conjugated linoleic acid isomers and mammary lipid...

Conjugated Linoleic Acid Isomers and Mammary Lipid Metabolism

Dr. Lance H. Baumgard1 and Dr. Dale E. Bauman2

1University of Arizona2Cornell University

Conjugated Linoleic Acids (CLA)• Many isomers (n=24) found in ruminant food products

– C18:2 cis-9, trans- 11– C18:2 trans-7, cis-9– C18:2 trans-10, cis-12– C18:2 cis-8, trans-10

• CLA has been shown to:– Anti-carcinogenic– Anti-atherogenic– Anti-diabetic– Enhanced immune system– Reduces severity of cachexia– Alleviates symptoms of lupus– Improved bone mineralization – Alters lipid metabolism

A B C

t10, c12 CLA c9, t11 CLA c9, c12 C18:2

Milk Fat CLA

• Early work focused on – Increasing CLA content– Identifying sources of variation– Determining composition of CLA isomers

• In attempt to enhance milk fat CLA it was serendipitously discovered that exogenous CLA causes severe milk fat depression (Chouinard et al., 1998)

Milk

fat (

%)

1.0

1.5

2.0

2.5

3.0

0 50 100 150

CLA-60 infused (g/d)Chouinard et al., 1998

Abomasal Infusion of CLADose Milk yield Milk fat Milk protein g/d days kg/d % g/d % g/d 01

100 1 16.0

15.0 3.31a

2.66b 540a

400b 3.15a

2.99b 790 760

02

31 60 92

5 21.5 20.4 20.9 18.3

2.81a

1.43b

1.38b

1.23b

599a

290b

295b

222b

3.31 3.37 3.53 3.46

696 675 717 627

03

15 18 29

3 26.9 27.5 29.4 26.8

3.34a

2.40b

2.36b

2.43b

883a

655b

691b

633b

3.14 3.03 3.04 3.15

831 826 882 829

1Loor and Herbein 1998; 2Chouinard et al., 1999; 3Chouinard et al., 1999

Which CLA Isomer is Responsible?

• CLA supplements contained many isomers

• Predominantly cis-9, trans-11 and trans-10, cis-12 CLA

• Diet-induced milk fat depression (MFD) associated with specific trans C18:1isomer

0

1

2

3

4

5

6/8 9 10 11 12 13/14

0

1

2

3

4

5

6/8 9 10 11 12 13/14

Double Bond Position

Com

posi

tion

(%)

Com

posi

tion

(%)

Normal Milk Fat

MF Depressed

Griinari et al. (1997)

C18:1 trans Fatty Acids

Hypothesis

• Rumen derived CLA or related metabolites containing a trans-10 double bond inhibit milk fat synthesis

linoleic acid(cis-9, cis-12 C18:2)

conjugated linoleic acid(cis-9, trans-11 CLA)

trans-11 C18:1

stearic acid (C18:0)

conjugated linoleic acid trans-10, cis-12 CLA

trans-10 C18:1


Rumen Biohydrogenation

linoleic acid(cis-9, cis-12 C18:2)

conjugated linoleic acid(cis-9, trans-11 CLA)

trans-11 C18:1


conjugated linoleic acid trans-10, cis-12 CLA

trans-10 C18:1


Griinari and Bauman, 1999

Change in rumen pH

Effects of CLA Isomers on Milk Fat %

1.5

2

2.5

3

3.5

-2 -1 1 2 3 4 5 6 7 8

Day

Milk

Fat

,per

cent

age

10,12 CLA9,11 CLAControl

Infusion

Baumgard et al., 2000 Am. J. Phys.

Milk Fat Response to trans-10, cis-12 CLA

300400500600700800

-1 1 2 3 4 5

Day of Infusion

Milk

Fat

Yie

ld (g

)

0

3.5

7

14

Dose (g/d)

Baumgard et al., 2001 J. Nutr.

trans-10, cis-12 CLA Dose Response

y = 0.24x2 - 6.99xR2 = 0.99

-60

-50

-40

-30

-20

-10

0

0 5 10 15trans-10, cis-12 CLA dose (g/d)

Perc

ent D

ecre

ase

in M

ilk F

at Y

ield

Baumgard et al., 2001Peterson et al., (in press)

trans-10, cis-12 CLA in Milk Fat

y = 152x2 - 176xR2 = 0.96

-60

-50

-40

-30

-20

-10

0

0 0.2 0.4 0.6 0.8

Milk fat trans- 10, cis- 12 CLA (g/100g fatty acids)

Perc

ent d

ecre

ase

in m

ilk fa

t yie

ld

Baumgard et al., 2001Peterson et al., (in press)

Effect of trans-10, cis-12 CLA (10 g/d) on Fatty Acid Composition

-2500

-2000

-1500

-1000

-500

0

Day Of Infusion

Dec

reas

e in

milk

fat s

ynth

esis

(m

mol

)

Total Decrease< C16

1 2 3 4

Milk Fat Yield -5% -18% -26% -45%


14:0/14:1

5

10

15

20

-1 1 2 3 4

16:0/16:1

5

10

15

20

25

-1 1 2 3 4

Treatmentcontrol cis-9, trans-11 CLA trans-10, cis-12 CLA

18:0/18:1

0.2

0.4

0.6

0.8

-1 1 2 3 4

Day of Infusion

trans 18:1/cis -9, trans -11 CLA

1234567

-1 1 2 3 4

Day of Infusion


-1000

-750

-500

-250

03.5 g/d 7.0 g/d 14.0 g/d

< C16C16:0 + C16:1> C16

Dec

reas

e in

Milk

Fat

Yie

ld (m

mol

)Effect of trans-10, cis-12 CLA on Milk Fatty Acid Composition


Milk Fat Yield -33%-25% -50%

Effect of Dose on ∆9-desaturase trans-10, cis-12 CLA, g/d

Substrate/product 0 3.5 7.0 14.0 SEM P Ratios 14:0/c9 14:1 16:0/c9 16:1 18:0/c9 18:1 t11 18:1/c9,t11 CLA

12.1a

18.5 0.4a

2.2a

12.7a

20.0 0.5a

2.6a

14.8a 19.2

0.6b 3.2b

20.2b

24.0 0.8c

4.1c

1.41.4

<0.1 0.1

<0.010.06

<0.01<0.01

Means within row with different superscripts differ (P < 0.05)


trans-10, cis-12 CLA in Milk Fat

-60

-50

-40

-30

-20

-10

0

0 0.2 0.4 0.6 0.8

Milk fat trans- 10, cis- 12 CLA (g/100g fatty acids)

Perc

ent d

ecre

ase

in m

ilk fa

t yie

ld

• diet-induced MFD

abomasal infusion

Circles are from Whitlock et al. (2002), Piperova et al. (2000) and Peterson et al., (2002)

Effects of CLA on Milk Fat Content During Late Lactation

1

2

3

4

-1 1 3 5 7 9 11Week of Treatment

%

CLACon

Perfield and Bauman 2002

Effect of trans-10, cis-12 CLA on Metabolic Flux Rates

100

600

1100

1600

Lipogenesis

nmol

es/1

00 m

g

CLACon

a

bb

a

CO2

Baumgard et al., (in press)

trans-10, cis-12 CLA and Mammary de novo Fatty Acid Synthesis

0

20

40

60

80

100

ACC FAS SCD

CLACon

a a

b b

a

b

mR

NA

exp

ress

ion


Effect of trans-10, cis-12 CLA on Mammary Triglyceride Synthesis

0

20

40

60

80

100

GPAT AGPAT

mR

NA

exp

ress

ion

CLACon

a a

bb


Effect of trans-10, cis-12 CLA on Mammary Fatty Acid Transport

0

20

40

60

80

100

LPL FABP

mR

NA

exp

ress

ion CLA

Con

a

b b

a


mRNA Abundance with Diet-Induced MFD

0%

25%

50%

75%

100%Fa

t (g/

d)

Prot

(g/d

)

KC

AS

AC

C

FAS

SCD

LPL

FAB

P

LIG

ASE

GPA

T

AG

PAT

Perc

ent o

f Con

trol

**

**

**

* P < 0.05

Peterson et al., 2002

Translocation SecretionCirculation Synthesis

DGAT

SFA (C16 - C18)

DesaturaseGPAT

FABP

UFALPL

Basal membrane ER- membrane Apical membrane

ACC FAS

FA synthesis de novo (C4 - C16)

TAG

synthesis

VLDL

TAGNEFA

Glycerol

Glucose

Glycerol

Glucose

Acetate

ßHBA

Common Coordination

• Central coordinator of lipid gene expression?– PPAR– SREBP– HNF

unchangedFeed intake

synthesis of protein and lactose unchanged or increased

Mammary epithelial cell apoptosis

plasma β-hydroxybutyrate unchangedKetogenesis

plasma concentration unchangedIGF-I

plasma glucose unchangedplasma concentration unchanged glucose response to insulin unchanged

Glucose homeostasisset-pointbasal insulinstimulated

plasma concentration unchangedLeptin

plasma non-esterified fatty acids unchangedresponse to β-adenergic stimulation unchanged

Lipolysisbasalstimulated

EffectProcess

Effects on Metabolic Processestrans-10, cis-12 CLA Inhibition of Milk Fat

Baumgard et al., 2000, 2001, 2002; Mackle et al., (in press)

CLA and Other Species

• Feeding CLA supplements decreases milk fat content in nursing women and lactating pigs

• Both species depend largely upon preformed fatty acid uptake and little on de novo fatty acid synthesis for milk fat production

Effective CLA Dose Body Fat vs. Lactation

Masters et al255571,2001.2 g/dWomen

Baumgard et al255273,5000.014%Cows

Blankson et alBerven et al

4NS

8484

234117

6,8003,400

6.8 g/d3.4 g/d

HumansHumans

Milk Fat

Body Fat

Baumgard et al5057810,0000.04 %Cows

Zambell et alNS94861,9501.9 g/dHumans

Ostrowska et al NS31

5050

71567

1,90015,400

0.125%1.000%

Pigs

West et al 6455

4242

1,0451,267

3542

1.0 %1.2 %

Mice

Ref% DecreaseDaysmg/kg0.75mg/din dietSpeciesFatDurationCLACLACLA

NS, not significant

CLA and Human Trials• Compared to rodent studies, CLA dose is

markedly lower (based upon % of diet and mg CLA/BW0.75) in human trials

• 20-25 g CLA/d is necessary to compare human trials to rodent experiments

• Due to high fat diets, humans rely little on de novo fatty acid synthesis– Therefore, if CLA reduces body fat by a similar

mechanism as in the mammary gland, CLA may be less effective in humans

Summary

• trans-10, cis-12 CLA markedly reduces milk fat synthesis

• cis-9, trans-11 CLA has little or no effect on mammary lipid metabolism

• Effects most aspects of lipid synthesis but especially pronounced on de novo fatty acid synthesis

• Solution to an old problem– MFD has been studied for over a century– Many theories have been proposed and

subsequently been proved inadequate

Summary Continued

• Little or no effect on other milk components• CLA reduces milk fat synthesis in pigs and

women• Mammary gland lipid metabolism is much

more sensitive (~ 20 x) to CLA than adipose lipid metabolism

• Other rumen derived conjugated dienes(trans-8, cis-10) and trienes (trans-10, cis-12, cis-15) probably reduce milk fat synthesis

Acknowledgements and Collaborators

– Dr. Bauman Dr. Yvan Chouinard– Debra Dwyer Dr. Bob Harrell– Ben Corl Dr. Tim Mackle– Jim Perfield II Dr. Mikko Griinari– Dr. Elvina Matitashvili Dr. Mark McGuire– Matt Madron Dr. Frank Dunshea– Dan Peterson Dr. Martin Auldist– Jane Kay

Any Questions?

conjugated linoleic acid isomers and mammary lipid...

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