Metabolism and Protective Properties of HDL

February 23, 2016

Khalid Al-Rasadi, BSc, MD, FRCPC

Head of Biochemistry Department, SQU

Head of Lipid and LDL-Apheresis Unit, SQUH

President of Oman society of Lipid & Atherosclerosis (OSLA)

DISCLOSURE

AstraZeneca

Pfizer

MSD

Abbott

Aegerion

Sanofi

Presentation Outlines

• HDL biogenesis and metabolism

• HDL function and cardioprotective properties

• HDL dysfunction

• Functional and compositional assessment of HDL

HDL biogenesis and metabolism

Structure of HDL

Surface monolayer ofphospholipidsand free cholesterol

Hydrophobic core of triglycerideand cholesteryl esters

apoA-I

apoA-II

Note: LFA1 denotes Lipid Free Apo Lipoprotein A1r HDL denotes Reconstituted HDL

HDL Consists of Heterogeneous

Particles, but Their Clinical Relevance

Remains to Be Established

HDL are Heterogenes (Contents)

PhospholipidsTriglycerids

Esterified cholesterol

Apo A1Free cholesterol

> 1000 different lipids(Phospholipid species, Cholesterol Ester, Trigylcerides, …)

70 different proteins(ApoA1, PON-1, ApoA2, ApoCIII,

ApoE, ApoH, ….)

CholesterolEster

PON-1

0

10

20

30

sn-2 Fatty Acid

sn-1 Fatty Acid

% O

ccu

rren

ce

16:0

PL

PO

PA

16:0

18:1

18:2

20:3

20:4

20:5

22:6

18:0

18:1

18:2

20:3

20:4

20:5

SL

18:2

18:1

18:2

20:4

20:5

18:1

18:2

Phospholipid Composition

PD

Discoidal

Spherical

Globular

Adapted from Barter PJ. Atheroscler Suppl. 2002;3:39-47.

HDL are Heterogeneous(Particle Shape)

HDL are Heterogeneous

(Surface Charge and Mobility)

Origin -Migration-Migration Pre--Migration

(E)HDL

Lipid-poor/Lipid-free apoA-I

Discoidal HDL

(A-I)HDL

(A-I/A-II)HDL

+Anode

-Cathode

HDL2aHDL2b HDL3cHDL3bHDL3a

HDL Particle Size/Electrophoretic Mobility

Apolipoprotein ContentParticle Shape

Discoidal

Spherical

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

Lipid-poor apoA-I

HDL Subpopulations

+other apos: A-IV, C, D, E, etc.

+non-apo proteins: inflam, thromb, etc.

--------------------------Alpha-migrating---------------------------Pre-beta-migrating--

Globular Discoidal

Globular

Adapted from Barter PJ. Atheroscler Suppl. 2002;3:39-47.

The HDL heterogeneity is the result of activity of several factors that

assemble and remodel HDL in plasma

apoA-I

(lipid-free)

Origin of ApoA-1

Chylomicrons

Lipolysis

Factors that assemble and remodel HDL in plasma

• ATP-binding cassette transporter (ABCA1)

• Lecithin—cholesterol acyltransferase (LCAT)

• Cholesteryl Ester Transfer Protein (CETP)

• Hepatic Lipase (HL)

• Phospholipid Transfer Protein (PLTP)

• Scavenger receptor class B type I (SR-B1)

Generates discoidal HDL by effluxing

phospholipids and cholesterol from cell

membranes to lipid-free/lipid-poor apoA-I

ABCA1

NC

Lipid-free/lipid-poor apoA-I

Discoidal HDL

• Generates most of the cholesteryl esters in plasma

•Activated by apoA-I and, to a lesser extent,

apoE and apoA-IV

•ApoA-II does not activate LCAT

• Remodels discoidal HDL into spherical HDL

LCAT

Spherical (A-I)HDL

UC

LCAT

Lyso-PCPC

LCAT Converts Discoidal HDL into Spherical HDL

Discoidal HDL

Spherical (A-I)HDL

UC

LCAT

Lyso-PCPC

LCAT Converts Discoidal HDL into Spherical HDL

Discoidal HDL

UC(concentration

gradient)

Peripheral cells

• Generates most of the cholesteryl esters in plasma

•Activated by apoA-I and, to a lesser extent,

apoE and apoA-IV

•ApoA-II does not activate LCAT

• Remodels discoidal HDL into spherical HDL

LCAT

Discoidal (A-I)HDL Discoidal (A-II)HDL

LCAT/LDL

Spherical (A-I/A-II)HDLClay et al. J. Biol Chem. 2000 275:9019-9025

Formation of (A-I/A-II)HDL by LCAT

• Transfers core lipids between HDL, VLDL and LDL

• Remodels HDL into large and small particles

• Mediates the dissociation of apoA-I from HDL

CETP

Barter et al; Biochem J. 1982; 2081:1, Swenson et al. J. Biol. Chem. 1988;263:5150, Tall. J. Lipid res. 1993; 34:1255

Role of CETP in plasma cholesterol transport

• Hydrolyses HDL phospholipids and triglycerides

• Remodelling is enhanced by CETP

• Mediates the dissociation of apoA-I from HDL

• Remodels HDL into small particles

Hepatic Lipase

TGCE

VLDL

Large Spherical HDLCE

TG

CETP

Lipid-free

apoA-I

HL

Small Spherical HDL

TGCE

CETG

•Transfers phospholipids between HDL and VLDL

•Remodels HDL into large and small particles

•Mediates the dissociation of apoA-I from HDL

PLTP

Scavenger receptor type B1 ( SRB1)

• SRB1 promotes the selective hepatic uptake of HDL cholesteryl esters

• SRB1 thus also remodels HDL

HDL function and cardioprotective properties

39

39

Mechanisms of Cellular Cholesterol Effluxto HDL particles

Extracellular space Cell membrane

FC

FC

FC

FC

ABCA1

Diffusion

SR-B1

Diffusion

SR-B1ABCG1

Diffusion

SR-B1ABCG1

Lipid-poor ApoA-I

Discoidal HDL

Small spherical HDL

Larger spherical HDL

LCAT

LCAT

40

Current Model of HDL-C Metabolism: Reverse Cholesterol Transport

BCE

Bile

SR-BILDLReceptor

Liver

FCCE LCAT

A-1

A-1

CEHL, EL

Macrophage

Mature HDL-C

Cuchel M et al. ATVB. 2003;23:1710–1712; Assmann G et al. Circulation. 2004;109(23 suppl 1):III-8–III-14.

FC

Nascent HDL

CE

TP

VLDL/LDL-C

Net transfer of cholesterol

FC

SR-BI

ABCG1

CE

Potential Antiatherogenic Actions of HDL

MCP-1 = monocyte chemoattractant protein-1

Adapted from Barter PJ et al. Circ Res. 2004;95:764-772.

Monocyte

Macrophage

Foam

cell

Vessel Lumen

Endothelium

IntimaCytokines

Adhesion

molecule

Oxidized LDL

LDL

LDL

HDL inhibits expression of endothelial cell adhesion

molecules and MCP-1

MCP-1

HDL inhibits

oxidation of LDL-C

HDL promotes efflux of

cholesterol from foam cells

Drew et al. Circulation 2009;119:2103-2111

ABCA1 RPKA

RPKA

PKA

PKA

R

R cAMP

cAMP

Lipid-free apoA-I

Ca2+

Ca2+Ca2+

Ca2+

InsulinAAAAA

AC AC

Adenylyl cyclase

cAMP

cAMP

cAMP

cAMP

cAMP

Insulin release

PKA

FOXO1

T24 S256

S319

HDL dysfunction

Functional and compositional

assessment of HDL

48

HDL: New Dimensions

QUANTITYHDL-C / Apo AI

QUALITYParticle structure

Lipidome, Proteome

Functionality

Kontush A, Chapman MJ. Pharmacol Rev. 2006.

Functional and Compositional Assessment of HDL

• Cholesterol efflux

• Antioxidant activity

• Anti-inflammatory activity

• Proteomics/lipidomics

Note: these are research tools w/o known clinical

relevance of application

Rosenson RS, Brewer HB Jr, Chapman MJ, Fazio S, Hussain MM, Kontush A, Krauss RM, Otvos JD, Remaley AT, Schaefer EJ.

Clin Chem. 2011;57(3):392-410.

Beyond HDL-C: Laboratory assessment of reverse cholesterol efflux (by plasma)

Study design: Endothelial effects of HDL -endothelial bioassays

Isolation of HDL2/3(by sequential ultracentrifugation)

Endothelial Function

(Endothelial cell NO production and vasoreactivity)

ESR spectroscopyOrgan chamber

ESR spectroscopy

Patients with acute coronary syndrome (n=25)Patients with stable coronary disease (n=25)

Healthy control subjects (n=25)

Vascular effects

Anti-oxidanteffects

(Endothelial cell superoxide production)

Anti-inflammatoryeffects

(Endothelial cell inflammatory activation)

Monocyte adhesion VCAM-1 expression

Effects on Re-Endothelialization

Carotid artery injury model in nude mice

Anti-thromboticeffects

Tissue factor Arterial thrombosis

HDL Proteome

Proposed Term Very Large HDL (HDL-VL)

Large HDL-V (HDL-L)

Medium HDL (HDL-M)

Small HDL (HDL-S)

Very Small HDL (VS-HDL)

Density range, g/mL 1.063-1.087 1.088-1.110 1.110-1.129 1.129-1.154 1.154-1.21

Size range, nm 12.9-9.7 9.7-8.8 8.8-8.2 8.2-7.8 7.8-7.2

Density gradient

ultracentrifugation HDL2b HDL2a HDL3a HDL3b HDL3c

Density range, g/mL 1.063-1.087 1.088-1.110 1.110-1.129 1.129-1.154 1.154-1.170

Gradient gel

electrophoresis HDL2b HDL2a HDL3a HDL3b HDL3c

Size range, nm 12.9-9.7 9.7-8.8 8.8-8.2 8.2-7.8 7.8-7.2

2D gel electrophoresis Alpha-1 Alpha-2 Alpha-3 Alpha-4 Preβ-1 HDL

Size range, nm 11.2-10.8 9.4-9.0 8.5-7.5 7.5-7.0 6.0-5.0

NMR Large HDL-P Medium HDL-P Small HDL-P

Size range, nm 12.9-9.7 9.7-8.8 8.8-8.2 8.2-7.8 7.8-7.2

Ion mobility HDL2b HDL2a + 3

Size range, nm 14.5-10.5 10.5-7.65

Separation of HDL by Physical Properties

Rosenson RS, Brewer HB Jr, Chapman MJ, Fazio S, Hussain MM, Kontush A, Krauss RM, Otvos JD, Remaley AT, Schaefer EJ.

Clin Chem. 2011;57(3):392-410.

Burning questions to resolve the puzzles with HDL?

• Can we relate HDL associated function to unique HDL subpopulations?

• Can distinct function of HDL subpopulations be accounted by specific protein components?

• Which of the known HDL functions protect against CVD?

Answers to these questions require much more research