Cardiac Biomarkers by sandip

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CARDIAC BIOMARKERSBY

Mr.Sandip Kanazariya

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BIOMARKER“A biomarker is a substance used

as an indicator of a biologic state”.

Morrow and de lomos three criteria for biomarkers– Accurate repeated measurements at

reasonable cost– Must provide additional information– Should aid treatment

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HEART FAILURE Heart failure a major and growing health problem appears to result not only from cardiac overload or injury but also from complex interplay among genetic, inflammatory and biological changes acting on cardiac myocytes, the cardiac interstitium or both.

ACUTE MYOCARDIAL INFARCTIONA sudden occlusion of a coronary artey by thrombus or by embolisation causes an acute myocardial infarction.

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CARDIAC BIOMARKERS• Cardiac biomarkers are protein molecules released

into the blood stream from damaged heart muscle

• Since ECG…… inconclusive ….biomarkers !!!!!?????myocardial injury

• These biomarkers have a characteristic rise and fall pattern

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CHARACTERISTICS OF AN IDEAL CARDIAC MARKER

• High cardiac specificity • Pharmacokinetics of cardiac biomarker• Easy diagnosis• Marker should play a designed role in the

treatment and management of clinical subject

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HISTORY OF CARDIAC BIOMARKERS

• 1954 - SGOT (AST)• 1955 - LDH • 1960 - CPK • 1972 - CPK isoforms by Electrophoresis • 1975 - CK - MB by immunoinhibition • 1975 - Myoglobin • 1985 - CK - MB Mass immunoassay • 1989 - Troponin T • 1992 - Troponin I

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CLASSIFICATION OF CARDIAC BIOMARKERS

• Biomarkers of myocardial injury– markers of myocardial necrosis– markers of myocardial ischemia

• Biomarkers of haemodynamic stress

• Inflammatory and prognostic Biomarkers

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BIOMARKERS OF MYOCARDIAL INJURY

• Markers of myocardial necrosis– Creatine kinase – MB– Myoglobin – Cardiac troponins

• Markers of myocardial ischemiaIschemia Modified Albumin (IMA)

Heart-type fatty acid binding protein (H-FABP)

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Zones of Ischemia Injury and Infarction with Transmural and Subendocardial Infarction

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CREATINE KINASECreatine kinase (CK/CPK) is an enzyme expressed in

a number of tissues.

Function: it catalyses the conversion of creatine to phosphocreatine degrading ATP to ADP

The CK enzyme consists of two subunits, B (brain type) or M (muscle type), Making three different isoenzymes: CK-MM, CK-BB and CK-MB

• CK-BB occurs mainly in tissues, rarely of any significance in the bloodstream

• Skeletal muscle expresses CK-MM (98%) and low levels of CK-MB (1%)

• The myocardium has CK-MM at 70% and CK-MB at ~30%

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CK-MB• High specificity for cardiac tissue

• Begins to rise 4-6 hours after onset of infarction

• Peaks at about 12 hours

• Returns to baseline at 24-36 hours

• Can be used to indicate early re-infarction if level normalizes and then increases again

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DETECTION • CK-MB now measured via a highly sensitive monoclonal antibody

assay

• Immunological Sandwich technique using two Abs for different epitopes of CK –MB molecule

• The first Ab is rendered immobile on a matrix (e.g. CrO2 particles)

• The second Ab conjugate to an enzyme (β- galacto- sidase)

• Separated bound sandwiches are reacted with their substrate (e.g. Chlorophenol β- Red Galactopyranoside)

• Liberated end product chlorophenol is measured spectrophotometrically and is proportionate to CK-MB amount (not the activity)

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DETECTION • The CK-MB isoforms may also be analyzed using high-

voltage electrophoresis

• The ratio of MB2/MB1 is calculated

• MB2 released from heart muscle and converted to MB1

• A level of MB2 > or = 1 and a ratio of MB2/MB1 > 1.5 indicates myocardial injury

• A result is positive if MB2 is elevated and the ratio is more than 1.5

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DRAWBACKS • False positive (for MI) CK-MB elevation can be seen in:

– Significant skeletal muscle injury– The MB fraction is determined to be expressed during

the process of muscle regeneration– Cardiac injury for reason other than MI

• Defibrillation • Blunt chest trauma• Cocaine abuse

The search for cardiac specificity continues…

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MYOGLOBIN• Small-size heme protein found in all tissues mainly assists in oxygen

transport

• It is released from all damaged tissues

• Increases often occur more rapidly than TI and CK

• Released from damaged tissue within 1 hour

• Normal value: 17.4-105.7 ng/ml

• Timing:– Earliest Rise: 1-3 hrs– Peak 6-9 hrs– Return to normal: 12 hrs

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CONDITIONS FOR MYOGLOBIN INCREASE

• Acute myocardial infarction

• Skeletal muscle damage, muscular dystrophy, inflammatory myopathies

• Renal failure, severe uremia

• Shock and trauma

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CLINICAL USEFULNESS OF MYOGLOBIN

• Rapid monitor of success of thrombolytic therapy

• Negative predictor of MI

DRAWBACKS

• Due to poor specificity, myoglobin levels do not always predict myocardial injury

• Not utilized often for AMI/cardiac damage

assessment because of its very rapid metabolism

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CARDIAC TROPONINS• Troponin is a complex of three regulatory proteins that

is integral to non-smooth muscle contraction in skeletal as well as cardiac muscle

• Troponin is attached to the tropomyosin sitting in the groove between actin filaments in muscle tissue

• Troponin has three subunits, TnC, TnT, and TnI– Troponin-C has calcium binding ability and has no

diagnostic value– Troponin-T binds the troponin tropomyosin complex, – Troponin-I is an inhibitory protein

Cardiac Troponin Release after MI

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TROPONIN I1. Cardiac Troponin I (cTnl) is a cardiac muscle protein

with a molecular weight of 24 kilo-Daltons.

2. The cTnl has a additional amino acid residues on its N-terminal that are not exist on the skeletal form.

3. The half life = 2~4 hours.

4. Serum increase = 2-8 hours

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TROPONIN T 1. Cardiac Troponin T (cTnT) is present in fetal skeletal

muscle.

2. In healthy adult skeletal muscle cTnT is absent.

3. The gene of cTnT may be re-expressed in skeletal muscle

disease. (Clin Chem. 1999;45:2129-2135)

4. Biological half life and early serum increases of cTnT are

similar to that of cTnI.

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TROPONIN LEVELS

• Less than 5% in cytosol

• Troponin levels begin to rise 2-3 hours after onset of myocardial injury

• Elevations in Troponin-I and Troponin-T can persist for up to 10 days after MI

• Remember, CK-MB returns to baseline by 48 hours

• Thus far, studies have failed to find a source of Troponin-I outside the heart, but have found some Troponin-T in skeletal muscle

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COMPARISON

NEJM 2002;Vol.346,No.26:2079-82

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COMPARISON

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Conditions commonly associated with cardiac troponin elevations

• Arrhythmias• Congestive heart failure• Coronary artery disease• Coronary vasospasm• Critically ill patient• Hypertension• Myocarditis• Pericarditis• Pulmonary embolism• Pulmonary hypertension, severe• Renal failure• Sepsis/septic shock• Sepsis-related myocardial dysfunction• Systemic inflammatory diseases• Trauma

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Increased Troponins

• Troponin T and I are not detected in healthy individuals

• Significant increase in Troponins reflects myocardial necrosis

• ACC/ESC has defined increase in Troponins as a measurement above 99th percentile value of reference group

• To reduce false-positive outcomes, CV of 10% at decision limit is recommended

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TROPONIN ASSAYS

• TropT (Roche Diagnostics, Germany) • Trop I (Siemens Healthcare Diagnostics)

• Troponin T – 99th percentile limits - 0.01 ng/mL – assay ranges - 0.01-25 ng/mL

• (Troponin I) – 99th percentile limits -0.04 ng/mL – assay range -0.04-40 ng/mL

• Reference limits based on the 99th percentile for a healthy population are 0.01 ng/mL (Troponin T) and 0.04 ng/mL (Troponin I)

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DIAGNOSIS BASED ON SENSITIVE TROPONIN I ASSAY

They used the concentration of 0.04 ng per mL as the upper reference limit and established the diagnosis of myocardial infarction if one value of more than 0.04 ng per mL was documented, combined with a rise or fall in the value of 30% or more within 6 hours after admission.

Patients with troponin rises benefit more from Glycoprotein IIb IIIa inhibitors such as,

• Abciximab• Eptifibatin• Clopidogrel

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IN VETERINARY USE• Most commonly used in dogs and

catsClinical conditions• Congestive heart failure• Percardial disease• Doxorubicin toxicity• Gastric dilatation and volvulus etc,.

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Fonflora et al study

120 animals were examined in emergency with cardiac troponin assays,

• First group= ctni less than .15ng/ml• Second group= ctni .15-1 ng/ml• Third group= ctni more than 1 ng/ml

Prognosis grave in second and third group

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BIOMARKERS OF MYOCARDIAL ISCHEMIA

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Ischemia Modified Albumin (IMA)

• A novel marker of ischemia, is produced when circulating serum albumin contacts ischemic heart tissues

• IMA can be measured by the albumin cobalt binding (ACB) assay that is based on IMA's inability to bind to cobalt

• Mechanism- due to structural change in the amino terminal end of albumin

• IMA levels rise within 6 hours

• remain elevated for 12 hours

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Drawbacks IMA levels raised in non- cardiac ischemia

Modification to n- terminal end may also be induced by extracellular hypoxia, acidosis etc,

Conclusion FDA in 2010 has approved a multimarker

approach for using the combination of ECG, the cTnI, and the IMA levels achieving a sensitivity of 95% for ACS

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Heart-type fatty acid binding protein (H-FABP)

• H-FABP is a very stable abundant [138] low molecularweight protein (14–15 kDa) in the cytoplasm of myocardial cells

• Appearing as early as 90 min after symptom onset and peaking within 6 h

• Parameters of kinetic release make it an ideal candidate both for early assessment or exclusion of AMI and for the measurement of a recurrent infarction

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• A study by Puls et al– the negative predictive value (NPV) of H-FABP was

an impressive 100%– its Positive predictive value was 41% which was

greater than that of both cTnT (29%) and NT-proBNP (19%).

• The myoglobin/heart FABP ratio has been used to differentiate between heart muscle and skeletal muscle injury

• Cardiodetect

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BIOMARKERS OF HAEMODYNAMIC STRESS

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NATRIURETIC PEPTIDES

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• The natriuretic peptides (NP) are a group of structurally similar but genetically distinct peptides.

• NPs are identified as regulatory diuretic-natriuretic substances responsible for salt and water homeostasis

• Lowers blood pressure.

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• The NP family includes ANP : -atrial natriuretic peptide (28 a.a.) N-terminal proANP (98 a.a.)

BNP : brain natriuretic peptide (32 a.a.) N-terminal proBNP (76 a.a.)

CNP : C-type natriuretic peptide (22 and 53 a.a.)

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Fig. Schematic representation of the ANP and BNP precursors with sequence numbering defining low-molecular-mass forms, N-terminal forms and high-molecular-mass precursors

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• ANP is released primarily in response to atrial wall stretching and intravascular volume expansion.

• BNP is mainly secreted by the ventricles

• CNP is found predominantly in the brain and also synthesized by vascular endothelial cells

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BRAIN NATRIURETIC PEPTIDE (BNP)

• originally isolated from porcine brain

• Subsequently also isolated from human heart

• Circulating levels of BNP are raised in patients with cardiovascular or renal disease

• More important than ANP in heart failure

• Greatest proportion of circulating BNP is thought to come from the ventricles (left)

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• Synthesis

Synthesis in myocytes

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Natriuretic peptide receptorThree receptors for natriuretic peptides :

Natriuretic peptide receptor -A Natriuretic peptide receptor -B Natriuretic peptide receptor -C

NPR-A and NPR-B • NPR-A and NPR-B are particulate guanylyl cyclases that catalyses

the conversion of GTP to c-GMP

• NPR-C They lack the guanyl cyclase domain and may influence the target cell function through inhibitory guanine nucleotide (Gi) protein, and they likely also act as clearance receptors for circulating peptides.

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RECEPTORS AND METABOLISM

NPR-A and NPR-B• NPR- A is the most abundant type in

large blood vessels• NPR-B predominate in the brain• Both receptors are present in the

adrenal glands and the kidney• Affinity for NPR-A : ANP > BNP > CNP for NPR-B : CNP > BNP > ANP

52Action of Atrial Natriuretic Peptide at Target Cells

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FUNCTIONS• ANP and BNP concentrations increase in

response to volume expansion and pressure overload of the heart

• ANP and BNP have been shown to be physiological antagonists of the effects of

(1) angiotensin II on vascular tone (2) aldosterone secretion (3) renal-tubule sodium reabsorption (4) vascular-cell growth

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Natriuretic Peptides

55Figure . Physiology of the natriuretic-peptide family

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• Clearance factors for natriuretic peptides NPR-C and neutral endopeptidase

Endopeptidase

• Neutral endopeptidase inactivates all three natriuretic peptides

• Present within renal tubular cells and vascular cells

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Conditions or factors commonly associated with B-typenatriuretic peptide or N-terminal-pro-B-type natriuretic

peptideelevations• Age• Arrhythmias• Cardiomyopathy: hypertrophic, ischemic, or dilated• Congestive heart failure• Coronary artery disease• Gender• Hypertension• Left ventricular diastolic dysfunction• Pulmonary embolism• Renal failure• Right heart failure• Right ventricular overloading: fluid, or pressure overloading• Sepsis or septic shock• Sepsis-related myocardial dysfunction

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Breathing Not Properly (BNP) study

• Prospective study of 1586 patients presenting to the emergency department with acute dyspnea

Outcomes of the study

• The predictive value of BNP much superior to previous standards including radiographic, clinical exam, or Framingham Criteria

• Bnp cut point was fixed at 100pg/ml and it showed 90% sensitivity and 80% specificity for diagnosing heart failure

Veterinary field

• Chronic left ventricular systolic and diastolic dysfunction

• Cardiac vs non- cardiac dyspnoea

• Recent development of ELISA kits for canine and feline NTproBNPby Guildhay Ltd (www. guildhay. Co.uk)

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Therapeutic potentialANP and BNP infusion• Decrease renin and aldosterone

concentration• Increase urinary sodium and water

excretion Neutralendopeptidase inhibitor• Nesiritide is a new drug that is a

synthetic BNP that vasodilates vessels and serves as a potent diuretic agent

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BNP for Rx of decompensated heart failure Nesiritide (h-BNP)

32 amino acid sequenceRecombinant technology using E-coli

DR I

MKRG

S SS

SGLG

FC CS SG

SGQVMK V L R

RH

KPS

NOTE: hBNP affects assay for BNP, but can still use proBNP or one of the proANP assays

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BIOMARKERS OF INFLAMMATION

PROGNOSTIC MARKERS AND MARKERS OF RISK

STRATIFICATION

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• C-reactive protein

• Myeloperoxidase

• Homocysteine

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C-REACTIVE PROTEIN

• CRP is an acute-phase protein produced by the liver

• Pentameric structure consisting of five 23-kDa identical subunits

• Plasma levels can increase rapidly to 10000x levels

• High-sensitivity CRP (hs-CRP) assays

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• CRP previously known to be a marker of high risk in cardiovascular disease

• More recent data may implicate CRP as an actual mediator of atherogenesis

Mechanism of CRP-mediated atherogenesis:• Once ligand-bound, CRP can:

– Activate the classical compliment pathway– Stimulate phagocytosis– Bind to immunoglobulin receptors– Endothelial dysfunction via ↑ NO synthesis– ↑LDL deposition in plaque by CRP-stimulated macrophages

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Clinical Uses – Screening for cardiovascular risk in

otherwise “healthy” individuals– Predictive value of CRP levels for

disease severity in pre-existing Coronary artery disease

Elevated levels predictive of• Long-term risk of first MI• Ischemic stroke

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Limitations of CRP• Low specificity• No evidence that lowering CRP levels decreases CV risk

Industry and FDA staff guidelines 2005 had given clinical cut off value as less than 1 mg/l as safe levels with hs-CRP tests

CRP Risk for CVD Less than 1.0 mg/L Low 1.0-2.9 mg/L Intermediate Greater than 3.0 mg/L High

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MYELOPEROXIDASE• MPO is an enzyme that aids white blood

cells in destroying bacteria and viral particles

• MPO catalyzes the conversion of hydrogen peroxide and chloride ions (Cl-) into hypochlorous acid

• MPO is released in response to infection and inflammation

• EPIC Norfolk StudySugiyama Am J Pathology 2001

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• MPO leads to oxidized LDL cholesterol – Oxidized LDL is phagocytosed by

macrophages producing foam cells• MPO leads to the consumption of nitric oxide

– Vasoconstriction and endothelial dysfunction

• MPO can cause endothelial denuding and superficial platelet aggregation

• MPO indicates activated immune cells– Activated immune cells and inflammation

lead to unstable plaque• Inflammatory plaque is inherently less stable

– Thin fibrous cap/fissured/denuded

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• In august 2005, FDA approved the first MPO assay, Cardio MPO tm developed by Prognostix,inc.

• The test is a sandwich enzyme immuno assay

• Normal plasma MPO levels are 51-633pmol/ml

Progression of Biomarkers in ACS

ACS, acute coronary syndrome; UA, unstable angina; NSTEMI, non–ST-segment elevation myocardial infarction; STEMI, ST-segment elevation myocardial infarction

Adapted from: Apple Clinical Chemistry March 2005

STEMIUA/NSTEMISTABLE CAD PLAQUE RUPTURE

MPOCRPIL-6

MPO ICAMsCD40LPAPP-A

MPOD-dimerIMAFABP

TnITnTMyoglobinCKMB

Inflammation has been linked to the development of vulnerable plaque and to plaque rupture

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HOMOCYSTEINE• Intermediary amino acid formed by the

conversion of methionine to cysteine

• Moderate hyperhomocysteinemia occurs in 5-7% of the population

• Recognized as an independent risk factor for the development of atherosclerotic vascular disease and venous thrombosis

• Can result from genetic defects, drugs, vitamin deficiencies

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• 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

• Normal levels less than 6micro mol/l

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• Elevated levels appear to be an independent risk factor, though less important than the classic CV risk factors

• Treatment includes supplementation with folate, B6 and B12

79Stefan Blankenberg, MD; Renate Schnabel, MD; Edith Lubos, MD, et al., Myeloperoxidase Early Indicator of Acute Coronary Syndrome and Predictor of Future Cardiovascular Events 2005

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BIOMARKERS OF INTEREST,ALTHOUGH THEY ARE NOT

CURRENTLY USED ROUTINELY1. Serum Amyloid Protein A2. sCD40 Ligand3. Cytokines4. Lipoprotein-Associated Phospholipase A25. Pregnancy-Associated Plasma Protein A6. Oxidized LDL7. Placental Growth Factor8. Matrix Metalloproteinases9. Monocyte Chemotactic Protein10. Tissue Plasminogen Activator Antigen (t-PA) and Plasminogen Activator

Inhibitor 1 (PAI-1)11. Secreted Platelet Granular Substances12. Isoprostanes13. Urinary Thromboxane14. Adhesion Molecules

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Serum Amyloid Protein A

• Serum amyloid protein A, an acute-phase protein and an apolipoprotein, has been used with hsCRP in cross-sectional studies.

• It can be synergistic with hsCRP326 but is much less commonly used.

• At present, no standardized assays, reference interval studies, nor consistent assay validations are available.

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sCD40 Ligand

• CD40 ligand is a transmembrane protein related to TNF.

• It has multiple prothrombotic and proatherogenic effects.

• What is usually measured is the soluble form of the receptor, sCD40 ligand, which has been shown to be a predictor of events after acute presentation.

• At present, standardized assays, reference interval studies, nor consistent assay validations are not available.

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Cytokines• A variety of stimulatory and inhibitory interleukins (TNF, IL-1,

IL-6, IL-8, IL-12, IL-18) are thought to help mediate the elaboration of CRP and the development of atherosclerosis and acute events.121 These cytokines may stimulate or inhibit leukocytes, often through T cell–mediated processes and effects on monocytes, which are indigenous to atherogenesis.

• In some studies, IL-6 is more prognostic than hsCRP.• These cytokines often have inhibitors and/or binding proteins

that modulate their effects. At present, standardized assays, reference intervals studies, and consistent assay validations are not available.

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Lipoprotein-Associated Phospholipase A2

• Phospholipase A2 (Lp-PLA2) is a phospholipase associated with LDL that is thought to be an inflammatory marker.

• It was previously known as platelet-activating factor (PAF) acetyl hydrolase.

• It is synthesized by monocytes and lymphocytes and is thought to cleave oxidized lipids to produce lipid fragments that are more atherogenic and that increase endothelial adhesion.

• An FDA-approved assay for this analyte includes obligatory reference intervals.

• It has been shown to be predictive of events in a primary prevention cohort, even when hsCRP is present in the model, suggesting that it measures something different from what is measured by the acute-phase reactants associated with hsCRP.

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Pregnancy-Associated Plasma Protein A

• Pregnancy-associated plasma protein A (PAPP-A) is a metalloproteinase that is thought to be expressed in plaques that may be prone to rupture.

• The literature in this regard is mixed at present concerning its use.

• At present, standardize assays, reference interval studies, and consistent assay validations are not available.

• Recent data suggest that heparin administration in MI patients is associated with increased PAPP-A concentrations; this may limit its prognostic role.

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Oxidized LDL• Oxidized LDL has been attributed a key role in the

development of atherosclerosis.• Several methods have been used to measure it, but they

yield potentially different data.• Some have correlated malondialdehyde LDL with the

development of atherosclerosis and short-term events.• Direct identification with antibodies suggests that

oxidized LDL may be released from vessels and may colocalize with lipoprotein a [Lp(a)] after acute events.

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Placental Growth Factor• Placental growth factor is an angiogenic factor related

to vascular endothelial growth factor (VEGF), which stimulates smooth muscle cells and macrophages.

• It also increases TNF and MCP-1. A novel assay for this analyte is thought to provide additional prognostic information on patients who present with ACS.

• At present, standardized assays, reference interval studies, and consistent assay validations are not available.

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Matrix Metalloproteinases• Matrix metalloproteinases (MMPs) can degrade the collagen matrix in

coronary artery or myocardium.• They are integral to remodeling of the coronary artery and/or the heart

after acute events.• Elaboration of MMP-9, a gelatinase, is thought to be important in plaque

destabilization; thus some have tried to measure it as a prognostic index.• Other MMPs participate in the elaboration of extracellular matrix in the

heart.• Many MMPs also have inhibitors [tissue inhibitors of metalloproteinase

(TIMPs)] that modulate their effects.• At present, standardized assays, reference intervals studies, and• consistent assay validations are not available.

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Monocyte Chemotactic Protein

• Monocyte chemotactic protein (MCP-1) is a chemokine that is thought to be responsible for the recruitment of monocytes into atherosclerotic plaque.

• It has been reported to be elevated in patients with ACS and to have long-term predictive value.

• However, at present, standardized assays, reference interval studies, and consistent assay validations are not available

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Tissue Plasminogen Activator Antigen (t-PA) andPlasminogen Activator Inhibitor 1 (PAI-1)

• t-PA is the body’s physiologic fibrinolytic activator.• PAI-1, its endogenous inhibitor, binds to t-PA.• Inhibition of fibrinolysis has been suggested to be a reason

for recurrent infarction; the fact that maximal inhibition usually occurs in the early morning hours provides a potential explanation for the circadian variability of AMI.

• It may also be the reason why persons with diabetes have such unstable disease; the growth factor properties of insulin stimulate increases in PAI-1.

• An accurate assessment of this system includes both t-PA and PAI-1, along with some assessment of bound versus free levels.

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Secreted Platelet Granular Substances

• Both platelet factor 4 (PF4) and beta thromboglobulin (BTG) are secreted when platelets aggregate.

• PF4 has a short half-life and is released by heparin.• BTG is not released by heparin and has a longer half-life.• Both markers have been used to assess platelet

aggregation.• BTG is by far the more reliable.• At present, standardized assays, reference interval

studies, and consistent assay validations are not available.

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Isoprostanes• Isoprostanes are the end breakdown products of lipid

peroxidation, and urinary levels have been used to assess the level of oxidative stress.

• It is thought that oxidation of LDL is essential for the development of atherosclerosis, and that HDL and other antioxidants work by antagonizing this oxidative stress.

• Urinary isoprostanes give some assessment of this critical process.

• The most commonly measured are F2-isoprostanes, but a large number of others are available for measurement.

• It does appear that they will eventually be helpful in assessing oxidative stress.

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Urinary Thromboxane

• Urinary thromboxane is the end metabolite of thromboxane A2, which is a measure of platelet aggregation.

• Urinary levels are elevated in patients with unstable coronary disease, in keeping with the known participation of platelets in the pathogenesis of CAD. This level is difficult to ascertain, and collecting urine in the acute situation is at times problematic.

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Adhesion Molecules• Adhesion molecules are a wide variety of molecules that

can potentially be measured as a way of assessing the adherence of leukocytes and/or platelets or other adhesive proteins to the endothelial matrix. Some are receptors.

Examples• include PECAM-1 (platelet-endothelial adhesion molecule

1), P-selectin, e-selectin, and VCAM-1 (vascular cell adhesion molecule 1).

• At times, the receptor itself is measured, but often it is a soluble portion that circulates that is measured.

• At present, standardized assays, reference interval studies, and consistent assay validations are not available

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Choline

• Choline is released after stimulation by phospholipase D and has been touted as a test of prognosis in patients with chest discomfort.


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Unbound Free Fatty Acid

• Unbound free fatty acid (uFFA)39 has also been touted as a marker of ischemia.

• Most fatty acid is bound, and ischemia is thought to increase the small unbound fraction.

• Initial studies have reported mixed results.• At present, standardized assays, reference

interval studies, and consistent assay validations are not available.

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Nourin

• Nourin I is a small protein released rapidly by “stressed myocytes.”

• It induces changes in a variety of inflammatory cytokines and attracts neutrophils.

• Preliminary studies have been done to attempt to validate its use.


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Copeptin• Copeptin, a 30 amino acid glycoprotein constituting the C-

terminal portion of arginine vasopressin, has been shown to be a prognostic biomarker in hemorrhagic and septic sepsis.

• More recently, data have shown that measurement of copeptin serves as a rapid and early rule-out biomarker for AMI at presentation in patients with symptoms suggestive of ACS with a normal cTn value.

• An assay measuring copeptin (CT-proAVP) has been described using the Brahms Kryptor Immunology Analyzer, Diamond Diagnostics Holliston, MA.

• Additional clinical and analytical validation studies will be necessary, especially head-to-head comparisons of copeptin versus the new hs-cTn assays.

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