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Ella Melissa L.

Dr. dr. Tinny E. H., SpPK(K)

 Heart disease is an affliction intimately tied to high technology
 Technology has had a causal role, in part by allowing people to
live longer, and in part by enabling a sedentary and overly
consumptive lifestyle.
 Diagnosis & treatment of heart disease also depend heavily on
advanced technology, including electrophysiologic, imaging,
catheterization, surgical and clinical laboratory modalities.
1. Measurement of proteins present in cardiac myocytes
indicates recent damage to cardiac muscle  used mainly for
the diagnosis and management of ischemic events (ACS).
 Although many different markers of ischemic damage have
been used in the past, at the time of writing the most
important markers by far are cardiac troponin I and cardiac
troponin T.
2. Measurement of substances that are damaging to the coronary
arteries, or at least have proven association with coronary heart
disease, is used to assess risk and select appropriate preventive

 The most important laboratory risk factors are lipids

 Of the many other substances that could be: homocysteine

(Hcy) and C-reactive protein (CRP)

2. Measurement of natriuretic peptides released from myocardium,

particularly B-type natriuretic peptide (BNP) and the related
inactive fragment, NT-pro-BNP, reflects the presence and
severity of heart failure
4. Laboratory testing has been less applicable to cerebrovascular
disease, but some new biomarkers may be considered
 Measurement of d-dimer is useful in ruling out pulmonary
5. Other potential biomarkers:
Choline, sCD40L, Ischemia Modified Albumin, Myeloperoxidase,
Oxidized LDL, LpPLA2, PAPP-A, PGF
 Biochemical markers of myocardial damage were essentially a
serendipitous discovery in the early 1950s, when LaDue and
coworkers were investigating the transaminase enzymes 
 The era of “cardiac enzymes”
 Transaminases have not endured as cardiac markers because of
their abundance in liver, skeletal muscle, and other tissues
 They were soon superseded for cardiac diagnosis by two other
enzymes, lactate dehydrogenase (LD) and creatine kinase (CK)
 LD is a zinc-containing enzyme that is part of the glycolytic
pathway and is found in virtually all cells in the body

 CK transfers high-energy phosphate between creatine and ADP,

mainly in muscle cells, but it is found in all types of muscle and in
brain and other tissues

 With both of these enzymes, improved cardiac specificity was

achieved through separation of isoenzymes

 As the subunit names imply, LD1 is relatively abundant in cardiac

muscle, whereas LD5 is more abundant in skeletal muscle.

 Patients with MI exhibit a characteristic pattern of “flipped” LD,

where the normal finding LD2 > LD1 is reversed
 CK is predominantly found as a dimer of catalytic subunits,
each with a molecular weight of about 40 kDa
 The two subunits are termed M (for muscle) and B (for brain)
 The three resulting isoenzymes are CK1 (BB), CK2 (MB) and
CK3 (MM)
 In the normal heart, an average of 15%–20% of the CK is CK-
MB; its distribution is not uniform, with CK-MB percentage
greater in the right heart than in the left heart (Marmor, 1980)
 A single study, however, suggests that CK-MB is not found in
normal myocardium, but appears only when the muscle becomes

 CK-BB is the dominant isoenzyme of CK found in brain, intestine,

and smooth muscle

 At the present time, there is no reference method for CK

isoenzyme analysis. The techniques most commonly used are
electrophoresis and various immunological methods.

 An L→P reference method, optimized for LD1, was developed by

the International Federation of Clinical Chemistry and Laboratory
Medicine (IFCC)
 Current routine methods for quantitation of total LD activity:
kinetic spectrophotometry to measure the interconversion of the
coenzyme NAD+ and NADH at 340 nm
 In the past, clinical laboratories would commonly analyze both
CK and LD isoenzymes to improve overall diagnostic
performance, especially because the two enzymes exhibit
different kinetics
 3 troponin subunits from complex that regulated the interaction
of actin & myosin and thus regulates cardiac contraction
 3 troponins:
◦ Troponin C : Ca2+ binding component
◦ Troponin I : the inhibitory component
◦ Troponin T : the tropomyosin-binding component
 Within the cardiac myocytes, cTnT and cTnI are bound
predominantly to muscle fibers

 this bound form is released slowly over the course of 1 to 2

weeks following myocardial infarction

 Thus, although cTnI and cTnT are relatively small proteins that
are rapidly cleared, their plasma levels fall slowly after cardiac

 A small fraction of cTn in the myocardial cell is free within the

cytoplasm; this averages 6% for cTnT and slightly lower (2%–5%)
for cTnI

 The free fraction allows early leakage from injured myocardial

cells and detection in a time frame similar to that of CK-MB, with
cTn reaching a peak at about 24 hours after MI
 Because of the slow release of fiber-bound cTn, the rapid
decline in circulating cTn right after its peak is typically
followed by a plateau or even a small secondary increase
 It is important that such an increase not be interpreted as
evidence of reinfarction
 In contrast to other cardiac markers, cTnT and cTnI are nearly
absent from normal serum
 With the best assays in wide use at present, which have
detection limits around 0.01 ng/mL, many healthy individuals
have undetectable levels, so the normal range is not well
 The 99th percentile of the healthy population is around 0.04
ng/mL, depending on the assay
 Method: sandwich imunokromatografi
 Kombinasi unik antara Ab monoklonal dan poliklonal
digunakan untuk identifikasi scr selektif & sensitif Troponin
 Sampel: serum, plasma dan whole blood.
 Antikoagulan: EDTA, sitrat. ‘No heparin’
 Bila sampel WB, harus segera dipakai < 4 jam
1. Kualitatif:
 Positif T C

 Negatif

 Gagal
2. Kuantitatif:
 Menggunakan easy reader.
 Konsentrasi hasil dalam ng/ml.
 Nilai normal:
 < 1 ng/ml (whole blood)
 < 0,8 ng/ml (plasma/ serum)
 Myoglobin is a heme-containing protein that binds oxygen
within cardiac and skeletal muscle; only a single form is common
to both muscle types
 Having a molecular weight of only 18 kDa, myoglobin apparently
leaks from damaged cells more rapidly than other proteins
 Following MI elevated myoglobin may be detectable in plasma
before CK-MB or cTn.
 Typically, myoglobin peaks about 6 hours after MI and
returns to baseline after 24 hours.
 Although myoglobin may offer some advantage in early
detection of myocardial damage, its value is limited by its
lack of specificity.
 Carbonic anhydrase III (CA III) is an enzyme present in skeletal
but not in cardiac muscle, hence it can serve as a sort of
“negative” cardiac marker
 It is released from damaged muscle at a fairly fixed ratio to
 Thus myoglobin is a more specific indicator of myocardial
damage when its ratio to CA III is also elevated
 Glycogen phosphorylase (GP) is a widely distributed enzyme that
catalyzes the first step in glycogenolysis
 A dimer of identical subunits, it has three characterized
isoenzymes: GPLL (liver), GPMM (muscle), and GPBB (brain)
 GPBB is also expressed in myocardium, as well as other tissues,
but it is not found in skeletal muscle
 The potential usefulness of GPBB is that it appears to be
released earlier than other markers, and may in fact be
released under conditions of reversible ischemia that do not
give rise to comparable elevations in other markers
(Rabitzsch, 1995; Krause, 1996)
 However, comparisons with modern cTn assays have been
limited and not particularly encouraging (Lang, 2000)
 Heart fatty acid–binding protein (HFABP) is a LMW (15 kDa)
protein that is a relatively early marker of myocardial damage,
with kinetics similar to that of myoglobin
 It is not cardiac specific and therefore does not seem to offer
advantages over myoglobin
 However, the ratio of myoglobin to HFABP is much lower in heart
than in skeletal muscle and may have diagnostic applicability
 Myosin makes up the thick filament of the muscle contractile
apparatus and is composed of a pair of heavy chains (200
kDa) and one pair each of type I and type II light chains (20–
26 kDa)
 Several of these components have been examined as cardiac
markers (Katus, 1988; Uji, 1991; Ravkilde, 1994; Ravkilde,
 It has not been possible to achieve complete cardiac
specificity with myosin, and it is not clear that it would offer
any important advantages
 Ischemia-modified albumin (IMA) is a unique type of cardiac
marker (FDA approved in 2003) that is not a protein released
from damaged myocytes (Bar-Or, 2000; Bar-Or, 2001;
Christenson, 2001; Bhagavan, 2003).
 Rather, the test detects a variant form of albumin with a reduced
affinity for metal ions near the N-terminus
 The variant is measured by a spectrophotometric determination
of Co++ binding
 The theoretical advantage of this test is that it detects ischemia
before irreversible cell damage occurs
 The change in albumin appears to occur within minutes of
ischemia and lasts for about 6 hours

 The test clearly is not specific for cardiac ischemia, but appears to
have a clinical sensitivity of 80%–90% for ACS at the time of
presentation — greater than that of the ECG (Roy, 2004; Sinha,
 Intermediary amino acid formed by the conversion of
methionine to cysteine
 Moderate hyperhomocysteinemia occurs in 5-7% of the
 Recognized as an independent risk factor for the
development of atherosclerotic vascular disease and venous
 Can result from genetic defects, drugs, vitamin deficiencies,
or smoking
 Homocysteine implicated directly in vascular injury including:
◦ Intimal thickening
◦ Disruption of elastic lamina
◦ Smooth muscle hypertrophy
◦ Platelet aggregation
 Vascular injury induced by leukocyte recruitment, foam cell
formation, and inhibition of NO synthesis
 Elevated levels appear to be an independent risk factor,
though less important than the classic CV risk factors
 Screening recommended in patients with premature CV
disease (or unexplained DVT) and absence of other risk
 Hcy has traditionally been measured by chromatographic
techniques (Vester, 1991; Ubbink, 1999; Frick, 2003; Arndt,
 A method using enzymatic adenosylation followed by
immunoassay has been automated on the Abbott IMx
analyzer (Shipchandler, 1995)
 And purely enzymatic methods suitable for automation have
also been introduced (Tan, 2000; Huijgen, 2004; Roberts,
 Present in two forms, atrial (ANP) and brain (BNP)

 BNP is used clinically because it exists in a much wider range

of concentrations in varying clinical settings, making it easier
to measure
 Both ANP and BNP have diuretic, natriuretic and hypotensive
 Both inhibit the renin-angiotensin system and renal
sympathetic activity
 BNP is released from the cardiac ventricles in response to
volume expansion and wall stress
 Approved by the FDA for diagnosis of cardiac causes of
 Currently measured via a rapid, bedside immunofluorescence
assay taking 10 minutes
 Especially useful in ruling out heart failure as a cause of
dyspnea given its excellent negative predictive value
 BNP test had the following
characteristics for diagnosis of HF:
sensitivity 90%, specificity 76%,
positive predictive value 79%, and
negative predictive value 89%
(Maisel, 2002).
 BNP levels decline when effective
therapy for HF is instituted, and so
the test may be used to monitor
the course of treatment (Cheng,
2001; Faggiano, 2009; Novo,
1. C-Reactive Protein
 Acute phase reactant
 CRP is considered to be marker of the atherosclerotic
process (chronic & acute atherosclerotic processes involve
an inflammatory component)
2. Choline
 Choline released after stimulation by phospholipase D during
ischemia and has been touted as a test of prognosis in ACS
patients with chest discomfort without increases in cTn
 At present, there is no standardized assay & no reference
interval studies or consistent assay validations
3. sCD40 Ligand
 A transmembrane protein related to tissue
necrosis factor (TNF)-alpha.
 It has multiple prothrombotic and
proatherogenic effects.
 Shown to be predictor of events after ACS
4. Myeloperoxidase
 Released when neutrophils
aggregate & thus may indicate an
active inflammatory response in
blood vessels.
 It has been shown to be elevated
chronically when chronic CAD is
 An assay has been cleared by the
FDA for prognostic use in high risk
patients with ACS.
5. Oxidized LDL
 Has been attributed a key role in the
development of atherosclerosis.
 Some have correlated malondialdehyde
LDL with the development of
 Direct identification with Ab suggests that
oxidized LDL may be released from
vessels & co-localize with Lpa after acute
6. Pregnancy-Associated Plasma Protein
 PAPP-A is metalloproteinase that is
thought to be expressed in plaques that
are prone to rupture.
 Present concerning its use in ACS.
7. Lipoprotein-Associated Phospholipase A2
 LpPLA2 is a phospholipase associated with
LDL & is thought to be an inflammatory
 It is synthesized by monocytes &
 It has been shown to improve prediction
of events in patients without prior AMI.
8. Placental Growth Factor
 Placental growth factor is an angiogenic
factor related to vascular endothelial growth
factor (VEGF), which stimulates smooth
muscle cells & macrophages.
 It also increases TNF & monocyte
chemoattractant protein-1.
 Placental growth factor is thought to provide
additional prognostic information for patients
who have ACS.