Diagnostic Value of Creatine Phosphokinase (CPK) lsoenzymes in the Absence of Elevated Total CPK Willie L. Ruff, PhD, Roger Worrell, MD, and Katherine Ng, MT Washington, DC

The value of creatine phosphokinase (CPK) isoenzymes, in the presence of normal total CPK, for assessing myocardial infarction is discussed. Forty-six patients in a coronary care unit were studied to ascertain the clinical usefulness of CPK isoenzymes for diagnosing myocardial infarction when the levels of total CPK were within normal range. The majority of patients studied had some cardiac abnormality on either ECG or x-ray examination. Approximately 50 percent of patients had chest pain, but only five of the 46 had a detectable CPK isoenzyme level on CPK electrophoresis. The results of this study suggest that CPK isoenzymes do not add anything of diagnostic value to the clinical picture if the total CPK is normal. The clinical usefulness of creatine phosphokinase (CPK) is well documented. 1-6 It is most useful as a laboratory aid for the diagnosis of acute myocardial infarction. Elevated levels of CPK are associated with, but not limited to, acute myocardial infarction. Increased levels of CPK have been noted in a number of clinical conditions (eg, progressive muscular dystrophy,7'8 polymyositis,7 muscle trauma,9 surgery,9 and intramuscular injections8'9). An elevation of total CPK in serum usually suggests muscle or skeletal disease. However, it may indicate central nervous system morbidity. To date, the most common use for CPK is in the diagnosis of acute myocardial infarction. Since CPK is elevated in a variety of clinical settings, further tests must be done to improve diagnostic accuracy and predictive value. A test that is being used with increasing frequency involves CPK isoenzymes MM, MB, and BB. Total CPK begins to rise four to six hours following a myocardial infarction, peaks at 18-30 hours, and returns to normal by 72 hours.8'0 Similarly, the CPK-MB isoenzyme rises and falls. Requests for reprints should be addressed to Dr. Willie L. Ruff, Clinical Laboratories, Howard University Hospital, 2041 Georgia Avenue, NW, Washington, DC 20060.

Characteristically, CPK-MB appears transiently in the serum after an acute myocardial infarction. Whereas total CPK remains elevated for approximately 60-72 hours following an infarction, CPK-MB disappears from the serum in about 30 hours.8 Subsequent reappearance of CPK-MB should lead to suspicion of extension of the infarct8 or reinfarction.4 Previously, it was suggested that CPK-MB may be present with a normal total CPK.8 The purpose of this study was to ascertain the value of CPK isoenzyme for assessing myocardial infarction in patients with normal total CPKs admitted to a coronary care unit.

Methods and Materials A 10 ml red top vacutainer tube of whole blood was collected from each patient upon admission to the coronary care unit. The specimens upon receipt at the laboratory were allowed to stand for 30 minutes at room temperature, with the red stoppers removed and Sur-Seps (General Diagnostics) added. Then, the samples were centrifuged for 14 minutes at room temperature in an IEC centrifuge (Damon Company). At the end of the centrifugation, the sera was poured into appropriately labeled 10 ml test tubes. Each sample was assayed for total CPK by the Bergmeyert1

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modification of the Rosalki12 procedure on the ABA-100 (Abbott Laboratories). This procedure was adapted to the ABA-100 (Bio-Dynamics/Boehringer Mannheim Corporation).13 CPK isoenzymes were assayed by the use of a CPK isoenzyme Kit (Dade Division, American Hospital Supply Corporation). In this procedure, the CPK isoenzymes are separated electrophoretically and visualized by incubating the cellulose support medium on an agar tray, using the Rosalki12 method for the determination of CPK activity. The records of the patients were examined for pertinent medical history, the presence of chest pain, ECG changes or signs of infarction, and other enzyme data. The results were tabulated.

Results A profile of 46 patients admitted to the Coronary Care Unit for evaluation and treatment for suspected myocardial infarction is given in Table 1. Nearly 50 percent of the patients complained of chest pain. The majority of the patients showed some cardiac abnormality on either ECG or x-ray. Only five of the 46 patients had a detectable MB fraction on CPK electrophoresis. The 1.6 percent MB reported for patient J.H. is not considered positive (to be positive, the MB fraction must exceed three percent of the total). The CPK values ranged from 16 to 1,449 mu/ml. Figure 1 shows the relationship between enzyme activity and time for total CPK in the peripheral circulation. The enzyme begins its ascent at about four to six hours after an infarction, reaches a maximum in 18-30 hours and returns to normal by 72 hours. A typical pattern for CPK isoenzymes is presented in Figure 2. The fastest moving peak, the BB fraction, migrates to the anode. The slowest 383

Table 1. Enzyme and Clinical Findings in Coronary Care Unit Patients

CPK (mu/ml)

MB (%)

-

0 4.7

1. S.G. 2. J.S.

-

31 487

3. R.M.

-

64

0

4. M.S.

-

152

0

5. D.L. 6. V.W.

-

188 73

0 0

19 18 16 110 33

0

0 0 0 0

18 21

0 0

68 87 164 27 58 43 57 996

0 0 0

7. 8. 9. 10. 11.

+

+

L.R. H.S. W.T. E.D. A.H.

+ +

12. W.M.

-

13. C.S. 14. S.N.

-

W.P. G.J. F.H. K.C. E.W. L.F. M.G. K.C.

-

15. 16. 17. 18. 19. 20. 21. 22.

-

-

+

+

+ +

-

23. B.S. 24. R.W. 25. A.T.

+

0 0 0 0 0

0 0

-

60 100 49

26. M.Y.

+

170

0

27. D.C. 28. R.S.

+

63 26

0 N 0

29. 30. 31. 32. 33. 34. 35. 36.

384

Chest Pain

+

W.G.

+

O.J. E.S. A.W. C.C. M.C. A.A. J.D.

+

+

+

-

39 43 47 78 29 80 49 93

0

0 0 0

0 0 0 0

Other ST-T changes, anteriolateral ischemia Left ventricular hypertrophy (LVH), anterior inferior wall ischemia Premature ventricular contractions (PVCs) Biventricular cardiomegaly ST-T changes secondary to lateral ischemia Congestive heart failure (CHF) History of myocardial infarction (Ml), 1977 Non-diagnostic ST-T wave changes Inferior wall ischemia Cardiomegaly x-ray findings X-ray findings

X-ray findings X-ray findings Lateral wall ischemia, PR interval (upper limit of normal)

Non-specific ST-T changes LVH and right atrial enlargment (RAE) Inferior lateral ischemia, angina Wenckebach phenomenon LVH Non-specific ST-T changes Old inferior wall infarct ST-depression ST-T changes, PVCs Atrial fibrillation, right bundle branch block (RBBB) Atrial flutter (AF) (2:1 AV Block), General Ml Anteriolateral ischemia, coronary by-pass Atrial fibrillation, supraventricular tachycardia CHF ST-depression V4-V6,? Anterior wall infarct or ischemia? Acute subendocardial Ml, PAT with varying block Anteriolateral ischemia, First-degree A-V block Old Ml? Old anterior septal Ml Anterior ischemia Left bundle branch block (LBBB) Arrhythmia Anteriolateral ischemia Normal Anteriolateral ischemia Anterior septal hemiblock, LAE, LVH Old anterior septal Ml Old anterior septal MI

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Table 1, Continued Chest Pain 37. R.C. 38. J.H. 39. I.B.

40. E.W. 41. Q.K. 42. M.H.

CPK (mu/ml)

+

169

-

37

MB (%)

Non-specific, ST-T lateral chest leads

0

APBS anteriolateral ischemia

+

182

+

26

7.7 0

-

58

0

43. J.H. 44. W.G.

+

88

1.6

+

1,449

11.5

45. G. E.

-

685

5.7

46. B.B.

+

23

moving peak, MM fraction, migrates to the cathode. The center peak, the MB fraction, is of intermediate migratory speed. With the methodology used in this study, the MB fraction is considered absent unless it exceeds three percent of the total activity. Thus, the presence of a small peak on an electrophoresis pattern does not mean that there is a clinically significant amount of MB present. The distribution of patients according to their total CPK enzyme level is illustrated in Figure 3. Approximately 60 percent of the patients had a CPK value of less than 84 mg/ml (normal, 75 m,/ml). About 25 percent had CPK values between 85 and 200 m,/ml. In about ten percent of the patients, the CPK values reached three to 20 times the upper limit of normal. Significant positive CPK-MB results are seen only in CPKs that are elevated three to 20 times the upper limit of normal. They are not seen in normal or slightly elevated CPKs.

Discussion Heart, skeletal muscle, and brain are rich sources of creatine phosphokinase. Other tissues contain some of that enzyme, but the concentrations are only a fraction of that found in heart and skeletal muscle.7 Normally, the amount of CPK in serum is small. A decrease of CPK activity from normal is usually diagnostically insignificant. However, an increase may be highly

Other

Acute anterior-inferior-lateral Ml Left axis deviation Multiple PVCs, CVH, right atrial enlargement (RAE) Acute infarct posterior wall RAE, left atrial enlargement (LAE), acute anterior septal Ml with lateral wall extension Sinus tachycardia anterior ischemia Atrial fibrillation CHF, ventricular premature beats, RAE

0

10

6-

4-

2-

III 0

6

12

24

18

30

36

Hours

Figure 1. The

relationship of CPK activity and time.

significant. When serum total CPK is elevated, its tissue origin is often unknown. The determination of the source of the abnormal enzyme level is crucial to diagnosis and management in such patients. Creatine phosphokinase can be separated into three bands electrophoretically, by column, etc. Electrophoretically, the band with the fastest migration (to the positive terminus) is the BB band, the band of intermediate migratory speed is MB, and the slower one is MM. Designation of the bands as BB, MB, and MM is done to

JOURNAL OF ThE NATIONAL MEDICAL ASSOCIATION, VOL. 71, NO. 4, 1979

show that all bands (isoenzymes) are composed of two polypeptide chains which are homologous, heterologous, or hybrids. Further, the letters B and M designate the tissue origin of the isoenzyme-B (brain) and M (muscle). The MB hybrid of CPK is largely derived from cardiac muscle. Hence, its appearance in serum is highly suggestive of myocardial infarction. However, it must be remembered that CPK-MB is seen in other disease states, eg, polymyositis,3 Duchenne muscular dystrophy,14'15 severe angina,1 and coronary insufficiency.' 385

Mm

BB MB

Figure 2. Electrophoresis pattern of a serum sample with a positive MB fraction.

35

in any of the patients with a normal total CPK. This is consistent with the fmdings of Hadden and Prentiss.9 The discrepancy in findings between the two laboratories may be due to sampling time and/or a time delay before assaying for the enzyme. CPK-MB rises four to six hours after an infarct, peaks at 18-30 hours and returns to normal by 72 hours.8'10 Hence, the sampling time is very important. If a patient has chest pain on one day and goes to the hospital on the following day, chances are good that the MB fraction may not be found. For not only is the sample time important, but also the size of infarct. CPK has a short half-life at ambient temperature. Hence, it must be assayed promptly or stored frozen. Incorrect storage of the samples before assaying them may lead to false negative results.

30

25

Literature Cited

C: co

02 20 0 4-

10

_ 5

25

25-54

55-84

+~~~~~~~~~~~~~

85-114 115-200 201-500 501-1,500

Normal 75 mu/ml + Presence of positive MB fraction -Absence of MB fraction Figure 3. Distribution of patients according to their total CPK results.

Of the 46 patients studied, none with normal total CPK levels had a myocardial infarction. The diagnosis of myocardial infarction was based on criteria of appropriate laboratory tests and clinical impression. Only when the 386

total CPK activity exceeded three times the upper limit of normal was a positive MB present. These findings do not agree with those of Wolf et al.8 No significant positive CPK-BM was found in this study

1. Galen RS: Isoenzymes and myocardial infarction. Diagnos Med 1(1):40-52, 1977 2. Michaelson SP, Karsh DL, Wolfson S, et al: Recurrent myocardial infarction with normal coronary arteriography. N Engi J Med 297(17):916-917, 1977 3. Nirenberg DW: False-negative CPKs. N EngI J Med 297(16):894, 1977 4. Roe CR, Lee LL, Wagner GS, et al: Combined isoenzyme analysis in the diagnosis of myocardial injury: Application of electrophoretic methods for the detection and quantitation of creatine phosphokinase MB isoenzyme. J Lab Clin Med 80(4):577-590, 1972 5. Rosalki SB: An improved procedure for serum creatine phosphokinase determination. J Lab Clin Med 69:696-705, 1967 6. Sobel BE, Roberts R, Larson KB: Estimation of infarct size from serum MB creatine phosphokinase activity: Applications and limitations. Am J Cardiol 37:474-485, 1976 7. Coodley EL: Enzymes in cardiac disease. In Coodley EL (ed): Diagnostic Enzymology. Philadelphia, Lea and Febiger, 1970, pp 39-71 8. Wolf PL, Williams D, Von des Muehil E: Practical Clinical Enzymology and Biochemical Profiling: Techniques and Interpretations. New York, John Wiley, 1973, pp 354-361. 9. Hadden DM, Prentiss T: Cardiac profiling by electrophoresis. Lab Management 15(5):21-24, 1977 10. Kachman JF, Moss DW: Enzymes. In Tietz NW (ed): Fundamentals of Clinical Chemistry, ed 2. Philadelphia, WB Saunders, 1976, pp 565-698 11. Bergmeyer HU: Methods of Enzymatic Analysis, ed 2. New York, Academic Press, 1974, p 789 12. Roberts R, Sobel BE: lsoenzymes of creatine phosphokinase and diagnosis of myocardial infarction. Ann Intern Med 79:741743, 1973 13. CPK: Instrument Application Sheet. Indianapolis, Bio-Dynamics/Boehringer Mannheim Corporation, 1975 14. Brownlow K, Elevitch FR: Serum creatine phosphokinase isoenzyme (CPK2) in myositis. JAMA 230:1141 -1144, 1974 15. Lum G: Diagnostic value of CPK isoenzymes. Lab Management 15(5):14-16, 1977

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