CLINICAL CHEMISTRY, Vol. 26, No. 9, 1980 1241

CLIN. CHEM. 26/9, 1241-1250 (1980)

Serum Enzymesand lsoenzymesin the Diagnosisand DifferentialDiagnosisof MyocardialIschemiaand NecrosisJohn A. Lott1 and John M. Stang2

Diagnosis of injury to the myocardium is facilitated by in-formation on the activities of creatine kinase (EC 2.7.3.2)MB isoenzyme (CK-MB) and lactate dehydrogenase (EC1.1.1.27) isoenzyme 1 in serum, these isoenzymes beingpresent in higher activities in the myocardium than in othertissues or in normal serum. The temporal relationships ofthese isoenzymes, total creatine kinase, total lactate de-hydrogenase, and aspartate aminotransferase (EC 2.6.1.1)are highly sensitive and specific for acute injury to theheart, particularly acute myocardial infarction. Chronicheart diseases, electric cardioversion for heart rhythmdisturbances, coronary catheterization, and exerciseusually do not produce increases of CK-MB, although ab-normal aspartate aminotransferase, creatine kinase,lactate dehydrogenase, and lactate dehydrogenase iso-enzyme 1 activities are seen in some individuals. Manyother causes of increased activities of these enzymes andisoenzymes in serum are unrelated to injury to the heart.Because CK-MB is present in the skeletal muscle in lowactivities, substantial injury to skeletal muscle can increaseCK-MB activities in the blood to abnormal values. Pulmo-nary embolism can mimic myocardial infarction in itsclinical presentation. In patients with an accurately knowntime of onset of symptoms and serial enzyme analysisevery 12 h during the first 48 h, acute myocardial infarctioncan be distinguished from pulmonary embolism by deter-minations of creatine kinase, CK-MB, aspartate amino-transferase, and lactate dehydrogenase isoenzyme 1 inserum.

AddItIonal Keyphrases: myocardial infarction . creatinekinase lactate dehydrogenase aspartate aminotrans-ferase #{149} variation, source of #{149} heart disease . anginapectoris #{149} myocardltis . anhyThme pulmonaiy embolism

various conditions reported to produce change in circulating“heart” enzymes drug overdose

IntroductionEvaluation of acute injury to the myocardium has been

facilitated by the development of highly sensitive and specificdeterminations of serum enzymes. The usefulness of thesetests is based on the repeated observation that cell injury re-leases enzymes into the bloodstream. Localized anoxia is themost prevalent cause of cell injury, and can lead to loss of cellmembrane integrity. Other causes of cell injury are chemicalpoisoning, trauma, excess heat or cold, starvation, dehydra-

tion, and bacterial toxins (1). Cell death is not a prerequisitefor the release of enzymes from the cytoplasm.

Changes in the activity in serum of aspartate aminotrans-ferase (AST), creatine kinase (CK) and its isoenzymes, andlactate dehydrogenase (LD) and its isoenzymes can reflectpathological changes in the heart.3 Several fortuitous cir-cumstances make this possible: (a) These enzymes are presentin extremely high activity in the cytosol of myocardial tissue,compared with their usual activities in serum. (b) The cyto-solic enzymes are released during injury and eventually enterthe vascular compartment. Mitochondrial and cell mem-brane-bound enzymes are released less readily, and mito-

chondrial AST and CK, for example, are seen infrequentlyafter myocardial injury (2, 3). (c) After release from tissue, theenzymes have a biological half-life in blood of hours to days,which permits post-injury detection. (d) CK-MB is presentin significant activities only in myocardium; its appearancein blood in amounts greater than 3 to 4% of the total CKusually points to myocardial injury.

Among diseases of the heart, serum enzyme analyses havetheir greatest application in the diagnosis of acute myocardialinfarction. Chronic diseases such as angina pectoris andcongestive heart failure usually are not associated with in-creased serum activities of enzymes from heart tissue.

Reference Values for Enzymes and lsoenzymesCrea tine kinase. Because of the great diversity of en-

zyme-assay methods in common use (4), there are no uniformserum reference values for AST, CK, LD, and their isoen-zymes. The reference values for CK-MB are of particular in-terest, this enzyme being currently the best serum marker ofinjury to the myocardium.

CK-MB activity in the myocardium is about 14% (5), 22%(6), or 42% (7) of the total CK activity; CK-MB in skeletalmuscle represents about 1% (8,9) or 2.7% (7) of the totalCKactivity. It is unlikely that the CK-MB in heart tissue variesas much as shown here; the use of fresh tissue and a stan-dardized methodology may yield a more definitive value.There are traces of CK-MB in many body tissues (6), butnearly all the CK-MB is present in the heart.

CK-MB is ordinarily present at very low activity in serum;whether it is found in the serum of healthy persons dependson the method used. Reported serum reference values forCK-MB by electrophoresis range from “absent or undetect-able” to “trace” (5, 10-18). Using agarose electrophoresis, wefound that the limit of detection for CK-MB to be about 3U/L. Electrophoresis followed by elution of the CK-MB bandand spectrophotometric quantitation is more sensitive than

Departments of Pathology’ and Medicine,2 Ohio State University,Columbus, OH 43210.

Direct correspondence to John A. Lott, Ph.D., Department of Pa-thology, Ohio State University Hospitals, Room N-329, 410 West 10thAvenue, Columbus, OH 43210.

Received Dec. 17, 1979; accepted April 9, 1980.

‘ Nonstandard abbreviations used: AK, adenylate kinase, EC2.7.4.3; ALT, alanine aminotransferase, BC 2.6.1.2; AST, aspartateaminotransferase, BC 2.6.1.1; CK, creatine kinase, EC 2.7.3.2;CK-MM, CK-MB, and CK-BB, isoenzymes of CK; LD, lactate de-hydrogenase, EC 1.1.1.27; LD-1, LD-2, etc., isoenzymes of LD; AMI,acute myocardial infarction; ECG, electrocardiogram.

00.51.02.04.08.0

16.0

530680840

1000172029005300

0.790.840.850.890.910.870.85

Hemolysis

nonevery slightslightmoderategrossgrossgross

1242 CLINICAL CHEMISTRY, Vol. 26, No. 9, 1980

Table 1. Reference Values for CK and CK-MBCK, total, CK-MB, Stated

Reference U/La U/I5 populatIon

Chromatographic methods5-90 2.9(2.1) Laboratory staff: 9

men21 74(32)

22 31.1 (16.6)b 0.8 (0.5) Blood donors: 25 men,

15 women

23 42-123 0-1.4 Laboratory staff: 24

217-1140 0-8.0 Patients without acute

myocardial

infarction: 2424 17-228 0-5.4 Asymptomatic men:

5314-296 0-4.5 Asymptomatic women:

5125 0-60 None Healthy young

persons: 6926 48 (20) 2.4 (0.7) Non-hospitalized

ambulatory

subjects: 19Immunoinhibition method

27 Up to 270 to 6C Healthy patients: 109men

Up to 150 to 6C Healthy patients: 127women

Radioimmunoassay method28 - l.2-l2.S’ Healthy adults: 51

‘Stated range, or mean (SD). CK-MM only. C CK-B’. d g/L.

electrophoresis alone; by this approach, no CK-MB was seenin healthy persons (19), and up to 2 U/L was seen in hospi-talized patients who did not have acute myocardial infarction(20).

CK-MB has been demonstrated many times in the serumof healthy persons by chromatographic methods (see Table1). The reference values are quite different, and CK-MB ac-tivity as a percentage of total CK activity also varies greatly.The variability may be related to analytical error, populationvariations, and differences in methods. Table 1 can be a guidefor individuals who use chromatographic methods, but ref-erence intervals are best determined locally. CK-MB can alsobe detected in normal serum by immunoinhibition (27) andradioimmunoassay (28).

Results from patients have been used to develop discrimi-nation values for CK-MB, to distinguish between individualswith and without acute myocardial infarction. The limits arerather arbitrary and are higher than CK-MB values in healthypersons: Patients with acute myocardial infarction reportedlywill have a CK-MB of >3% (29), >4% (23, 30), >5% (32), >3U/L (31), or >5 U/L (14,26).

Reporting CK-MB as a percentage of total CK has meritbecause after massive skeletal muscle injury, both total CKand CK-MB may be increased in serum, but the percentagethat is CK-MB typically will be less than 3-4% of total CK andthe source of the increased serum enzymes is skeletal muscle.For optimum aid to interpretation, CK-MB should be re-ported both as a percentage of CK and in units per liter. Theformer points to the source, the latter to the amount of enzymereleased.

For total CK, healthy subpopulations exhibit strikinglydifferent values. In testing medical students, we observed themeans in men to be 1.5-fold the mean in women, probably

because of a larger muscle mass in men (33). Exercise is alsoan important variable (see below).

Lactate dehydrogenase. Reference ranges for LD isoen-zymes as determined by electrophoresis and staining with dyehave been reviewed and summarized (34). Again, as othershave shown (35, 36), the reference values are method- and (or)population-dependent. The values we obtained for 49 ap-parently normal volunteer blood donors after agarose elec-trophoresis and fluorescence detection agreed well with thoseof DiGiorgio (34). None of our blood donors showed the ab-normal LD-1 > LD-2 pattern which is seen after acute myo-cardial infarction.

Interference from Hemoglobin, and Stability ofSamples

The quality of serum samples determines whether validenzyme and isoenzyme results can be obtained.

The erythrocytes are rich in adenylate kinase (AK) butpractically free of CK; however, AK will be measured as CKonce the AK inhibitor in the CK reagent is overcome. Weadded saline-washed and lysed erythrocytes to serum andfound that for 0.5 g hemoglobin per liter there was little in-crease in “CK” (CK + AK), but with 1.0,2.0,4.0,8.0, and 16.0g of hemoglobin per liter, the “CK” activities increased pro-portionately from 40 to 340 U/L. With the ACT (CorningMedical, Medfield, MA 02052) agarose electrophoreticmethod, hemolysis up to 16.0 g/L does not prevent detectionof CK-MB from heart or skeletal muscle; the AK fromerythrocytes migrates cathodal to CK-MM and does not ob-scure CK-MB if present.

Hemolysis is intolerable in the measurement of serum LD;however, hemolysis alone will not produce the LD-1 > LD-2“flip.” We added saline-washed and lysed erythrocytes toserum with the following results:

Hemoglobin, LD, total LD-1/LD-2gIL UIL ratio

LD isoenzymes were determined by agarose (Corning) elec-trophoresis. Our findings were consistent with the observationthat LD-2 exceeds LD-1 in erythrocytes (37).

Serum samples for CK analysis should be stored in the darkat 4 or -20 #{176}Cif analysis is delayed. Full activity is restoredby N-acetyl-L-cysteine after storage for 7 days at 4 #{176}Cor onemonth at -20 #{176}C(38). Native CK-MB is very stable in serumstored at 4 #{176}C;we saw no loss in activity during storage for aslong as 11 days (39).

LD is unstable in serum, losing activity during storage at25, 4, or -20 #{176}C;the least loss occurs at 25 #{176}C(40). If imme-diate analysis is impossible, the serum should be stored atroom temperature but analyzed within 48 h, at which time theloss in LD activity will be about 6% or less (40). We deter-mined the stability of LD isoenzymes by prompt analysis offresh sera and re-analysis after storage at 25 and 4 #{176}C.Thedata in Table 2 give the ratio (times 100) of the enzyme’spercentage of total LD on the day of collection to the percentof total on the day of re-analysis. LD-5 is the most labile iso-enzyme, and is lost faster at 4 #{176}Cthan at 25 #{176}C.Only 25 #{176}Cshould be the temperature at which serum samples are storedfor LD isoenzyme analysis, and the test should be performedwithin 24 h of collection.

We observed no loss of activity for AST, a very stable en-

CLINICAL CHEMISTRY, Vol. 26, No. 9, 1980 1243

Table 2. StabilIty of LD Isoenzymes after Storage at 25 #{176}C(n =

sosnzymss of LO

12) or 4 #{176}C(n = 11)

1 2 3 4 5

Percentage ratio (SD)5

After 1 day at 25 #{176}C 102 (7.7) 98(12.1) 101 (4.6) 103 (7.0) 99(20.4)After 2 days at 25 #{176}C 106 (8.0) 102 (2.8) 102 (3.0) 93 (9.0) 87 (12.9)

After 1 day at 4#{176}C 109(21) 109(7) 98(7) 91(7) 89(12)After3daysat4#{176}C 119(29) 115(11) 97(6) 76(8) 82(12)

CV,%(n=51) 5 2 2 4 9

‘llsoenzyme’s % of totaI LO (on day of collection/on day of reassay)] X100.

zyme, during daily analysis of poois with 25 U/L (normal)activity and sixfold normal activity when they were stored for9daysat4or-20#{176}C.

Choice of Method for CK-MBThe interpretation of an abnormal CK-MB activity de-

pends on the method used. Electrophoresis is technicallysimple but is less sensitive than chromatography (41, 42).Although electrophoresis is at best semi-quantitative, thisdoes not diminish its clinical usefulness. One may actually seethe isoenzyme bands, and can readily note an unsatisfactoryseparation of the isoenzymes. This is not true for chroma-tography, where CK-MM can merge into CK-MB on an un-satisfactory column or in a sample with a very high CK-MMactivity. In some chromatographic methods CK-MB andCK-BB are eluted together, which is undesirable. Chroma-tography is more time consuming than electrophoresis, butwith good technique it can be more precise than electropho-resis (39).

Immunoinhibition methods for CK-MB are technicallysimple, but have a lower sensitivity and specificity for acutemyocardial infarction than electrophoresis or chromatogra-phy. The problem may lie in the quality of the antibody andthe complete inhibition of the enzymatic activity of CK-M.We found that in specimens with low activities of CK-MB,

extraordinarily precise and stable spectrophotometry is re-quired to produce reliable results (16). Becauskimmunoin-hibition (anti-M) methods do not distinguish betweenCK-MB and CK-BB, CK-BB could be interpreted as CK-MB(43).

Radioimmunoassay methods for CK isoenzymes are beingdeveloped and may be the methods of the future (44-46).Reportedly, they are 10-fold more sensitive than chroma-tography in detecting CK-MB after acute myocardial in-farction.

Wagner puts the issue of which method to use into a clinicalperspective:”.. . one of the attractive aspects of the [electro-phoretici CK isoenzyme technique is that it does not requirethe observation of either a change in ratio of different isoen-zymes or an elevation above the upper limit of normal, butrather the detection of the appearance of an MB band” (47).Despite the lower sensitivity of electrophoresis, it provideseminently useful clinical data.

Changes in Serum Enzymes as Related to theDiagnosis of Heart Disease

Effect of Exercise

Serum AST, CK, and LD activities always must be con-sidered in the context of the physical activity of the individual.Increased serum CK as a result of exercise is well recognized(19,48-55). Increases in AST (50-53) and LD (49,51-53) havealso been seen, but the changes were generally smaller thanfor CK. Extreme physical exertion during a 160-km marathon

#{149}increasedserum CK as much as 40-fold and LD as much as12-fold the upper reference limit in 20 runners of ages 27-63years (19). CK is the most sensitive indicator of micro traumato skeletal muscle.

When the effects of physical stress and acute myocardialinfarction need to be differentiated in patients, assay of theisoenzymes of CK and LD can resolve the problem. CirculatingCK-MB has not been observed in persons after exercise, eitherby electrophoretic (54), chromatographic (55), or immu-noinhibition methods (56). Mild exercise may produce onlya “trace” (57) of CK-MB in blood; only extreme exercise willresult in as much as threefold the upper reference CK-MBactivities (19). In 13 well-conditioned collegiate varsityswimmers whose blood was sampled during their period ofactive training, we saw traces of CK-MB in six, 4 U/L in one,and none in the rest; assay was by electrophoresis. Values forAST and LD were normal or only slightly increased in all 13;LD isoenzymes were always normal. A serum LD-1 > LD-2has been seen in athletes after marathon running (19, 54).

Extreme physical stress probably causes release of enzymesfrom the heart in the absence of acute myocardial infarction.The LD-1 > LD-2 pattern is convincing evidence of this, asis the appearance of CK-MB. However, in the presence of asharply increased CK, CK-MB from skeletal muscle may alsobe detected, even with the less-sensitive electrophoreticmethods, but the C K-MB will usually be less than 3% of thetotal CK.

Effect of InjectionsIntramuscular injections can produce CK increases in blood

(11,56,58-61). The CK is from skeletal muscle, and there isno injury to the myocardium; CK-MB remains normal.

Changes during Ischemic Heart DiseaseAcute myocardial infarction. Correct diagnosis of acute

myocardial infarction is an important clinical challenge andmay determine the survival of the patient. The history maybe noncontributory and electrocardiographic changes may benonspecific or absent, particularly with a small infarct; con-sequently, the clinician relies on laboratory determinationsof serum enzyme activities to help with the diagnosis. Afterinfarction, enzymes originating from the cytosol of myocardialtissue are detected in increased activity in the peripheralblood. The ideal enzyme test would be positive when an infarcthas occurred (sensitive) but negative in the absence of one(specific). Further, the test should promptly become positiveafter the onset of infarction and remain positive for a pro-longed period, so that an abnormal activity would still bedetected in patients admitted after a delay. Assays for CK-MBapproximate this ideal, if the testing is done at frequent in-tervals within the first 48 h after onset of symptoms.

Five recent articles review the use of serum enzyme data forthe diagnosis of acute myocardial infarction (62-66). Here,we summarize and condense the reports that have appeared

Table 3. SensitIvity and Specificity of Diagnostic Tests for Acute Myocardial infarction

PatIent populatIon SensitIvity, SpecIfIcIty,Reference Total Proven Infarct Test %

Konttinenand 61 27 CK 100 -

Somer, 1973 CK-MB (a) 97 -

(14) LD1 abnormal (a) 91 -

Wagner et al., 328 91 ECG 66 1001973(6?) CK 98 85

CK-MB (a) 100 99LD1 > LD2 (a) 90 95

Blomberg, et al., 212 51 ECG 63 1001975(15) CK 100 85

CK-MB (a) 94 100Galen, et al., 94 46 EGG 72 100

1975(29) CK-MB(a) 100 93

LD1 > LD2 (a) 78 99

Varat and Mercer, 100 47 CK 94 721975(23) CK-MB(b) 100 92

Roarketal., 151 80 ECG 78 1001976(13) CK 100 65

CK-MB (a) 96 100

Neufeld et al., 400 CK-MB (a) 100 98

1977 (68)Flanker,1978 (69) - 69 CK-MB (c) 93 -

Kraftetal., 201 87 GK 93 76

1978 (25) CK-MB (b) 98 97

LD 91 80

LD1 abnormal 86 90

Grandeetal., 401 192 ECG 82 100

1978(70) AST 97 88

CK 97 78

CK-MB (a) 100 98

LD 97 81

von Arnim 143 34 ECG 74 100

etal.,1978 CK 94 57

(71) CK-MB (C) 70 95

Gann etal., 98 44 ECG 70 100

1978(18) CK-MB(a) 100 86LD1 > LD2 (a) 70 94

Leungand Henderson, 228 101 LD1/LD2>0.76(a) 100 91

1979(72)

BillstrOmet 370 173 CK 100 86

al.,1979 (43) CK-MB (c) 99 100

Obzansky and 71 28 CK 100 88

Loft,1980 CK-MB (a) 96 98

(16) CK-MB (c) 79 93

LD1 > LD2 (a) 61 98

Straussand Roberts 67 29 CK 100 60

1980(73) CK-MB(d) 100 100

LD 100 39

AST 89 48

AVERAGES, 2525 1499 ECG (7) 73 100

above studies CK (11) 98 75(no.of studies) CK-MB (a) (9) 98 97

CK-MB (b)(2) 99 95CK-MB (c)(4) 85 96

LD1 > LD2 (a)(4) 75 97

(a). electrophoresis; (b). chromatography; (c), lmmunolnhibltion; (d). electrophoresis followed by elution and fluorescence analysis.

1244 CLINICALCHEMISTRY, Vol.26,No.9,1980

No. of patients

Peak CK, U/L5

Totaltime CK was

abnormal

Peak CK-MB, %

Totaltime CK-MB

was abnormal

AMI wIthuneventful

course

43

25-68

24-30 h

2-18%

max. 20 h

AMI wIthtachyarrhythmlas

12

51-195

4 days or more

5-40%

max. 36 h

AMI withcardlogenlc

shock

5

195-330

“ >4days

29-41%

>4 days

AMI withsuspected

relnfarctlon

38

28-245

variable

6-38%

variable

CLINICALCHEMISTRY, Vol.26,No. 9,1980 1245

since CK-MB was recognized to be an important marker ofmyocardial injury (see Table 3).

Electrocardiograms, although specific, average only 73%sensitivity (27% false-negative results). This agrees very wellwith the reported 30% of patients who had autopsy-proveninfarctions but normal electrocardiograms (74), From thesummary at the end of Table 3-if we assume that it is validto combine and average results from different reports-CK-MB determination by electrophoresis or column chro-matography is the most efficient (75) of the listed proceduresfor the diagnosis of acute myocardial infarction.

Immunoinhibition procedures for CK-MB (16, 69, 71) areboth less sensitive and less specific for acute myocardial in-farction than are electrophoretic and chromatographicmethods. The report by Billstr#{246}met al. (43) is an exceptionto this.

An abnormal result for an LD-1 test, i.e., where LD-1 ex-ceeds LD-2 (“flipped” LD-1 and LD-2 bands) is not as sen-sitive for acute myocardial infarction as CK-MB; however, a

falsely positive LD-1 test rarely occurs, making the test veryspecific. The test may have greater sensitivity and slightly lessspecificity for acute myocardial infarction if test results areinterpreted as abnormal when the ratio of LD-1 to LD-2 is 0.76or more (72). A normal CK value cannot be used to rule outacute myocardial infarction when the time of infarction isunknown; but because AST can remain abnormal for about96 h and LD for more than 120 h after infarction, they are ofvalue in diagnosing acute myocardial infarction when the timeof onset is unknown or when hospital admission is delayed(68).

Note that a normal value for CK should not be used to ex-clude analysis of CK-MB. Documented cases of acute myo-cardial infarction show that the CK value remained within thereference range but CK-MB was increased (17, 66). Never-

theless, an abnormal CK-MB and a normal CK should bequestioned for possible laboratory error or an artifactualCK-MB. When CK-MB is released from the myocardium,about fivefold as much CK-MM is released with it; further,the half-life of CK-MB is shorter than that of CK-MM (76).Thus it is unusual to have an increased CK-MB and a normalCK in a patient with suspected myocardial infarction.

In many patients with acute myocardial infarction, thereis a point at which there is an acute onset of symptoms. The“time window” during which CK, CK-MB, and LD-1 are ab-normal differs for each enzyme, but is brief in “uncompli-cated” infarction. In six patients with proven infarction whoseblood was frequently sampled, serum CK and CK-MB activitytook 2 to 12 h after the episode to increase. CK-MB persistedfor not less than 24 h or more than 72 h. CK was increased foran average of 60 h, peaking at 16 to 20 hand persisting for anaverage of 24 h after the disappearance of CK-MB (67). Othersfound that CK-MB was normal from 0 to 4 h after infarction,

and that CK-MB disappeared after 36 h (71). We found in allof 11 patients for whom the time of infarction was reasonablywell established that both CK and CK-MB became abnormalwithin the first 48 h (16).

The typical time course for CK and LD isoenzymes is forCK-MB to peak first, with LD-1 exceeding LD-2 3 to 5 h later(77). The average peak times after infarction in 47 patientswere 6 h for CK-MB, 18 h for CK, 24 h for AST, and 48 h forLD (15). This is the classical temporal sequence of the enzymeabnormalities after acute myocardial infarction, but myo-cardial infarction extension and actual re-infarction willchange it. New infarctions can be detected or excluded readilyby enzyme tests in patients where re-infarction is suspectedor in those with post-infarction symptoms (73). The classicaltemporal sequence of enzyme and isoenzyme abnormalitiesis highly sensitive and specific for acute myocardial infarction;however, in one report (68) 25% of the patients with acutemyocardial infarction did not follow this pattern (see Table4).

The pattern of CK-MB release is different in patients withcardiogenic shock after acute myocardial infarction, wherethere may be infarction of 30-40% of the myocardium. Peakenzyme activities are higher and the high activities persistlonger. Progressive myocardial damage may be occurring,which leads to a vicious cycle of progressive compromise ofcardiac function and worsening ischemia. The prognosis isgrave (68, 78).

Blood should be sampled at admission and at 8- to 12-hintervals for at least 48 h after an episode that may indicateacute myocardial infarction. The minimum number of sam-ples for enzyme analysis is two, obtained 12 and 24 h after theonset of symptoms (17). If blood is collected only every 24 h,then there is a chance of missing an increased CK-MB in thepatient. If the time of onset of symptoms is accurately known,

and if during a 48-h period blood samples taken every 12 hindicate that CK-MB, CK, and LD-1 are always normal, acutemyocardial infarction can be excluded. Exceptions to this areextremely uncommon (see Table 3). Further, if the enzymevalues peak within the first 48 h and then return to normal,collections beyond 48 h are unnecessary for making the initialdiagnosis. Subsequent samples are necessary only if compli-cations of the clinical course, such as extension of the infarctor re-infarction, are suspected.

Angina pectoris. Angina pectoris is believed to result fromtransient ischemia of the myocardium that falls short ofproducing infarction. Marked coronary atherosclerosis is

usually present in patients with angina pectoris, but, by def-inition, myocardial infarction is absent. Angina pectoris is thusa clinical syndrome, not an anatomic disorder. The imbalancebetween oxygen need and supply to the myocardium is be-lieved to be the source of the pain, although the exact causeof pain is unknown. Undoubtedly, some patients with occlu-

Table 4. Time Course of Enzyme Abnormalities after Acute Myocardial Infarct (AMI)DIagnosIs

‘Reference ranges: CK, 0-20 U/L; CK-MB. absent by electrophoresis. (Adapted from ref. 68 with permission.)

1246 CLINICAL CHEMISTRY, Vol. 26,No.9,1980

sions of the smaller coronary branches are diagnosed as havingangina pectoris rather than acute myocardial infarction. Ina recent report, five patients with chest pain, abnormalCK-MB activities of 6-17%, peaks of CK-MB at 8 h, and dis-appearance of CK-MB in 10-14 h were diagnosed as having“acute coronary ischemia” rather than acute myocardial in-farction. Acute myocardial infarction was ruled out on thebasis of a normal value for serum myoglobin and other find-ings (79). Given that all these patients had abnormal CK-MBand increasing, then decreasing, CK-MB activities after theonset of chest pain, some myocardial injury was likely.

Isolated cases of angina pectoris in large series have also hadabnormal CK-MB activities (25,29,80); minor necrosis cannotbe ruled out in these patients. Activities of LD isoenzymes inserum reportedly are normal in angina pectoris (81). Instancesof abnormal serum enzyme activities in angina pectoris areso infrequent that an increased CK-MB must be consideredrepresentative of myocardial injury in the patient with chestpain and suspicious symptoms.

Changes during Non-lschemic Heart DiseaseCongestive heart failure. Congestive heart failure usually

is the result of decreased pumping capacity or an increasedpressure/volume load on the heart. The consequence is aninadequate blood supply to metabolizing tissue. Although theonset of congestive heart failure may be related to progressivecoronary atherosclerosis, the morphological changes seen aregenerally away from the heart and secondary to circulatoryfailure.

Increased CK-MB actually has been seen in a few patientsin large series diagnosed as having congestive heart failure (25,29, 32). Such results may have signaled injury to the heart.

The enzyme abnormalities commonly seen in congestiveheart failure are increased activities of AST, alanine amino-transferase (ALT), LD, and LD-5, probably all originatingfrom liver as result of passive congestion (81, 82). In thepresence of hepatic necrosis secondary to passive liver con-gestion, AST and ALT values may be sharply increased,whereas CK may be normal. The AST activities in such casesmay be much higher than those seen after acute myocardialinfarction (83).

CK-MB activity was increased in 19 of 129 patients withcongestive heart failure, particularly in patients with in-flammatory heart disease (84). There was no correlation be-tween the amount of CK-MB activity and the severity ofventricular dysfunction. In acute myocardial infarctioncomplicated by congestive heart failure, there may be acombined pattern of increased heart enzymes and liver en-zymes in blood.

Myocarditis. Abnormal serum AST, CK, CK-MB, and LDactivities usually are present in myocarditis during the courseof active inflammation (85,86). CK-MB and LD-1 values canbe markedly abnormal, and the pattern of enzyme abnor-malities mimics acute myocardial infarction.

In one case of infectious myopericarditis, we initially sawincreased enzyme activities (CK was fourfold, AST threefold,and LD twofold the upper reference value) and increasing,then decreasing, activities of CK, CK-MB, and LD, and theLD-1 > LD-2 pattern. The similarity to the clinical pictureof acute myocardial infarction was striking, in that the patientalso presented with acute-onset chest pain. The early elec-trocardiogram suggested acute inferior myocardial infarction;later tracings pointed to pericarditis. Acute myocardial in-farction in this 34-year-old man was ruled out on the basis ofa normal myocardial scintigram. A fourfold inciease in thetiter of Epstein-Barr virus supported a diagnosis of nonbac-terial infectious myocardial inflammation.

Arrhythmias. Heart rhythm disturbances usually do not

lead to the release of enzymes into the blood. If tachycardiaoccurs, AST, CK, and LD activities may increase (87-89).Tachycardia may subject the myocardium to unusual stress;for example, in the presence of atherosclerosis, the decreaseddiastolic time caused by tachycardia may compromise coro-nary filling, such that oxygenation of the heart maybe inad-equate (90). Not surprisingly, CK-MB activity is abnormalin a small percentage of patients with tachycardia (23, 68).

Effects of Medical Procedures on Enzyme ReleaseTherapeutic electric countershock. Serum CK increases

in about half of patients who are given electrical countershockto correct heart-rhythm disturbances. The powerful con-tractions of the intercostal muscles caused by the shock causea release of CK (91, 92). CK and LD increased slightly in 6%to 18%, respectively, of patients so treated, but CK-MB andLD-1 remained normal. In no cases did the patient’s valuesexceed the laboratory’s reference limits (93). Other authorsfound that CK rose above pre-treatment activities in six of 12patients after countershock, but only two patients had post-shock CK activities that slightly exceeded the reference range(94). Some found increased CK (to sixfold the upper referencelimit) in 15 of 30 patients after countershock, the rest re-maining within the reference range. Two patients had CK-MBactivity about twice the reference limits; however, theirCK-MB activities were much less than what was seen in acomparative population with acute myocardial infarction (95).Cardioversion by countershock does not obscure the diagnosisof acute myocardial infarction when based on serial deter-minations of enzymes, including CK-MB, because after theprocedure the enzyme increases are very transient, andCK-MB usually is not increased.

Cardiac catheterization and coronary arteriography.These two commonly performed diagnostic procedures usuallydo not produce serum CK-MB abnormalities unless acutemyocardial infarction occurs (67, 96). CK activity increasedin 46% of patients studied (67), but the increase seen in an-other study was solely the result of injections (97) or possiblytrauma at the site of insertion of the catheter (98). CK-MBremained normal in all 39 patients in one study (99) and in 94of 100 patients in another (55) after catheterization of thecoronary arteries. Of the six patients affected in the latterstudy, one had an intra-myocardial injection, one an en-domyocardial biopsy, one experienced arrhythmia requiringcountershock, and three had unexplained increases of CK-MB. Cardiac catheterization usually is not associated withmyocardial injury. When CK-MB is seen, some injury to themyocardium must be considered.

Exercise testing in patients with suspected coronary arterydisease. Serum enzyme changes are negligible in patientsundergoing exercise-stress testing. Small increases in LD-5have been seen, although CK and LD-1 were normal imme-diately after exercise (100). Generally, enzyme activities peak12 to 18 h after exercise, and samples should thus be collectedat that time. In another report, CK was increased 19 h afterexercise, and more patients with abnormal electrocardiogramshad increases in CK (101). AST, CK, LD, and a-hydroxy-butyrate dehydrogenase (EC 1.1.1.27) were unchanged 20 hafter exercise in 28 patients with coronary artery disease. Tenof these patients had chest pain and electrocardiographicchanges. CK-MB remained normal in all (102). Exercisetesting does not confound a diagnosis of acute myocardialinfarction; this is important because the test itself may pre-cipitate bona fide infarction.

Miscellaneous Causes of Serum EnzymeAbnormalities Which May Involve the Heart

Poisoning. Overdoses of ethanol, hypnotics, sedatives,tranquilizers, and antidepressants, and exposure to CO can

CLINICAL CHEMISTRY, Vol. 26.No. 9,1980 1247

lead to marked increases in serum enzymes; in some patients,toxic effects on the heart can be demonstrated.

Alcohol in overdose can produce myopathic changes leadingto a seven- to 23-fold increase in serum CK (103-106), a160-fold increase in CK-MB (107), and two- to fivefold in-creases in AST and LD (104, 105). Patients with deliriumtremens had the highest values for CK (103) and CK-MB(107), and their high activities were not believed to be relatedsolely to muscular activity. The enzyme release is the directtoxic effect of alcohol on muscle cells, both skeletal andmyocardial, and a possible change in membrane permeability.Cardiomyopathy and reversible right bundle-branch blockhave been described in a patient poisoned with ethanol (104);CK-MB increased by as much as 160-fold in another groupalso points to heart injury (107).

Normal serum ALT, AST, and CK activities were observedin one group of five patients in coma and overdosed withamitriptyline, a barbiturate, or methaqualone, but a secondgroup of 14 patients poisoned with a barbiturate, carbromal,methaqualone, or ethchlorvynol showed as much as 30-foldnormal CK and fourfold normal AST and ALT activities. Themagnitude of the increases correlated roughly with the du-ration of coma (108). Cardiac injury was ruled out on the basisOf clinical, biochemical, and electrocardiographic evidence,but because isoenzyme studies were not done, myocardialinjury could not be excluded in this study (108). Increased CK,but no CK-MB, was seen in four patients (109) and a 20-foldincrease of CK was seen in one patient after barbiturate poi-soning (110). Glutethimide poisoning in two patients pro-duced up to 120-fold increases in CK and traces of CK-MBin blood. The increase in enzyme activities was attributed tohypothermia during coma (111), and the CK-MB probablycame from skeletal muscle. Quinidine produced a 22-fold in-crease in CK and a twofold increase in AST in another patient;CK-MB was normal, and there was probably no myocardialinjury (112). CO is a potent muscle poison (113) and increasesof 1000-foldnormal have been seen in both CK and CK-MB;cardiac injury is likely (43). Imipramine caused myocardialdamage in a 2’/2-year-old child who ingested 1 g of the drug;bizzare electrocardiographic changes and fivefold normal CKactivities were seen. Recovery was complete except for apersistent partial right bundle-branch block (114).

Trauma. Trauma to skeletal muscle, particularly if massive,can lead to increased activities of AST, CK, CK-MB, and LDin serum. We have seen automobile accident cases with in-creased CK-MB, although the amount of CK-MB was usuallyless than 2% of the total CK activity. We and others (115) haveobserved CK activities of 100-fold normal and an intenseCK-MB band on the electrophoretogram in patients withmultiple trauma not involving the thorax; the CK-MB wasabout 1% (our observation) or less than 5% (115) of the total

CK. Traumatic injury to the heart, whether accidental or after

cardiac surgery, can result in an increased CK-MB in serumin the same range as seen after acute myocardial infarction (9,77, 116). As much as 124 U of CK-MB per liter has been de-tected in blood in 23 of 53 patients after traumatic, acutecerebrovascular, or infectious brain damage, and 15 of the 23had electrocardiographic evidence of myocardial injury (117).These authors propose that acute brain injury may lead tosecondary acute myocardial infarction.

Hypothermia-hyperthermia. Profound hypothermia cancause release of C K-MB to the extent of 3 to 32% of total CK,coming from both heart and skeletal muscle (118) because ofdiffuse injury of muscle. Hypothermia induced in dogs wasshown to cause a 10-fold increase in serum CK (118).

Patients with fever usually have normal values for serumenzymes. However, the syndrome of malignant hyperpyrexiamay cause 125-fold increases in CK, with CK-MB as high as

6% of total CK (119). This is a well-defined disorder, and the

very high temperatures can be produced by muscle relaxantsand certain anesthetic agents in genetically predisposed in-dividuals. It is also seen in some patients without any familyhistory of the disease (120-124). There is undoubtedly damageto the myocardium, and the prognosis is grave in a fulminantcase.

Reye ‘s syndrome. Patients with Reye’s syndrome are be-lieved to have myocardial injury. Increased AST, CK, CK-MB,and LD have been seen in patients with this disorder (125-127), and a mitochondrial band migrating cathodally toCK-MM has been seen (127). Up to 70-fold normal CK ac-tivities were present in serum, and the patients who suc-cumbed had the highest values (125, 126).

Pulmonary embolism. Pulmonary embolism often must bedistinguished from acute myocardial infarction, because the

clinical presentation may be similar; however, serum enzymetests are not very helpful in making a diagnosis of pulmonaryembolism. Wacker’s triad of an increased LD and bilirubinand normal AST obtained in only 12% of a series of cases(128). LD was increased in 87% of patients with pulmonaryembolism, but there were many false positives, so the LD testis nonspecific (128).

Lung tissue contains high activities of LD-3 (37), thoughthe isoenzyme pattern in serum after pulmonary embolismdoes not commonly show an increase in LD-3. Rather, whatis seen is an increased LD-1, possibly from hemolyzed eryth-rocytes, or an increased LD-5 (129, 130). Although CK-BB isthe predominant isoenzyme in lung, the total CK activity inlung tissue is only about 0.3% of the total CK in skeletal muscle(6). Increases in serum CK activity after pulmonary embolismwould not be expected, in light of the low activity of CK in lungand the instability of CK-BB in blood (76). Patients withpulmonary embolism do not show increases in CK if otherevents such as acute myocardial infarction and intramuscularinjections have not occurred (129). If the cardiac enzymes are

persistently normal after acute onset of symptoms, acutemyocardial infarction may be ruled out in these patients.

Myopathic disorders. Duchenne’s muscular dystrophyoften presents with greatly increased serum CK and CK-MB;myocardial involvement in early disease is believed to be ab-sent (8, 131), and the CK-MB may come from immatureskeletal muscle. Polymyositis-dermatomyositis is associatedwith a mean CK-MB of 3.4% (9) and 6 to 39% (132) of totalCK. None of these patients had acute myocardial infarction,and the differential diagnosis in such patients is rarely aproblem. “Increased” CK-MB was reported in a patient withRocky Mountain spotted fever (133), and one case of typhoidfever seen here had 240-fold normal CK and about 1% CK-MBin serum. Both cases were believed to have no cardiac in-volvement.

We thank H.-D. Gruemer, M.D., and Carl E. Speicher, M.D., forhelpful criticisms of the manuscript, and Mrs. Cindy Berner for sec-retarial assistance.

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