Atherosclerosis

What Matters Most in Its Etiology ?

Michael P.C. Ip, Ph.D.
MetroHealth Medical Center
Case Western Reserve University

Presented in Singapore
SACB-NACCCA Joint Conference 6/17/2000

Atherosclerosis, Lipids and

Cardiovascular Disease (CVD)
The knot between atherosclerosis, lipids, & CVD

ÿ Atherosclerosis - closely related to Hyperlipidemia

ÿ Excess lipids or fatty substances in the blood is an

important risk factor in developing atherosclerosis and
CVD

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What is the Scope of the Problem ?
ÿ Immense
ÿ Steadily aging population in the US
Atherosclerosis – age is a definite risk
factor Men age >65
ÿ Ischemic heart disease 192 of every 1,000
Stroke 40 of every 1,000
Peripheral Artery Disease 69 of every 1,000

Heart facts
• Heart Disease - Nation’s # 1 killer
• Claims half-million lives every year
– high cholesterol is a well established risk
factor
– few denies that excess LDL-cholesterol
increase the risk of a heart attack

2
Monitoring Cholesterol in the
US (1980s)
• NIH (CCLBC 1983):
hypercholesterolemia must be treated
• 5% of the adult population (9 million
people) now take cholesterol-lowering
drugs in the hope of warding off stroke
or heart disease

More Heart Facts

(after 2 decades of lipid monitoring)
• Currently most heart attack victims
have cholesterol that qualify as
normal
– heart attack rate has fallen by half in recent
decades
– average cholesterol levels has fallen
slightly

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80 % of
patients who
eventually
suffer from
MI have the
same
cholesterol Patients who later suffered
levels as myocardial infarction (MI)
those who
Patients
will not
who
do not

Review Objectives
ÿ Identify the major factors important to
etiology of atherosclerotic lesions
ÿ Knowing the dynamics of their possible
interactions
ÿ Current strategies in reducing CVD risk

Department of Pathology
MetroHealth Medical Center, Cleveland, Ohio

4
 What is New on the Understanding
of Atherosclerosis ?
• A dynamic, inflammatory process that is
eminently modifiable
• Not just the work of ↑ total cholesterol
and LDL-cholesterol

Atherosclerosis
Foam Fatty Imtermediate Fibrous Complicated
Cells Streak Lesion Atheroma Plaque Lesion/Rupture

Thrombogenic
Lipid Core

From 1st decade From 3rd decade From 4th decade

Smooth muscle
Growth mainly by lipid accumulation and collagen
Thrombosis

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 Stable plaque with
core of lipids

Coronary Artery Disease begin with atherosclerosis

LDL Cholesterol
Platelets

Smooth
muscle cells
Endothelial
cells Endothelial injury by homocysteine, Monocytes
endotoxin ………. (rush to sites of injury, causing
further inflammation…..…)

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 2
Endothelial
cells express
selectin,
MCP-1

1 Endothelial cells
proliferate ……..
in an effort to heal the
lesion, causing plaque
to form on vessel
lining.

• Endothelial cells
are activated.
• Express surface
bound selectin &
adhesion molecules
Captured to attract & capture
monocytes circulating
• inflammatory cells
• • Facilitate their
migration into
subendothelial
space

Smooth muscle cells Inflammatory cells, predominantly

macrophages, some T cell and mast
cells, tend to accumulate at the shoulder
region of the plaque.
Weissberg, Heart 2000; March 83:247-252

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 Fibrotic cap •• Smooth muscle cells contain
large amount of contractile
built by proteins
VSMC ••
Vascular Smooth Its temporal function can be
Muscle Cells altered (via influence of
extracellular proteins)
(VSMC)
• Can develop an abundance of
synthetic organelles
• Facilitate their
• Take on repairative role to
migration into
“stabilize” the plaque
subendothelial
• space
Build a stable fibrous cap
(collagen, elastin & other
Stable Plaque matrix material) over the lipid
core

• VSMC are the only cells capable of synthesising the cap.

• Plays pivotal role in maintaining plaque stability and protecting
against plaque rupture and consequent thrombosis.
Weissberg, Heart March 2000; 83:247-252

Vessels can change their shape to accommodate an atheroma.

atheroma. Assessment
of severity of stenosis by coronary angiography may be misleading (Cleveland
Clinics J Med 1999)

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 Intravascular Ultrasound Atheroma within artery

Unstable Plaque – Inflammation & Rupture •• Composition of fibrotic cap - the

Platelets rapidly
accumulates at site of •
Preponderance of erosion or rupture;
inflammatory cells
intravascular thrombosis
out-strips repairative
ability of VSMC
can occur
• (INF- γ) inhibits
most important determinant to
plaque stability
• Inflammatory cells can weaken
fibrotic cap
- metalloproteinase degrades
cap matrix protein
- proinflammatory cytokines
Facilitate their VSMC
proliferation & collagen
migration into
synthesis
subendothelial
- Activated
space macrophages can
induce VSMC apotosis
• VSMC in mature plaque more
susceptible to apotosis

• Plaques with a large lipid pool and a thin fibrotic cap are
prone to rupture

Weissberg, Heart March 2000; 83:247-252

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 What makes a Plaque Vulnerable ?
• Large lipid core (>50% of plaque volume ?)
• High density of macrophages relative to
VSMC at cap
• High tissue factor content
• Thin cap in which the collagen structure is
disorganized

Thrombosis caused by
cap erosion. Mass of
thrombus completely
occludes a human
coronary artery. The
plaque itself is intact.

Thrombosis caused by
disruption. Thrombosis
may initiate from within
the core, and extend
into the lumen.

Davis, Heart March 2000; 83:361-366

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 Thrombosis Caused by
Disruption

• The typical lesion of

unstable angina
• Occlusion is incomplete

• Predominant cause of
(>85%) of major
coronary thrombi in
white males with
↑ LDL-Cholesterol
↓ HDL-Cholesterol

Davis, Heart March 2000; 83:361-366

Much is known on dynamic process of Atherosclerosis

Subclinical plaque rupture & repair of cracks and fissures in the
fibrotic cap is an on-going process. The size of the plaque grow
bigger, but the patient can be completely nonsymptomatic.

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 About Lipid Internalization - Macrophages do not take up
LDL readily unless it is oxidized
Paradox about LDL-Receptors
ÿ Foam cells develop in
patients and in animals
that totally lack LDL-
receptors
Transgenic Apo E
deficient mice
High blood cholesterol
Short life-span
Atherosclerotic lesion in adults
Incubating macrophages or SMC with very high levels of LDL - does not
increase LDL internalization

Steinberg Circulation 1997; 95:1062-1071

Chemically
modified by
treating LDL
in vitro with
acetic anhydride

Continue to take up acetyl LDL

until cells are engorged with stored cholesterol esters
Steinberg Circulation 1997; 95:1062-1071

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 Evidence that Oxidation of
LDL Occurs in vivo
ÿ LDL extracted from atherosclerotic lesions
are oxidatively modified
ÿ Atherosclerotic lesions show +ve
immunohistochemistry with Antibodies
generated from Ox-LDL

Circulating Circulating Monocytes Circulating LDL

T-Lymphocytes
1. Adhesion to aortic endothelium
a) Changes in monocytes
b) Changes in endothelial 6. Increased rate
8. Adhesion & of entry,
penetration cells
2. Penetration into intima decreased
3. Phenotypic modulation rate of
in subendothelial space exit
4. Replication

Arterial T-Cells Arterial Tissue Macrophages Intimal LDL

7. Oxidation
5. Uptake - modified LDL
Modified LDL
Foam Cells - Arterial macro-
Fatty Streak } macrophages / SMC-derived

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 Triglyceride-Rich Lipoprotein
• Triglyceride-rich lipoproteins
may also be very atherogenic
- chymomicrons (floats) - VLDL
- chymomicron remnants - VLDL remnants
- IDL
• Hyperglyceridemia may predict
the presence of small, dense LDL
DM & Type III
particles Hypercholesterolemia
High risk of CVD

Triglycerides in Blood
Macrophages Express a Distinct Receptor for Triglyceride-Rich Lipoproteins (TRLP)

VLDL in a normal person does not bind to TRLP / apo B48

receptors, so VLDL is lipolysed and removed by the liver
Miller, Hospital Practice, Sept 99, 34:67-73

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 In Patients with Hypertriglyceridemia, VLDL
Binds to TRLP / apo B48 Receptors

Rapid accumulation of lipids within macrophages and monocytes lead to

formation of foam cells. The binding to endothelial cells also impairs their
ability to release tissue plasminogen activator to mediate fibrinolysis.
Miller, Hospital Practice, Sept 99, 34:67-73

Lipoprotein Heterogeneity & Atherogenicity

Genetic Disposition & Environmental Related

• LDL-Cholesterol has its Polyacrylamide Gradient

own subclasses Gel Electrophoresis

• By electrophoresis it
can be separated into
its own classes
• What matters is the
marked atherogenic
potential of some
lipid subfractions

Superko, Sci & Med 1997

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• 56 y.o man non-smoker,
lean, ran marathon.
LDL
• His dad died of CVD at Larger diameter smaller
age of 58, smoker, over- LDL in size
wt, never exercised.
• This physically fit
56 y.o man who had
perfectly normal serum
lipids
LDL-chol 121 mg/dL, HDL-chol
42 mg/dL, Trig 109 mg/dL

Had AMI
Superko, Sci & Med 1997

100
This trait cannot be reliably detected
by routine blood test
80
of Population

60 Patients with triglycerides

>200 mg/dL are almost
always associated with
20 Pattern A pattern B. But it will result
Pattern B in 30-50% error if
%

triglyceride (when within

20 physiological range) is used
to predict LDL pattern
0
0 50 100 150 200 250 300
Triglyceride (mg/dL)

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 IIIa + IIIb
• 36%
4 months treatment with
niacin converted the Diameter
ο
lipoprotein to pattern A 251 A
and improved other
pattern B abnormality
(the ↑ IDL & post-
prandial lipidemia).

• TG, LDL-cholesterol, and IIIa + IIIb

HDL were not much 15%
changed with niacin Diameter
ο
therapy 268 A

Big and bad Small and bad

LDL Patterns

A B

Diameter
ο
Diameter
ο
268 A 251 A

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 Local Inflammation Smooth Muscle Cells

Endothelial Cell

Bacterium Endothelial Damage

Endotoxin ÿ Atheroma
Foam
LDL, TRLR Macrophage Formation
Cell

Intima

Adventitia

Benitez, Hospital Practice, Sept 99

Far-reaching effect of bacteria-induced systemic

inflammation on plaque stability
Systemic Inflammation
NO
CRP Endotoxin,
Fibrogens Inflammatory Vessel
Cytokines Constriction

TNF-α, Adhesion
Molecules Superimposed
Thrombosis
Coagulation Rupture
Factors Matrix Metallo-
proteinases
Interferon-γ
x Atheroma

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 ↓NO, ↑O2- …)
Endothelial Dysfunction – contributes to vascular resistance (↓
Blood passing through an atherosclerotic plaque does not flow smoothly. The
turbulent stress may tear the fatty deposit and the intima itself ……

Endothelial Dysfunction
Relating to Reduced
Nitric Oxide Factor

Exposing the atheromatous intima may active the platelets. Blood components
enter fissure. Platelets adhere to exposed collagen. Aggregation of platelets
form plugs. Fibrin formation initiated, and hemostatic plug is produced.

stenosed Stress Induced

Artery
Endothelial Damage
Tend to occur at vessel
bifurcation sites where blood
flow is most turbulent
- Non-Q-wave Infaract
- Myocardial Infarction
Non-Q wave Infarct - Stroke

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For coronary artery
disease, is previous
cardiac injury
important ?????
- compromized ventricular
function
- presence of residual
ischemia
- presence of life -
threatening ectopic
ventricular activity Tissue
injury

How is life after some tissue injury ???

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Potential Problems of Rebuilding

Normal Left ventricle with

muscle bundles held in
alignment by fibrillar
collagen.

Collagen disruption may

potentially allow bundles to
misalign

Cardiac Instability May Ensue

Counting our Knowledge on

Atherosclerosis – What Matters Most ????

ÿ Plaque build-up - multi-step processes

ÿ LDL-cholesterol - important component
ÿ But macrophages at intima with little oxidised
LDL-chol or triglyceride rich lipoprotein
do not develop readily into foam cells

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Atherosclerosis Reviewing The Other Players
Window of Opportunities for Intervention
ÿ Oxidative stress – balance between
oxidant and antioxidant
ÿ Oxidative damage
- activated macrophages can all generate reactive O2 ,
endothelial cells, superoxide
smooth muscle cells
ÿ
Antioxidants
- vitamin C - vitamin E
- β-carotene - selenium

Principal Antioxidants and their Interactions

ÿ Vitamin C
Regenerates active α-tocopherol (vitamin E) by reducing
its radical form

ÿ Vitamin E
Transport and storage depends on selenium; absorption is
reduced when vitamin A and β-carotene levels are high
ÿ β-Carotene
Conversion to vitamin A requires vitamin E

ÿ Selenium
Synergistic with vitamin E

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 Atherosclerosis
The Other Players
ÿ Homocysteine
- a class of in-born error of metabolism
- if untreated, patients may develop CHD during their teens or
during infancy
ÿ Diabetes Mellitus
- National Institute of Diabetes and Digestive and Kidney Disease
(NIDDK) and NHLBI emphasised that CVD and Type 2 diabetes
may be prevented or at least postponed by lifestyle changes that
maintain normal wt and physical activity

Change in Olive
Oil
Lifestyle ???

23
 Fish Oil Puzzle
Eskimo diet:
- mainly of fish and seal meat
- high in saturated fat
- plasma lipids are low
- ischemic heart disease is rare

Fish Oil has high levels of

n-3 Fatty Acids

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 Diet & Infaraction Trial Study
(DART)

Weighing the Facts

ÿ Sizable amount of n-3

fatty acids reduce
platelet adhesion, and
aggregation, but only
moderately.

ÿ Even small doses of

aspirin have a much
greater inhibitory effect
on platelet function.

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 Gauging the Complexity of Atherosclerosis
Lipid
Disorders
↑ TRL & Endocrine
↑ LDL Remnants Disorders
Ox-LDL Obesity

Smoking S LDL
↓ HDL
Diabetes
LP(a)
mellitus
Hypertension
Cytokine &
Atherosclerosis Growth Factor
Physical Genetic
Disorder
Inactivity Deposition
Platelet
Stress & Activation
Homocysteinemia
Depression Fibrinogen
Infection & Activation
↑ PAI-1
Slide courtesy: Dr. Tao Wang Inflammation
Otsuka Zmerica Pharmaceuticals Thrombogenic Disorders

Summary
ÿ Nationally average cholesterol levels has fallen slightly
ÿ
Heart attack rate has fallen by half in recent decades
ÿ
LDL-Cholesterol, Triglyceride-Rich Lipoprotein
are heterogeeous, so is their atherogenicity
ÿ
LDL dense and small is bad
ÿ “Meaningful” atherogenic direct LDL-cholesterol
monitoring is important for treatment

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 Summary
ÿ
Oxidized LDL enters macrophages and form foam
cells, so will trig-rich lipoproteins, if the stage is
properly set
ÿ Oxidant-antoxidant balance important
ÿ
DM is a CHD risk, so is hyperhomocysteinemia,
infections, stress, thrombogenic disorders,
hypertension, smoking, endocrine disorders.

Summary
ÿ Atherosclerosis is a dynamic, inflammatory process
that is eminently modifiable
ÿ
The interplay of these factors makes the study
of atherosclerosis difficult
ÿ This complexity also creates tremendous
opportunities for pathological modifications

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 Join NACCCA
ÿ North American Chinese Clinical Chemists
Association
ÿ
Speakers Aw, Wu, Chan, Zhou, Yeung, Ip are
NACCCA members
ÿ Membership application
www.naccca.org

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