Ways Of Inducing Regrowth

Question 1

In CNS stumps of damaged axons…

Answer 1

Form short neurons eventually die and degenerate

Question 2

Describe the PNS environment

Answer 2

Has a more favourable environment mainly due to Schwann cells that are potent promotors of neurotic outgrowth

Question 3

Describe Schwann cells in re growth

Answer 3

Produce growth promoting factors such as NGF
Contain cell adhesion molecules (laminitin and fibronectin) in their basal laminae (ECM) that promote axon growth
Proliferate and form empty endoneural tubes in which growth cones can act as feelers. These tubes do not exist in the CNS

Question 4

Describe the CNS environment

Answer 4

Less favourable environment, mainly because central myelin is a potent inhibitor of axon outgrowth

Question 5

Oligodendrocytes expresss.

Answer 5

Molecules in the adult that block axon re growth. CNS white matter is selectively inhibitory for axonal growth - thought to be the reason why myelination occurs late in development
Additionally, laminitis and fibronectin are no longer expressed and thus important adhesion molecules are absent from the e generating environment

Question 6

Describe myelin associated inhibitors

Answer 6

Nogo (Nogo-A NI-250)

    Myelin-associated                MAG (myelin-associated glycoprotein)
    Inhibitors (MAIs)  
                                                 OMgp (oligodendrocyte myelin glycoprotein)

Question 7

Describe Nogo

Answer 7

Located in the endoplasmic reticulum of the oligodendrocytes but not in Schwann cells.

It has three isoforms:
- Nogo-A (NI-250) —» unique to oligodendria
- Nogo-B (NI-35) —» absent in myelin
- Nogo-C —» absent in myelin

It has 2 inhibitory domains:
- amino Nogo (only found in Nogo-A)
- Nogo-66

Nogo-A is the most inhibitory because it has 2 inhibitory domains

Question 8

Convergence towards ROCK activation induces…

Answer 8

Actin depolymerisation (inhibiton of axon growth )

Question 9

Describe chondroitin sulphate proteoglycans

Answer 9

Family of molecules with a protein core and negatively charged GAGs attached (electrostatically repellent for growth cones).

Released by hypertrophic, reactive phenotypic astrocytes found in the glial scar.

CSPGs are re-expressed after injury in the brain and spinal cord.

CSPGs bind to many receptors (LAR, PTPsigma, NGR) ultimately activating ROCK resulting in actin depolymerization so inhibition of axon growth.

Question 10

Glial scarring after injury

Answer 10

After injury:

CSPG expression is rapidly upregulated by reactive astrocytes.

This forms an inhibition gradient.

Highest concentration is at the centre of the lesion and it diminishes towards the penumbra of the lesion

Question 11

Summary of the differences in environments on the CNS in a developing, mature and injured neuron.

Answer 11

Both MAIs and CSPGs are involved in regenerative failure.

MAIs are constitutively expressed.

CSPGs are strongly upregulated by injury.

Question 12

Secondary changes occurring after CNS injury

Answer 12

Astrocyte proliferation
Activation of microglia
Formation of a glial scar
Inflammation
Invasion by immune cells
Proliferation of oligodendrocyte precursor cells.

These changes are thought to render the environment inhospitable for regeneration.

Question 13

Intrinsic factors

Answer 13

It has been shown that a central axon can regenerate in a peripheral nerve.

BUT the rate of elongation is still inferior to a peripheral axon that is navigating the same path. …. So, the environment is not the only factor explaining the differences of regeneration between PNS and CNS.

This fact emphasizes a difference between the intrinsic ability of the
peripheral and central nerves themselves.

Explanation:
As opposed as PNS neurons, epigenetic changes leading to neuronal expression of regeneration-associated genes (RAGs) upon axon injury is limited in CNS.
Growth-associated protein 43 (GAP-43) is expressed at high levels in both types of neurons in the embryo. Most adult CNS neurons lose these proteins. However, they are retained throughout life by neurons in the PNS.
High expression of proteins suppressing axonal growth in CNS neurons (PTEN, SOCS3, EFA-6).

Question 14

Why CNS regeneration is poor

Answer 14

1. Extrinsic factors (environment):
   - Low concentration of neurotrophins (no Schwann cells)
   - No endoneural tube formation (no Schwan cells)
   - Myelin is a potent inhibitor of axon growth (constitutively expressed MAIs: NogoA, MAG, OMgp)
   - Formation of glial scar (secretion of CSPGs by reactive astrocytes upregulated by injury)
   - MAIs & CSPGs bind to receptors on CNS axons which activate ROCK —» actin depolymerisation
   - Inflammation
   - Immune reactions
2. Intrinsic factors:
   - Limited epigenetic changes favouring axonal grow upon axon injury (no phenotype change).
   - High expression of proteins inhibiting axon growth.

Question 15

Example of in vivo work to help axonal re growth in CNS

Answer 15

Neural grafts

Question 16

Describe neural grafts

Answer 16

Provide a source of depleted substance.
Stimulate neuron growth and promote survival of neurons.
Replace lost structure in brain and spinal cord.

Question 17

Materials used for neural grafting

Answer 17

Tissue from the foetal CNS
Tissue from the PNS
Peripheral autonomic neurons
Tissue from outside the nervous system
Isolated, cultured, or genetically engineered cells.

Question 18

Describe foetal CNS tissue

Answer 18

Foetal tissue develops and integrates within a host organism following grafting (foetal brain and spinal cord).
Functional improvements in animals with neurological deficits.
Ongoing clinical trials are examining the effectiveness of neural grafting with human foetal tissue for the treatment of people with Parkinson’s disease.
—»The number of foetal nerve cells needed for neural grafting may be of
critical importance, especially when the graft recipient’s brain is relatively large.
—» Identification of the region of the foetal brain required for grafting is difficult.

Question 19

Describe the use of peripheral nerve for graft

Answer 19

If a section of peripheral nerve is attached to the cut end of the optic nerve and its other end was inserted into superior colliculus, then regeneration of the optic nerve occurs:
- Retinal ganglion axon regenerates and reinnervates the superior colliculus.
- New synapses are formed.
- These new synapses are functional (if retinal neurons are activated by light, postsynaptic neurons in the colliculus respond).
- Administration of growth factors during nerve grafts can enhance regeneration

Question 20

Peripheral autonomic tissue

Answer 20

Share common heritage with nervous tissue.
Synthesise neurotransmitters and other chemical substances that are similar to those found in the CNS (adrenal medulla ——» dopamine)
These neurons are easily accessible and exhibits a great potential for regrowth

Question 21

Isolatated and cultured cells

Answer 21

Most cells survive for only a limited period in primary culture, either replicating a limited number of times or not at all.
Some cells can continue to replicate and thus can potentially survive indefinitely.
Sustained self-propagating cells in culture are known as continuous cell lines (CCL).
Experiments have shown that neuronal CCLs can be successfully grafted into the CNS and may attenuate functional problems produced by lesions in the CNS.
But…uncontrolled growth may lead into tumour formation.

Question 22

Genetically engineered cells

Answer 22

Cells may be designed to synthesise a specific chemical substance or to perform a specific function before being implanted into a recipient.
Cells derived from foetal CNS tissue, the prospective host, primary cell cultures, or a CCL can be genetically engineered.
Two types of non-neuronal cells have been used for genetic engineering and neural grafting: astrocytes and fibroblast cells.
Fibroblasts have been genetically engineered to synthesise an enzyme important for the production of the brain chemical L-dopa, which is a precursor of the brain chemical dopamine.
When used for neural grafting, these genetically engineered cells enhanced survival and growth of neurons, improved CNS function, or both.

Question 23

Clinical trials

Answer 23

Infusion trophic factors (neurotrophins NGF, BDNF) into injury site.
Inhibition of RhoA (and consequently ROCK) by Cethrin.
Insertion of conduits rich in matrix molecules (laminin).
Schwann cells grafting into the site of injury.
Administration of antibodies to the central inhibitory environmental factors to neutralise MAIs.
Transplantation of foetal neurons or neural stem cells into site of injury.
Immunosuppression.