The Players - Apolipoproteins

 Apo AI (liver, small intestine)

 Structural; activator of lecithin:cholesterol acyltransferase
(LCAT)
 Apo AII (liver)
 Structural; inhibitor of hepatic lipase; component of ligand for
HDL binding
 Apo A-IV (small intestine)
 Activator of LCAT; modulator of lipoprotein lipase (LPL)
 Apo A-V (liver)
 Direct functional role is unknown; regulates TG levels.
 Apolipoproteins

 Apo B-100 (liver)

 Structural; synthesis of VLDL; ligand for LDL-
receptor
 Apo B-48 (small intestine)
 Structural; synthesis of chylomicrons; derived from
apo B-100 mRNA following specific mRNA editing
 Apo E (liver, macrophages, brain)
 Ligand for apoE receptor; mobilization of cellular
cholesterol
 Apolipoproteins

 Apo C-I (liver)

 Activator of LCAT, inhibitor of hepatic TGRL
uptake
 Apo C-II (liver)
 Activator of LPL, inhibitor of hepatic TGRL uptake
 Apo C-III (liver)
 Inhibitor of LPL, inhibitor of hepatic TGRL uptake
 Lipoprotein Classes

Chylomicrons, LDL HDL

VLDL, and
their catabolic
remnants
> 30 nm 20–22 nm 9–15 nm
D<1.006 g/ml D=1.019-1.063g/ml D=1.063-1.21 g/ml

Doi H et al. Circulation 2000;102:670-676; Colome C et al. Atherosclerosis 2000;

149:295-302; Cockerill GW et al. Arterioscler Thromb Vasc Biol 1995;15:1987-1994.
Lipids Online
TG Rich: Chylomicrons
Surface Monolayer
Phospholipids (5%)
Free Cholesterol (1%)
Protein (1%)

Hydrophobic Core
Triglyceride (93%)
Cholesteryl Esters (1%)

Cholesterol and
Atherosclerosis, Grundy)
 CE Rich: LDL
Surface Monolayer
Phospholipids (25%)
Free Cholesterol (15%)
Protein (22%)

Hydrophobic Core
Triglyceride (5%)
Cholesteryl Esters (35%)

Cholesterol and
Atherosclerosis, Grundy)
 LDL Metabolism

LDL is removed by
apoB100 receptors
which are mainly
expressed in the
liver

Hepatic Lipase
Cholesteryl ester
transfer protein
Cholesterol and
Atherosclerosis, Grundy)
 LDL Uptake by Tissues

X X
Defects in the LDL receptor leads to familial hypercholesterolemia

Cholesterol and
Atherosclerosis, Grundy)
 CE Rich: HDL
Surface Monolayer
Phospholipids (25%)
Free Cholesterol (7%)
Protein (45%)

Hydrophobic Core
Triglyceride (5%)
Cholesteryl Esters (18%)

Cholesterol and
Atherosclerosis, Grundy)
 HDL Subpopulations
Particle Shape Apolipoprotein Composition
Discoidal

Spherical
A-I HDL A-I/A-II A-II HDL
HDL

Particle Size Lipid Composition

TG, CE, and PL

HDL2b HDL2a HDL3a HDL3b HDL3c

Rye KA et al. Atherosclerosis 1999;145:227-238.

Lipids Online
 HDL Maturation
HDL is secreted in a
discoidal form from the liver
and gut.

As it acquires cholesterol
from tissues in the
circulation, it matures into a
spherical form through the
action of lecithin:cholesterol
acyl transferase

Cholesterol and
Atherosclerosis, Grundy)
 HDL Metabolism
Nascent HDL (lipid-poor apoA-I) is produced by the liver and intestine
 HDL Metabolism
Free cholesterol is acquired from peripheral tissues
 HDL Metabolism
LCAT converts free cholesterol to cholesteryl esters
 HDL Metabolism
A variety of enzymes interconvert HDL subspecies
 HDL Interconversions

Cholesterol and
Atherosclerosis, Grundy)
 HDL Interconversions

Cholesterol and
Atherosclerosis, Grundy)
 HDL Metabolism
Cholesteryl esters can be selectively taken up via SR-BI
 HDL Metabolism
HDL particles can be taken up by a receptor-mediated process
 HDL Metabolism
Lipid-poor apoA-I can be removed by the kidney
Cholesterol Homeostasis
 LDL Cellular Metabolism
LDL are taken up by the LDL Receptor into clathrin-coated pits

Cholesterol and
Atherosaclerosis, Grundy)
 LDL Cellular Metabolism
LDL dissociates from the receptor; the receptor recycles to the membrane

Cholesterol and
Atherosaclerosis, Grundy)
 LDL Cellular Metabolism
In the lysosome, lipids are deseterified; proteins are hydrolyzed

Cholesterol and
Atherosaclerosis, Grundy)
 LDL Cellular Metabolism
Increase in free cholesterol regulates decrease cholesterol synthesis
and uptake; increase cholesterol esterification

Cholesterol and
Atherosaclerosis, Grundy)
Role of LXR and FXR

When cholesterol accumulates

in cells, cholesterol is oxidized
to create oxysterols
Role of LXR and FXR

Oxysterols activate LXR

through LXR/RXR
heterodimers to activate genes
such as the CYP7A1 enzyme
that catalyzes the rate-limiting
step in bile acid biosynthesis
Role of LXR and FXR

In the intestine, LXR also

activates ABC-1 to remove
cholesterol
 Role of LXR and FXR

In the intestine, FXR

activates expression of I-
BABP, a protein that
increases the transport of
bile acids back to the liver
from the intestine, reducing
their excretion.
 Role of LXR and FXR

The FXR receptor is activated

by bile acids. In the liver,
activation of FXR-RXR
heterodimers by bile acids
results in the feedback
inhibition of CYP7A expression
and reduced biosynthesis of
bile acids.
 Cholesterol Recycling

Cholesterol and
Atherosaclerosis, Grundy)
Hepatic Cholesterol Metabolism
Reverse Cholesterol Transport -
Peripheral Cells
Aqueous Diffusion:
Slow, unregulated, dictated
by membrane composition

Von Eckardstein et al, ATVB 2001

Reverse Cholesterol Transport -
Peripheral Cells
SR-BI: Binding of HDL to SR-
BI leads to reorganization of
cholesterol within the plasma
membrane and facilitates
cholesterol efflux

Von Eckardstein et al, ATVB 2001

Reverse Cholesterol Transport -
Peripheral Cells
ABC1: Fast and involves the
translocation of cholesterol from
intracellular compartments to the
plasma membrane via signal
transduction processes

Von Eckardstein et al, ATVB 2001

Reverse Cholesterol Transport -
Intravascular and Liver

TALL et al, ATVB 2000

Development of Athersoclerosis
 Evolution and Progression of
Coronary Atherosclerosis

O cclusive
Thr om bus

I nt im al I njur y Lipid- Rich Plaque Fibr om us cula r

Fat t y St r eak Plaque Dis r upt ion Thr om bus Lysis Response O cclusion

Ath e ro g e n ic Ris k F a c to rs Th ro m b o g e n ic Ris k F a c to rs

20 40 50 60
0
Ag e (y e a rs )
Adapted from Fuster, 1992
 Endothelial Dysfunction

 Increased endothelial
permeability to lipoproteins and
plasma constituents mediated
by NO, PDGF, AG-II,
endothelin.
 Up-regulation of leukocyte
adhesion molecules (L-selectin,
integrins, etc).
 Up-regulation of endothelial
adhesion molecules (E-selectin,
P-selectin, ICAM-1, VCAM-1).
 Migration of leukocytes into
artery wall mediated by oxLDL,
MCP-1, IL-8, PDGF, M-CSF.

Ross, NEJM; 1999

 Formation of Fatty Streak

 SMC migration stimulated by

PDGF, FGF-2, TGF-B
 T-Cell activation mediated by
TNF-a, IL-2, GM-CSF.
 Foam-cell formation
mediated by oxLDL, TNF-a,
IL-1,and M-CSF.
 Platelet adherence and
aggregation stimulated by
integrins, P-selectin, fibrin,
TXA2, and TF.

Ross, NEJM; 1999

 Formation of Advanced, Complicated Lesion

 Fibrous cap forms in response

to injury to wall off lesion from
lumen.
 Fibrous cap covers a mixture of
leukocytes, lipid and debris
which may form a necrotic core.
 Lesions expand at shoulders by
means of continued leukocyte
adhesion and entry.
 Necrotic core results from
apoptosis and necrosis,
increased proteolytic activity
and lipid accumulation.

Ross, NEJM; 1999

 Development of Unstable Fibrous Plaque

 Rupture or ulceration of fibrous

cap rapidly leads to thrombosis.
 Occurs primarily at sites of
thinning of the fibrous cap.
 Thinning is a result of continuing
influx of and activation of
macrophages which release
metalloproteinases and other
proteolytic enzymes.
 These enzymes degrade the
matrix which can lead to
hemorrhage and thrombus
formation

Ross, NEJM; 1999

 Plaque Rupture with Thrombus

Thrombus Fibrous cap

1 mm
Lipid core
Illustration courtesy of Frederick J. Schoen, M.D., Ph.D.
Lipids Online
 Growth Factors and Cytokines
Involved in Atherosclerosis
Growth Factor/Cytokine Abbr. Source Target
Epidermal growth factor EGF P EC, SMC
Acidic fibroblast growth factor aFGF EC ,M, SMC EC
Basic fibroblast growth factor bFGF EC ,M, SMC EC, SMC
Granulocyte macrophage colony stimulating factor GM-CSF EC ,M, SMC, T EC, M
Heparin-binding EGF-like growth factor HB-EGF EC ,M, SMC SMC
Insulin-like growth factor-I IGF-I EC ,M, SMC, P EC, SMC
Interferon  IFN- T, M SMC
Interleukin–1 IL-1 P, EC, M, SMC, T EC, M, SMC
Interleukin-2 IL-2 T EC, M, T
Interleukin-8 IL-8 EC ,M, SMC, T EC, T
Macrophage colony stimulating factor M-CSF EC ,M, SMC, T M
Monocyte chemotactic protein-1 MCP-1 EC ,M, SMC M
Platelet-derived growth factor PDGF EC ,M, SMC, P EC, M, SMC
RANTES SIS T M, T
Transforming growth factor- TGF- M EC
Transforming growth factor- TGF- EC ,M, SMC, T, P M, SMC
Tumor necrosis factor- TNF- EC ,M, SMC, T EC
Tumor necrosis factor- TNF- T EC, M, SMC
Vascular endotholelial growth factor VEGF EC ,M, SMC EC
Role of Lipoproteins in
Atherosclerosis
 CHD
CHD Mortality
Mortality isis Correlated
Correlated with
with Plasma
Plasma
Cholesterol
Cholesterol Levels
Levels
CHD Death Rate/1000

18 Six Year CHD Mortality from

16 MRFIT
14
12
10
8
6
4 Borderline
2 Desirable High HIGH
0
140 160 180 200 220 240 260 280 300
P las ma Choles terol (mg/dl)
LaRosa et al, 1990
 Role of LDL in Atherosclerosis
LDL Readily Enter the Artery Wall Where They May be Modified

Vessel Lumen
LDL

Endothelium
Oxidation of Lipids LDL Hydrolysis of Phosphatidylcholine
and ApoB to Lysophosphatidylcholine
Aggregation Other Chemical Modifications

Modified LDL
Modified LDL are Proinflammatory Intima

Steinberg D et al. N Engl J Med 1989;320:915-924.

Lipids Online
 Role of LDL in Atherosclerosis

Vessel Lumen
Monocyte

LDL

Endothelium
MCP-1 LDL

Modified LDL

Intima

Navab M et al. J Clin Invest 1991;88:2039-2046.

Lipids Online
 Role of LDL in Atherosclerosis

Monocyte
Vessel Lumen

LDL

Endothelium
MCP-1
LDL
Intima
Modified LDL

Modified LDL Promote

Differentiation of
Monocytes into
Macrophage Macrophages

Steinberg D et al. N Engl J Med 1989;320:915-924.

Lipids Online
 Role of LDL in Atherosclerosis

Monocyte Vessel Lumen

Adhesion LDL
Molecules

Endothelium
MCP-1
LDL

Cytokines
Modified LDL

Macrophage
Intima

Nathan CF. J Clin Invest 1987;79:319-326.

Lipids Online
 Role of LDL in Atherosclerosis

Monocyte Vessel Lumen

Adhesion LDL
Molecules

Endothelium
MCP-1
LDL

Modified LDL
Taken up by
Macrophage

Intima
Foam Cell Macrophage

Steinberg D et al. N Engl J Med 1989;320:915-924.

Lipids Online
 Role of LDL in Atherosclerosis

Monocyte Vessel Lumen

Remnant Lipoproteins

Adhesion Endothelium
MCP-1
MCP-1
Molecules
Remnants

Modified Intima
Cytokines Remnants
Growth Factors
Metalloproteinases

Macrophage
Cell Proliferation
Foam Cell Matrix Degradation

Doi H et al. Circulation 2000;102:670-676.

Lipids Online
 HDL is Protective

120 110
186 events in 4,407
per 1,000 (in 6 years)

100 men (aged 40–65 y)

80
Incidence

60

40 30
21
20

0
< 35 35–55 > 55
HDL-C (mg/dL)

Assmann G, ed. Lipid Metabolism Disorders and Coronary Heart

Disease. Munich: MMV Medizin Verlag, 1993
Lipids Online
 HDL Prevent Foam Cell Formation

Monocyte Vessel Lumen

LDL

Adhesion Endothelium
Molecules
MCP-1
LDL

Cytokines Modified LDL

Foam
Cell
Macrophage

HDL Promote Cholesterol Efflux Intima

Miyazaki A et al. Biochim Biophys Acta 1992;1126:73-80.

Lipids Online
 HDL Inhibits Oxidative Modification
of LDL

Monocyte
Vessel Lumen

LDL

Adhesion Endothelium
Molecules
MCP-1
LDL
HDL Inhibit
Oxidation
Cytokines Modified LDL of LDL

Foam
Cell
Macrophage

Intima
HDL Promote Cholesterol Efflux

Mackness MI et al. Biochem J 1993;294:829-834.

Lipids Online
 HDL Inhibits Expression of
Adhesion Molecules

Monocyte
HDL Inhibit Adhesion Molecule Expression

LDL Vessel Lumen

Adhesion Endothelium
Molecules
MCP-1
LDL
HDL Inhibit
Oxidation
Cytokines Modified LDL of LDL

Foam
Cell
Macrophage

Intima
HDL Promote Cholesterol Efflux

Cockerill GW et al. Arterioscler Thromb Vasc Biol 1995;15:1987-1994.

Lipids Online
 Suggested Risk Factors for CVD
 LDL Oxidation  Endothelial Injury  Plaque Instability
LDL-C Triglycerides/VLDL Plasma Metaloproteinase-9
Anti-OxLDL Non-HDL-C
OxLDL apoA-1/apoB  Thrombi Formation
LDL Oxid. Lag Time HDL-2/HDL-3 Factor VII
Negative LDL LDL size Fibrinogen
HDL-C Postprandial TG PAI-1
Paraoxonase IDL Factor VII
PAF acetylhydrolase Chylo. Remnants Tissue Plasminogen Activator
F2-Isoprostanes Blood Pressure D-Dimer
TBARS Homocysteine Plasmin-Antiplasmin Complex
ORAC Prothrombin Fragment 1+2
 Endothelial Dysfunction
Breath Ethane Platelet Activation
 Inflammatory Response von Willibrand’s Factor
P-Selectin
C-Reactive Protein
sICAM-1
IL-6
sVCAM-2
Lp-PLA2
Assymetric Dimethyl Arginine
Nitrate/Nitrite
Diet, Lipoproteins and CVD
 Seven
Seven Countries
Countries Study:
Study: CHD
CHD Events
Events are
are
Correlated
Correlated with
with Saturated
Saturated Fat
Fat
5
R = 0 .8 4
CHD De a th s a n d MI/1 0 0

4
N
U
3
W
2 C
S
M
G
D
1 B
Z
V
K
0
0 5 10 15 20
% Calories from S aturated F at
Keys, 1970
 Changes in Lipids with Step 1 and Step 2 Diets
Tot al Chole s t er ol Tr iglycer ides
DAI RY DELTA DAI RY DELTA
0
20
-5

³ TG , m g/ dl
³ TC, m g/ dl

15
- 10
10
- 15
5
- 20
0
St ep 1 St ep 2 St ep 1 St ep 2
LDL Chole s t er ol HDL Chole s t er ol
DAI RY DELTA DAI RY DELTA
0 0

-4
³ LDL- C, m g/ dl

³ HDL- C, m g/ dl -2
-8
-4
- 12

- 16 -6

St ep 1 St ep 2 St ep 1 St ep 2
 Regression Equations Have Been Developed to
Predict Average Lipid Responses to Dietary Changes

 Keys (1965) TC = 1.35* (2*S - P) + 1.52*Z

 Hegsted (1965) TC = 2.16*S - 1.65*P + 0.067*C - 0.53
 Mensink (1992) TC = 1.51*S - 0.12*M - 0.60*P
 Hegsted (1993) TC = 2.10*S - 1.16*P + 0.067*C
 Yu (1995) TC = 2.02*c12:0-c16:0 - 0.03*c18:0
- 0.48*M - 0.96*P
 Howell (1997) TC = 1.918*S - 0.900*P
+ 0.0222*C
 Newer Equations Can Accurately Predict Population
Response to Changes in Dietary Fat

Ob s e rv e Ho we ll He g s te d Me n s in k Ke y s
³ Chole s te rol (m g/dl)

25
20
15
10
5
0
-8 .7 % Milk fa t -1 3 .1 % Milk fa t
% Kcal Reduction in Milkfat
 Dietary Mechanisms to Lower LDL

 Reduce cholesterol intake

 Increase ACAT activity (SFA)
 Inhibit cholesterol absorption (plant sterols)
 Inhibit bile acid uptake (soluble fibers)
 Inhibit HMGCoA-reductase (tocotrienols)
 Inhibit FXR activation (guggelsterone)
 Thrombogenic Risk Factors May be as
Important as Lipid Risk Factors
 Fibrinogen: Upper tertile for fibrinogen
associated with 2.3-fold increase in risk for
myocardial infarction.
 Factor VII: 25% increase in factor VIIc is
associated with a 55% increase in risk of a fatal
CHD events within 5 years.
 Changes in Hemostasis Factors with
Step 1 and Step 2 Diets
Fa c tor VII Fibrinoge n
DAIRY DELTA DAIRY DELTA
0
15

³Fibrinogen, mg/dl
-2 12
³Factor VII, %

9
-4
6

-6 3
0
Ste p 1 Ste p 2 Ste p 1 Ste p 2
 Dietary Components and CHD Risk
Summary of the Nurses’ Health Study
Fruit (3.8 serv vs 0.6 serv)
Vegetables (6.8 serv vs 1.5 serv)
Vit E (Supplement vs no Supplement)
Margarine (<1 tsp/mo vs >4 tsp/d)
Alcohol (1 drink/d vs none)
Nuts (5 servings/wk vs almost never)
Folic Acid (>545 ug/d vs <190 ug/d)
Fiber (23g/d vs 12 g/d)
Whole grains (>1.7 serv vs <0.25 serv)
Saturated Fat (10.7% vs 18.8%)
Total Fat (29.1% vs 46.1%)
Eggs (<1/wk vs >1/d)

-6 0 -5 0 -4 0 -3 0 -2 0 -1 0 0 10 20

Pe rc e nt Cha n ge in CHD Ris k