Unloading Adaptation• Experimental models of decreased use

– Immobilization– Hindlimb suspension– Spaceflight– (Denervation)

• Factors contributing to atrophy• Clinical consequences of immobilization

Immobilization• Mechanical fixation

– External (cast)– Internal (pins)– Mixed (bone-mounted external clamps)

• Posture• Muscle activity

– Animal models: length-dependent activity– Human/clinical

Fournier study• ‘Residual’ muscle activity depends

on length• Muscle mass preserved at long

length• Reduced activity (short) without

extra atrophy

Lieber study• External Fixator

– Immobilize only one joint– No wiggling

• Quadriceps– Vasti: single joint knee

extensors– Rectus femoris: biarticular KE

and hip flexor

Muscle-specific atrophyVastus Medialis Rectus Femoris

Dark: fastLight: slow

Use and mechanics influence atrophy• RF is relatively spared (biarticular)• Fiber type

– Slow fibers in slow VM sensitive– Fast fibers in fast VL sensitive

Ubiquitin/Proteasome• Predominant pathway for protein degradation• Anti-ribosome• Ubiquitin• Poly-Ub• Proteasome

EM of proteasome

Pollard & Earnshaw, 2008

“Atrogene” signaling• MuRF + Atrogin/MafBx

– Muscle specific E3 ligases– Seem to drive atrophy

Transgenic HSP70 expression reduces immobilization-atrophy Senf & al., 2008

FOXO1/3a

Akt

HSP70

MuRF Atrogin

Protein Degradation

Growth Factors

“Stress”

Unloading• Reduce force, maintain mobility• Spaceflight

– Maintains mobility, decreases ROM– Inertial loading– Rapid loss of bone and muscle

• 6° head-down bed rest– Space-mimetic– Cardiovascular & hemodynamic

• Hindlimb suspension

Space: Loss of function• Rapid loss of strength (20%

3 weeks)• Slower, variable loss of

mass ~15% 5 weeks

Adams & al., 2003

Spaceflight muscle disruption• SLS-1 (1991)• 9 days• 25% atrophy• Expanded

interstitia

Riley & al., 1996

Ground control 9 days SLS-1 + 3h

Spaceflight muscle disruption• Sarcomere disruption• Z-disk streaming

Spaceflight: fiber adaptation• Sandona & al 2012

– Mice Drawer System (MDS)– 91 days on ISS

• Fiber properties• Transcriptional

profiling

Image: NASA

Muscle-specific atrophy• EDL: fast muscle

doesn’t care (much)• Soleus: postural

muscleA few type 2b fibersA few type 1 fibers

No atrophyAtrophy

Spaceflight-induced genes• “Stress Response”

– PERK– HSP70– NFkB

• Atrophy– MuRF– Atrogin

• Channels

Fold induction with 90 day spaceflight

Ubiquitin ligases

6° head-down bedrest• 30-90 days

– Blood draws– Biopsies/scans

• Space-mimetic– Fluid shift– Cardiorespiratory

• Similar magnitude muscle/bone strength loss

Photo: NASA Ames

Muscle atrophy during bedrest• Nitrogen balance

– Net amino acid intake-excretion– Protein accretion

estimate

• Strength loss:selective

Stein & Schulter 1997

Negative nitrogen balanceatrophy

-60

-40

-20

0

20

40

600 5 10 15 20 25

Knee Ext

Knee Flex

Weeks (16 wk bed+recovery)

Ste

rngt

h C

hang

e (%

)

Muscle-specific atrophy• By MRI volume

Miokovic, & al.,2012

Acute ‘atrophy’ with bed rest• 24 hours BR/HDT• 0.5, 2, 5 hour upright• 15% apparent

atrophy overnight• Apparent

hypertrophy inneck muscles

• Full recovery in0.5-2 hours

• Fluid shiftConley & al., 1996

Calf, horizontal

Calf, head-down

Neck, horizontalNeck, head-down

Hindlimb suspension• Rodent model

– Capture tail in low stress mesh/friction tape– Suspend by runner system– Hindlimbs just elevated

• Fluid shift• Unload, esp anti-grav• Stretch flexors

Shimano & Volpon, 2007

ControlPair-fed

Suspended

Suspendedand casted

Time (weeks)

Suspension Atrophy• Young rats (~100g)• Soleus

– 40% atrophy– 100% loss-of-growth– Mass preserved by

casting

• Protein accretion– Control: +13%/-8%/day– Suspended:+11%/-28%

Goldspink & al., 1986

Atrogene signaling during HS• Rat Medial Gastroc

– Rapid muscle mass loss– Preceded by MuRF/MAFbx

• Transgenic MAFbx– Smaller cells

Bodine & al., 2001

Proteolytic systems during HS• Lysosomes

– Acidic, autophagic compartment– Cathepsin proteases

• Calpains– Calcium-activated cytosolic

Taillandier & al 1996Enns & al., 2007

Calpain action during HS• cp mice express calpain

inhibitor• Doesn’t (much) change

loss of mass• Substantial sparing of

force production

Salazaar & al., 2010

Calpain Targets• Structural: Desmin, nebulin, utrophin• Suspension disrupts

sarcomere structure• Calpastatin (cp)

mice retain struct &force capacity

• Calpains ‘release’sarcomere matrix tofacilitate digestion

Salazaar & al., 2010

Summary• Models of decreased use• Atrophy rules

– Immobility, inactivity atrophy– Strength loss precedes mass loss– Large fibers are more sensitive

• Active degradation pathways– Proteasome (MuRF/MAFbx)– Lysosomes (cathepsin)– Calpains (sarcomere stability)