Traditionally, it has been thought that the mammalian central nervous system (CNS)
does not regenerate. Possibly due to the inhibitory extracellular environment post-injury
as well as the limited intrinsic characteristics of adult post-mitotic neurons (Smith
et al., 2015);
Figure 1
. Modulation of several molecular mechanisms promotes some degree of cell survival
and axon regeneration in the adult CNS, but much remains to be elucidated. Of particular
note, activating inflammatory mechanisms has been shown to either promote or limit
axon regeneration in vivo (Filbin, 2006). The optic nerve crush injury model has been
extensively used as a means of studying axon growth in the damaged CNS and has facilitated
significant strides in our understanding of possible mechanisms required to enhance
regeneration of the damaged adult CNS. While this model represents an initial step
in guiding further research into axon regeneration, it is important to note that neurological
conditions, such as stroke, are characterized by significant axonal damage and models
that mimic this axon-specific damage in the brain are currently being explored.
Figure 1
Schematic outlining the impact of damage and regeneration to axons in the central
nervous system.
Neurons in the normal nervous system form circuits and are supported by glial cells
(A). However, the central nervous system is susceptible to both physical damage and
neurodegeneration, leading to neuronal death, including axonal degeneration. The environment
post injury is not permissive to regeneration. The formation of the glial scar creates
a physical barrier blocking axon regeneration at the injury site (B). Therapeutic
agents (e.g., granulocyte-macrophage colony-stimulating factor) may be able to promote
axon regeneration and repair of damaged axons (C).
Stroke is modelled in animals in order to gain a better understanding of the mechanisms
mediating neurodegeneration, which will guide the development of novel therapeutic
approaches. One of the most well characterized models of stroke is the middle cerebral
artery occulsion (MCAO) model. However, there are challenges associated with reproducibility
of this model in animals. The MCAO model results in significant hemipsheric brain
damage, resulting in significant loss of brain tissue in human stroke cases. This
significant loss of brain tissue correlates with high mortality rates in humans and
in mouse models and the investigation of axonal damage mechanisms is understandably
very complex. In addition, the damage induced by MCAO is particularly variable between
operated animals. Conversely, the endothelin-1 (ET-1) model of stroke can be used
to generate a focal and region-specific stroke in animals. Importantly, ET-1 can indeed
target axonal projections, dependent upon location in the brain that the ET-1 is introduced.
Using the ET-1 mouse model, we have demonstrated targeted damage of axonal projections
in the corpus collasum, with associated subcortical white matter (SWM) damage, culminating
in forelimb motor deficits (Theoret et al., 2016). We have also reported that granulocyte-macrophage
colony-stimulating factor (GM-CSF) plays a critical role in attenuating ET-1-induced
motor deficits through activation of the mTOR signalling pathway (Theoret et al.,
2016). However, the ET-1 mouse model of stroke has a number of limitations in terms
of its utility toward evaluating axon regeneration. Given the well established use
of the optic nerve crush injury model in analysis of axon regeneration, this may be
highly amenable to further evaluate the potential regenerative impact of GM-CSF.
The optic nerve has been well-characterized in terms of morphology and functionality.
All the axons in the optic nerve arise from a single neuronal cell population called
the retinal ganglion cells (RGCs) which can easily be experimentally manipulate. Moreover,
because of its accessibility and anatomy, the eye is suitable for effective application
of potential therapeutics such as growth-promoting factors like GM-CSF. Potential
therapeutics can readily diffuse to the retinal ganglion cells without the need to
overcome the blood-retinal barrier. The optic nerve crush model involves the use of
forceps to crush and damage axonal connections in the optic nerve, this can be accompanied
with administration of therapeutic agent(s), such as GM-CSF. Interestingly, and with
particular relevance to the potential application of GM-CSF in brain pathologies,
GM-CSF readily crosses both the blood-brain and blood-spinal barriers (McLay et al.,
1997), making it an attractive therapeutic option for brain injury.
GM-CSF is a pro-inflammatory cytokine, composed of four alpha helices that bind to
the alpha subunit of the GM-CSF transmembrane receptor. The alpha-subunit of the GM-CSF
receptor is specific for binding GM-CSF, whereas the beta-subunit is involved in signal
transduction. GM-CSF is part of the haematopoietin receptor superfamily and plays
a prominent role in promoting proliferation, differentiation and maturation of hematopoietic
precursors cells (Burgess and Metcalf, 1980). GM-CSF also regulates various cellular
processes through the activation of intracellular signalling mechanisms such as MAPK,
PI3K/Akt and JAK/STAT signalling pathways (Schallenberg et al., 2009; Legacy et al.,
2013; Theoret et al., 2016). The binding of GM-CSF to the alpha subunit of the GM-CSF
receptor leads to the phosphorylation of the beta subunit of the receptor by JAK2,
a tyrosine kinase. GM-CSF has been shown to act as a neuronal growth factor and as
a neuroprotective agent both in vitro and in vivo (Schabitz et al., 2008; Legacy et
al., 2013; Hanea et al., 2016; Theoret et al., 2016). The intracellular MAPK-ERK1/2
pathway activation has been implicated in GM-CSF's role in protecting RGCs from cell
death both in vitro and in vivo (Schallenberg et al., 2009). In addition, GM-CSF has
been shown to stimulate neurite growth in retinal explants and cultured RGCs (Legacy
et al., 2013; Hanea et al., 2016). Overall, these studies suggest that GM-CSF might
be a novel therapeutic agent for neural repair following traumatic injury to the CNS.
The binding of GM-CSF to the alpha subunit of the GM-CSF receptor leads to the phosphorylation
of the beta subunit of the receptor by JAK2, a tyrosine kinase. Interestingly, the
expression of GM-CSF alpha-receptor is up-regulated following ischemic brain injury
(Schabitz et al., 2008; Theoret et al., 2016). Furthermore, intravenous injection
of GM-CSF in the MCAO rat-model resulted in significant reduction in infarct volume
(Schabitz et al., 2008). Further research is needed to better understand the role
of GM-CSF on axon regeneration and the intracellular signalling mechanisms that induce
its proposed neuroprotective effects.
Translation of the therapeutic potential of GM-CSF from the laboratory to the clinic
is slowly progressing; for instance, Jim et al. (2012) reported that GM-CSF improves
cognitive function in cancer patients. Although the mechanism by which GM-CSF reduces
cognitive decline is not well understood; it is hypothesized that GM-CSF treatment
may result in increased angiogenesis, neurite outgrowth or neuronal survival in brain
regions known to be involved in cognitive function. Stroke in humans is often associated
with cognitive dysfunction; this is particularly important because depending on location
of a stroke, patients may experience significant cognitive decline. Further research
is necessary to fully elucidate the mechanisms involved and the role of GM-CSF in
post-stroke humans.
These key findings regarding the neuroprotective role of GM-CSF and its ability to
enhance neurite outgrowth suggest that GM-CSF might be an ideal therapeutic agent
following CNS injury. However, there seems to be many downstream intracellular effectors
of GM-CSF in the CNS. There is a critical need to define the intracellular mechanisms
that contribute to the effects of GM-CSF following CNS injury and to clarify whether
there is cross-talk between these mechanisms in order to fully understand the role
of GM-CSF in the CNS. Overall, these studies indicate that GM-CSF may be a novel therapeutic
target to promote regeneration in the injured CNS and further research is necessary
to clarify the impact of this compound on both axon regeneration and plasticity in
the CNS.