Muscle Diseases Techniques 1.1 : Functional assessments to evaluate therapeutic efficacy

Question 1

What is DMD caused by?

Answer 1

Complete lack of dystrophin.

Question 2

Where is dystrophin found?

Answer 2

Around the sarcolemma.

Question 3

Which parts of the body are more affected in DMD?

Answer 3

Lower extremity and torso more affected than upper limbs.

Question 4

What is becker’s disease?

Answer 4

Like DMD but less severe, some dystrophin present.

Question 5

What two structures does dystrophin connect?

Answer 5

Connects muscle fibre to the lamina, disperses forces evenly.

Question 6

What kind of protein can dystrophin act like (name 2`)?

Answer 6

Anchoring protein and possibly signalling.

Question 7

What happens to mice with dystrophin knockouts regarding lifespan?

Answer 7

Die prematurely.

Question 8

What happens to other proteins with dystrophin absence?

Answer 8

Compensatory upregulation of the dstrophin like protein, utrophin.

Question 9

What are mice with upregulated utrophin called?

Answer 9

mdx mice

Question 10

What are mice with a double knockout of dystrophin and utrophin called?

Answer 10

dko mice.

Question 11

Is the muscle an endocrine organ?

Answer 11

Yes.

Question 12

What is the aim of treating DMD?

Answer 12

Improving both muscle size and functionality. Pure size increase is useless.

Question 13

Are muscles from mdx mice bigger than wild type mice? How do their strengths compare?

Answer 13

mdx mice muscles are larger, but they have the same strength.
So overall, mdx mice muscles are weaker and fragile given no dystrophin, and lower cross sectional force.

Question 14

What are some types of non/minimally invasive analysis techniques?

Answer 14

Running, swimming, climbing etc.

Question 15

What is the vertical hang test, and is it non-invasive? Name a pro and con of this.

Answer 15

It is, and is the latency to fall from a hanging wire.
Cant assess specific muscles, is a crude assessment
Is simple and a natural exercise

Question 16

What is the grip strength test, and is it non-invasive? Name a pro and con of this.

Answer 16

It is.
Mouse holds onto a bar, and lifted from its tail by the assessor.
Assesses muscle strength, and simple
Can be inaccurate as assessors hold mice differently, cant assess specific muscles

Question 17

What is the difference between in vitro, in situ and in vivo?

Answer 17

In vitro - isolated from the body - ie cell bath
In situ - Still in the body, as real life as pissble
In vivo - assessing entire muscle group

Question 18

Name an in vitro technique on muscle, a pro and con.

Answer 18

Muscle cut out, hooked to a force gauge.
pro - directly assesses functionality free of neurovascular influence
con-less physiological assessment

Question 19

Name an in situ technique on muscle, a pro and con.

Answer 19

Muscles stimulated individually
pro - preserves neurovascular supply, is more physiological
con - technically difficult, menial to carry out, expensive equipment

Question 20

What is a problem with individual muscle stimulation in situ?

Answer 20

Can be minimally invasive by anaesthetising, but cant do this too often as it might interfere.

Question 21

Name a technique at the cellular level. What can it assess (name 2). Name a pro and con

Answer 21

Single fibres are assessed, hooked to a force gauge.
Assesses Ca2+ release and uptake
Vmax (velocity of shortening)
pro - cellular level assessment, valuable info
con - technically difficult and expensive

Question 22

What happens to CK (creatine kinase) levels in damaged muscles and in DMD?

Answer 22

Elevated, indicating muscle trauma.

Question 23

What does a drug lowering CK levels indicate?

Answer 23

Means its successful in lowering trauma. If CK levels remain, drug most likely isnt working.

Question 24

How can dye be used to assess damage and drug vbiability?

Answer 24

Damaged muscles let in more dye, more damaged is more dyed.

If a drug works, muscles its affecting must be less dyed.

Question 25

How can fibrosis levels be used to assess damage levels and drug viability?

Answer 25

Muscles can be dyed for collagen, more damaged muscles are more fibrotic.
If the drug is working, should mean less dye positive fibres.

Question 26

How can Ca2+ reuptake levels be used to assess damage and drug viability?

Answer 26

Ca2+ reuptake is lowered in damaged fibres. Drug must increase reuptake of Ca2+.

Question 27

What 4 factors indicate a drug is working?

Answer 27

Lowering CK levels
Lowering dye penetrating membrane
Lowering collagen/fibrosis
Improving Ca2+ reuptake

Question 28

How can dystrophin’s ability to disperse forces be tested?

Answer 28

By electrical stimulation after streching. Isometric contraction force decreases with stretch.

Question 29

What can be used to assess membrane integrity?

Answer 29

Dye.

Question 30

What happens when cells are ruptured in situ, regarding Ca2+ levels? Why is this a problem?

Answer 30

Caq2+ enters the cell from the extracellular matrix. Usually kept very low.

Question 31

What can be said of isolated dystrophic muscle fibres regarding architecture?

Answer 31

Are wavy and have strange branches. Branches are weak points especially when stretched.

Question 32

What can be said of the position of the mucei of dystrophic muscle fibres?

Answer 32

They are centrally nucleated, suggesting they are regenerating.

Question 33

Does muscle damage occur when healthy muscles do isometric contraction? What about dystrophic muscles?

Answer 33

No it doesn’t. It does in dystrophic muscles, are inherently fragile, and tear apart.

Question 34

Why do dystrophic muscles tear apart when contracting? What is a possible cause of this (aside from the lack of dystrophin)?

Answer 34

Some parts of the fibre contract, while others relax, tearing it apart. A failure of the beta plate (neuromuscular junction) could cause this.

Question 35

What is the mechanical hypothesis for dystrophic necrosis?

Answer 35

Loss of dystroglycan integrity leads to membrane rupture and protein accumulation.

Question 36

What happens to the force deficit if dystrophic muscles are stretched then stimulated?

Answer 36

Increases.

Question 37

What is the calcium hypothesis for dystrophic necrosis?

Answer 37

Influx of Ca2+ intracellularly overwhelms the cell’s buffer systems.
High Ca2+ levels for too long activates damaging pathways.
Linked to mechanical hypothesis.

Question 38

What are the steps of muscle regeneration, and how is it different in DMD?

Answer 38

Contraction induced damage -> Ca2+ homeostasis lost -> muscle degeneration -> muscle regeneration
Normally regeneration is effective
In DMD, works well initially, but trauma is too rleentless and eventually wanes.

Question 39

What happens if Ca2+ influx is quickly buffered? What happens in DMD?

Answer 39

Damage can be averted, and the muscle fibre preserved.

In DMD, cant be buffered, and once Ca2+ reaches a threshold, damaging pathways are activated.

Question 40

What effect does the Ca2+ influx have on satellite cells?

Answer 40

If it is unbuffered for a periodof time, satellite cells are activated.