Cholesterol Cholesterol Absorption and Absorption and...

Cholesterol Synthesis and Absorption Markers

Thomas Dayspring MD, FACP, FNLA1

Thomas Dayspring, MD, FACP, FNLAThomas Dayspring, MD, FACP, FNLA

Diplomate of the American Board of Clinical Diplomate of the American Board of Clinical LipidologyLipidologyFellow of the National Lipid AssociationFellow of the National Lipid Association

North American Menopause Society Certified North American Menopause Society Certified Menopause PractitionerMenopause Practitioner

Thomas Dayspring, MD, FACP, FNLAThomas Dayspring, MD, FACP, FNLA

Diplomate of the American Board of Clinical Diplomate of the American Board of Clinical LipidologyLipidologyFellow of the National Lipid AssociationFellow of the National Lipid Association

North American Menopause Society Certified North American Menopause Society Certified Menopause PractitionerMenopause Practitioner

Cholesterol Cholesterol Absorption and Absorption and Synthesis BiomarkersSynthesis Biomarkers

Cholesterol Cholesterol Absorption and Absorption and Synthesis BiomarkersSynthesis Biomarkers

Director of Cardiovascular Education Director of Cardiovascular Education The Foundation for Health Improvement and Technology (FHIT) The Foundation for Health Improvement and Technology (FHIT)

Richmond, VARichmond, VA

Disclosures (Last 12 months)Disclosures (Last 12 months)� Consultant or Advisory Board

• Abbott Labs• Amarin• Genetech (The Roche Group)• Genzyme• Glaxo Smith Kline (GSK)• Health Diagnostic Labs • Kowa & Eli Lilly• Merck• Omthera

�� Lecture BureauLecture Bureau•• AbbottAbbott•• GSKGSK•• HD LabHD Lab•• KowaKowa/Lilly/Lilly•• Merck Merck

12

3 5

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8

11 13

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14 15

17

Steranes are aromatic triterpenes(6 isoprene units arranged as tetracyclic rings)

Isoprene unit

A

Steranes, Sterols, SteroidsSteranes, Sterols, SteroidsSteranes, Sterols, SteroidsSteranes, Sterols, Steroids►►Sterols, also Sterols, also known as known as steroid steroid alcohols alcohols are are steranessteraneswith a with a hydroxyl group at the 3hydroxyl group at the 3--positionposition of the Aof the A--ring. ring. They are They are a group of compounds which a group of compounds which are essential components of are essential components of cell membranes in both plants cell membranes in both plants and animals. and animals.

Neil HAW & Huxley RR. Atherosclerosis Supplements 3 (2002) 11/15

►►Endogenous or exogenous sterols that are not cholesterol are Endogenous or exogenous sterols that are not cholesterol are referred to as referred to as noncholesterol sterols or Xenosterolsnoncholesterol sterols or Xenosterols

►►Plant sterols (Plant sterols (phytosterolsphytosterols) which are structurally similar to ) which are structurally similar to cholesterol, are not synthesized by the human body and are cholesterol, are not synthesized by the human body and are minimally absorbed from the gutminimally absorbed from the gut

SterolHO

Sterane(gonane)



CholesterolCholesterol

►►Cholesterol Cholesterol occurs as a component of plant membranes occurs as a component of plant membranes and as part of the surface lipids of leaves where it is and as part of the surface lipids of leaves where it is sometimes the major sterol. sometimes the major sterol.

►►The The quantity of cholesterol is generally small when quantity of cholesterol is generally small when expressed a percent of total lipid. While cholesterol expressed a percent of total lipid. While cholesterol averages perhaps 50 mg/kg total lipid in plants, it can be as averages perhaps 50 mg/kg total lipid in plants, it can be as high as 5 g/kg (or more) in animalshigh as 5 g/kg (or more) in animals

Behrman, E.J., Gopalan, V.J. Chem. Educ., 2005, 82 (12), p 1791

Cholesterol , from the Greek chole (bile) and stereos (solid)

followed by the chemical suffix -ol for an alcohol,

►►In In opposition to a opposition to a widespread belief, plants do widespread belief, plants do contain cholesterol both in a contain cholesterol both in a free or esterified free or esterified form. form.

OHOH

CHCH33

CHCH33

1122

33

44

55

991010 88

66

77

1111

12121313

1414 1515

16161717

1818 2020

21212222

2323

2424

2525

2727

2626

1919

3 h3 hydroxy ydroxy cholesterolcholesterol

CampesterolCampesterolOHOH

SitosterolSitosterolOHOH

StigmasterolStigmasterolOHOH

OHOH AvenosterolAvenosterol

Cholesterol Cholesterol vsvs PhytosterolsPhytosterols

The majority of the differences are in the “R” tail with plant sterols The majority of the differences are in the “R” tail with plant sterols having an extra methyl (campesterol) or ethyl (sitosterol) group at having an extra methyl (campesterol) or ethyl (sitosterol) group at

the Cthe C--24 position and different levels of desaturation24 position and different levels of desaturation

The more carbon atoms and desaturation, the The more carbon atoms and desaturation, the less less the intestinal absorptionthe intestinal absorption

CholesterolCholesterol

3(β)-cholest-5-en-3-ol

CycloalkenesCycloalkenes

CholesterolCholesterol SitosterolSitosterol CampesterolCampesterol

Sterol and Stanol StructuresSterol and Stanol Structures

OHOH OHOH OHOH

Sterols have a double bond at the ∆∆5 position

CholestanolCholestanol SitostanolSitostanol CampestanolCampestanol

OHOH OHOH OHOH

Saturation of the Saturation of the ∆∆ 5 double bond of sterols by enzymes in the liver results in 5 5 double bond of sterols by enzymes in the liver results in 5 αα stanolsstanols



Cholesterol

OHOH

OHOHHH

OHOH

HH

Cholesterol MetabolismCholesterol MetabolismCholesterol MetabolismCholesterol Metabolism

(Cholest-5-en-3 β-ol)

Cholesterol

(5β-Cholestan-3 β-ol)

5α-Cholestanol

(5α-Cholestan-3 β-ol)

5β-Coprostanol

OO

OOHH

OOHH

5α-Cholestanone

(5α-Cholestan-3-one)

(Cholest-4-en-3-one)

5β-Cholestanone

(5β-Cholestan-3-one)

Cholestenone

http://en.wikipedia.org/wiki/Coprostanol

CoprostanolCoprostanol is is formed by the formed by the conversion of conversion of

cholesterol cholesterol in in the the gut of most higher gut of most higher

animals by animals by intestinal bacteriaintestinal bacteria..

The The general general scheme for its scheme for its

production via a production via a ketone ketone

intermediate intermediate

NonchNoncholesterol Sterols/Stanolsolesterol Sterols/Stanols

Klaus von Bergmann et al. Am J Cardiol 2005;96 (supp l):10D–14D

SterolsCholesterol Cholesterol Campesterol Sitosterol BrassicasterolStigmasterol

Sitostanol Campestanol

Stanols

236.5 236.5 0.474 0.326 0.047 0.011

0.012 0.003

Serum concentration of cholesterol and noncholester ol sterols/stanols in patients with mild hypercholeste rolemia

Serum concentration (mg/dL)

Cholesterol SynthesisCholesterol Synthesis►► In the whole animal, and presumably in humans, most In the whole animal, and presumably in humans, most

cholesterol is synthesized and utilized in the extrahepatic cholesterol is synthesized and utilized in the extrahepatic organsorgans

►► The CNS contains The CNS contains as much as 25% of the total amount of as much as 25% of the total amount of unesterified cholesterol unesterified cholesterol in the entire body, in the entire body, and that and that is is mostly mostly produced produced via via local de novo synthesislocal de novo synthesis

►► Under dietary conditions Under dietary conditions equivalent to those found in equivalent to those found in Western Western humans, the humans, the extrahepatic tissues probably account extrahepatic tissues probably account for > 80% for > 80% of whole of whole animal sterol synthesis in virtually every animal sterol synthesis in virtually every species that species that has been has been studied.studied.

►► Most of the cholesterol carried in LDL is taken up into the Most of the cholesterol carried in LDL is taken up into the liver (liver (indirect reverse cholesterol transportindirect reverse cholesterol transport))

Dietschy JM, Turley SD & Spady DK. J Lip Res 1993;34: 1637-1659



Cholesterol SynthesisCholesterol Synthesis

AcetateAcetate AcetoacetylAcetoacetyl--CoACoA

HMGCoAHMGCoA

HMG synthaseHMG synthase

MevalonateMevalonate IsopentylIsopentyl--pyrophosphatepyrophosphate

DimethylallylDimethylallyl--PPPP

FarnesylFarnesyl--pyrophosphatepyrophosphate

condensationcondensation

isomerizationisomerization

SqualeneSqualeneSqualene Squalene synthasesynthase

LanosterolLanosterol CholesterolCholesterol1919--20 reactions including 20 reactions including Desmosterol, LathosterolDesmosterol, Lathosterol

4 ringed structure4 ringed structure

IsoprenoidsIsoprenoids

Dayspring T in Chap 14 Davidson, Dayspring T in Chap 14 Davidson, TothToth, Maki Therapeutic Lipidology 2008, Maki Therapeutic Lipidology 2008

HMG CoA reductaseHMG CoA reductase

7-dehydrodesmosterol7-dehydrocholesterol

Lathosterol Desmosterol

Cholesterol

Lanosterol Bloch Cholesterol Synthetic PathwayKandutsch-Russell

Cholesterol Synthetic Pathway

► Cellular cholesterol precursors are also found in normal human plasma, at concentrations roughly 1:1000 of that of cholesterol

► These values are highly heritable and could potentially be used to predict individual responsiveness to the cholesterol-lowering regimen

Lusa S et al. JBC 2003;278:19844-51

EsterificationEsterification : joining of an sterol: joining of an sterol--alcohol alcohol with a long chain fatty acidwith a long chain fatty acid

Free Cholesterol * (unesterified)

Esterified cholesterol or Cholesteryl Ester (CE) Esterified cholesterol or Cholesteryl Ester (CE) (Non(Non--polar, hydrophobic or water insoluble)polar, hydrophobic or water insoluble)

*A chemical compound possessing both hydrophilic and lipophilic

hydrophilic properties.

Usually esterified with oleic or palmitic acid creating cholesteryl oleate or palmitate

The ester bond is formed between the carboxylate group of a fatty acid

and the hydroxyl group of cholesterol.

Cholesteryl esters have a lower solubility in water than cholesterol.

OO

CCOHOH

CHCH33

CHCH33

11

22

33

44

55

99

1010 88

66

77

1111

12121313

1414 1515

16161717

1818 2020

21212222

2323

2424

2525

2727

2626

1919

O

Lipoprotein

Free or unesterified cholesterol



Cholesterol Absorption and SynthesisCholesterol Absorption and Synthesis

►►Normal human serum contains small amounts of the Normal human serum contains small amounts of the cholesterol precursors squalene, cholestenol, cholesterol precursors squalene, cholestenol, desmosteroldesmosterol and and lathosterollathosterol which which reflect cholesterol reflect cholesterol synthesis synthesis especially in ratios to serum cholesterolespecially in ratios to serum cholesterol

►►Small concentrations of Small concentrations of cholestanol, campesterol and cholestanol, campesterol and sitosterolsitosterol are also detectable in serum the ratios of which are also detectable in serum the ratios of which are are related to cholesterol absorptionrelated to cholesterol absorption

►►The two groups (synthesis and absorption markers) of The two groups (synthesis and absorption markers) of xenosterols are xenosterols are nnegatively (inversely) related to each egatively (inversely) related to each other in the general populationother in the general population

Miettinen TA et al. Atherosclerosis 2010;210:592-59 7

Plasma Absorption / Synthesis Markers Ranges

Normal Hypo

Campesterol (µg/mL)

Campesterol ratio 10 2

mmol/mol cholesterol

Sitosterol (µg/mL)

Cholestanol (µg/mL)

Sitosterol ratio 10 2


Desmosterol ratio 10 2


Desmosterol (µg/mL)

Cholestanol ratio 10 2


Ste

rol A

bsor

ptio

n M

arke

rs

Hyper

> 4.44

> 241

> 3.18

> 169

> 3.48

> 195

> 1.28

> 65

< 2.102.1 - 4.3

115 - 240

1.43- 3.17

76 - 168

2.02 - 3.47

117 - 194

0.5 - 1.27

< 114

< 1.42

< 75

< 2.01

< 116

< 0.49

< 3031.0 - 64

Based on quintile cut-points from 500 patientsUnless campesterol or sitosterol are in the ranges seen with phytosterolemicpatients (100-300 µg/mL) one cannot state with assu rance the phytosterols are causal of atherosclerosis

Ste

rol

Syn

thes

is

Mar

kers

Note that the concentrations are reported as absolute

values or as ratios of cholesterol

Cholesterol Synthesis and Cholesterol Synthesis and Absorption RelationshipsAbsorption Relationships

Relationship between absorbed dietary cholesterol a nd the rate of sterol synthesis in freshly isolated blood monocyte s

Donald J. McNamara et al, JCI 1987;79:1729-39

9876543210

2

4

6

8

10(8) (38)

(30)(18)

(14) (16)(17)

(7)

(2)

r = -0.931 p < 0.01

Absorbed Dietary Cholesterol (mg/kg/day)

Mon

onuc

lear

Cel

l Ste

rol

Syn

thes

is (p

mol

/h/1

06

cells

(5)

The rates of incorporation of [2The rates of incorporation of [2--1414C]acetate into sterols in freshly C]acetate into sterols in freshly isolated blood isolated blood mononuclear leukocytes or MNL mononuclear leukocytes or MNL ((picomolespicomoles per hour per hour

per 10per 1066 cells) plotted against the mass of absorbed dietary cells) plotted against the mass of absorbed dietary cholesterol (milligrams per kilogram per day).cholesterol (milligrams per kilogram per day).

When individual data are considered rather than means of group data, the degree of suppression of MNL sterol synthesis was found to be linearly related to the mass

of cholesterol actually absorbed by

the volunteers (milligrams per

kilogram per day)



Using Sterol BiomarkersUsing Sterol Biomarkers

►►The The use of use of surrogate markers like noncholesterol sterols surrogate markers like noncholesterol sterols is is qualitative in nature and cannot qualitative in nature and cannot replace or replace or yield the quantitative yield the quantitative data derived by data derived by isotopic methodsisotopic methods. .

►►HoweverHowever, compared with the more , compared with the more complicated and complicated and exact exact isotopic measures of isotopic measures of cholesterol synthesis cholesterol synthesis and absorption, the and absorption, the assessment of assessment of noncholesterol sterols is affordable noncholesterol sterols is affordable and can be and can be performed on large performed on large populations. populations.

►►This This has led to the wide use of has led to the wide use of noncholesterol sterols to noncholesterol sterols to measure and characterize cholesterol metabolismmeasure and characterize cholesterol metabolism

MacKay DS & Jones PJH. Current Opin Lipidol 2012;23:2 41-247

Cholesterol Absorption and BalanceCholesterol Absorption and Balance

Intestinal Brush Border

Dietary Cholesterol & Plant Sterols

500 of each mg/day

Biliary Cholesterol Transport

Synthesized cholesterol (400 mg/day)Synthesized cholesterol (400 mg/day)

Secretion of 24 Secretion of 24 gmsgms/day of bile salts of /day of bile salts of which 2 which 2 gmgm is cholesterol is cholesterol

The intestine absorbs about 50% of cholesterol that is presented to it, but that can vary individually from 20-80%

Cholesterol AbsorptionCholesterol Absorption

Bosner MS et al. J Lipid Res 1999;40:302-308

Cholesterol absorption measured in Cholesterol absorption measured in 100 healthy patients using dual isotope 100 healthy patients using dual isotope

tracer technique. tracer technique.

0

10

20

30

40

Num

ber o

f Sub

ject

s

Decile Percent Cholesterol Absorption1st 2nd 3rd 4th 5th 6th 7th 8th 9th 10th

HyperHyper--absorbersabsorbersHypoHypo--absorbersabsorbers

While intestinal absorption of bile acids While intestinal absorption of bile acids is essentially is essentially complete under normal conditions, complete under normal conditions, cholesterol absorption cholesterol absorption in healthy in healthy adult volunteers is variable, with 29adult volunteers is variable, with 29––81% (mean 56%) 81% (mean 56%) absorbed absorbed

in in the small intestine. This range of variability has been observed in the small intestine. This range of variability has been observed in many studies many studies where where cholesterol absorption ranged from 25 to 75cholesterol absorption ranged from 25 to 75% % J Lip Res 1998;39:2415-22 ..

The majority The majority absorb about absorb about 55% of dietary 55% of dietary

sterolssterols



Diet and Cholesterol AbsorptionDiet and Cholesterol Absorption

Sehayek E et al. J Lip Res 1998;39:2415-22

NNeither either dietary dietary cholesterol nor dietary cholesterol nor dietary fat fat significantly altered significantly altered

% cholesterol % cholesterol absorption.absorption.

RRegardless of egardless of diet type, the diet type, the individuals within individuals within each each dietary dietary group group differed differed

markedly markedly in the percentage in the percentage dietary cholesterol dietary cholesterol

absorptionabsorption. .

10

20

30

40

50

60

70

80

Cho

lest

erol

Abs

orpt

ion

(%)

60±12 58±1056±13

LF LC HF LC HF HC

Variability in dietary cholesterol absorption rates according to diets.

Displayed figures are the group mean ± SD values (measured during 3 rd week of the diet)

L = Low H = High F = Fat C = Cholesterol

LF = low fat diet

LC = low cholesterol diet

HF = high fat diet

HC = high cholesterol diet


Chylomicron & HDL Transport



500 of each mg/day





Of the absorbed cholesterol 75% is from bile and 25% from diet

Understanding Cholesterol ConsumptionUnderstanding Cholesterol Consumption

�� The average diet consists of 200 The average diet consists of 200 –– 600 mg of cholesterol600 mg of cholesterol�� Based on numerous studies, evidence has accumulated to Based on numerous studies, evidence has accumulated to

suggest that suggest that the amount of dietary cholesterol (consumption) the amount of dietary cholesterol (consumption) has no substantial effect on CAD riskhas no substantial effect on CAD risk except in certain except in certain circumstancescircumstances

�� Canadian guidelines do not specify an upper limit for dietary Canadian guidelines do not specify an upper limit for dietary cholesterol citing there is little relationship between dietary cholesterol citing there is little relationship between dietary cholesterol and CVDcholesterol and CVD

Kidambi S et al. Future Lipidology. 2006;1:357-368




Chylomicron & HDL Transport



500 of each mg/day


Ileal Bile Salt Reabsorption

Fecal Sterols Fecal Sterols (400 mg/day)(400 mg/day)



Portal venous return is Portal venous return is > 95% of biliary secretion> 95% of biliary secretion


Of the absorbed cholesterol 75% is from bile and 25% from diet

Lacteal Plasma

Enterocyte

Steroidogenic tissue

Hepatocyte

Bile ductule

Small Intestinal Lumen

Adipocyte

Peripheral Cell De novo SynthesisFoam Cell

Hepatocyte & Enterocyte Sterol Absorption

NPC1L1

NPC1L1ABCA1

ABCA1

SR-B1

Noncholesterol sterols

Myocyte

LDL receptors

Cholesterol

TG

ABCG5 G8

Phytosterols

Micelle Cholesterol

ABCG5 G8

Noncholesterol sterols

Cholesterol

SR-B1

ABCB11Bile Acids

ABCA1 = ATP binding cassette transporter A1ABCG5, ABCG8 = ATP binding cassette transporter G5 & G8ABCB11 = ATP binding cassette transporter B11NPC1L1 = Niemann Pick C1 like 1 proteinSR-B1 = Scavenger receptor B1

Phytosterols

Bile acids

Genetic Expression of NPC1L1 and ABCG5, ABCG8 help

regulate cholesterol homeostasis

Ge L, Wang J et al. Cell Metab 2008;7:508-519

NPC1L1 Mediated Internalization of CholesterolNPC1L1 Mediated Internalization of CholesterolNPC1L1 protein

recycles between the plasma membrane (PM) and endocytic recycling

compartment (ERC).

When the extracellular cholesterol

concentration is high, cholesterol is

incorporated into the PM and is sensed by cell surface-localized

NPC1L1.

NPC1L1 and cholesterol are then internalized

together through clathrin/AP2-mediated

endocytosis and transported along

microfilaments to the ERC in vesicles.

The ERC is where massive amounts of cholesterol and NPC1L1 are stored. of NPC1L1 and eventually decreasing cholesterol absorption. When the intracel lular cholesterol level is low, ERC-localized NPC1L 1

moves back to the PM along microfilaments in order to absorb cholesterol.

Endocytic Recycling

Compartment

OH

Cholesterol NPC1L1 AP2Clathrin

Extracellular

Intracellular

High Cholesterol

NPC1L1 AP2 Clathrin Complex

Clathrin

AP2

Low Cholesterol



PhytosterolemiaPhytosterolemiaPhytosterolemiaPhytosterolemia►►Mutations Mutations in in either ABCG5 either ABCG5 (sterolin(sterolin--1) or ABCG8 1) or ABCG8

(sterolin(sterolin--2) cause 2) cause the autosomal recessive the autosomal recessive ddisease isease of sitosterolemia of sitosterolemia

►►Phytosterolemia leads to increased atherosclerosisPhytosterolemia leads to increased atherosclerosis, , formation of tuberous and formation of tuberous and tendinoustendinous xanthomasxanthomas, , and and macrothrombocytopeniamacrothrombocytopenia, with rare cases of , with rare cases of endocrine disruption or cirrhosis of the liver also endocrine disruption or cirrhosis of the liver also reportedreported

SolcaSolca C et al. C et al. JJ. . Lipid ResLipid Res. . 2013. 54: 3972013. 54: 397––409.409.

Phytosterols Phytosterols –– Injurious?Injurious?Phytosterols Phytosterols –– Injurious?Injurious?►► It It is not is not clear whetherclear whether, when , when ABCG5, ABCG8 ABCG5, ABCG8

function is impaired, any function is impaired, any subsequent subsequent pathophysiological pathophysiological issues that arise are because issues that arise are because of of cholesterol cholesterol toxicity or xenosterol toxicity (or bothtoxicity or xenosterol toxicity (or both).).

►► In sitosterolemic individualsIn sitosterolemic individuals, as well as in several in , as well as in several in vivo and vivo and in vitro in vitro studies, phytosterols can affect the studies, phytosterols can affect the activity of activity of several metabolic several metabolic pathways.pathways.

SolcaSolca C et al. C et al. JJ. . Lipid ResLipid Res. . 2013. 54: 3972013. 54: 397––409.409.

Sterol Biomarkers of Sterol Biomarkers of Synthesis and Synthesis and

Absorption and CV RiskAbsorption and CV Risk



Cofan M et al. Nutrition, Metabolism & Cardiovascula r Diseases 2011;21: 651e657

In both dyslipidemic and healthy populations, In both dyslipidemic and healthy populations, the Metabolic the Metabolic Syndrome Syndrome is associated is associated with with increased plasma increased plasma lathosterol, a lathosterol, a

cholesterol synthesis marker, and decreased plasma cholesterol synthesis marker, and decreased plasma sitosterol, a sitosterol, a marker of marker of cholesterol absorption. cholesterol absorption.

Cholesterol Homeostasis Markers in Cholesterol Homeostasis Markers in Metabolic Syndrome PatientsMetabolic Syndrome Patients

Chan DC et al. Obes Res 2003;11:591-596

Associations between plasma campesterol concentration and plasma lathosterol (A) and VLDL-apoB secretion (B) rate.

500

R = -0.366

VLD

L ap

oB s

ecre

tion

(mg/

kg/d

ay R = -0.359450

400

350

300

250

200

150

100

50

00 100 200 300 400 500 600 700 0 100 200 300 400 500 600 700

30

25

20

15

10

5

0

Campesterol (mmol/mol cholesterol)Campesterol (mmol/mol cholesterol)

Lath

oste

rol (

mm

ol/m

olch

oles

tero

l

A B

The plasma The plasma campesterol:cholesterolcampesterol:cholesterol ratio ratio was significantly was significantly lower (51%, lower (51%, p p << 0.01), whereas the 0.01), whereas the ratios ratios of of lathosterol:cholesterollathosterol:cholesterol and and lathosterol:campesterollathosterol:campesterol were 24were 24% (% (p < p < 0.05) and 64% higher 0.05) and 64% higher ((p p << 0.01), respectively, 0.01), respectively, in the in the lowlow--cholesterol absorption compared with the cholesterol absorption compared with the highabsorptionhighabsorption groupgroup. .

Moreover, the subjects in the Moreover, the subjects in the lowlow--cholesterol absorption cholesterol absorption group had significantly higher group had significantly higher plasma plasma VLDLVLDL-- and IDLand IDL--apoB pool sizes (37% and 27%, respectivelyapoB pool sizes (37% and 27%, respectively, , p < p < 0.01) and VLDL0.01) and VLDL--apoB secretion apoB secretion

rate (30%, rate (30%, p p 0.050.05) compared with the high) compared with the high--absorption group.absorption group.

LOWER Absorption HIGHER LOWER Absorption HIGHER

Syn

thes

is

VLD

L-P

Phytosterols and Family History of Phytosterols and Family History of Coronary Heart DiseaseCoronary Heart Disease

Sudhop T et al. Metabolism 2002;51:1519-1521

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

Without History With History Without History With HistoryCampesterol Sitosterol

Pla

sma

conc

entr

atio

n m

g/dL

Pla

sma

conc

entr

atio

n m

g/dL

Individual concentrations of plant sterols in patie nts with and without family history of CHD

Mean Mean



FFramingham ramingham OOffspring ffspring SStudy (tudy ( FOSFOS) ) Cholesterol Absorption/Synthesis Markers are Cholesterol Absorption/Synthesis Markers are

Associated with Prevalent CVDAssociated with Prevalent CVD

Matthan N, et al. J Lipid Res 2009;50:1927-1935

Case control study in 155 cases and 414 controls. Multivariable analysis adjusted for Case control study in 155 cases and 414 controls. Multivariable analysis adjusted for diastolic blood pressure, LDLdiastolic blood pressure, LDL--C, HDLC, HDL--C, triglycerides; diabetes and hypertension C, triglycerides; diabetes and hypertension medication; age, sex, systolic blood pressure, body mass index, and smoking.medication; age, sex, systolic blood pressure, body mass index, and smoking.

Absorption Absorption MarkersMarkers

Synthesis Synthesis MarkersMarkers

Squalene

Desmosterol

Lathosterol

Campesterol

Sitosterol

Cholestanol

Latho/Camp

Latho/Sito

-1 10 2 3 4 5 6

Matthan N, et al. J Lipid Res 2009;50:1927-1935

“In conclusion, present results indicate that alterations in “In conclusion, present results indicate that alterations in cholesterol homeostasis, cholesterol homeostasis, namely high cholesterol namely high cholesterol absorption absorption andand low cholesterol synthesis, are associated low cholesterol synthesis, are associated with increased CVD risk with increased CVD risk in a subset of men and women in a subset of men and women with similar plasma LDLwith similar plasma LDL--cholesterol cholesterol concentrations”concentrations”

“Additionally“Additionally, the cholesterol homeostasis markers , the cholesterol homeostasis markers appear to be appear to be better predictors of disease than traditional better predictors of disease than traditional lipid risk factors lipid risk factors in this study in this study population”population”

PROPROspectivespective CACArdiovascularrdiovascular MMunster Study unster Study ((PROCAMPROCAM): Elevated ): Elevated Phytosterols and CHDPhytosterols and CHDPROPROspectivespective CACArdiovascularrdiovascular MMunster Study unster Study ((PROCAMPROCAM): Elevated ): Elevated Phytosterols and CHDPhytosterols and CHD

Assmann G et al. Nutrition, Metabolism & Cardiovascu lar Diseases (2006) 16, 13e21

Hazard ratios for development of coronary events according to

sitosterol concentration (mmol/L) among men in

different categories of 10-year global coronary risk (hazard ratio of 1 = global risk < 10% and sitosterol ≥ 5.25 mmol/L).

The participants in the category with low global risk ( < 10%)

were divided into groups with low (≤ 5.25 mmol/L, 39 cases, 140 controls) and high (> 5.25 mmol/L, 17 cases, 46 controls)

sitosterol concentrations.

20

15

10

5

0<10% 10.0 – 19.9% ≥20%

Global Risk (in 10 Years)Sitosterol ( µmol/L)

≤5.25

>5.25

Hazard Ratio for development of coronary events for sitosterol concentration by Global Risk

1.49

4.15

17.23

5.762.89

P=0.326P=0.383

P=0.032

At medium level of global risk (10.0 - 19.9%), low s itosterol concentrations were observed in 29 cases and 53 controls and high sitosterol lev els in 18 cases and 24 controls, while at high global risk ( ≥ 20%), low sitosterol levels occurred in 38 cases a nd 47 controls while

high sitosterol levels were measured in 18 cases an d 8 controls.

1



PROPROspectivespective CACArdiovascularrdiovascular MMunster Study unster Study ((PROCAMPROCAM): Elevated ): Elevated Phytosterols and CHDPhytosterols and CHDPROPROspectivespective CACArdiovascularrdiovascular MMunster Study unster Study ((PROCAMPROCAM): Elevated ): Elevated Phytosterols and CHDPhytosterols and CHD

Assmann G et al. Nutrition, Metabolism & Cardiovascu lar Diseases (2006) 16, 13e21

►►The PROCAM study was conducted before statins were The PROCAM study was conducted before statins were available, and few participants were receiving other lipidavailable, and few participants were receiving other lipid--modifying medications when their blood samples were modifying medications when their blood samples were drawn drawn

►►Two potential explanations for these data are:Two potential explanations for these data are:

►►That sitosterol is somehow involved in the disease process That sitosterol is somehow involved in the disease process

►►Sitosterol is a surrogate for some other factor or condition that is Sitosterol is a surrogate for some other factor or condition that is involved in atherogenesis.involved in atherogenesis.

i.e. hyperabsorption of cholesterol

Phytosterols and AtherogenesisPhytosterols and Atherogenesis

►►HHyperabsorption of phytosterols can produce yperabsorption of phytosterols can produce premature CAD and aortic stenosis in individuals with premature CAD and aortic stenosis in individuals with defects in ABCG5/8 raises the defects in ABCG5/8 raises the possibility that increased possibility that increased serum levels of phytosterols can contribute to CAD in the serum levels of phytosterols can contribute to CAD in the general populationgeneral population..

►►Variation in the plasma concentration of plant sterols is Variation in the plasma concentration of plant sterols is highly heritable and that genetic polymorphisms in highly heritable and that genetic polymorphisms in ABCG8 contribute to this variability. ABCG8 contribute to this variability.

Manoj D. Patel, Paul D. Thompson. Atherosclerosis 20 06;186:12–19

Genser B et al. European Heart Journal (2012) 33, 44 4–451

►► The The metameta--analyses analyses were not supportive of were not supportive of any relationship between any relationship between serum concentrations of sitosterol and campesterol (both absolute serum concentrations of sitosterol and campesterol (both absolute concentrations and concentrations and ratios to cholesterol) and risk of CVD. ratios to cholesterol) and risk of CVD.

►► Our Our systematic review and metasystematic review and meta--analysis did not reveal any evidence of analysis did not reveal any evidence of an an association between association between serum concentrations of plant sterols and risk of serum concentrations of plant sterols and risk of CVDCVD..

►► Our Our systematic review and metasystematic review and meta--analysis analysis have several limitationshave several limitations. . ----------further research will be necessary to further research will be necessary to assess whether assess whether plant sterols are plant sterols are causally involved in causally involved in atherogenesis atherogenesis ------ does not address the vascular does not address the vascular effects or safety of longeffects or safety of long--term consumption of plant sterolterm consumption of plant sterol--enriched foods.enriched foods.



Genser B et al. European Heart Journal (2012) 33, 444–451

Serum levels of Serum levels of sitosterol in CVD sitosterol in CVD

cases and controls cases and controls (ratios (ratios to total to total cholesterol).cholesterol).

1.50-.5-1 -.25-.75

Study ID SMD (95% CI)

Case Control Rajaratnam 2000 Rajaratnam 2001 Gylling 2009 D+L Subtotal I-V Subtotal

Cohort Strandberg 2006 D+L Subtotal I-V Subtotal

Cross Sectional Wilund 2004 (M) Wilund 2004 (F) Fassbencer 2008 Shay 2009 (no statin) Silbernagel 2009 D+L Subtotal I-V Subtotal

Nested Case Control Pinedo 2007 Escurriol 2009 Mathan 2009 Mathan 2010 D+L Subtotal I-V Subtotal

D+L Overall I-V Overall

0.50 (0.12, 0.89) 0.31 (-0.26, 0.89) 0.54 (0.16, 0.93) 0.48 (0.24, 0.73 0.48 (0.24, 0.73)

0.28 (0.06, 0.51) 0.28 (0.06, 0.51) 0.28 (0.06, 0.51)

-0.10 (-0.23, 0.03) -0.08 (-0.22, 0.07) -0.26 (-0.44, -0.07) -0.20 (-0.45, 0.06) -0.16 (-0.42, 0.09) 0.06 (-0.04, 0.15) -0.10 (-0.20, 0.00) -0.06 (-0.12, -0.00)

0.00 (-0.12, 0.12) -0.09 (-0.23, 0.05) 0.29 (0.10, 0.47) -0.31 (-0.49, -0.13) -0.03 (-0.23, 0.18) -0.03 (-0.11, 0.04)

0.01 (-0.09, 0.12) -0.02 (-0.06, 0.03)

Higher Levels in Controls Higher Levels in Cases Standardized Mean Difference (CVD cases – controls)

►► Why is cholesterol the preferred substrate and why are Why is cholesterol the preferred substrate and why are phytosterols poor substrates for esterification by ACAT and phytosterols poor substrates for esterification by ACAT and LCAT?LCAT?

►► Why did humans evolve the ABCG5 and ABCG8 Why did humans evolve the ABCG5 and ABCG8 nonesterified sterol efflux transporters ?nonesterified sterol efflux transporters ?

►► Why are these sterol efflux transporters expressed at the Why are these sterol efflux transporters expressed at the two critical locations (gut lumen/enterocyte interface and two critical locations (gut lumen/enterocyte interface and hepatobiliary interface?hepatobiliary interface?

Phytosterols “Good or Evil” EnigmaPhytosterols “Good or Evil” Enigma

A teleology is any philosophical account that holds that A teleology is any philosophical account that holds that final final causes exist causes exist in nature, meaning that design and purpose analogous to that found in in nature, meaning that design and purpose analogous to that found in

human actions are inherent also in the rest of naturehuman actions are inherent also in the rest of nature..http://en.wikipedia.org/wiki/Teleologyhttp://en.wikipedia.org/wiki/Teleology

Current Opin Lipidol 2012;23:241-247

►► The measurement The measurement and reporting of and reporting of noncholesterol sterols (NCSs) noncholesterol sterols (NCSs) is is very heterogeneous and standardization would allow better comparison very heterogeneous and standardization would allow better comparison or metaor meta--analyses of results across different assessment methods and analyses of results across different assessment methods and improve the use of NCSs in both research and clinical settings.improve the use of NCSs in both research and clinical settings.

►► Noncholesterol Noncholesterol sterols are still a valuable research tool for the overall sterols are still a valuable research tool for the overall assessment of cholesterol assessment of cholesterol metabolism and metabolism and may may have clinical potential in have clinical potential in the personalization of diet and the personalization of diet and medicine. They are an affordable medicine. They are an affordable means means of of estimating cholesterol estimating cholesterol metabolism metabolism

►► Cholesterol Cholesterol metabolism of different disease states metabolism of different disease states can be can be phenotypedphenotypedwith NCSs and this may help with NCSs and this may help in identifying in identifying appropriate hypolipidemic appropriate hypolipidemic treatmentstreatments. .



Principles of Sterol Testing: Review

�Markers of absorption and synthesis are reported as absolute values and as ratios adjusted for cholesterol. They have the same meaning: some believe the ratios are best used if patient is on a cholesterol lowering drug

�Absorption markers are sitosterol and campesterol which are phytosterols and cholestanol, a metabolite of cholesterol found in some dietary products

�The synthesis markers are desmosterol and lathosterol

�As a very general rule typically hyperabsorbers tend to be hyposynthesizers and vice versa. Hypoabsorption is desirable and requires no treatment per se

�The pattern associated with the most CV risk is hyperabsorption with hyposynthesis

�The disease phytosterolemia is very rare and the sitosterol, campesterol levels will be in the 100-300 µg/mL

�Although elevations of phytosterols are associated with CV risk, one cannot state with assurance the phytosterols are causal of atherosclerosis

Two Hearts Peter Max

Thank You For Thank You For Your AttentionYour Attention

See my 5 part audiovisual Lecture Series on Sterols at www.lecturePad.org

Cholesterol Cholesterol Absorption and Absorption and...

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