Health and Exercise Science Research · Health and Exercise Science Research Citrulline 1. Emphasize the importance of skeletal muscle size and quality for healthy ageing. 2. Briefly

Health and Exercise Science Research


Citrulline

1. Emphasize the importance of skeletal muscle size and quality for

healthy ageing.

2. Briefly review best protein nutrition practice for ageing muscles.

3. Evaluate the biological rationale and existing scientific evidence

base for emerging nutrients and/or neutraceuticals related to

promoting the health of ageing muscles.

4. Highlight gaps in research knowledge for the nutrition industry and

academia to exploit with regards to the active ageing market.

This presentation aims to…

Weak │ Promising │ Strong

Resveratrol Vitamin D

Arginine

Leucine Β-HMB Creatine

N-3 PUFA

Vitamins C and E

Sodium nitrate

Phosphatidic acid

Ursolic acid

Alcohol


The importance of muscle size and quality extends

beyond performance nutrition

Strength sports Aesthetics

PERFORMANCE

NUTRITION

ACTIVE

NUTRITION

HEALTH AND

WELLNESS

Power : mass

Healthy ageing Healthy weight loss Child growth ↓ Diabetes risk

Better choices, more often

↓ Obesity risk

Wolfe RR. The underappreciated role of muscle in health and disease. Am J Clin Nutr, 2006


Mu

scle

ma

ss (

AU

)

Age (years) 50

Healthy

Disuse-

accompanied

90

Ageing is associated with a gradual loss of skeletal muscle

mass that is exacerbated by periods of muscle disuse

“Threshold of disability”


65 YEARS OLD

PLUS

LATE 30’S &

EARLY 40S

MIDDLE AGED

The market space of active ageing extends beyond 65

years plus

MUSCLE

RECONDITIONING


6

“The progressive decline in skeletal muscle mass (myopenia) and strength (dynapenia) that

predisposes reduced muscle function with advancing age”

Report of the International working group on sarcopenia

Fielding et al 2011 Age and Ageing 12(4): 249-256

What is the definition of sarcopenia?


The cause of sarcopenia is clearly multifactorial,

interconnected and complex

Exercise

Insulin Resistance/Blood Flow

Injury/Illness

DNA Damage/Apoptosisis

Hormonal Changes

Anabolic Resistance

Inactivity

Motor Units

Oxidative Stress

Nutrition

Satellite Cells Low grade inflammation

AGE 25 AGE 65

Morley 2012 Fam Practice


What is muscle protein synthesis and why should the nutrition

industry care in terms of healthy ageing

Amino Acids

BLOOD

Amino Acids

MUSCLE PROTEINS

(MYOFIBRILLAR AND MITOCHONDRIAL)

MPS MPB MUSCLE

Mitochondrial Proteins

PB

PS

Collagen Proteins

PB PS

PS


0 5 10 15 20 25 30 35 40 450.00

0.05

0.10

0.15Young

Elderly

Dose of EAA

** **

Myofibrillar protein FSR (%/h)

Cuthbertson et al. 2005 FASEB J

1

2 3

The response of muscle protein synthesis to amino acid

feeding is impaired in older adults


10

Low-grade inflammation impairs the stimulation of muscle

protein synthesis in response to food intake

• 2 groups of rats: low grade inflamed (LGI)

group and a control non-inflamed (NI) group

• Muscle (gastrocnemius) protein synthesis

• Fasted and fed state

Non inflammed Low grade inflammed

0

2

4

6Fasted Fed

a

b

a a

Muscle protein synthesis (%/day)

Balage et al. 2010 J Nutr Biochemistry


INGESTED PROTEIN

SOURCE

The magnitude of the “muscle protein synthetic” response to

ingested protein is regulated on several levels of physiology

MUSCLE PROTEIN SYNTHESIS

Myofibrillar │ mitochondrial │ collagen

3. Amino acid uptake by skeletal

muscle

Amino acid transporters

Leucine mTORC1

p70S6K1 4E-BP1

MAP4K

LAT1

4. Anabolic cell

signaling

a. Protein digestibility

b. Amino acid absorption kinetics

1. Blood amino acid

availability

c. Splanchnic

retention of

amino acids

a. Blood flow

b. Capillary recruitment (via insulin)

c. Microvascular perfusion

2. Amino acid delivery to skeletal

muscle

↑ Insulin response

11


Witard et al 2014 Am J Clin Nutr

Dose of whey protein (g)

Muscle protein synthesis (%/h)

0 10 20 40

0.00

0.02

0.04

0.06

0.08

0.10

aa

bb

0 10 20 40

0.00

0.03

0.05

0.08

0.10 c

b

aa

Yang et al 2012 Br J Nutr

Dose of whey protein (g)


The ‘optimum’ protein dose for maximal stimulation of

muscle protein synthesis increases with advancing age


The potential drawback associated with recommending

increased daily protein intakes in older adults

Leidy et al. 2007 Obesity

High altitude

exposure

Anorexia

Reduced energy intake

↓ appetite regulating

hormones

15% 30%

0

20

40

60

80

100

15% 30%

0

20

40

60

80

100

Hunger (VAS: 1-100) Fullness (VAS: 1-100)

Protein content of diet

*

*

─ Older (64 years old) women

─ 9 wk dietary control

─ En% protein - 15% vs 30%


Maximise anabolic potency Enhance anabolic sensitivity

Daily protein intake Meal-by-meal

protein intake

Prior exercise Coingesting other nutrients

Anti-inflamm.

nutrients Vitamin

D

Optimising the response of MPS

Nitrates Neutra-

ceuticals Alcohol

How can industry and academia collaborate to

overcome age-related anabolic resistance?

β-HMB

Creatine Antioxidants Leucine

Ca+ Free acid

Arginine

Citrulline

Sodium nitrate

Phosphatidic acid

Ursolic

acid

Curcumin

Vitamins C and E

N-3 PUFA

Resveratrol


Fish oil supplementation potentiates the response of muscle

protein synthesis to simulated feeding in older adults

Basal Fed Basal Fed

0.00

0.02

0.04

0.06

0.08

0.10

Mixed muscle protein synthesis (%/h)

Before n-3 PUFA

supplementation

After n-3 PUFA

supplementation

* • Healthy active young men (n=5) and women (n=4)

• 8 weeks of fish oil derived n-3 PUFA supplementation

- 4 g/day fish oil capsules

- 1.86 g/day EPA and 1.50 g/day DHA

• Basal and stimulated measurements of MPS

Smith et al. 2011 Clinical Science,


Kalmorat & Gray 2013 Biochem and Biophys

• C2C12 myotubes

• Incubated with ethanol (control),

EPA or DHA

• Stimulated by leucine

• Protein synthesis and protein

breakdown

EPA is the “anabolic” component of fish oils

Protein synthesis rate

(Phenylalanine kinetics)

Control EPA DHA

0.0

0.5

1.0

1.5*

Protein breakdown rate

(Phenylalanine kinetics)

Control EPA DHA

0

5

10

15

20

*


Resveratrol may protect against the age-related loss of

muscle loss via anti-inflammatory properties

Wang et al. 2014 Int Immunopharmacology

• C2C12 myotubes (a mouse cell line)

• Treated with TNF-α

• Incubated with or without resveratrol

• Direct and indirect markers of

muscle atrophy

0

5

10

15

20

25

TNF-α:

Resveratrol:

_ +

_ _

_

+

+

+

Myotube diameter (μm)


Myotubes Myoblasts Osteoblasts

0

20

40

60

80Pre

Post

Tracer uptake of 25(OH)D

Abboud et al. 2013 Endocrinology

Vitamin D is taken up and stored in skeletal muscle cells

*

*


Control Ins+Leu Ins+Leu+VitD

0.0

20.0

40.0

60.0

80.0

100.0

120.0

140.0

a

b

c

Salles et al. 2013 Mol. Nutr. Food Res

Protein synthesis rate

(A.U.)

• C2C12 myotubes

• Control (unstimulated cells)

• Ins+Leu (stimulated with leucine and insulin only)

• Ins+Leu+VitD (cultured in 1,25(OH)2D3 for 72 h)

Vitamin D has been shown to enhance the MPS response

to an anabolic stimulus


Curcumin may protect against muscle loss in older adults

by reducing muscle inflammation and oxidative stress

Ono et al. 2015 Exp Phys

• Old mice injected with

streptozotocin to induce T1D

• 2 weeks of diet with or without

curcumin (1500 mg· kg-1 · day)

• Measurements of skeletal muscle

weight and myocyte cross-

sectional area

Con

trol

Con

trol+

Cur

cum

inT1D

T1D+C

urcu

min

0

50

100

150

200

250

a a

bc

Rectus femoris weight (mg)


INGESTED PROTEIN SOURCE

Biological rationale behind the potential for nitrates to enhance

the MPS response

(MYOFIBRILLAR) MUSCLE PROTEIN SYNTHESIS

a) Protein digestibility b) Amino acid absorption kinetics

1. Blood amino acid availability

2. Amino acid delivery to skeletal muscle

3. Amino acid uptake by skeletal muscle

mTORC1

p70S6K1 4E-BP1

MAP4K

LAT1

a) Blood flow b) Capillary recruitment c) Microvascular perfusion

Amino acid transporters

Leucine

4. Anabolic cell signaling


Dietary nitrate supplementation

(MYOFIBRILLAR) MUSCLE PROTEIN SYNTHESIS

↑ Nitric oxide availability

↑ insulin delivery

↑ muscle perfusion and capillary action

Rationale behind the potential for nitrates (arginine, citrulline

and sodium nitrate) to enhance the MPS response


Kouw et al. 2016 Am J Phys,

Sodium nitrate and protein coingestion: findings from

pharmacological and physiologically relevant studies

0.00

0.02

0.04

0.06

0.08

*

Fasted Protein Protein +

sodium nitrate

Myofibrillar-MPS

Dillon et al. 2011 Am J Phys,

Faste

d yo

ung

Faste

d old

AA +

SNP y

g

AA +

SNP o

ld

0.00

0.02

0.04

0.06

0.08

0.10

*Mixed-MPS


Maximise anabolic potency Enhance anabolic sensitivity

Daily protein intake Meal-by-meal

protein intake

Prior exercise Coingesting other nutrients

Anti-inflamm.

nutrients Vitamin

D

Optimising the response of MPS

Nitrates Neutra-

ceuticals Alcohol

Take home messages:

Optimising nutrition for older muscles

β-HMB

Creatine Antioxidants Leucine

Ca+ Free acid

Arginine

Citrulline

Sodium nitrate

Phosphatidic acid

Ursolic

acid

Curcumin

Vitamins C and E

N-3 PUFA

Resveratrol


What opportunities exist for industry-academic partnerships

in context of healthy musculoskeletal ageing?

1. Can fish oil supplementation enhance the response of MPS to

protein ingestion in active older adults? Protection against muscle

disuse?

2. Can vitamin D supplementation enhance the response of MPS to

protein ingestion in active older adults?

3. Can the anti-inflammatory and antioxidant nutrients such as

resveratrol and curcumin protect ageing muscles against

sarcopenia?

4. Is there a role for other neutraceutical compounds such as

phosphatidic acid and ursolic acid in reducing the age-related

loss of muscle mass?


26

Acknowledgements


Oliver Witard

Thanks for listening..……

………. any questions?

[email protected]

@OllyWitard

27

mailto:[email protected]





Additional slides

28


29 Report of the European working group on sarcopenia in older people

Cruz-Jentoft 2010 Age and Ageing 39: 412-423.

What criteria should be used to diagnose sarcopenia?

2. Measure grip strength

Normal ♂ ≥ 30 kg

♀ ≥ 20 kg

Low ♂ ≤ 7.25 kg/m2

♀ ≤ 5.67 kg/m2

Normal ♂ ≥ 7.25 kg/m2

♀ ≥ 5.67 kg/m2

No sarcopenia Sarcopenia

1. Measure gait speed

No sarcopenia

Older adult (> 65 years)

3. Measure muscle mass

Low ♂ ≤ 30 kg

♀ ≤ 20 kg

Sarcopenia

≤ 0.8 m/s > 0.8 m/s


30

What are prevalence estimates of sarcopenia?

1. On average, 5-13% of older adults aged 60-70 years are affected by

sarcopenia.

2. On average, 11-50% of older adults aged 80+. Briefly review best

protein nutrition practice for ageing muscles.

3. What about 40 and 50 year olds?


What is mTOR and why should we care in terms of healthy

ageing?

MYOFIBRILLAR PROTEIN SYNTHESIS

mTORC1

p70S6K1 4E-BP1

AMPK

REDD1/2

Akt MAP4K3

↓ energy status Hypoxia

EAA

(Leucine) Exercise

TSC1

TSC2

BNIP-3

Insulin

hVps34

Drummond et al 2009 J Appl Phys


What is the biological rationale for leucine fortification?

MYOFIBRILLAR PROTEIN SYNTHESIS

mTORC1

p70S6K1 4E-BP1

AMPK

REDD1/2

Akt MAP4K3

↓ energy status Hypoxia

EAA

(Leucine) Exercise

TSC1

TSC2

BNIP-3

Insulin

hVps34

Drummond et al. 2009 J Appl Phys


Fortifying a suboptimal protein dose with leucine “rescues”

a submaximal response of muscle protein synthesis

Bukhari et al. 2015 Am J Phys

• Study cohort of older women (~65

years old)

• Acute bout of single-leg resistance

exercise

• Meal-like whey protein dose vs.

leucine enriched suboptimal EAA

dose

WP L-EAA

0.00

0.04

0.08

0.12FedFasted




• Active young (~26 years) men

• Leucine (3.42 g L-leucine + ~400 mL

water)

• HMB (3.42 g free-acid form ß-hydroxyl-ß-

methylbutyrate)

• HMB (3 g calcium form ß-hydroxyl-ß-

methylbutyrate)

Leucine and HMB ingestion stimulates muscle protein

synthesis at rest in young men

Wilkinson et al. 2013 J. Phys.

Wilkinson et al. 2017 J. Phys.

Bas

al

Leuc

ine

HM

B-free

acid

HM

B-calcium

0.00

0.02

0.04

0.06

0.08

0.10 *


Myofibrillar-MPS

• Physically-active young adults

• Rex + EE + HIIT

• 25 g whey protein or 25 g whey protein + 1.5 g/ kg

BM alcohol

• Resting and post-exercise measurements of MPS

Parr et al. 2014 PlosOne

Alcohol consumption impairs the MPS response to protein

ingestion during exercise recovery

Rest PRO ALC+PRO

0.00

0.02

0.04

0.06

0.08

a

b

c


36

LBM (Kg)

Hoffman et al. 2012 J Int Soc Sp Nutr Joy et al. 2014 Nutr and Metab

Phosphatidic

acid Placebo

Pre Post Pre Post

40

50

60

70

80

LBM (Kg) Phosphatidic

acid

Placebo

Pre Post Pre Post

40

50

60

70

80*

Mixed findings from studies examining the ergogenic role

of phosphatidic acid on muscle growth


0 1 2 4-1.00

0.00

1.00

2.00

3.00

4.00

*

Focal adhesion

Kinase (FAK) activity

Week

The incorporation of EPA/DHA into the muscle phospholipid

membrane upregulates anabolic cell signaling

0 1 2 4

0.0

2.0

4.0

6.0

8.0

10.0

Week

EP

A +

DH

A (

% o

f to

tal f

att

y a

cid

s)

a

c

A

a,bb

0 1 2 4

0.0

2.0

4.0

6.0

8.0

10.0

12.0

14.0

Week

n-3

PU

FA

(%

of to

tal fa

tty

ac

ids

)

a a,b

b

c

B

McGlory et al. 2014 Prostaglandins Leukot. Essent Fatty Acids,

Basal Fed Basal Fed

0.00

0.50

1.00

1.50

2.00

2.50

*

mTORC1

phosphorylation

Pre

supplementation

Post

supplementation

Smith et al. 2011 Clinical Science


Fish oil supplementation enhances exercise training

induced strength gains in older women

Rodacki et al. 2012 Am J Clin

• Healthy older (~64 years old) women

• 90 d strength training with or without fish

oil supplementation

• 2 g/day fish oil capsules

• Pre and post measurements of muscle

mass, strength and power

Knee flexor peak torque

(Nm)

Pre Post

0

10

20

30

40Placebo

Fish oil

**

*


Mu

sc

le P

rote

in B

ala

nc

e

Time

Meal (and/or

exercise

Meal (and/or

exercise)

Meal (and/or

exercise)

Redrawn from Phillips et al. 2005 JACN

Area of B > Area of A = Decrease in muscle mass

A A A

B B

+

_

Muscle loss with ageing is underpinned by a negative

net muscle protein balance

B

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Health and Exercise Science Research · Health and Exercise Science Research Citrulline 1. Emphasize the importance of skeletal muscle size and quality for healthy ageing. 2. Briefly