Importance of lipid profile change in cardiovascular disease

1. LIPID PROFILE AND
INVESTIGATIONS OF
MYOCARDIAL INFARCTION
DR. RHUTA SHAH
M.D.(Pathology)
Consultant Histopathologist & Cytopathologist
@MahaGujarat Hospital
@P.D. Patel Ayurvedic Hospital
Ethical Committe Member@ Dinsha Patel College of Nursing

2. What is Lipid Profile?
Lipid profile is a group of tests that are
often ordered together to determine risk of
coronary artrey disease.

3. Tests included in Lipid profile
Total cholesterol
Triglycerides
Phopholipids
Chylomicrons
Cholesterol:HDL ratio
LDL:HDL ratio
VLDL
HDL,LDL
ApoA,B
Lipoprotein A
S.Homocystine

7. LIPID PROFILE
Adult Values (mg/dl)
TYPE DESIRABLE BORDER LINE HIGH RISK
Chylomicrons <28 28 >28
Cholesterol <200 200-240 >240
Triglycerides <250 250-400 >400
Phospholipids <150 150-400 >400
LDL <130 130-160 >160
HDL >60 35-60 <35
Chol:HDL <4 4-6 >6
LDL:HDL <3 3 >3
ApoA1 100-130 Normal Values
ApoB 80-100

8. Prerequisite in blood collection &
Storage
Individual should be on usual diet
No vigorous exercise within 24hrs
No physical stress within 24hrs
No alcohol intake within 24hrs
12hrs fasting or minimum 9hrs fasting
Sample should be taken in sitting position
Tourniquet should be released in 1min
For routine test serum is preferred
For ultracentrifugation/electrophoresis- Plasma
For lipoproteins- Plasma
For storage – Serum
Preferred anticoagulant- EDTA
Measurement must be made in different samples taken 1week apart

9. Factors affecting concentration of plasma
lipids & lipo proteins
Age: increases with age.
Sex: females have lowered levels than males.
Seasonal variations: slightly higher in winter.
Life style:sedentary life style & increse intake of
saturated fats & cholesterol increse plasma lipid values.
Secondary causes: thyroid, hepatic & kidney diseases.
Meditation: decreses level.
Medication: OCpills, post menopausal estrogen, anti HT
increses level.
Stress: increses level.
During collection & storage.
Trauma, acute bacterial or viral infection & pregnancy.

10. Estimation of plasma lipids
Cholesterol: By enzymatic method
Cholesterol ester + H2O
cholesteryl ester hydrolase
Cholesterol + free fatty acid.

11. cholesterol + o2
cholesterol oxidase
cholest-4-en-3- one +H2O2
H2o2 + phenol + 4- aminoantipyrine
peroxidase
Quinoneimine dye +2H2O
The absorbance of quineiemine dye produced
in reaction measured at 500 nm.

12. Triglycerides: By enzymatic method
Triglyceride lipase glycerol + fatty
acid
Glycerol + ATP glycerokinase glycerophosphate + ADP
Glycerophosphate + NAD glycerophosphate
dehydrogenase
Dihydroxyacetonephosphate + NADH + H+
NADH formation can be measured
spectrophotometrically at 340 nm.

13. Triglyceride blanks
It is used to measure free glycerol in
plasma which can interfere with the
estimation of TG value.
Normal value : <1.5 mg /dl
A blank assay without the addition of
lipase provides measure of pre-existing
glycerol & TG value can be corrected
accordingly.

14. Increased free glycerol:
After extremely vigorous exercise
In patients with uncontrolled diabetes
After chance contamination with glycerol
lubricant used as stoppers in some blood
collection tubes
Recent ingestion of glcerol containing
medications
In hyperglycerolemia which arises secondary to
mutation in the glycerol kinase gene on chr.
XP21.3

15. Phospholipids: by enzymatic method
Phospholipids phospholipaseD choline
Choline choline oxidase Betaine + H2O2
Estimation of phospholipids is of value in :
obstructive jaundice, Tangier disease, abeta or
hypolipoproteinemia, & LCAT deficiency.

16. Estimation of lipoprotein & lipoprotein
cholesterol
HPLC
Ultracentrifugation
Gel filteration
Electrophoresis
Polyanion precipitation methods

17. Electrophoretic method
Most commonly used support medium agarose
gel
Chylomicrons, if present remain at the origin
HDL migrates the fastest
VLDL slowest
LDL intermediate between HDL & VLDL.
- Ch VLDL LDL HDL +

18. Polyanion precipitation methods
Some lipoproteins are precipitated with
polyanions such as heparin sulfate,
dextran sulphate, phosphotungstate, and
others in the presence of divalent cations
such as Ca++,Mg++and Mn++.

19. Methods for measuring HDL-C
values
Currently, homogeneous assays are the
most popular method for measuring HDL-
C .
Assay kit contains 2 reagents
First reagent forms a stable complex with
non-HDL proteins,preventing them from
participating in the reaction.
Second reagent releases HDL-C that is
then measured enzymatically.

20. Methods for LDL-C
measurement
Friedwald calculation
LDL-C=(TC)-(HDL-C)-(PLASMA TG/5)
Concentration expressed in mg/dl
LDL-C concentrations can also be
measured by homogeneous assays when
triglycerides conc. Is high because these
are not interefered by triglycerides.

21. Ultracentrifugation
This take advantage of 2 properties of
lipoproteins:
1. Lipoproteins have lower densities than the
other plasma macro molecules.
2. Each class of lipoprotein has different
densities.
Ultracentrifugation methods are largely
of historical & academic interest &are seldom
used clinically.

23. Standing plasma test
Chylomicrons,if present in appreciable quantities are
detected using standing plasma test.
2ml plasma is placed in a10*75mm test tube and allowd
to stand in refrigerator at 4degree celsius undisturbed
overnight.
Chylomicrons accumulate as a floating “creamy layer”
and can be detected visually.
The presence of chylomicrons in fasting plasma is
considered abnormal
icro

24. Apolipoprotein analysis
Studies have indicated that ApoA-1 & ApoB may be
better determinants of atherosclerotic disease than lipid
or lipoprotein determination.
ApoA-1 is present in HDL while ApoB is present in VLDL
,IDL,LDL.
Low ApoB & high ApoA-1 levels is good for health.
Measured by immunoassay or immunonephalometry.
These tecniques rely upon measurement of the turbidity
caused by the apolipoprotein –antibody complexes.
Limitation-inherent turbidity of lipemic samples , or even
non-lipemic samples due to repeated freezing and
thawing may interfere with the readings.

25. Dry chemistry analyzer
They use reagent impregnated strips to
which 10-30microlit of sample is applied .
Sample diffuse to 3 different reagent
impregnated zones , where it dissolves the
reagent & allow the enzymatic reaction.
At the end of incubation period, light
source illuminates the strip & reflection of
reaction mixture is measured.

27. INVESTIGATIONS OF
MYOCARDIAL INFARCTION
Patients with ischemic heart disease fall
into two categories:
Patients with stable angina secondary to
chronic heart disease.
Patients with acute coronary
syndromes(ACS).
ACS is further divided into two groups:
1.STEMI
2.NSTEMI

29. Pathophysiology of acute
myocardial infarction
STEMI generally occurs when coronary blood
flow decreases abruptly after a thrombotic
occlusion of a coronary artery previously
affected by atherosclerosis.
Slowly developing high grade coronary artery
stenosis do not usually precipitates STEMI
because of the development of a rich collateral
network over time.
Most common artery involved is left anterior
descending branch of left coronary artery(40-
50%)

31. Diagnostic evaluation of Myocardial
infarction
Clinical presentation:
Pain: Deep, visceral,heavy ,squeezing,crushing.
Area involved is central portion of chest and or
epigastrium and on occasion it radiates to arm.
Pain may radiate as high as the occipital area
but not below the umbilicus.
Pain is not relieved by rest which differentiates it
from angina pectoris.

33. Physical findings
Pallor associated with perspiration &
coolness of extremities occurs commonly.
The combination of substernal chest pain
persisting for >30 min & diaphoresis
strongly suggests STEMI.
3rd & 4th heart sounds may be present
,decreased intensity of 1st heart
sound,paradoxical splitting of 2nd heart
sound.

34. LABORATORY FINDINGS
ECG:
During the initial stage of the acute phase
of infarction, total occlusion of an
epicardial artery produces ST-segment
elevation.
Later on Q-wave evolves inthe ECG and
ultimately diagnosed as Q-wave M.I.

36. Serum cardiac markers
Two types:
Enzymes – creatine kinase,lactate
dehrogenase(LDH), aspartate
transaminase(AST).
Protein :troponin , myoglobin,c-reactive
protein, myosin.

37. ENZYMES
With myocardial ischemia cardiac
enzymes are released only if there is cell
death.
Therefore these enzymes are marker of
myocardial infarction & not of ischemia.
Reversible ischemia does not lead to
release of cardiac enzymes.

38. AST&LDH
Lacks specificity and therefore are of
academic interest only not used clinically
now.
Increases in 8-12hrs
Peak-12-36hrs
Falls to normal value in 3-4 days

39. Creatine kinase
Has two dimers:
1.muscle(M)
2.brain(B)
It has 3 isoenzymes:
CK-MM- skeletal muscle
CK-BB- brain & lung
CK-MB-specific to myocardium

40. Diagnostic values
CK-MB >5ng/ml
CK-MB:CK ratio >2.5
CK-MB2:CK-MB1 ratio >1.5
CK-MB rises with in 2-4hrs
Peaks in 24hrs
Returns to normal value in 72hrs

41. PROTEIN MARKERS
Myoglobin :heme containing protein which binds
oxygen with in cardiac & skeletal muscle.
Its small size make it leaks from cell quickly after
cell injury & makes it the earliest marker to rise
after M.I.
Rises with in 2-3hrs
Peak-6-8hrs
Return to normal value in 18-24hrs
Normal value-(85-90ng/ml)
Cleared mainly by renal filtration and excretion

43. An alternative approach
Measure myoglobin:carbonic anhydrase III
ratio
CAIII is a negative marker of myocardial
infarction so if myoglobin increased but not
CAIII it is diagnostic of cardiac injury

44. Troponins
They are replacing CK-MB as the
diagnostic test of choice for M.I. & even
recent guidelines on markers of cardiac
injury had recommended its use as “gold
standard” for diagnosis of M.I.
Normally absent in plasma
For assays of troponin better to use
heparanized plasma.

45. Troponins are the compounds which binds
to tropomyosin & generate excitation-
contraction coupling.
Has 3 subunits , named after their function:
cTn T(tropomyosin binding)
cTn I (inhibitory)
cTn C(calcium binding)-not specific

47. cTn T & cTn I bound predominantly to
muscle fibres.
Released slowly over a period of 1-2 wks
following M.I.
Although both are small molecules &
rapidly cleared ,their plasma values fall
slowly after cardiac injury.
Rises with in 4-6hrs
Peak-20-24hrs
Remain elevated for:
cTnT – 10-14days
cTnI – 7-10 days

48. False positive increase:
Pericarditis
Myocarditis
Pulmonary embolism
Chronic renal failure
sepsis

49. C-reactive protein
Used to predict the risk of myocardial infarction
in patient with angina.
Risk of new infarcts in who recover from
infarction.
S.C-RP >3mg/dl-high risk of CVS disease.
S.C-RP 1-3mg/dl –moderate risk.
S.C-RP <1mg/dl – low risk.
CRP related to cardiac risk has been given the
name “high sensitivity CRP”(hsCRP).

51. Other applications of cardiac
markers
Cardiac markers can serve as non-invasive
indicator of reperfusion,either spontaneously or
as a result of therapy.
Due to “washout phenomenon” a bolus of
myocyte proteins is released in minutes
following reperfusion ,so blood sampled in the
next 60-120 min should exihibit an increase in
marker concentration above a defined threshold.
Myoglobin was found to perform better than CK-
MB or cTn.

52. Cardiac markers are also useful for
detecting reinfarction in patients of
myocardial infarction.
CK-MB is superior to cTn because of its
rapid decline after MI.
cTn is the marker of choice in situations of
MI following surgical
procedures(perioperative MI).

53. New tests of myocardial injury
1.Markers of activation of coagulation
system:
P-selectin(indicative of platelet activation)
Fibrin monomers
Thrombus precursor proteins

54. 2.Glycogen phosphorylase
Present in brain & myocardium
Isoenzyme form BB is considered
Released earlier than other markers &
also increased in reversible ischemia.

55. 3.Heart fatty acid binding
protein(HFABP)
Low molecular weight protein .
Early marker of myocardial damage ,with
kinetics similar to those of myoglobin.

56. 4.Ischemia modified
albumin(IMA)
Postulated to arise from interaction of
albumin with free radicals at sites of tissue
ischemia.
The theoritical advantage of this test is
that it detects ischemia prior to irreversible
damage.
Change occurs with in minutes of ischemia
& lasts for about 6hrs.

57. 5.Homocysteine
Increased level are related to increased risk of
coronary artery disease
Cellular effects that have been documented in
experimental systems include:
Endothelial injury
Alteration of NO metabolism
Platelet activation
Smooth muscle proliferation
Toxicity of Hcy can also be attributed to it’s
enzymatic conversion to Hcy thiolactone which
can modify LDL and enhance its uptake by
macrophages.

58. CARDIAC IMAGING
Two dimensional echocardiography:
Abnormalities of wall motion are present
almost universally.
Doppler echocardiography is useful in the
detection & quantitation of ventricular
septal defect & mitral regurgitation, two
serious complication of STEMI.

59. Radionucleotide imaging
techniques
Myocardial perfusion scanning:
201Thallium or 99mTc-sestamibi which are
distributed in proportion to myocardial blood flow
& concentrated by viable myocardium ,reveal a
defect(“cold spot”) in most patients during the
first few hrs after development of a transmural
infarct.
Although it is very sensitive it cannot distinguish
acute infarcts from chronic scars.

62. Radionucleotide
ventriculography
It is carried out with 99mTc-labelled red
blood cells .
It demonstrates wall motion disorders and
reduction in the ventricular ejection
fraction in patients with STEMI

64. Take home message
High fat containing diet & unhealthy life style causes
dyslipidemia & disturbs normal lipid profile which
increases the chances of an attack of acute myocardial
infarction.
Adults age 20 yrs or older should go for a routine lipid
profile at least once in a 5 years .
It is better to opt for a healthy lifestyle & a healthy diet to
prevent the risk of coronary artery disease.

65. THANK
YOU

66. References :
www.medscape.com
Harrison 17th edition
Henry’s clinical diagnosis & management
by laboratory methods 21st edition