Lipoprotein

Lipoproteins

Presented by: Farhad Jahanfar

Lecture Outline

� What are lipoproteins? What do they do

� Basic structure of lipoproteins

� Lipoprotein metabolism

� Cholesterol homeostasis

� Development of atherosclerosis

� Role of lipoproteins in atherosclerosis

� Role of diet in atherosclerosis prevention

Lipoprotein Overview

What are lipoproteins?

� Lipoproteins are protein-lipid complexes.

The Players – Lipids

Triacylglycerol

Phospholipids

CholesterolCholesteryl esters

What are lipoproteins?

� Hydrophobic lipids (TG, CE) in core;

� Hydrophilic lipids (UC, PL) on surface

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

What do lipoproteins do?

� Serve to transport lipid-soluble compounds between tissues Substrates for energy metabolism (TG) Essential components for cells (PL, UC) Precursors for hormones Precursors for eicosanoids Lipid soluble vitamins Precursors for bile acids

Lipoprotein Classes

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.

HDLHDLLDLLDLChylomicrons,Chylomicrons,VLDL, and VLDL, and

their catabolic their catabolic remnantsremnants> 30 nm> 30 nm 20–22 nm20–22 nm 9–15 nm9–15 nm

D<1.006 g/ml D=1.019-1.063g/ml D=1.063-1.21 g/ml

Lipids Online

Surface MonolayerSurface Monolayer Phospholipids (5%)Phospholipids (5%)Free Cholesterol (1%)Free Cholesterol (1%)Protein (1%)Protein (1%)

Hydrophobic CoreHydrophobic CoreTriglyceride (93%) Triglyceride (93%) Cholesteryl Esters (1%)Cholesteryl Esters (1%)

TG Rich: Chylomicrons

Cholesterol and Atherosclerosis, Grundy)



TG Rich: VLDL




CE Rich: LDL




CE Rich: HDL


Chylomicron Metabolism


Long-chain fatty acids are re-esterified into triacylglycerols in the gut and transferred; chylomicrons which contain apoB48 are synthesized and secreted into the blood via the lymphatic circulation



ApoC’s, apoE and cholesteryl esters are acquired from HDL in circulation.

ApoA-I and apoA-IV may be acquired from either the intestine or from HDL in circulation.



ApoC-II activates lipoprotein lipase which catalyses the hydrolysis of triacylglycerols



Apolipoproteins are transferred back to HDL



The chylomicron remnant is taken up by the apoB48/remnant receptor in the liver

Lipoprotein Metabolism

� Exogenous/chylomicron pathway (dietary fat)

� Endogenous pathway (lipids synthesized by the liver)

� HDL metabolism (apolipoprotein transfer, cholesteryl ester transfer, reverse cholesterol transport

VLDL Biogenesis


Microsomal TG transfer protein (MTP)

Facilitates the translocation, folding of apoB and addition of lipids to lipid binding domains

TG and cholesterol are synthesized in the liver as VLDL which contains apoB-100

VLDL Metabolism


Apo C’s and apoE and cholesteryl ester are acquired from HDL in circulation

Fatty Acid Transport


ApoC-II activates lipoprotein lipase which catalyses the hydrolysis of TG

VLDL Metabolism


Apolipoproteins are transferred back to HDL

The end product is a VLDL remnant (IDL)

VLDL Remnant Uptake


The remnant particle (IDL), if it contains apoE, can be taken up by the apoE/remanant receptor

VLDL Conversion to LDL


Further action on IDL by hepatic lipase loses additional apolipoproteins (apoE) becomes and is converted to LDL

LDL Metabolism


Hepatic LipaseCholesteryl ester transfer protein

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

LDL Uptake by Tissues


Defects in the LDL receptor leads to familial hypercholesterolemia

X X

Corneal arcus

Tendon xanthoma

Tendon xanthoma

Lipoprotein Metabolism

� Exogenous/chylomicron pathway (dietary fat)

� Endogenous pathway (lipids synthesized by the liver)

� HDL metabolism (apolipoprotein transfer, cholesteryl ester transfer, reverse cholesterol transport

HDL Subpopulations

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

Apolipoprotein CompositionApolipoprotein Composition

A-I HDLA-I HDL A-I/A-II A-I/A-II HDLHDL

A-II HDLA-II HDL

Particle ShapeParticle Shape

DiscoidalDiscoidal

SphericalSpherical

Lipid CompositionLipid Composition

TG, CE, and PLTG, CE, and PL

Particle SizeParticle Size

HDLHDL2b2b HDLHDL2a2a HDLHDL3a3a HDLHDL3b3b HDLHDL3c3c

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

HDL MetabolismNascent HDL (lipid-poor apoA-I) is produced by the liver and intestine

HDL MetabolismFree cholesterol is acquired from peripheral tissues

HDL MetabolismLCAT converts free cholesterol to cholesteryl esters

HDL MetabolismA variety of enzymes interconvert HDL subspecies

HDL Interconversions


HDL Interconversions


HDL MetabolismCholesteryl esters can be selectively taken up via SR-BI

HDL MetabolismHDL particles can be taken up by a receptor-mediated process

HDL MetabolismLipid-poor apoA-I can be removed by the kidney

Cholesterol Homeostasis

Hepatic Cholesterol Metabolism


Hepatic Cholesterol Synthesis

Cholesterol and Atherosaclerosis, Grundy)

Rate LimitingOnly pathway for cholesterol degradation

Energetically expensive; prefer to conserve what is already made/acquired – LDL receptor pathway

LDL Cellular Metabolism


LDL are taken up by the LDL Receptor into clathrin-coated pits



LDL dissociates from the receptor; the receptor recycles to the membrane



In the lysosome, lipids are deseterified; proteins are hydrolyzed



Increase in free cholesterol regulates decrease cholesterol synthesis and uptake; increase cholesterol esterification

↓↓XXX

Sterol Regulatory Element Binding Proteins and Cellular Cholesterol Metabolism

SREBP Cleavage Activating Protein


Intestinal Cholesterol Metabolism

Schmitz et al, JLR 2001



Lipids are absorbed from the intestine via a micellar transport process



Liberated unesterified cholesterol and plant sterols are transported back into the lumen via ATP-binding cassette (ABC) proteins G5 and G8 (heterodimers)

Defects in ABCG5 or ABCG8 leads to sitosterolemia

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



Reverse Cholesterol Transport - Peripheral Cells

Von Eckardstein et al, ATVB 2001

Aqueous Diffusion:Slow, unregulated, dictated by membrane composition



SR-BI: Binding of HDL to SR-BI leads to reorganization of cholesterol within the plasma membrane and facilitates cholesterol efflux



ABC1: Fast and involves the translocation of cholesterol from intracellular compartments to the plasma membrane via signal transduction processes

Reverse Cholesterol Transport - Intravascular and Liver

TALL et al, ATVB 2000

Development of Athersoclerosis

Evolution and Progression ofCoronary Atherosclerosis

I ntimal InjuryFatty Streak

Lipid- RichPlaque

PlaqueDisruption Thrombus Lysis Response

FibromuscularOcclusion

OcclusiveThrombus

020 40 50 60

Age (years )

Atherogenic Ris k Fac tors Thrombogenic Ris k Fac tors

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 mmLipid core

Illustration courtesy of Frederick J. Schoen, M.D., Ph.D.

Lipids Online

Growth Factors and CytokinesInvolved in Atherosclerosis

Growth Factor/Cytokine Abbr. Source Target

Epidermal growth factor EGF P EC, SMCAcidic fibroblast growth factor aFGF EC ,M, SMC ECBasic fibroblast growth factor bFGF EC ,M, SMC EC, SMCGranulocyte macrophage colony stimulating factor GM-CSF EC ,M, SMC, T EC, MHeparin-binding EGF-like growth factor HB-EGF EC ,M, SMC SMCInsulin-like growth factor-I IGF-I EC ,M, SMC, P EC, SMCInterferon λ IFN-λ T, M SMCInterleukin–1 IL-1 P, EC, M, SMC, T EC, M, SMCInterleukin-2 IL-2 T EC, M, TInterleukin-8 IL-8 EC ,M, SMC, T EC, TMacrophage colony stimulating factor M-CSF EC ,M, SMC, T MMonocyte chemotactic protein-1 MCP-1 EC ,M, SMC MPlatelet-derived growth factor PDGF EC ,M, SMC, P EC, M, SMCRANTES SIS T M, TTransforming growth factor-α TGF-α M ECTransforming growth factor-β TGF-β EC ,M, SMC, T, P M, SMCTumor necrosis factor-α TNF-α EC ,M, SMC, T ECTumor necrosis factor-β TNF-β T EC, M, SMCVascular endotholelial growth factor VEGF EC ,M, SMC EC

Role of Lipoproteins in Atherosclerosis

CHD Mortality is Correlated with Plasma Cholesterol Levels

LaRosa et al, 1990

140 160 180 200 220 240 260 280 300

Plasma Cholesterol (mg/dl)

02468

1012141618

CH

D D

eath

Rat

e/10

00 Six Year CHD Mortality from MRFIT

DesirableBorderline

High HIGH

Role of LDL in Atherosclerosis

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

EndotheliumEndothelium

Vessel LumenVessel LumenLDLLDL

LDL Readily Enter the Artery Wall Where They May be ModifiedLDL Readily Enter the Artery Wall Where They May be Modified

LDLLDL

IntimaIntima

Modified LDLModified LDL

Modified LDL are ProinflammatoryModified LDL are Proinflammatory

Hydrolysis of PhosphatidylcholineHydrolysis of Phosphatidylcholineto Lysophosphatidylcholineto Lysophosphatidylcholine

Other Chemical ModificationsOther Chemical Modifications

Oxidation of LipidsOxidation of Lipidsand ApoBand ApoB

AggregationAggregation

Lipids Online


LDLLDL

LDLLDL

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


Vessel LumenVessel Lumen

IntimaIntima

MonocyteMonocyte


MCP-1MCP-1

Lipids Online


LDLLDL

LDLLDL




IntimaIntima

MonocyteMonocyte


Modified LDL PromoteModified LDL PromoteDifferentiation ofDifferentiation ofMonocytes intoMonocytes intoMacrophagesMacrophages

MCP-1MCP-1

MacrophageMacrophage

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LDLLDL

LDLLDL

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


Vessel LumenVessel LumenMonocyteMonocyte



MCP-1MCP-1

AdhesionAdhesionMoleculesMolecules

CytokinesCytokines

IntimaIntima

Lipids Online


LDLLDL

LDLLDLEndotheliumEndothelium



MCP-1MCP-1

AdhesionAdhesionMoleculesMolecules


Foam CellFoam Cell

Modified LDL Modified LDL Taken up by Taken up by MacrophageMacrophage

IntimaIntima

Lipids Online





MCP-1MCP-1AdhesionAdhesionMoleculesMolecules

Foam CellFoam Cell

IntimaIntimaModifiedModifiedRemnantsRemnantsCytokinesCytokines

Cell ProliferationCell ProliferationMatrix DegradationMatrix Degradation

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

Growth FactorsGrowth FactorsMetalloproteinasesMetalloproteinases

Remnant LipoproteinsRemnant Lipoproteins

RemnantsRemnants

Lipids Online

HDL is Protective

110

3021

0

20

40

60

80

100

120

< 35 35–55 > 55

Inci

den

ceper

1,0

00 (

in 6

yea

rs)

HDL-C (mg/dL)

Assmann G, ed. Lipid Metabolism Disorders and Coronary Heart Disease. Munich: MMV Medizin Verlag, 1993

186 events in 4,407 men (aged 40–65 y)

Lipids Online

HDL Prevent Foam Cell Formation

LDLLDL

LDLLDL

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






CytokinesCytokines

IntimaIntimaHDL Promote Cholesterol EffluxHDL Promote Cholesterol Efflux

Foam Foam CellCell

Lipids Online

HDL Inhibits Oxidative Modificationof LDL

LDLLDL

LDLLDL

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






CytokinesCytokines

Foam Foam CellCell

HDL Promote Cholesterol EffluxHDL Promote Cholesterol EffluxIntimaIntima

HDL InhibitHDL InhibitOxidationOxidation

of LDLof LDL

Lipids Online

HDL Inhibits Expression ofAdhesion Molecules

LDLLDL

LDLLDL

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



MonocyteMonocyte




CytokinesCytokines

IntimaIntima

HDL InhibitHDL InhibitOxidationOxidation

of LDLof LDL

HDL Inhibit Adhesion Molecule ExpressionHDL Inhibit Adhesion Molecule Expression

Foam Foam CellCell

HDL Promote Cholesterol EffluxHDL Promote Cholesterol Efflux

Lipids Online

Suggested Risk Factors for CVD� LDL Oxidation

LDL-C

Anti-OxLDL

OxLDL

LDL Oxid. Lag Time

Negative LDL

HDL-C

Paraoxonase

PAF acetylhydrolase

F2-Isoprostanes

TBARS

ORAC

Breath Ethane

� Endothelial Injury

Triglycerides/VLDL

Non-HDL-C

apoA-1/apoB

HDL-2/HDL-3

LDL size

Postprandial TG

IDL

Chylo. Remnants

Blood Pressure

Homocysteine

� Thrombi Formation

Factor VII

Fibrinogen

PAI-1

Factor VII

Tissue Plasminogen Activator

D-Dimer

Plasmin-Antiplasmin Complex

Prothrombin Fragment 1+2

Platelet Activation

� Inflammatory Response

C-Reactive Protein

IL-6

Lp-PLA2

� Endothelial Dysfunction

von Willibrand’s Factor

P-Selectin

sICAM-1

sVCAM-2

Assymetric Dimethyl Arginine

Nitrate/Nitrite

� Plaque Instability

Plasma Metaloproteinase-9

Diet, Lipoproteins and CVD

Seven Countries Study: CHD Events areCorrelated with Saturated Fat

0 5 10 15 20

% Calories from S aturated F at

0

1

2

3

4

5

CH

D D

ea

ths

an

d M

I/10

0 R = 0.84

V

MC

DG

SW

B

Z

UN

E

K

Keys, 1970

St ep 1 St ep 2

- 20

- 15

- 10

- 5

0

³TC,

mg/

dl

Total Cholest erol

DAI RY DELTA

St ep 1 St ep 2

- 16

- 12

- 8

- 4

0

³LDL

-C,

mg/

dl

LDL Cholest erolDAI RY DELTA

St ep 1 St ep 20

5

10

15

20

³TG

, m

g/dl

Tr iglycer idesDAI RY DELTA

St ep 1 St ep 2

- 6

- 4

- 2

0

³HDL

-C,

mg/

dl

HDL Cholest erolDAI RY DELTA

Changes in Lipids with Step 1 and Step 2 Diets

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

-8.7% Milkfat -13.1% Milkfat

% Kcal Reduction in Milkfat

0

5

10

15

20

25

³ C

hole

ster

ol (

mg/

dl)

Obs erve Howell Hegs ted Mens ink Keys

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)

� 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.

Thrombogenic Risk Factors May be as Important as Lipid Risk Factors

Changes in Hemostasis Factors withStep 1 and Step 2 Diets

Step 1 Step 2

-6

-4

-2

0

³Fac

tor V

II, %

Factor VII

DAIRY DEL TA

Step 1 Step 20

3

6

9

12

15

³Fib

rinog

en, m

g/dl

FibrinogenDAIRY DEL TA

Dietary Components and CHD RiskSummary of the Nurses’ Health Study

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)

Eggs (<1/wk vs >1/d)

Saturated Fat (10.7% vs 18.8%)Total Fat (29.1% vs 46.1%)

-60 -50 -40 -30 -20 -10 0 10 20

Percent Change in CHD Ris k

Fruit (3.8 serv vs 0.6 serv)

Vegetables (6.8 serv vs 1.5 serv)

Thanks

Lipoprotein

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