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Repair And Regeneration > Week 12 > Flashcards

Week 12 Flashcards

1
Q

Why do we use animal models?

A
  1. Understand the pathophysiology of spinal cord injury (SCI)
  2. To test new compounds to limit the impact of injury
  3. To find new strategies for treatment
  4. To improve current therapies
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2
Q

How do we model spinal cord injury?

A
  1. In-vitro models of cellular neural tissue trauma

2. In Vivo models of spinal cord injury

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3
Q

What are the usefulness of in vitro models of mechanical injury?

A
  1. Explore mechanisms of cell death
  2. Screening pharmacological agents
  3. Investigating mechanisms of SCI in transgenic mixe
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4
Q

What are most commonly used for in vitro models of mechanical injury?

A
  1. Contusion and compression
  2. Stretch
  3. Transection
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5
Q

What are the main materials used in in-vitro studies?

A
  1. Primary cultures from DRG
  2. Dissociated primary mixed cell cultures
  3. Reconvened primary cell culture
  4. Use organotypic cultures
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6
Q

What are the factors that can influence the success of the culture?

A
  1. Origin of the cells or tissue
  2. Cell plating density
  3. Medium composition
  4. Substrate in which the cells are grown
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7
Q

Example of a contusion of an organotypic spinal cord slice

A

Step 1: laminectomy and extraction of the cord
Step 2: tissue placed on tissue chopper and cut at a thickness of 400 micrometer
Step 3:spinal cord slices are separated in dissecting medium and placed on millipore membrane
Step 4: slices cultured in an incubator at 37 degree Celsius for 7 days
Step 5: injury device (pin drop, made in house) used to injure spinal cord slices

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8
Q

What is an example of a cut injury in a mixed myelinating culture ?

A
  1. Isolate spinal cord injury
  2. Stains across days - stain neurites in the culture
    - use anti-PLP DM20 antibody
  3. Myelination process increases across time in the culture
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9
Q

Cut injury in a mixed myelinating culture

A
  1. Cutting the culture with a scalpel blade
  2. Induces the formation of a reactive astrocytes in the lesion
  3. Cells with a rounded or elongated morphology migrated into the lesion
  4. At post-lesion 10, astrocytes became predominantly elongated in the morphology and strongly expressed GFAP and chondroitin sulfate proteoglycan
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10
Q

What is anti-cs-56 antibody?

A

Inhibitory to axonal regrowth

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11
Q

Main In Vivo models and aims

A

Most use rats or mice and employ lower thoracic injuries

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12
Q

Why is lower thoracic injuries used?

A
  1. Easier to perform
  2. Very easy results
  3. Animals will be paralysed from hind-limb and you can follow recovery from these animals
  4. Less complications
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13
Q

How are contusion and compression done?

A
  1. Have platform on top of the injury

2. Do contusion on dorsal part of spinal cord

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14
Q

Define hemisection

A

Cut half of the spinal cord

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15
Q

Main In-Vivo models and aims

A
  1. The model do NOT attempt to reproduce the event leading to injury
  2. Paralyse animals
  3. SCI is some under highly controlled conditions - sterile conditions
  4. Don’t model accidents because during accidents things aren’t sterile
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16
Q

What is the aims of in-Vivo models?

A

Reproduce:

  1. The pathophysiology of SCI
  2. Main outcomes of SCI
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17
Q

Assessing the behavioural outcome to trauma

A

Assess paralysis in rodents
Assess gait and motor coordination by using BMS scale
Injury was initially symmetrical but the injury progressed and became asymmetrical
BMS scale from 0 (full paralysis) to 9 (proper regain of proper gate)

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18
Q

What is used to support synapse formation and function?

A

Specific formulation of nutrients

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19
Q

In Alzheimer’s disease

A

Memory performance declines as the number of synapses fall

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20
Q

Why is the formulation of nutrients that support the synapses important?

A

It may be useful for the repair of spinal cycle

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21
Q

What can contusion devices model?

A

Different degrees of injury

22
Q

The compression model

A
  1. Balloon inserted within spinal column and inflate progressively with time
  2. Clip compression
  3. Static weight compression
23
Q

What are examples of secondary injury?

A
  1. Loss of neurons/ axons
  2. Demyelination
  3. Inflammation
  4. Reactive Oxidative Damage and the astrocytic glial scar
  5. cyst formation
24
Q

What are examples of Primary Injury?

A
  1. Loss of neurons/axons

2. Demyelination

25
Q

What can a bolus injection of DHA in mice in tail vein, 30 min after spinal cord compression injury counteract?

A

Deleterious effects of early trauma, inflammation, and scar formation and improve functional recovery

26
Q

What is contusion/compression model?

A

A rod of specific weight usually 10g is dropped from a precise height above the surface of cord to induce SCI of defined severity

27
Q

Application of hydrostatic oressure in a sealed vesselor as a pressure pulse

A

Prolonged hydrostatic pressure models are less efficient than when a transient
pressure pulse is applied using a fluid percussion device
• CNS tissue is not deformed significantly under hydrostatic pressure, therefore new
systems have been developed to allow deformation of the tissue in reaction to the
pressure (this is an important mechanism of mechanical injury, seen in clinical
compression trauma)

28
Q

What is transection

A 
A plastic stylet scrapes adherent cells
from a culture dish
• A rotating scribe can be used to increase
throughput
• Injury severity can be controlled by
adding 1 to 6 scribes to vary the
proportion of cells injured
29
Q

What is transection used for?

A

the study of primary axotomy –stab wounds, penetrating skull

fractures, gunshots

30
Q

Dissection of the neuraxis from embyronic day 13.5

A
  1. Myelinating spinal cord culture prepared by enzymatic and mechanical dissocation of tissue from fetuses of time-mated mice
  2. At the appropriate stage of gestation, the pregnant mouse was killed with an overdose of halothane and CO2, followed by cervical dislocation.
  3. The inclusion of the caudal part of the myelencephalon with the spinal cord in the cultures increased the amount of tissue from which cells could be harvested
  4. the myelencephalon extending 2–3 mm rostral to the cervical flexure, and approximately 8–10 mm of spinal cord caudal to the cervical flexure, were removed from the fetus into sterile DMEM
  5. spinal cords were transferred to the dry lid of the Petri dish, minced with a scalpel blade, and then collected into a bijou containing 1 mL of HBSS (calcium/magnesium‐free) for up to six spinal cord
31
Q

BMS score

A
  1. 0 - no hind limb movement
  2. 1-7 - recovery of hind limb joints
  3. 8-9 - recovery of plantar placement
  4. 10-11 Recovery of plantar stepping
  5. 12-14 - Recovery of coordination
  6. 15-18 Recovery of paw position and toe clearance
  7. 19-21 - Trunk stability and tail position improve3
32
Q

What has recent clinical trials demonstrate?

A

The efficacy of Fortasyn connect in Alzheimer’s disease

33
Q

What is Fortsayn connect?

A

Specific multi-nutrient combination containing DHA, EPA, choline, uridine monophosphate, phospholipid and various vitamins

34
Q

Examined the effect of Fortasyn Connect in a rat compression model of spinal cord injury

A

for 4 or 9 weeks following the injury, rats were fed either a control diet or a diet enriched with low, medium, or high doses of Fortasyn® Connect. The medium-dose Fortasyn® Connect-enriched diet showed significant efficacy in locomotor recovery after 9 weeks of supplementation, along with protection of spinal cord tissue (increased neuronal and oligodendrocyte survival, decreased microglial activation, and preserved axonal integrity)

35
Q

What is Fortasyn Connect?

A
  1. A specific formulation of nutrients designed to support synapse formation and function in Alzheimer’s disease
36
Q

What is Souvenaid?

A

A new approach to management of early AD

37
Q

What does Fortasyn Connect contain?

A

Contains all the precursors and supporting nutrient to build new neuronal membranes

  1. CDP-choline pathway
38
Q

What are examples of the precursors?

A
  1. Phospholipid
  2. Choline
  3. Omega 3 fatty acid
  4. b-vitamin
  5. Antioxidant
39
Q

What is Walking trace analysis?

A
  • Record the footprint of animals by putting ink fore paw and hind paw
  • As animal is walking on a white sheet of paper u can get prints of animal and measure the gait parameters from these prints
40
Q

What is cat walk?

A
  • Have a plate which has a green light which cant escape from the plate
  • You place an animal in the array – natural behavior for rodent to walk between walls? – don’t like open spaces
  • Paws in contact with the plate allows light to escape and we can record these prints
  • Have camera – put field in software – can extract 40-55 parameters which can describe the gait of the rodent
41
Q

What is digigait?

A
  1. Extract paw from animal when it walks
  2. Treadmill
  3. Impose a speed for the animal
  4. camera send signal to software - extract and digitise the paw
42
Q

What is the digigait, the catwalk or the footprint based on?

A

The use of central pattern generator

43
Q

What is the beam walking taks?

A
  • When rat is walking onto beams of different thickness to look into the balance
44
Q

What is the staircase test?

A

Looking at skilled falling movements

use staircase field with a pellet and the animal is being asked to retrieve the pellet

45
Q

What can contusion devices model?

A

Different degrees of injury

46
Q

What is balloon compression?

A

1.insertion of a catheter with a small,
inflatable balloon affixed to its end (Figure 2C) into the epidural or
subdural space.

2.The balloon is then filled with saline or air to a fixed
volume that compresses the spinal cord

3.The balloon is deflated after a specific duration

4.Preparation may involve laminectomy at the desired
lesion site, or merely laminotomy at a location caudal to this site.
Injury can be graded by varying the inflation volume, duration of
compression or both

47
Q

What is clip compression?

A

1.The procedure involves performing a
laminectomy at the desired level of the spine

2.The clip is closed at
specific force around the spinal cord, producing an acute injury

3.eft to compress the cord for a select amount of time, usually
for at least 1 min, to produce an injury

4.A force of 50 and 35 g is used
to produce a severe and moderate injury, respectively. The duration of
clip application can also be varied to produce injuries of varying
severity.

48
Q

What is the hemisection miodel?

A
  1. To study the mechanism related to axotomy (acute stage)
  2. To test treatment for regeneration of severed axon (later stages)
  3. complete transection of multiple tracts is necessary to prove axonal regeneration and/or sprouting of damaged axons
    (compared to compensatory plasticity of spared neuronal circuitry)
49
Q

In vitro models

A

preferred method for the study of acute and sub-acute pathophysiology after trauma

50
Q

In vivo model

A

Mimic similar aspect of pathophysiological and clinical development of post trauma

51
Q

What is the clinical SCI often the result of?

A
  1. Sudden impact of the bone structures transmitted to the spinal cord (contusion)
  2. Followed by sustained compression from the dislocation or burst vertebrae
  3. sometimes also transection of axons by penetrating bone fragments

Repair And Regeneration (16 decks)

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