the severity of lower extremity muscle ischemia was sufficient to produce detectable myocyte necrosis. We did not obtain additional samples to determine IMA concentration >3 h after tourniquet release. However, previous studies have shown that IMA concentrations return to baseline within 6 – 12 h after cardiac ischemia during cardioversion and percutaneous coronary interventions, respectively [16,17]. Our IMA results are different than those obtained by other investigators using alternative experimental models of skeletal muscle ischemia. Apple et al. reported lower IMA concentrations in 19 runners upon completion of a marathon compared to before and 24 –48 h after the race, despite an 18 fold increase in median post-race myoglobin concentration [7]. A similar pattern of post-exercise decrease in IMA was reported by Roy et al. [8]. Twenty-three patients with symptomatic lower extremity peripheral vascular disease underwent treadmill exercise testing. Mean IMA concentrations were statistically lower immediately after subjects
267
experienced peak exercise-induced claudication pain compared to baseline and one hour recovery time points. Unlike both of these models, which combined skeletal muscle mechanical work and ischemia, our model involved isolated muscle ischemia. Thus, comparisons of results may not be appropriate. Other investigators have identified a likely explanation for immediate post-exercise skeletal muscle ischemia declines in IMA. Suspected temporal lactate interference in the ACB\ test for IMA was confirmed by the addition of increasing concentrations of lactic acid to pooled serum producing a linear decrease in IMA concentrations [6]. In the present study, post-tourniquet release samples could not be drawn until patients arrived in recovery (median of 12 min) for practical reasons. Thus, peak systemic lactic acid concentrations and associated interference with IMA measurements may have been averted. The ABC test for ischemia modified albumin measures unbound cobalt after incubation with patient serum. The amount of unbound cobalt is directly correlated to the concentration of ischemia modified albumin [1– 4] but is also inversely correlated to the concentration of unmodified albumin [6]. Therefore, changes in serum albumin concentration during periods of muscle ischemia, such as decreased intravascular volume during a marathon race [7] or increased intravascular volume due to intravenous fluid infusion during surgery, as in our study, will affect cobalt binding independent of changes in the concentration of ischemia modified albumin. In the present study, noncardiac ischemic increases in IMA could be a result of both skeletal muscle ischemia modification of albumin as well as decreased total albumin due to dilution from IV fluids. While the experimental design does not permit accurate determinations of the contribution from each mechanism to the total change in IMA, our results indicate that postarthroscopic knee surgery increases in IMA are nonspecific, and if an elevated IMA were obtained in an orthopedic patient with signs and or symptoms suggestive of post-operative ischemia, the positive predictive value of an elevated IMA is likely to be very low. The optimum IMA cutoff point for ruling out cardiac ischemia concurrent with, or following, regional skeletal muscle ischemia has not been determined, and additional studies are required to evaluate the ACB\ test performance in surgery and trauma patients. In addition, the literature lacks evaluations of IMA in patients with common medical conditions, such as congestive heart failure, diabetes mellitus, renal failure, and hypertension. Finally, albumin concentration is an important confounding variable when measuring single or serial IMA concentrations in patients suspected of cardiac ischemia. References [1] Bar-Or D, Lau E, Winkler JV. A novel assay for cobalt – albumin binding and its potential as a marker for myocardial ischemia—a preliminary report. J Emerg Med 2000;19:311 – 5.
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