Discussion of Cardiac Enzymes - dated as no real discussion of highly sensitive Troponin Assays

1. Biochemical Detection of AMI
Frank W. Meissner, MD, FACP, FACC, FCCP

2. Historical Perspective
●AST, previously SGOT, used since 1954
when elevated levels were described in
4/5 patients with clinical MI
●Abnormal levels of LDH were reported
in the next 3 years

3. ●1959, elevations creatine kinase (CK)
found patients => muscular dystrophy
●1960, elevated CK in patients with MI
●1962, WHO listed enzymatic abnl as
diagnostic criteria for MI
●1966, CK isoenzymes identified
Historical Perspective

4. Enzyme Characteristics
●CK catalyzes transfer of high-energy
phosphate groups
●Found predominately in tissues that
consume large amounts of energy
●2 subunits termed M and B with 3
distinct isoenzymes
■Subforms of both MM and MB have been
described

5. ●CK is inactivated and proteolyzed in
lymph
●CK is not excreted with the urine and is
not influenced by renal or hepatic flow
●Plasma levels of CK depend on tissue
injured, blood flow and local lymphatic
perfusion
Enzyme Characteristics

6. Distribution CK Tissue
Isoenzymes
●MM CK predominant isoenzyme of muscle
actual concentration variable
●Substantial proportion skeletal muscle may
contain 1- 3 % of MB CK
●Skeletal muscle damaged regenerates B
chains in response to injury & elevations
MB CK can be seen

7. ●Severe & continuing muscle injury can
result in substantial incr plasma MB CK
●In absence of overt pathologic process
of skeletal muscle detectable quantities
of MB are not seen in response to
modest skeletal muscle injury
Distribution CK Tissue
Isoenzymes

8. ●Contains ~ 1600 IU/g CK activity
compared with 225-12,000 IU/g in
skeletal muscle
●Contains ~ 15 -30 % MB CK but may
increase in response to injury
●Approximately 85% of CK is depleted in
response to injury with ~ 15% reaching
the circulation
Human Myocardium

9. Rates of Release
●Serum CK activity exceeds normal
range within 4-8 hours following onset
of infarction
●Time to peak varies from 8-58 hours
(mean peak ~ 24 hours)
●Peak levels will occur earlier in patients
with recanalization

10. Relationship Total CK & MB CK
●Released equally from myocardium
during infarction
●MB CK should be ~ 15% of total CK
○noncardiac release of MM CK in patients
with large infarctions thereby resulting in a
lower relative % of MB CK ( 1-21% )
○any elevation of MB CK above baseline
associated with a rising/falling pattern is
compatible with infarction

11. Noncardiac Sources of MB
CK
●skeletal muscle
●uterus
●diaphragm
●gastrointestinal tract
●prostate
●thyroid
●urethra

12. Techniques Measurement of MB
CK
●MM CK is stable for 48 hrs at room
temperature compared with ~ 2 hrs for
MB and BB ( ~ 24 hrs with refrigeration)
○Electrophoretic method
○Column methods
○Immunoassay

13. Immunoassay Procedures
●Antibodies M subunit protein - inhibits
CK-M activity
●CK-B activity proportional MB CK
activity
●Combines enzymatic assay & detects
conversion creatine phosphate + ADP
to ATP
●False positive results:macrokinases,
incr BB CK, muscle adenylate kinase

14. CK-MB Confirmation Test
●monoclonal antibody to MB CK
●performed routinely when MB activity is
elevated on screen
●usually 24 hour turnaround time

15. False Positive MB CK
●skeletal muscle injury or myopathy
○polydermatomyositis
○Duchenne’s muscular dystrophy
●renal failure
○myopathy related to parathyroid hormone &
calcium abnl
○abnormal clearance related to
reticuloendothelial dysfunction

16. False Positive MB CK
●hypothyroidism
○skeletal muscle myopathy
○alterations in clearance
●tumors containing MB or BB CK (rare)

17. Summary
●laboratory artifacts can occur each the
method used to measure MB CK
●MB CK levels can be elevated with
normal total CK level
●elevation of MB CK with a rising/ falling
pattern
●MB CK > 7.5 ng/ml is highly suggestive
of MI with the confirmation test

18. MB CK Subforms
●Quantifies 2 subforms of CK MB~ 6min
●MB2 is present in myocardial tissue is
converted to MB1 on release into blood
●diagnostic criteria for MI are CK-MB2
level > 1.0 U/L & ratio MB2/MB1 >1.5
within 6 hours
●ratio of MB2/MB1 higher reperfused
infarction ( > 3.8 within 2 hrs)

19. Use of Subforms of MB CK to Dx AMI
●NEJM, Sept 94- Puleo et al evaluated
1110 patients who came to ER with
chest pain
●121 patients had myocardial infarction
based on MB CK level > 14 U/L
●92 MI patients presented within 6 hours
and had complete CK testing

20. ●Subform assay positive in 88/92
patients within 6 hr interval: sensitivity
95.7% ; specificity 93.9%
●Subform activity elevated at 1.22 hrs
●Sensitivity for the conventional MB
assay was only 48.2%
Use of Subforms of MB CK to Dx AMI

21. Summary
●Potential earlier diagnosis of AMI in ER
with potential change in triage policies
●Negative subform assay is not reliable
unless sample obtained 6 hours after
onset of symptoms
●Assay would be falsely positive in
patients with skeletal muscle injury

22. Troponin Complex
●Troponin complex located on thin
filament of contractile apparatus has 3
subunits
○troponin T (tropomyosin-binding unit)
○troponin I (actomyosin-ATP inhibiting
subunit)
○troponin C (calcium-binding subunit)

23. Cardiac Troponin T
●Unique myocardial antigen released
into circulation during ischemia
●Early rise in TnT 2ndary cytoplasm
leakage with second peak reflecting
damage to subcellular apparatus
●Immunologically specific assay utilizing
monoclonal anti-human cardiac TnT
antibody

24. ●Cardiac troponins not found in serum of
healthy people
●Appears in circulation within 3.5 hours
with a plateau from the 2nd to the 5th
day
●100% sensitivity for detecting MI from
10 hours - 5 days (levels may be
increased up to 14 days)
Cardiac Troponin T

25. Reperfusion & Troponin T
●Pxts without thrombolytic therapy or
spontaneous early reperfusion there is
no distinct peak on day 1
●peak in TnT levels is found at a median
time 14 hours with early reperfusion (<
3.5 hours)

26. Prognostic TnT Unstable
Angina
●NEJM,July 1992- Hamm et al screened
25 pts: accelerated angina & 84 pts with
acute angina @ rest: CK,CK MB & TnT
q8 hrs
●25 patients had Class 1 or 2 angina
none had measurable troponin T
●33/84 patients with rest angina had
measurable troponin T (median,
0.50ug/L)

27. NEJM, July 1992
●10/33 patients with positive troponin T
had acute myocardial infarctions within
2 - 10 days of hospitalization
●CK MB activity elevated in only one of
patients
●Incidence MI & death significantly
higher in group with positive troponin T

28. Clinical Use Guidelines
●Late diagnosis of myocardial infarction
●Assessment of patients myocardial
ischemia & skeletal muscle injury
●Diagnosis of perioperative MI in CABG
○TnT levels do not exceed 2.5ug/l in
uncomplicated surgery
●Blunt chest/heart trauma
●Myocarditis

29. Measurement Cardiac Troponin
I
●NEJM March 1994, Adams et al, Tn-I a
sensitive & specific marker
perioperative infarction vascular surgery
patients
○100 patients without perioperative infarction
detected by echo, 19 patients had + MB CK
○1/ 100 had slight elevation troponin I

30. Lactate Dehydrogenase
Enzymes
●Ubiquitous in many tissues & catalyzes
reduction of pyruvate to lactate
●Most tissues contain all 5 isoenzymes
●LD 1 - heart/ LD 5 - skeletal muscle/liver
●LD rises @ 10 hours & peaks 24-48 hours
●LD 1/ LD 2 ratio of 1 suggestive acute
infarction
●False positives skeletal muscle injury

31. Myoglobin
●Heme protein functions as O2 reservoir
●Found in skeletal & cardiac muscle
●Peaks in 10 hrs is rapidly cleared by
kidneys within 24 hours
●Comparable to CK MB subforms in
detecting recanalization

32. Summary
●Single set ER cardiac enzymes not
sensitive enough to exclude MI
●CK MB subforms < 6 hours incr sensitivity
●False positive MB - skeletal muscle injury
●MI > 24 hours total LDH if elevated
isoenzymes
●Cardiac troponin T late infarction or
skeletal muscle injury