Invest Ophthalmol Vis Sci. 2014 Mar 28;55(3):1953-64. doi: 10.1167/iovs.13-13630.
MMPs in the neuroretina and optic nerve: modulators
of glaucoma pathogenesis and repair?
Lies De Groefa, Inge Van Hovea, Eline Dekeystera, Ingeborg Stalmansb, Lieve Moonsa
a
Laboratory of Neural Circuit Development and Regeneration, Animal Physiology and
Neurobiology Section, Department of Biology, KU Leuven, Leuven, Belgium
b
Laboratory of Ophthalmology, Department of Neurosciences, KU Leuven, Leuven, Belgium
Corresponding author:
Prof. Dr. Lieve Moons
Research Group Neural Circuit Development and Regeneration
Animal Physiology and Neurobiology Section
Department of Biology
KU Leuven
Naamsestraat 61, Box 2464
B-3000 Leuven, Belgium
Tel: (32)-16-32.39.91
Fax: (32)-16-32.42.62
Email: lieve.moons@bio.kuleuven.be
1
Abstract
Multiple studies in glaucoma patients and in animal models of spontaneous and
experimentally induced glaucoma, reported changes in the expression and activity of several
matrix metalloproteinases (MMPs) in the retina, optic nerve, aqueous humor and trabecular
meshwork. These data have led to the hypothesis that MMPs might be involved in glaucoma
onset and/or disease progression. However, reports are conflicting and research aiming at
providing a clear definition of their causative role is lacking.
In glaucoma, MMPs are thought to act at two different levels. In the trabecular meshwork,
they fine-tune the aqueous humor outflow rate and intraocular pressure, in the neuroretina and
optic nerve, however, their role during glaucoma disease progression is much less clear. This
review provides a comprehensive overview of the research conducted on the expression and
function of MMPs in the retina and optic nerve, and on the elucidation of their potential
involvement during glaucoma pathogenesis. Additionally, we describe the insecure balance
between detrimental and potential beneficial MMP activities during central nervous system
recovery and how MMP-based therapies could help to overcome the current pitfalls in the
development of retinal ganglion cell neuroprotection and axon regeneration approaches for
the treatment of glaucoma.
2
1. Introduction
1.1. MATRIX METALLOPROTEINASES
At present, more than 20 human matrix metalloproteinases (MMPs) have been identified and
divided into collagenases (MMP-1, -8 and -13), gelatinases (MMP-2 and -9), stromelysins
(MMP-3, -10 and -11), and a heterogeneous group of MMPs (MMP-7, MMP-12, MMP-20,
MMP-26 and MMP-28), based on their preferential substrates. The membrane-type or MTMMPs (MT1- to MT6-MMP) are considered a separate class, regardless of their substrate
preference 1.
All of these zinc (II)-dependent proteases are characterized by a conserved domain structure,
including a catalytic domain, which is shielded by an auto-inhibitory pro-domain to keep the
MMP in an inactive state 2-4. Besides activation of MMPs via cleavage/disruption of this prodomain, also transcriptional, posttranscriptional and epigenetic mechanisms contribute to a
strict control of MMP activity, as do compartmentalization, substrate availability and
inhibition of the MMPs
4, 5
. The latter is ensured by -amongst others- the TIMPs (tissue
inhibitors of MMPs, TIMP1-4), that effectuate a local, reversible inhibition of MMPs in
tissues 2, 5.
MMPs were named after their ability to cleave and remodel the extracellular matrix (ECM),
however, they clearly have a much broader degradome, also comprising proteinases, growth
factors, cytokines, cell surface receptors, cell adhesion molecules, and even DNA repair
enzymes and mediators of apoptosis. Essentially all MMPs have been linked to disease
development, including neurodegenerative disorders such as multiple sclerosis, Huntington’s
disease, Alzheimer’s disease, Parkinson’s disease, etc.
4, 6-8
. Despite their detrimental impact
during central nervous system (CNS) pathology, MMPs are increasingly recognized as
essential players during CNS development and in the healthy adult brain. Indeed, a well3
balanced MMP/TIMP expression has been shown to confer to neurogenesis, neurite
outgrowth, myelinogenesis, angiogenesis, cell survival and adult neural progenitor migration
7-13
. Importantly, these processes do not only occur during development, but also after CNS
injury, thus promoting a reparative role for MMPs during recovery from CNS injury. Taken
together, there is ample evidence corroborating MMPs as fine regulators of CNS physiology
and pathology, and any disturbance of their activity may result in profound implications on
cell-cell and cell-ECM interactions.
1.2. GLAUCOMA
Worldwide, more than 60 million people are affected by glaucomatous optic neuropathies,
making this multifactorial neurodegenerative disease the second most important cause of
irreversible blindness
14
. Glaucomatous neurodegeneration, characterized by progressive
degeneration of the optic nerve and apoptosis of retinal ganglion cell (RGC) somata, is
thought to be inflicted by a combination of genetic predisposition and age-related and
environmental stressors 15, 16. The most prevalent and important risk factor, as well as the sole
target for clinical intervention, is elevated intraocular pressure (IOP). Although many patients
benefit from IOP lowering therapies, some patients continue to lose vision in spite of all
current treatments
17, 18
. Furthermore, despite intensive research efforts, the precise cellular
and molecular events translating elevated IOP and other contributing stressors into
progressive RGC death, and the time course of glaucoma pathogenesis, still remain largely
elusive. Expansion of our knowledge about glaucoma pathogenesis and new insights into
contributing molecules/processes, seem of utmost importance for the development of
innovative therapies that can preserve or restore vision.
The complex etiology of the disease is reflected in the divergent array of glaucoma models,
both genetic and experimentally induced, that are available to study its pathogenesis in
4
laboratory animals
19, 20
. Although each of these models mimics only a certain aspect of the
disease, they do provide valuable insights in the underlying causes and mechanisms leading to
RGC degeneration. While ocular hypertension-induced glaucoma models and optic nerve
crush/transection are representative for the large cohort of patients with axonal damage
inflicted by elevated IOP (or trauma), N-methyl-D-aspartate (NMDA)- and kainic acid (KA)mediated models and ischemia-reperfusion injury represent excitotoxicity-induced RGC
damage and RGC death caused by vascular insufficiency, respectively. Of note, although
covering the variety in initial insults contributing to glaucoma pathogenesis, this richness of
models might also be (partially) responsible for the seemingly contradicting results that are
sometimes emerging from different studies investigating glaucoma pathology.
2. Current knowledge on MMP expression and activities in the
posterior segment of the glaucomatous eye
In many cases, glaucomatous damage to the optic nerve and retina is caused by a pathological
IOP elevation. MMPs are the major matrix degrading enzymes and have been described as
important modulators of IOP in the healthy human eye, functioning in a feedback mechanism
that continuously remodels the trabecular meshwork ECM composition in order to maintain a
stable aqueous humor outflow resistance and IOP
21
. However, IOP homeostasis is only one
aspect of glaucoma in which MMPs are involved. Regardless of the primary cause of the
retinal damage, glaucomatous optic neuropathies share a common endpoint involving
progressive atrophy of the optic nerve and apoptosis of RGC somata. Both in the retina and
optic nerve, MMPs are known to be expressed by various glial as well as neuronal cell types
(Tables 1 and 2). It is therefore very likely that MMPs, besides their role as trabecular ECM
remodelers in the anterior segment of the eye, also contribute to the degenerative events
5
manifesting in the posterior segment.
2.1. MMP EXPRESSION AND ACTIVITIES IN THE GLAUCOMATOUS RETINA
The gelatinases, and notably MMP-9, are by far the best studied MMPs in the retina. This
might be largely due to the fact that gelatin zymography allows straightforward quantification
of gelatinase activity, as such circumventing sensitivity and specificity issues related to antiMMP antibodies required for Western blot. Limited research has also been performed on the
expression of MMP-1, MMP-3, MT1-MMP and TIMP-1 in the healthy and glaucomatous
retina, but data about other MMPs/TIMPs are virtually absent, as apparent from Table 1.
MMP expression in the retina of glaucoma patients
Thus far, no more than 4 studies have investigated MMP localization in the human retina 22-25
(Table 1). These failed to detect any MMP-3 or MMP-9 expression in the healthy retina 24, yet
found MMP-2 to be expressed in RGC somata and their axons 23. MMP-1 was observed both
in nuclear and plexiform layers
outer segments
25
24
, while MT1-MMP was only present in the photoreceptor
and TIMP-2 expression was observed within the inner and outer nuclear
layers of the healthy retina 24. In contrast, TIMP-1 expression, which was undetectable in the
healthy retina, appeared to be upregulated in RGC somata and axons of POAG patients 23.
MMP expression in the retina in animal models of glaucoma
In the retina, like in any other tissue, the status and composition of the ECM is believed to
modulate the synthesis and release of MMPs, and vice versa. This MMP-ECM interplay
affects cell-cell and cell-ECM interactions, which may ultimately determine cell survival
versus death
26
. Several studies in rodent models of glaucoma, induced by ischemia-
reperfusion injury 27-31, NMDA- and KA-mediated excitotoxicity 32, 33, optic nerve transection
6
34
, or ocular hypertension 26, revealed a positive correlation between RGC death and MMP-9
activity in the RGC layer (Table 1), whereas MMP-9 activity negatively correlated with
laminin immunostaining in the inner limiting membrane. Increased MMP-9 activity in the
RGC layer plays a key role in the promotion of RGC death, as it induces degradation of
laminin, resulting in abrogation of integrin-mediated survival signalling pathways and
detachment-induced apoptosis of RGCs (‘anoikis’)
26, 28, 35
(Figure 1). Indeed, MMP-9
deficient mice subjected to an ischemia-reperfusion injury model are protected from RGC
27
death and laminin degradation
, providing the ultimate evidence for MMP-9 as a crucial
regulator of RGC death. In contrast, MMP-2 deficiency did not protect from RGC death after
ischemia-reperfusion injury
27
and MMP-2 activity/expression was reported to remain
unchanged after excitotoxic injury, ischemia-reperfusion injury or optic nerve transection 22, 30,
32-34
. However, the latter is still under debate, as others revealed increased MMP-2 activity
within the first hours post ischemia-reperfusion or excitotoxic injury 31, 33 (Table 1).
Co-localization studies by means of immunohistochemistry confirmed reactive astrocytes as
the major source of MMP-9 expression after retinal ischemia-reperfusion injury or KAmediated excitotoxicity in mice
29, 33
, and excluded endothelial cells, microglia and RGCs 29.
In contrast, in situ zymography pointed out that RGCs exhibit increased MMP-9 activity after
ischemia-reperfusion injury or NMDA-mediated excitotoxicity in rats
28, 32
, while the equine
retina revealed MMP-9 expression localizing to the RGC layer, inner plexiform layer and
photoreceptor segments
24, 36
(Table 1). The cellular source(s) of the observed MMP-9
elevation in the glaucomatous retina thus remain obscure and more research with carefully
validated anti-MMP-9 antibodies is required. For MMP-2, constitutive expression has been
located to many cell types. In mice, MMP-2 was found in RGCs, Müller cells and astrocytes
33
. Also in the monkey retina, MMP-2 has been detected in RGC somata and their axons,
whereas MMP-2-positive astrocytes were only found in the optic nerve head 22. In the equine
7
retina, MMP-2 expression was observed in both the inner and outer segments of the
photoreceptors and in the adjacent outer limiting membrane 36 (Table 1).
Notably, while MMP-9 expression is low in the healthy retina, prominent expression of its
major inhibitor, TIMP-1, is seen in RGC somata and their axons in the nerve fiber layer
26
22, 23,
(Table 1). Moreover, the increase in MMP-9 activity in response to elevated IOP was
repeatedly reported to be accompanied by an increased TIMP-1 expression, which was found
to display a positive correlation with the degree of IOP exposure 26, 32, 37, 38. While constitutive
TIMP-1 expression was suggested to support synaptic plasticity at the RGC terminals and
maintenance of an intact ECM (i.e. by keeping MMP activity within limits) 22, the increase in
TIMP-1 levels might facilitate its neuroprotective effects on RGCs, either via inhibition of
detrimental MMP-9 activity, or via MMP-independent anti-apoptotic actions
26
, as shown
elsewhere in the CNS 39, 40 (Figure 1). Expression of TIMP-2, which seems rather confined to
the outer retina, was reported to remain unchanged 24, 31, 33, 36 (Table 1).
Several theories, that have in common the inflammatory cytokine interleukin-1 (IL-1), exist
about how MMP-9 activity is induced in experimental models of RGC degeneration (Figure
1). Glaucomatous injury in the retina stimulates the production of IL-1β by astrocytes, which
could upregulate MMP-9 synthesis in astrocytes (and/or RGCs)
increases nitric oxide production
30, 33, 41
30
. In addition, IL-1 also
, which may activate extracellular pro-MMP-9 via S-
nitrosylation and oxidation of the enzyme’s cysteine switch
32, 33, 42
. Alternatively, MMP-9
expression might be augmented directly or indirectly, via IL-1 signalling cascades, by
increased retinal glutamate levels, which can be induced by various stimuli, including injury,
ischemia and elevated IOP
26, 33
. Finally, in addition to IL-1, other cytokines such as TNF-α
and TGF-β2 are likely to contribute to the upregulation of MMP expression in the
glaucomatous retinas as well. In rodent models for glaucoma, as well as in human patients,
elevated levels of TNF-α and decreased levels of TGF-β2 have been repeatedly noted in the
8
glaucomatous retina, and these cytokines are well known as, respectively, inducer and
inhibitor of MMP transcription
26, 43-46
. However, in contrast to reported observations in the
optic nerve (head) (see below), a causal relationship between altered TNF-α or TGF-β2
expression and increased MMP transcription has not yet been described in the glaucomatous
retina.
Of note, alternative/additional mechanisms of action of MMP-9, besides anoikis, have never
been excluded. These could comprise a direct contribution of MMP-9 to apoptotic signalling
cascades, MMP-9-dependent cleavage of precursors of neurotoxic proteins, MMP-9-mediated
increases in Ca2+ influx or dysfunctional retrograde transport of cell survival factors
secondary to the loss of ECM attachment 27, 32, 47.
Limited attention has been devoted to expression of other MMPs, i.e. MMP-3 and MT1-MMP,
in the healthy and glaucomatous retina so far. A microarray analysis on the retina of rats
subjected to an ocular hypertension glaucoma model, revealed 3.5-fold upregulated MMP-3
mRNA levels at 35 days post induction of ocular hypertension
37
. Likewise, MMP-3 mRNA
levels were strongly increased after optic nerve crush and optic nerve transection in rats, i.e.
over 50-fold and over 250-fold, respectively 43. The cellular origin of MMP-3 expression has
not yet been discovered. Nevertheless, these data, suggesting MMP-3 to be an important
player during glaucomatous damage, together with the described pro- and anti-apoptotic
actions of MMP-3 in the CNS
48, 49
, indicate that MMP-3 is worthwhile investigating. For
MT1-MMP, knowledge is restricted to expression studies in the healthy retina, where MT1MMP has been reported in the nerve fiber layer of newborn (P0) mice and adult rabbits
50, 51
and in the photoreceptor segments of equine retina36.
Overall, the expression of MMP-2, -3, and -9 in the healthy and glaucomatous retina hints that
MMPs might be important players in retinal health and disease. However, largely due to a
9
lack of functional studies, it is currently impossible to define their exact or causative role.
Whereas MMP-9 has a clear negative impact on RGC survival, the role of MMP-2 in
glaucomatous damage in the retina remains obscure. Although MT1-MMP, MMP-1 and
MMP-3 were shown to be expressed in the retina, the latter even being upregulated upon
axonal damage, we can only speculate about their function. For TIMP-1, a neuroprotective
role in the CNS has been suggested, both via modulation of MMP activity as well as via
MMP-unrelated pathways, but whether a similar function exists in the retina remains currently
elusive.
2.2. MMP EXPRESSION AND ACTIVITIES IN THE GLAUCOMATOUS OPTIC
NERVE (HEAD)
As RGC axons exit the primate (or canine) eye to the optic nerve, their sole support and
protection is the lamina cribrosa, making the optic nerve head (ONH) a fragile site in an
otherwise rigid corneoscleral shell
52
and the major site of impact of the mechanical stress
generated by elevated IOP. The lamina cribrosa is composed of cribriform plates that are
aligned to form channels for the RGC axons, and an ECM with collagenous columns, to
which glial cells (GFAP+ astrocytes and GFAP- lamina cribrosa cells) are anchored by a
basement membrane 53. Upon exposure to excessive mechanical stress, characteristic changes
take place in the lamina cribrosa: the cribriform plates collapse and the glial cells shift their
production of ECM components to those that are characteristic for the glaucomatous
phenotype and increase their secretion of MMPs
22, 53-57
. As a consequence, the ECM is
remodeled, which adversely affects the capacity of the lamina cribrosa to support RGC axons
and predisposes RGCs to axonal compression, arrest of axoplasmic flow and apoptosis 53, 55, 56.
Although the rodent ONH only has a rudimentary lamina cribrosa, it does possess a glial
10
lamina with a collagenous composition and a ultrastructural organisation that closely
resembles that of the primate. Indeed, the ONH still appears to be the site of early pressureinduced optic nerve injury in rodents, and the above mentioned mechanisms of pathological
remodelling seem conserved 58.
MMP expression in the ONH of glaucoma patients
Despite the restricted number of studies investigating in vivo MMP/TIMP expression and
activities in the human glaucomatous optic nerve, we do have some basic indirect insights into
the actions of MMPs that contribute to optic nerve axonal degeneration in glaucoma patients.
Human ONH glia were reported to increase their secretion of cytokines, such as transforming
growth factor-β1 (TGF-β1) 53, 57, 59 and tumor necrosis factor-α (TNF-α) 44, 45, 54, 60, in response
to glaucomatous damage, which may act in an auto- or paracrine way to promote other
stretch-induced reactions, including stimulation of MMP-2 secretion, leading to progressive
ECM remodeling (Figure 2). Indeed, increased immunoreactivity for MMP-2 has been
observed in the lamina cribrosa of glaucomatous patients and in vitro in ONH glial cells
undergoing mechanical stretching, corroborating ONH glial cells -next to the RGC axons- as
the source of MMP-2 53, 54, 57, 60 (Table 2). In addition to MMP-2, also MT1-MMP and TIMP3 were reported to be upregulated in an in vitro culture of human lamina cribrosa cells
exposed to cyclic mechanical stretching 57.
In the healthy human optic nerve, quiescent astrocytes express low levels of MMP-1, -2, and
MT1-MMP, as well as TIMP-1 and -2
23, 60
. In the glaucomatous optic nerve, astrocytes
undergo a transition from a quiescent to a reactive phenotype, which involves increased
expression of MMP-1 and MT1-MMP, changes in cell shape and relocation in the nerve
bundles
23, 60
. The increased expression of MT1-MMP on their plasma membrane was
suggested to allow astrocytes to detach and migrate from the underlying basement membrane,
11
by cleavage of ECM and cell surface adhesion molecules at their migration front and/or by
altering their cytoskeleton (Figure 2). Of note, the membrane-bound MT1-MMP can not only
directly exert these actions, but can also indirectly activate MMP-2 expressed by RGC axons
and ONH glial cells, assuring that MMP-2 is activated at the cell-ECM interface
23
.
Subsequently, MMP-1 is believed to permit migration of astrocytes throughout the ECM of
the lamina cribrosa into the nerve bundles, where MMP-1, if not counterbalanced by TIMP-1,
will continue to degrade the ECM around the axons and consequently interfere with neuronal
survival 23 (Figure 2). Notably, a similar neurotoxic effect of MMP-1, released by astrocytes,
was observed in organotypic rat spinal cord and human brain astrocyte cultures, via
destruction of the ECM and/or activation of cytokines 61.
MMP expression in the OHN in animal models of glaucoma
Several lines of evidence, obtained from research in various animal models of glaucoma,
support the above-mentioned modes of action of MMPs in the glaucomatous ONH. TGF-β1
receptor, MMP-2 and TIMP-1 were shown to be augmented in the optic nerve during the
onset stage in an IOP-induced glaucoma model in rats
62
, and in the healthy monkey optic
nerve, low levels of MMP-1, -2, and MT1-MMP, as well as TIMP-1 and -2, expressed by
quiescent astrocytes, were revealed
22
. In the latter study, Agapova et al. confirmed their
previous work on human samples. In addition, by comparing MMP expression in the optic
nerve of monkeys with ocular hypertension-induced glaucoma to monkeys subjected to optic
nerve transection, they were able to prove that the observed ECM remodeling and elevated
MMP-1 and MT1-MMP expression in the optic nerve during glaucoma, occur in response to
elevated IOP and are not secondary to axonal loss 22.
In addition to these animal studies, Hughes et al. investigated the involvement of MMPs in
Wallerian degeneration induced by ONC in rats and found MMP-9 expression in optic nerve
12
astrocytes, which is in contrast to the expression pattern found in the glaucomatous monkey
ONH, where MMP-9 was expressed by oligodendrocytes
22, 63
. MMP-3, in accordance to
expression studies in monkey, was observed in glial cells and blood vessels and upregulated
in glia upon glaucomatous damage to the optic nerve
22, 63
. MMP-7 and MMP-12, finally, are
both expressed by astrocytes upon optic nerve crush in rats 63 (Table 2).
Overall, even when integrating the findings from human and animal studies, the available data
about the expression of MMPs in the glaucomatous optic nerve are sparse and puzzling (Table
2). There is a general consensus that elevated IOP induces an increase in MMP-1 and MT1MMP expression levels, which might in turn negatively impact on the optic nerve
microenvironment. However, reports disagree on the effect of ocular hypertension-induced
mechanical stretching on MMP-2 and TIMP expression in the optic nerve head: while some
describe an increase in MMP-2 and TIMP-1 levels
expression of MMP-2, TIMP-1 and TIMP-2
22, 23, 63
53, 57, 60, 62
, others note constitutive
, or even decreased levels of TIMP-1 and
TIMP-2 due to axon loss 22.
3. Can MMP modulation facilitate neuroprotective and
regenerative glaucoma therapies?
Current glaucoma therapies are all directed towards a sustained reduction of IOP, and slow
down or halt glaucomatous disease progression in many but not all patients 17, 18. Nonetheless,
glaucoma typically leads to irreversible loss of vision because of the inability of adult
mammals to repair or regenerate damaged RGCs. An effective treatment for glaucoma, able to
restore vision, would involve neuroprotection and/or replacement of damaged RGCs in the
retina, as well as induction of their axonal regenerative capacity in the optic nerve 64. Here, we
13
review current evidence for a carefully balanced MMP activity as crucial modulator of the
receptiveness of the retinal and optic nerve extracellular environment to neuroprotection,
stimulation of axon regeneration and cell transplantation therapy.
3.1. CAN MMPS MODULATE RGC NEUROPROTECTION IN THE RETINA?
MMP-9 deficiency was already mentioned earlier in this review for its neuroprotective effects
in a rat ischemia-reperfusion model for glaucoma
27
(Figure 1). In retinal excitotoxicity
models in rats and mice, the broad-spectrum MMP inhibitor GM6001 prevented pathological
remodeling of the inner limiting membrane and detachment-induced apoptosis of the RGCs,
confirming that MMPs, most likely MMP-9, indeed contribute to RGC death 28, 32, 33. Specific
MMP-9 inhibitors, or agents acting on downstream targets in the MMP-9-induced RGC
apoptotic pathway, might thus serve as neuroprotective agents in retinal neurodegenerative
diseases, including glaucoma, retinal artery/vein occlusion and ischemic optic neuropathy
32
.
Also TIMP-1 expression has been shown to be upregulated in the retina of rats exposed to
ocular hypertension26,
37, 38
. This TIMP-1 elevation is likely to reflect the canonical co-
regulation of MMPs and TIMPs, designed to keep pro-apoptotic MMP-9 activity within limits
26
, but, in addition, TIMP-1 might also promote RGC survival by directly suppressing
apoptosis signalling pathways, in an MMP-independent manner 26, 39, 40 (Figure 1).
3.2. CAN MMPS MODULATE AXONAL REGENERATION IN THE OPTIC NERVE?
Upon injury in the mammalian CNS, the ECM transforms from a growth-permissive to an
inhibitory environment, thereby largely preventing CNS neurons from regenerating axons to
their former targets. This failure of the mammalian CNS to regenerate axonal projections is
multifactorial. In addition to the diminished intrinsic growth capacity of adult CNS neurons 65,
14
the injured CNS becomes deprived of neurotrophic factors and/or their receptors 66, and glial
scar- and myelin-derived inhibitory molecules form a physicochemical barrier for
regenerating axons 67, 68.
MMPs have been suggested as key facilitators of successful axonal regeneration for several
reasons (Figure 3). First of all, MMPs add to the clearance of cellular and matrix debris at the
site of injury. Second, all major constituents of the glial scar are substrates of at least one
MMP
68, 69
and increased MMP activity reduces glial scarring and associated inhibitory
molecules. In particular, the chondroitin sulphate proteoglycans (CSPGs) have been denoted
for their strong inhibitory impact on axonal regeneration
67
and are subject to cleavage by
MMPs. Indeed, the repulsion of neurite growth cones and ‘masking’ of neurite-promoting
laminin by CSPGs, can be (partially) abrogated by MMPs 68, 70, 71. Third, MMPs are also able
to degrade myelin-derived inhibitory ligands released by degenerating CNS axons, such as
neurite outgrowth inhibitor (Nogo), myelin associated glycoprotein (MAG), oligodendrocyte
myelin glycoprotein (OMgp) and myelin basic protein (MBP)
8, 67, 68, 72, 73
. Forth, besides
disarming inhibitory ECM and signalling molecules, MMPs can also unmask, activate or
release others that have a beneficial effect on CNS repair. As such, they can indirectly provide
neurotrophic support to regenerating axons via the release of sequestered growth factors (e.g.
release of basic fibroblast growth factor (bFGF) from ECM heparin sulphate proteoglycans)
or the conversion of inactive pro-growth factors to their active forms (e.g. nerve growth factor
(NGF), brain-derived neurotrophic factor (BDNF))
8, 68
. Finally, as the directed outgrowth of
axons requires extensive motility and infiltration within the nervous tissue, MMPs can invoke
‘focalised’ proteolysis at the growth cone
74
thereby reorganizing the ECM to facilitate
attachment and motility of the growing axon 75 (Figure 3).
Although thus far only described in the brain and spinal cord, it can be assumed that these
MMP actions can be extrapolated to the entire CNS, including the retina and optic nerve.
15
Indeed, stimulation of axonal regeneration after optic nerve transection in rats, by intravitreal
transplantation of a peripheral nerve segment, coincides with significantly enhanced MMP
activity in the regenerating optic nerve as compared to the non-regenerating optic nerve
67
.
More specifically, MMP-2 and MMP-9, and to a lesser extent also MMP-1 and MMP-3, are
upregulated in the proximal optic nerve stump and at the site of the optic nerve injury, while
TIMP-1 and -2 are suppressed. This immediate local upregulation (coinciding with post injury
inflammation) of MMP-2 and -9 by reactive astrocytes is thought to clear tissue debris, in
concert with invading macrophages (Figure 3). The later burst in gelatinase activity (i.e. 8
days post injury), which co-localizes with ECM compounds of the glial scar, might serve to
break down and dissolve the glial scar, thereby clearing the path for regenerating axons
67, 76
(Figure 3). Remarkably, gelatinase activity only rarely co-localizes with regenerating optic
nerve axons, in contrast to regenerating peripheral nerves, where growth cones secrete MMP2 (and MMP-3) to facilitate the progression of dorsal root ganglia cell axons on the peripheral
nerve basal lamina 74, 77. Instead, reactive astrocytes at the site of injury appear to be the major
source of gelatinases in the CNS 67, 76. Ahmed et al. hypothesized that the numerous collateral
sprouts in the proximal stump of the transected optic nerve and the growth cones at the front
of the regenerating axons, release a -thus far unknown- factor that conditions the surrounding
astrocytes to increase MMP and downregulate TIMP expression
67
(Figure 3). The result is a
resolution of glial- and myelin-derived growth inhibitory ligands, and RGC axon regeneration
is less hampered
67
. Also other experimental approaches that succeeded to induce axonal
regeneration in the optic nerve, attributed at least part of their success to MMP-related
activities. Transplantation of neural tube-derived chicken embryonic stem cells at the site of
optic nerve transection in rats induced the activation of MMP-2 and MT1-MMP in optic nerve
astrocytes and resulted in regrowth of axons across the lesion site due to degradation of
CSPGs
16
78
. Similarly, transplantation of olfactory ensheating cells was reported to promote
neurite outgrowth in vitro in a rat retinal cell culture, via secretion of MMP-2 and degradation
of CSPGs
79
. Moreover, endogenous MMP-2, produced by retinal astrocytes, seems to be
essential to neurite outgrowth in an in vitro mouse retinal explant model 80.
Taken together, MMPs can potentially play part in many different aspects of successful
axonal regeneration. In the optic nerve, it is already evident that they can add to the
conversion of the nerve environment from a repressive to a facilitative substratum for axon
growth. However, many other aspects of RGC axon regeneration to which MMPs could
theoretically contribute, remain unexplored. Recently, substantial progress was made in
initiating robust axonal regeneration of RGCs in rodents via induction of controlled ocular
inflammation and PTEN (phosphatase and tensin homolog) deletion
81-85
. Given these new
insights, it might be a good time to re-assess the potential involvement of MMPs in these
models of RGC axonal regeneration.
3.3. CAN MMPS MODULATE CELL THERAPY IN THE RETINA?
Besides the prevention of neurite outgrowth, glial scar-derived growth-inhibitory molecules
also hamper successful cell transplantation therapies by blocking donor-host integration in the
retina
86-89
. However, increased MMP expression and subsequent decreased deposition of
inhibitory ECM molecules in the retina, was reported to create a more permissive
environment for regeneration and cell integration. Indeed, elevated expression of MMP-2, -9
and MT1-MMP in the mouse retina, decreases glial barrier formation at the outer limiting
membrane and facilitates incorporation of grafted photoreceptor sheets
89
. Accordingly,
controlled release of MMP-2 directly at the site of injury, enhances progenitor cell integration
and retinal repopulation after subretinal transplantation in rodents
86, 90-93
. Unfortunately,
focus has been on cell therapies for the ‘outer’ retina, i.e. for photoreceptor degeneration 94-98,
and little evidence has been provided thus far that intravitreally transplanted stem/progenitor
17
cells can survive and integrate in the ‘inner’ retina.
Of note, transplantation of progenitor/stem cells in the inner retina could be used to replace
lost RGCs and ultimately, but rather unlikely, to rewire the entire complex retinotopic circuit.
But, alternatively, progenitor/stem cells could also be delivered to provide neuroprotection to
the remainder RGCs or to transform the host retina into a state that is more permissive to
endogenous attempts of repair.
Anyway, regardless of the envisioned strategy, the improvement of the integration efficiency
of the transplanted cells is one of the many challenges for the successful development of cell
therapy in the glaucomatous retina
87
. Nonetheless, given the modest successes with human
Müller stem cell and rat oligodendrocyte precursor cell transplantations in rat glaucoma
models 99, 100, the recent insights in the nature of the barriers to inner retinal engraftment
87, 88
and the lessons learned from research in the outer retina, it might be worthwhile to explore
whether MMPs could be facilitators of host-donor integration in the inner retina.
4. Concluding remarks and future directions
Overall, our knowledge on the involvement of MMPs during the onset and disease
progression of glaucoma is limited. While MMPs are recognized beneficial players in IOP
homeostasis in the anterior segment of the eye
21
, we are still in the dark about their nature
and the processes they contribute to, in the posterior segment. In the optic nerve, MMP-1 and
MT1-MMP are believed to be involved in the onset of glaucoma, and upregulated MMP-3
and -9 expression in the retina hints that MMPs might be important players in glaucoma
pathogenesis as well. However, because of the shortage of functional studies, it is currently
impossible to define their exact or causative role.
Nevertheless, the scientific community progresses to understand the complex spatial and
temporal regulation of beneficial and detrimental effects of MMPs during and after CNS
18
injury and is starting to explore the potential of MMP-based approaches to facilitate CNS
repair. In the retina and optic nerve, in particular the gelatinases are emerging as modulators
of the receptiveness to neuroprotection, stimulation of axon regeneration and cell
transplantation therapy. However, more in-depth studies of MMP activity in time and space
are imperative to disentangle the complex ‘protease web’ of which they are part and to
identify the myriad of functions fulfilled by MMPs 101. Such studies should consist of systemwide, in vivo approaches, including conditional and/or cell-specific MMP gene deletion,
MMP downregulation via RNAi, highly specific inhibitors or blocking antibodies/nanobodies,
or alternatively, MMP upregulation via cell-specific viral vector-mediated gene delivery or
recombinant MMP, all in combination with high-content proteomic profiling techniques.
Indeed, essential to the design of future MMP-based therapies, is the identification of direct
MMP substrates and downstream pathways underlying time and space restricted MMP
functions. This can be achieved by means of degradomics studies after genetic or
pharmacological MMP inhibition, mice deficient in particular MMP substrates, in vitro
cleavage, cleavage site protection/mutagenesis or yeast-two-hybrid assays. These approaches
should point out MMP drug targets exacerbating pathology as well as anti-targets that provide
protection or resolution, and will not only allow the development of a novel generation of
highly specific MMP inhibitors and MMP inducers/activators, but more likely, they will result
in alternative therapies targeting upstream or downstream factors of MMP-modulated
networks.
19
Conflicts of interest statement
The authors have no conflict of interest.
Acknowledgements
The authors acknowledge the Research Foundation Flanders (FWO-Vlaanderen, Belgium,
G.05311.10) and the KU Leuven Research Council (KU Leuven, Belgium, BOF-OT/10/033).
Lies De Groef is a research fellow of the Flemish government agency for Innovation by
Science and Technology (IWT-Vlaanderen, Belgium), Eline Dekeyster is a research fellow
and Ingeborg Stalmans is a senior clinical research fellow of FWO-Vlaanderen.
20
References
1.
Klein T, Bischoff R. Physiology and pathophysiology of matrix metalloproteases.
Amino Acids 2010.
2.
Ethell IM, Ethell DW. Matrix metalloproteinases in brain development and
remodeling: synaptic functions and targets. J Neurosci Res 2007;85:2813-2823.
3.
Page-McCaw A, Ewald AJ, Werb Z. Matrix metalloproteinases and the regulation of
tissue remodelling. Nat Rev Mol Cell Biol 2007;8:221-233.
4.
Rosenberg GA. Matrix metalloproteinases and their multiple roles in
neurodegenerative diseases. Lancet Neurol 2009;8:205-216.
5.
Sternlicht MD, Werb Z. How matrix metalloproteinases regulate cell behavior. Annu
Rev Cell Dev Biol 2001;17:463-516.
6.
Miller JP, Holcomb J, Al-Ramahi I, et al. Matrix metalloproteinases are modifiers of
huntingtin proteolysis and toxicity in Huntington's disease. Neuron 2010;67:199-212.
7.
Rivera S, Khrestchatisky M, Kaczmarek L, Rosenberg GA, Jaworski DM. Metzincin
proteases and their inhibitors: foes or friends in nervous system physiology? J Neurosci
2010;30:15337-15357.
8.
Yong VW. Metalloproteinases: mediators of pathology and regeneration in the CNS.
Nat Rev Neurosci 2005;6:931-944.
9.
Fujioka H, Dairyo Y, Yasunaga K, Emoto K. Neural functions of matrix
metalloproteinases: plasticity, neurogenesis, and disease. Biochem Res Int 2012;2012:789083.
10.
Milward EA, Fitzsimmons C, Szklarczyk A, Conant K. The matrix metalloproteinases
and CNS plasticity: an overview. J Neuroimmunol 2007;187:9-19.
11.
Tonti GA, Mannello F, Cacci E, Biagioni S. Neural stem cells at the crossroads:
MMPs may tell the way. Int J Dev Biol 2009;53:1-17.
12.
Zhang H, Adwanikar H, Werb Z, Noble-Haeusslein LJ. Matrix metalloproteinases and
neurotrauma: evolving roles in injury and reparative processes. Neuroscientist 2010;16:156170.
13.
Verslegers M, Lemmens K, Van Hove I, Moons L. Matrix metalloproteinase-2 and -9
as promising benefactors in development, plasticity and repair of the nervous system. Prog
Neurobiol 2013;105:60-78.
14.
Quigley HA, Broman AT. The number of people with glaucoma worldwide in 2010
and 2020. Br J Ophthalmol 2006;90:262-267.
15.
Calkins DJ, Horner PJ. The cell and molecular biology of glaucoma: axonopathy and
the brain. Invest Ophthalmol Vis Sci 2012;53:2482-2484.
16.
Leske MC, Heijl A, Hyman L, Bengtsson B, Dong L, Yang Z. Predictors of long-term
progression in the early manifest glaucoma trial. Ophthalmology 2007;114:1965-1972.
17.
Heijl A, Leske MC, Bengtsson B, Hyman L, Hussein M. Reduction of intraocular
pressure and glaucoma progression: results from the Early Manifest Glaucoma Trial. Arch
Ophthalmol 2002;120:1268-1279.
18.
Iwase A, Suzuki Y, Araie M, et al. The prevalence of primary open-angle glaucoma in
Japanese: the Tajimi Study. Ophthalmology 2004;111:1641-1648.
19.
Bouhenni RA, Dunmire J, Sewell A, Edward DP. Animal models of glaucoma. J
Biomed Biotechnol 2012;2012:692609.
20.
Johnson TV, Tomarev SI. Rodent models of glaucoma. Brain Research Bulletin
2010;81:349-358.
21.
De Groef L, Van Hove I, Dekeyster E, Stalmans I, Moons L. MMPs in the Trabecular
Meshwork: Promising Targets for Future Glaucoma Therapies? Investigative Ophthalmology
& Visual Science 2013;54:7756-7763.
21
22.
Agapova OA, Kaufman PL, Lucarelli MJ, Gabelt BT, Hernandez MR. Differential
expression of matrix metalloproteinases in monkey eyes with experimental glaucoma or optic
nerve transection. Brain Res 2003;967:132-143.
23.
Agapova OA, Ricard CS, Salvador-Silva M, Hernandez MR. Expression of matrix
metalloproteinases and tissue inhibitors of metalloproteinases in human optic nerve head
astrocytes. Glia 2001;33:205-216.
24.
Webster L, Chignell AH, Limb GA. Predominance of MMP-1 and MMP-2 in
epiretinal and subretinal membranes of proliferative vitreoretinopathy. Exp Eye Res
1999;68:91-98.
25.
Smine A, Plantner JJ. Membrane type-1 matrix metalloproteinase in human ocular
tissues. Curr Eye Res 1997;16:925-929.
26.
Guo L, Moss SE, Alexander RA, Ali RR, Fitzke FW, Cordeiro MF. Retinal ganglion
cell apoptosis in glaucoma is related to intraocular pressure and IOP-induced effects on
extracellular matrix. Invest Ophthalmol Vis Sci 2005;46:175-182.
27.
Chintala SK, Zhang X, Austin JS, Fini ME. Deficiency in matrix metalloproteinase
gelatinase B (MMP-9) protects against retinal ganglion cell death after optic nerve ligation. J
Biol Chem 2002;277:47461-47468.
28.
Santos ARC, Corredor RG, Obeso BA, et al. Beta1 Integrin-Focal Adhesion Kinase
(FAK) Signaling Modulates Retinal Ganglion Cell (RGC) Survival. PLoS One 2012;7:e48332.
29.
Zhang X, Cheng M, Chintala SK. Optic nerve ligation leads to astrocyte-associated
matrix metalloproteinase-9 induction in the mouse retina. Neurosci Lett 2004;356:140-144.
30.
Zhang X, Chintala SK. Influence of interleukin-1 beta induction and mitogen-activated
protein kinase phosphorylation on optic nerve ligation-induced matrix metalloproteinase-9
activation in the retina. Experimental Eye Research 2004;78:849-860.
31.
Zhang X, Sakamoto T, Hata Y, et al. Expression of matrix metalloproteinases and
their inhibitors in experimental retinal ischemia-reperfusion injury in rats. Exp Eye Res
2002;74:577-584.
32.
Manabe S, Gu Z, Lipton SA. Activation of matrix metalloproteinase-9 via neuronal
nitric oxide synthase contributes to NMDA-induced retinal ganglion cell death. Invest
Ophthalmol Vis Sci 2005;46:4747-4753.
33.
Zhang X, Cheng M, Chintala SK. Kainic acid-mediated upregulation of matrix
metalloproteinase-9 promotes retinal degeneration. Invest Ophthalmol Vis Sci 2004;45:23742383.
34.
Sun MH, Chen KJ, Tsao YP, et al. Down-regulation of matrix metalloproteinase-9 by
pyrrolidine dithiocarbamate prevented retinal ganglion cell death after transection of optic
nerve in rats. Curr Eye Res 2011;36:1053-1063.
35.
Halfter W, Willem M, Mayer U. Basement Membrane-Dependent Survival of Retinal
Ganglion Cells. Investigative Ophthalmology & Visual Science 2005;46:1000-1009.
36.
Hofmaier F, Hauck SM, Amann B, Degroote RL, Deeg CA. Changes in matrix
metalloproteinase network in spontaneous autoimmune uveitis model. Invest Ophthalmol Vis
Sci 2011.
37.
Ahmed F, Brown KM, Stephan DA, Morrison JC, Johnson EC, Tomarev SI.
Microarray analysis of changes in mRNA levels in the rat retina after experimental elevation
of intraocular pressure. Invest Ophthalmol Vis Sci 2004;45:1247-1258.
38.
Guo Y, Cepurna WO, Dyck JA, Doser TA, Johnson EC, Morrison JC. Retinal cell
responses to elevated intraocular pressure: a gene array comparison between the whole retina
and retinal ganglion cell layer. Invest Ophthalmol Vis Sci 2010;51:3003-3018.
39.
Magnoni S, Baker A, Thomson S, et al. Neuroprotective effect of adenoviral-mediated
gene transfer of TIMP-1 and -2 in ischemic brain injury. Gene Ther 2007;14:621-625.
40.
Tan HK, Heywood D, Ralph GS, Bienemann A, Baker AH, Uney JB. Tissue inhibitor
22
of metalloproteinase 1 inhibits excitotoxic cell death in neurons. Molecular and Cellular
Neuroscience 2003;22:98-106.
41.
Kido N, Inatani M, Honjo M, et al. Dual effects of interleukin-1beta on N-methyl-Daspartate-induced retinal neuronal death in rat eyes. Brain Res 2001;910:153-162.
42.
Gu Z, Kaul M, Yan B, et al. S-Nitrosylation of Matrix Metalloproteinases: Signaling
Pathway to Neuronal Cell Death. Science 2002;297:1186-1190.
43.
Agudo M, Perez-Marin MC, Lonngren U, et al. Time course profiling of the retinal
transcriptome after optic nerve transection and optic nerve crush. Mol Vis 2008;14:1050-1063.
44.
Tezel Gln, Li LY, Patil RV, Wax MB. TNF-α and TNF-α Receptor-1 in the Retina
of Normal and Glaucomatous Eyes. Investigative Ophthalmology & Visual Science
2001;42:1787-1794.
45.
Tezel Gln, Carlo Nucci LCNNO, Giacinto B. TNF-α signaling in glaucomatous
neurodegeneration. Progress in Brain Research: Elsevier; 2008:409-421.
46.
Nakazawa T, Nakazawa C, Matsubara A, et al. Tumor necrosis factor-alpha mediates
oligodendrocyte death and delayed retinal ganglion cell loss in a mouse model of glaucoma. J
Neurosci 2006;26:12633-12641.
47.
Chintala SK. The emerging role of proteases in retinal ganglion cell death. Exp Eye
Res 2006;82:5-12.
48.
Kim EM, Hwang O. Role of matrix metalloproteinase-3 in neurodegeneration. J
Neurochem 2011;116:22-32.
49.
Kim E-M, Shin E-J, Choi JH, et al. Matrix Metalloproteinase-3 Is Increased and
Participates in Neuronal Apoptotic Signaling Downstream of Caspase-12 during Endoplasmic
Reticulum Stress. Journal of Biological Chemistry 2010;285:16444-16452.
50.
Gariano RF, Hu D, Helms J. Expression of angiogenesis-related genes during retinal
development. Gene Expr Patterns 2006;6:187-192.
51.
Takano A, Hirata A, Inomata Y, et al. Intravitreal plasmin injection activates
endogenous matrix metalloproteinase-2 in rabbit and human vitreous. Am J Ophthalmol
2005;140:654-660.
52.
Sigal IA, Ethier CR. Biomechanics of the optic nerve head. Exp Eye Res 2009;88:799807.
53.
Kirwan RP, Crean JK, Fenerty CH, Clark AF, O'Brien CJ. Effect of cyclical
mechanical stretch and exogenous transforming growth factor-beta1 on matrix
metalloproteinase-2 activity in lamina cribrosa cells from the human optic nerve head. J
Glaucoma 2004;13:327-334.
54.
Akhter N, Nix M, Abdul Y, Singh S, Husain S. Delta-Opioid Receptors Attenuate
TNF-alpha-Induced MMP-2 Secretion from Human ONH Astrocytes. Invest Ophthalmol Vis
Sci 2013.
55.
Dahlmann-Noor AH, Vijay S, Limb GA, Khaw PT. Strategies for optic nerve rescue
and regeneration in glaucoma and other optic neuropathies. Drug Discov Today 2010;15:287299.
56.
Hernandez MR. The optic nerve head in glaucoma: role of astrocytes in tissue
remodeling. Progress in Retinal and Eye Research 2000;19:297-321.
57.
Quill B, Docherty NG, Clark AF, O'Brien CJ. The Effect of Graded Cyclic Stretching
on Extracellular Matrix–Related Gene Expression Profiles in Cultured Primary Human
Lamina Cribrosa Cells. Investigative Ophthalmology & Visual Science 2011;52:1908-1915.
58.
Morrison JC, Cepurna Ying Guo WO, Johnson EC. Pathophysiology of human
glaucomatous optic nerve damage: Insights from rodent models of glaucoma. Experimental
Eye Research 2011;93:156-164.
59.
Zode GS, Sethi A, Brun-Zinkernagel AM, Chang IF, Clark AF, Wordinger RJ.
Transforming growth factor-beta2 increases extracellular matrix proteins in optic nerve head
23
cells via activation of the Smad signaling pathway. Mol Vis 2011;17:1745-1758.
60.
Yan X, Tezel G, Wax MB, Edward DP. Matrix metalloproteinases and tumor necrosis
factor alpha in glaucomatous optic nerve head. Arch Ophthalmol 2000;118:666-673.
61.
Vos CM, Sjulson L, Nath A, et al. Cytotoxicity by matrix metalloprotease-1 in
organotypic spinal cord and dissociated neuronal cultures. Exp Neurol 2000;163:324-330.
62.
Johnson EC, Jia L, Cepurna WO, Doser TA, Morrison JC. Global Changes in Optic
Nerve Head Gene Expression after Exposure to Elevated Intraocular Pressure in a Rat
Glaucoma Model. Investigative Ophthalmology & Visual Science 2007;48:3161-3177.
63.
Hughes PM, Wells GM, Perry VH, Brown MC, Miller KM. Comparison of matrix
metalloproteinase expression during Wallerian degeneration in the central and peripheral
nervous systems. Neuroscience 2002;113:273-287.
64.
Bull ND, Martin KR. Optic nerve restoration: new perspectives. J Glaucoma
2007;16:506-511.
65.
Yang P, Yang Z. Enhancing intrinsic growth capacity promotes adult CNS
regeneration. Journal of the Neurological Sciences 2012;312:1-6.
66.
Goldberg JL, Barres BA. The Relationship between Neuronal Survival and
Regeneration. Annual Review of Neuroscience 2000;23:579-612.
67.
Ahmed Z, Dent RG, Leadbeater WE, Smith C, Berry M, Logan A. Matrix
metalloproteases: degradation of the inhibitory environment of the transected optic nerve and
the scar by regenerating axons. Mol Cell Neurosci 2005;28:64-78.
68.
Pizzi MA, Crowe MJ. Matrix metalloproteinases and proteoglycans in axonal
regeneration. Exp Neurol 2007;204:496-511. Epub 2006 Dec 2020.
69.
Muir EM, Adcock KH, Morgenstern DA, et al. Matrix metalloproteases and their
inhibitors are produced by overlapping populations of activated astrocytes. Molecular Brain
Research 2002;100:103-117.
70.
Cua RC, Lau LW, Keough MB, Midha R, Apte SS, Yong VW. Overcoming neuriteinhibitory chondroitin sulfate proteoglycans in the astrocyte matrix. Glia 2013;NA.
71.
Ferguson TA, Muir D. MMP-2 and MMP-9 increase the neurite-promoting potential
of schwann cell basal laminae and are upregulated in degenerated nerve. Mol Cell Neurosci
2000;16:157-167.
72.
Chantry A, Gregson N, Glynn P. Degradation of myelin basic protein by a membraneassociated metalloprotease: neural distribution of the enzyme. Neurochem Res 1992;17:861867.
73.
Gijbels K, Proost P, Masure S, Carton H, Billiau A, Opdenakker G. Gelatinase B is
present in the cerebrospinal fluid during experimental autoimmune encephalomyelitis and
cleaves myelin basic protein. J Neurosci Res 1993;36:432-440.
74.
Zuo J, Ferguson TA, Hernandez YJ, Stetler-Stevenson WG, Muir D. Neuronal matrix
metalloproteinase-2 degrades and inactivates a neurite-inhibiting chondroitin sulfate
proteoglycan. J Neurosci 1998;18:5203-5211.
75.
Murphy G, Gavrilovic J. Proteolysis and cell migration: creating a path? Curr Opin
Cell Biol 1999;11:614-621.
76.
Duchossoy Y, Arnaud S, Feldblum S. Matrix metalloproteinases: potential therapeutic
target in spinal cord injury. Clin Chem Lab Med 2001;39:362-367.
77.
Demestre M, Wells GM, Miller KM, et al. Characterisation of matrix
metalloproteinases and the effects of a broad-spectrum inhibitor (BB-1101) in peripheral
nerve regeneration. Neuroscience 2004;124:767-779.
78.
Charalambous P, Hurst LA, Thanos S. Engrafted chicken neural tube-derived stem
cells support the innate propensity for axonal regeneration within the rat optic nerve. Invest
Ophthalmol Vis Sci 2008;49:3513-3524.
79.
Pastrana E, Moreno-Flores MT, Gurzov EN, Avila J, Wandosell F, Diaz-Nido J.
24
Genes associated with adult axon regeneration promoted by olfactory ensheathing cells: a new
role for matrix metalloproteinase 2. J Neurosci 2006;26:5347-5359.
80.
Gaublomme D, Buyens T, De Groef L, et al. Matrix metalloproteinase 2 and
membrane-type 1 matrix metalloproteinase co-regulate axonal outgrowth of mouse retinal
ganglion cells. J Neurochem 2013;(in revision).
81.
Ahmed Z, Aslam M, Lorber B, Suggate EL, Berry M, Logan A. Optic nerve and
vitreal inflammation are both RGC neuroprotective but only the latter is RGC axogenic.
Neurobiol Dis 2010;37:441-454.
82.
Kurimoto T, Yin Y, Omura K, et al. Long-distance axon regeneration in the mature
optic nerve: contributions of oncomodulin, cAMP, and pten gene deletion. J Neurosci
2010;30:15654-15663.
83.
Lorber B, Berry M, Douglas MR, Nakazawa T, Logan A. Activated retinal glia
promote neurite outgrowth of retinal ganglion cells via apolipoprotein E. J Neurosci Res
2009;87:2645-2652.
84.
Lorber B, Guidi A, Fawcett JW, Martin KR. Activated retinal glia mediated axon
regeneration in experimental glaucoma. Neurobiol Dis 2012;45:243-252.
85.
Yin Y, Cui Q, Gilbert HY, et al. Oncomodulin links inflammation to optic nerve
regeneration. Proc Natl Acad Sci U S A 2009;106:19587-19592.
86.
Jiang C, Klassen H, Zhang X, Young M. Laser injury promotes migration and
integration of retinal progenitor cells into host retina. Mol Vis 2010;16:983-990.
87.
Johnson TV, Bull ND, Martin KR. Identification of barriers to retinal engraftment of
transplanted stem cells. Invest Ophthalmol Vis Sci 2010;51:960-970.
88.
Kinouchi R, Takeda M, Yang L, et al. Robust neural integration from retinal
transplants in mice deficient in GFAP and vimentin. Nat Neurosci 2003;6:863-868.
89.
Tucker B, Klassen H, Yang L, Chen DF, Young MJ. Elevated MMP Expression in the
MRL Mouse Retina Creates a Permissive Environment for Retinal Regeneration. Invest
Ophthalmol Vis Sci 2008;49:1686-1695.
90.
Suzuki T, Mandai M, Akimoto M, Yoshimura N, Takahashi M. The simultaneous
treatment of MMP-2 stimulants in retinal transplantation enhances grafted cell migration into
the host retina. Stem Cells 2006;24:2406-2411.
91.
Tucker BA, Redenti SM, Jiang C, et al. The use of progenitor cell/biodegradable
MMP2-PLGA polymer constructs to enhance cellular integration and retinal repopulation.
Biomaterials 2009;31:9-19.
92.
Yao J, Tucker BA, Zhang X, Checa-Casalengua P, Herrero-Vanrell R, Young MJ.
Robust cell integration from co-transplantation of biodegradable MMP2-PLGA microspheres
with retinal progenitor cells. Biomaterials 2011;32:1041-1050.
93.
Zhang Y, Klassen HJ, Tucker BA, Perez MT, Young MJ. CNS progenitor cells
promote a permissive environment for neurite outgrowth via a matrix metalloproteinase-2dependent mechanism. J Neurosci 2007;27:4499-4506.
94.
Bartsch U, Oriyakhel W, Kenna PF, et al. Retinal cells integrate into the outer nuclear
layer and differentiate into mature photoreceptors after subretinal transplantation into adult
mice. Exp Eye Res 2008;86:691-700.
95.
Klassen H, Kiilgaard JF, Zahir T, et al. Progenitor cells from the porcine neural retina
express photoreceptor markers after transplantation to the subretinal space of allorecipients.
Stem Cells 2007;25:1222-1230.
96.
Lamba DA, Gust J, Reh TA. Transplantation of human embryonic stem cell-derived
photoreceptors restores some visual function in Crx-deficient mice. Cell Stem Cell 2009;4:7379.
97.
MacLaren RE, Pearson RA, MacNeil A, et al. Retinal repair by transplantation of
photoreceptor precursors. Nature 2006;444:203-207.
25
98.
Gonzalez-Cordero A, West EL, Pearson RA, et al. Photoreceptor precursors derived
from three-dimensional embryonic stem cell cultures integrate and mature within adult
degenerate retina. Nat Biotech 2013;31:741-747.
99.
Bull ND, Irvine KA, Franklin RJ, Martin KR. Transplanted oligodendrocyte precursor
cells reduce neurodegeneration in a model of glaucoma. Invest Ophthalmol Vis Sci
2009;50:4244-4253.
100. Bull ND, Limb GA, Martin KR. Human Muller stem cell (MIO-M1) transplantation in
a rat model of glaucoma: survival, differentiation, and integration. Invest Ophthalmol Vis Sci
2008;49:3449-3456.
101. Morrison CJ, Butler GS, RodrÃ-guez D, Overall CM. Matrix metalloproteinase
proteomics: substrates, targets, and therapy. Current Opinion in Cell Biology 2009;21:645653.
26
Figures
Figure 1.
27
Figure 2.
28
Figure 3.
29
Legends to figures
Figure 1. Schematic representation of known MMP and TIMP activities in the
glaucomatous retina.
Proposed MMP-9 working mechanisms (depicted in blue): Various insults, including elevated
IOP and ischemia, result in the elevation of retinal glutamate levels and stimulate the
production of IL-1 by astrocytes, inducing an upregulation of MMP-9 synthesis in astrocytes
and/or RGCs. In addition, IL-1 also increases the production of nitric oxide, which can
activate extracellular pro-MMP-9. Finally, MMP-9 expression increases as elevated IOP
results in a diminished deposition of TGF-β2, a known MMP inhibitor, in the ECM. This
increased MMP-9 activity in the RGC layer abrogates laminin-integrin signalling, mediated
by FAK, PI3K and PKB kinases, resulting in decreased expression of the anti-apoptotic
protein Bcl-xL and detachment-induced apoptosis of RGCs.
Proposed TIMP-1 working mechanisms (depicted in red): Ocular hypertension-induced
elevation of MMP-9 activity in the RGC layer, is accompanied by elevated TIMP-1
expression in RGCs. This TIMP-1 elevation is likely to act in a dual way: first of all, TIMP-1
will inhibit MMP-9 and as such keep pro-apoptotic MMP-9 activity within limits, second,
TIMP-1 might also promote RGC survival by directly suppressing pro-apoptosis signalling in
an MMP-independent manner.
NFL: nerve fiber layer, GCL: ganglion cell layer; IPL: inner plexiform layer, INL: inner
nuclear layer; OPL: outer plexiform layer, ONL: outer nuclear layer, PRL: photoreceptor
layer,
FAK:
focal
adhesion
kinase,
PI3K:
phosphatidylinositide
3-kinase,
PIP2:
phosphatidylinositol 4,5-bisphosphate; PIP3: phosphatidylinositol 3,4,5-triphosphate; PKB:
protein kinase B, Bcl-xL: B-cell lymphoma-extra large; NO: nitric oxide; IL-1: interleukin-1;
30
TGFβ2: transforming growth factor-β2.
31
Figure 2. Schematic representation of suggested MMP and TIMP activities in the
glaucomatous optic nerve.
In response to the mechanical stress generated by elevated IOP, lamina cribrosa cells and
astrocytes increase their secretion of TGF-β1 and TNF-α, which act in an auto- or paracrine
manner to induce MMP-2 expression and ECM remodeling in the optic nerve head (1).
Moreover, astrocytes also express MMP-1 and MT1-MMP while undergoing a transition from
a quiescent to a reactive phenotype. This transition involves detachment from the basement
membrane and relocation throughout the axon bundles, processes which are both facilitated
via proteolysis of the ECM and cell surface adhesion molecules by MMP-1 and MT1-MMP
(2). If not counterbalanced by TIMP-1, expressed in RGC axons and astrocytes, MMP-1 will
continue to degrade the ECM, resulting in a lamina cribrosa extracellular environment that is
no longer able to support axonal/neuronal survival (3).
32
Figure 3. Schematic representation of suggested MMP and TIMP activities in the
regenerating optic nerve.
MMP-2, -9, and to a lesser extent MMP-1 and -3, are upregulated at the site of injury and in
the proximal stump of the optic nerve, presumably by (an) unknown factor(s), released by the
numerous collateral axonal sprouts that stimulate astrocytes to increase their MMP expression.
Initially, MMP-2 and -9 contribute to the removal of tissue debris during early post-injury
inflammation (1). Later on during the recovery phase, a well-balanced MMP activity might
promote axonal regeneration via degradation of glial scar-inhibitory ligands, such as CSPGs
(2); via degradation of myelin-derived inhibitory ligands, such as Nogo, MAG, OMgp and
MBP (3); via unmasking, release and/or activation of growth-promoting molecules such as
bFGF, pro-NFG and pro-BDNF (4); and/or via focalized ECM proteolysis at the growth cones
of regenerating axons (5).
33
Legends to tables
Table 1. MMP expression and activity in the healthy versus glaucomatous retina.
This table provides an overview of reported MMP expression/activity in the healthy and
glaucomatous retina, reported from in vivo studies, with corresponding references. Expression
data in human patients and data obtained in animal models of glaucoma are categorized under
different subheadings, with the latter being subdivided according to the type of initial insult
leading to the glaucomatous retinal damage. Of note, optic nerve ligation has been grouped
under the subheading ‘ischemia-induced animal models for glaucoma’, however, as not only
the ophthalmic artery but also the optic nerve is ligated during this procedure, RGC
degeneration in this model is likely due to a combination of ischemia and axonal compression.
NFL: nerve fiber layer, OLM: outer limiting membrane, GCL: ganglion cell layer, IPL: inner
plexiform layer, OPL: outer plexiform layer, INL: inner nuclear layer, IHC :
immunohistochemistry, ISZ: in situ zymography, ISH: in situ hybridisation, WB: western blot,
qRT-PCR: quantitative RT-PCR, POAG: primary open angle glaucoma.
34
Table 2. MMP expression and activity in the healthy versus glaucomatous optic nerve.
This table provides an overview of MMP expression/activity in the healthy and glaucomatous
optic nerve, reported from in vivo studies, with corresponding references. It illustrates the
variety in species, glaucoma models and experimental details that have contributed to a very
rich data set about MMP expression/activity, but that have also hampered a straightforward
interpretation of these data due to conflicting outcomes. Expression data in human patients
and data obtained in animal models of glaucoma are categorized under different subheadings.
IHC : immunohistochemistry, ISH: in situ hybridisation, qRT-PCR: quantitative RT-PCR,
POAG: primary open angle glaucoma, NTG: normal tension glaucoma.
35
Table 1.
Expression/activity in the retina
Species/glaucoma model
Technique(s)
Reference
Human patients
MMP-1
nuclear and plexiform layers
human
IHC
24
MMP-2
RGCs and their axons in the NFL
human
IHC
23
MMP-3
no detectable immunoreactivity
human
IHC
24
MMP-9
no detectable immunoreactivity
human
IHC
24
MT1-MMP
photoreceptor outer segments
human
WB
25
TIMP-1
no detectable immunoreactivity
human
IHC
24
► glaucoma:
human (POAG)
IHC
23
human
IHC
24
rat (hypertonic saline
episcleral vein injection)
microarray,
qRT-PCR
37
increased MMP-9 expression in apoptotic cells in GCL, correlating
with the degree of IOP exposure
rat (hypertonic saline
episcleral vein injection)
IHC
26
► glaucoma:
rat (hypertonic saline
episcleral vein injection)
microarray,
qRT-PCR
38
GCL
► glaucoma: increased immunoreactivity, correlating with the degree of IOP
exposure
rat (hypertonic saline
episcleral vein injection)
IHC
26
► glaucoma:
increased TIMP-1 expression
rat (hypertonic saline
episcleral vein injection)
qRT-PCR
32
► glaucoma:
increased TIMP-1 expression
rat (hypertonic saline
episcleral vein injection)
microarray,
qRT-PCR
37
► glaucoma:
increased TIMP-1 expression
rat (hypertonic saline
episcleral vein injection)
microarray,
qRT-PCR
38
WB, IHC
34
microarray,
qRT-PCR
43
WB, IHC
34
WB
30
zymography
31
TIMP-2
RGC somata and axons, no immunoreactivity in astrocytes
inner and outer nuclear layers
Ocular hypertension-induced glaucoma models in rodents
MMP-3
► glaucoma:
MMP-9
► glaucoma:
TIMP-1
increased MMP-3 expression
decreased MMP-9 expression
Optic nerve injury-induced glaucoma models in rodents
MMP-2
► glaucoma:
MMP-2 expression remains unchanged
rat (optic nerve
transection)
MMP-3
► glaucoma:
increased MMP-3 expression
rat (optic nerve crush,
optic nerve transection)
MMP-9
► glaucoma:
increased MMP-9 expression in RGC layer
rat (optic nerve
transection)
Ischemia-induced glaucoma models in rodents
MMP-2
MMP-9
36
► glaucoma:
MMP-2 expression remains unchanged
mouse (optic nerve
ligation)
► glaucoma:
increased pro-MMP-2 expression
rat (ischemia-reperfusion
injury)
dot-like staining pattern in GCL
► glaucoma: increased eGFP+ area and intensity
mouse (optic nerve
ligation)
MMP-9-eGFP
reporter mice
27
► glaucoma:
increased MMP-9 expression in RGCs
mouse (ischemiareperfusion injury)
IHC, ISZ
28
► glaucoma:
increased MMP-9 expression in reactive astrocytes
mouse (optic nerve
ligation)
WB, IHC
29, 30, 33
TIMP-1
TIMP-2
► glaucoma:
increased pro-MMP-9 expression in interphotoreceptormatrix
rat (ischemia-reperfusion
injury)
zymography,
IHC
31
► glaucoma:
TIMP-1 expression remains unchanged
rat (ischemia-reperfusion
injury)
reverse
zymography
31
► glaucoma:
TIMP-1 expression remains unchanged
mouse (optic nerve
ligation)
WB
33
► glaucoma:
TIMP-2 expression remains unchanged
mouse (optic nerve
ligation)
WB
33
► glaucoma:
TIMP-2 expression remains unchanged
rat (ischemia-reperfusion
injury)
reverse
zymography
31
rat (intravitreal NMDA
injection)
zymography
32
RGCs, Müller glia, astrocytes
► glaucoma: slightly increased MMP-2 protein levels measured via WB, yet no
change in activity levels measured via zymography
mouse (intravitreal KA
injection)
WB,
zymography,
IHC
33
► glaucoma:
rat (intravitreal NMDA
injection)
zymography,
ISZ
32
Excitotoxicity-induced glaucoma models in rodents
MMP-2
MMP-9
► glaucoma:
MMP-2 activity remains unchanged
increased in expression of pro- and active MMP-9 in RGCs
Miscellaneous
RGCs and their axons in the NFL
monkey
IHC
22
photoreceptor inner and outer segments; weak immunoreactivity in OLM
horse
IHC
36
MMP-9
GCL, IPL and photoreceptor segments
horse
IHC
36
MT1-MMP
photoreceptor inner segments
horse
IHC
36
NFL
mouse (P0)
ISH
50
inner retinal layer
rabbit
IHC
51
TIMP-1
RGC somata and axons in NFL, no immunoreactivity in astrocytes in NFL
monkey
IHC, ISZ
22
TIMP-2
photoreceptor inner and outer segments
horse
IHC
36
MMP-2
37
Table 2.
Expression/activity in the optic nerve
Species/glaucoma model
Technique(s)
Reference
Human patients
cytoplasm of few glial cells; faint staining of glial processes around axons and in
pial septae
► glaucoma: increased number of MMP-1+ glial cells
human (PAOG, NTG)
IHC
60
few astrocytes
► glaucoma: increased immunoreactivity in astrocytes and ECM in cribriform
plates; axons; small vessels and pial septae
human (POAG)
IHC
23
few glial cells; faint staining of glial processes around axons
increased intensity of immunostaining and number of MMP-2+ cells;
increased intensity of immunostaining of glial processes around axons and along
pial blood vessels
► intracytoplasmatic immunostaining of glial cells is more intense in areas with
preserved axons
human (POAG, NTG)
IHC
60
astrocytes; axons, higher immunoreactivity in unmyelinated (pre)laminar regions
than in myelinated postlaminar region
► glaucoma: MMP-2 expression remains unchanged
human (POAG)
IHC
23
few glial cells; faint staining of glial processes around axons
► glaucoma: increased immunoreactivity in astroglial cells, glial processes around
axons and along pial septae
► immunoreactivity is more intense in areas with preserved axons
human (POAG, NTG)
IHC
60
perivascular cells
human (POAG)
IHC
23
MMP-7
no detectable immunoreactivity
human (POAG)
IHC
23
MMP-9
no detectable immunoreactivity
human (POAG)
IHC
23
MMP-12
no detectable immunoreactivity
human (POAG)
IHC
23
MT1-MMP
few astrocytes; granular immunostaining associated with blood vessels; no
immunoreactivity detected in axons or ECM
► glaucoma: increased immunoreactivity in astrocytes
human (POAG)
IHC
23
TIMP-1
astrocytes and axons
► glaucoma: TIMP-1 expression remains unchanged
human (POAG)
IHC
23
TIMP-2
astrocytes and axons
► glaucoma: TIMP-2 expression remains unchanged
human (POAG)
IHC
23
MMP-1
MMP-2
► glaucoma:
MMP-3
Ocular hypertension-induced glaucoma models in monkey
MMP-1
few astrocytes; small blood vessels
► glaucoma: increased immunoreactivity in reactive astrocytes and ECM; many
quiescent astrocytes in myelinated nerve express mRNA
monkey (laser scarification
of trabecular meshwork)
ISH, IHC
22
MMP-2
few astrocytes; axons
► glaucoma: decreased MMP-2 immunoreactivity in nerve bundles due to loss of
axons; MMP-2 expression in astrocytes remains unchanged
monkey (laser scarification
of trabecular meshwork)
ISH, IHC
22
MMP-3
small bloodvessels
monkey (laser scarification
of trabecular meshwork)
IHC
22
MMP-9
some oligodendrocytes in the nerve bundles in the myelinated nerve
► glaucoma: MMP-9 expression remains unchanged
monkey (laser scarification
of trabecular meshwork)
IHC
22
MT1-MMP
low immunoreactivity/mRNA in few astrocytes and around small blood vessels
► glaucoma: increased immunoreactivity/mRNA in reactive astrocytes
monkey (laser scarification
of trabecular meshwork)
ISH, IHC
22
TIMP-1
astrocytes and axons
► glaucoma: decreased immunoreactivity due to loss of axons
monkey (laser scarification
of trabecular meshwork)
ISH, IHC
22
TIMP-2
astrocytes and axons
► glaucoma: decreased immunoreactivity due to loss of axons
monkey (laser scarification
of trabecular meshwork)
ISH, IHC
22
38
Ocular hypertension- and axonal injury-induced glaucoma models in rodents
MMP-2
MMP-3
► glaucoma:
increased MMP-2 expression
rat (hypertonic saline
episcleral vein injection)
microarray,
qRT-PCR
62
► glaucoma:
MMP-2 expression remains unchanged
rat (optic nerve crush)
qRT-PCR,
zymography
63
► glaucoma:
increased MMP-3 expression, primarily associated with GFAP+
rat (optic nerve crush)
qRT-PCR, IHC
63
increased MMP-7 expression, primarily associated with GFAP+
rat (optic nerve crush)
qRT-PCR, IHC
63
increased pro-MMP-9 expression, primarily associated with GFAP+
rat (optic nerve crush)
qRT-PCR, IHC,
zymography
63
increased MMP-12 expression, primarily associated with GFAP+
rat (optic nerve crush)
qRT-PCR, IHC
63
increased TIMP-1 expression
rat (hypertonic saline
episcleral vein injection,
optic nerve transection)
microarray,
qRT-PCR
62
astrocytes
MMP-7
► glaucoma:
astrocytes
MMP-9
► glaucoma:
astrocytes
MMP-12
► glaucoma:
astrocytes
TIMP-1
39
► glaucoma: