REVIEWS

MATRIX METALLOPROTEINASES AS
MODULATORS OF INFLAMMATION
AND INNATE IMMUNITY
William C. Parks*‡, Carole L. Wilson§ and Yolanda S. López-Boado||
As their name implies, matrix metalloproteinases are thought to be responsible for the turnover
and degradation of the extracellular matrix. However, matrix degradation is neither the sole nor
the main function of these proteinases. Indeed, as we discuss here, recent findings indicate that
matrix metalloproteinases act on pro-inflammatory cytokines, chemokines and other proteins to
regulate varied aspects of inflammation and immunity.

*University of Washington,
Harborview Medical Center,
Department of Medicine,
Box 359640, 325 9th Avenue,
Seattle, Washington 98104,
USA.
‡
Departments of Medicine
and §Pathology, University of
Washington School of
Medicine, Seattle,
Washington 98195, USA.
||
Department of Internal
Medicine (Molecular
Medicine), Wake Forest
University School of
Medicine, Winston-Salem,
North Carolina 27157, USA.
Correspondence to W.C.P.
e-mail:
parksw@u.washington.edu
doi:10.1038/nri1418
If inflammation and immunity are defined broadly,
including all of the processes that are associated with
defence against microorganisms and other environ-
mental insults, then a wide variety of cell types and pro-
teins participate in these processes. Although leukocytes
are the paradigmatic inflammatory cell type, they are
not the only cells that function in defence and immu-
nity. For example, epithelial cells, although specialized to
have distinct functions in different tissues, respond sim-
ilarly to injury and infection, and regulate leukocyte
influx by equivalent mechanisms. Overall, the pro-
grammes that regulate repair, defence, inflammation
and immunity, regardless of the cell type invoked, might
have co-evolved, particularly with respect to the type of
protein that functions in all of these processes. As dis-
cussed in this review, new views on the function of
matrix metalloproteinases (MMPs) indicate that this
family of enzymes regulates various inflammatory and
repair processes and therefore might represent an early
step in the evolution of the immune system.
The MMP family
The MMP family currently comprises 25 related, but dis-
tinct, vertebrate gene products, of which 24 are found in
mammals (TABLE 1). MMPs are secreted or anchored to
the cell surface, thereby confining their catalytic activity to
membrane proteins and proteins in the secretory path-
way or extracellular space. To be classified as an MMP, a
protein needs to have at least the conserved PRO-DOMAIN
and CATALYTIC DOMAIN (FIG. 1). The pro-domain of a typical
MMP is ~80 amino acids and contains the consensus
sequence PRCXXPD (where X denotes any amino acid).
The exception is MMP23, in which the crucial cysteine
residue is found in a distinct amino-acid sequence1. The
catalytic domain of a typical MMP contains a zinc ion
(Zn2+) in the active site (the reason for the prefix ‘met-
allo’) that is ligated to three conserved histidine residues
in the sequence HEXXHXXGXXH. The glutamic acid
residue (E) in this catalytic motif provides the nucleo-
phile that severs peptide bonds. The backbone structures
of the MMP catalytic domain, including a characteristic
MET TURN that is caused by a conserved methionine
residue downstream of the zinc-binding site2, are similar
to those of the astacin-, reprolysin- (also known as ADAM)
and serralysin-family metalloproteinases. Together,
these four families comprise the metzincins CLAN of the
metalloendopeptidase superfamily. Similar to the diverse
functions of MMPs, these other metalloproteinases
also participate in a range of processes, such as matrix
synthesis, cytokine activation and ligand shedding.
With the exception of MMP7, -23 and -26, MMPs
have a flexible proline-rich HINGE REGION and a carboxy
(C)-terminal HEMOPEXIN-LIKE DOMAIN, which functions in
substrate recognition (FIG. 1). Other domains found in
MMPs are restricted to subgroups of enzymes. For exam-
ple, four membrane-type MMPs (MMP14, -15, -16 and
-24) have transmembrane and cytosolic domains,
whereas MMP17 and MMP25 have C-terminal hydro-
phobic extensions that function as GLYCOSYLPHOSPHATIDYL-
INOSITOL (GPI)-ANCHORING SIGNALS. The two gelatinases

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Table 1 | Mammalian matrix metalloproteinases

Designation* Common name Other name(s) Substrates‡ References§
MMP1 Collagenase-1 Fibroblast collagenase, Type I and II fibrillar collagens?|| 111–114
interstitial collagenase
ColA¶ 115
ColB¶ 115
MMP2 Gelatinase A 72-kDa gelatinase, CCL7 27
72-kDa type IV collagenase CXCL12 67
MMP3 Stromelysin-1 Transin-1 E-cadherin 100
Laminin, type IV collagen 101
PRO-DOMAIN Latent TGF-β1 81
The matrix metalloproteinase MMP7 Matrilysin PUMP Pro-α-defensins 51
(MMP) pro-peptide region FAS ligand (CD95L) 53
(or pro-domain) contains ~80 Latent TNF 102
amino acids, typically with a Syndecan-1 66
hydrophobic residue at the E-cadherin 46
amino terminus. It also contains Elastin 35
the highly conserved sequence
PRCXXPD, where X denotes MMP8 Collagenase-2 Neutrophil collagenase Mouse CXCL5 68
any amino acid. The thiol group MMP9 Gelatinase B 92-kDa gelatinase, Zona occludens 1 103
of the cysteine residue in this 92-kDa type IV collagenase α1-Antiproteinase 99
sequence ligates with the zinc Latent TGF-β1 23
ion that is held by the histidine Latent VEGF 75
residues in the catalytic domain Fibrin 104
of the MMP. In this state, the NG2 proteoglycan 105
enzyme is stable and inactive
and is known as a zymogen.
MMP10 Stromelysin-2 Transin-2 ND
MMP11 Stromelysin-3 ND
CATALYTIC-DOMAIN
MMP12 Metalloelastase Latent TNF 106
The typical matrix
metalloproteinase catalytic MMP13 Collagenase-3 Rat collagenase Type I and II fibrillar collagens?|| 111–114
domain contains ~160–170
MMP14 MT1-MMP Membrane-type MMP ProMMP2 32
residues, including the binding
Fibrillar collagens 31,33,34
sites for the structural (calcium
Fibrin 34,107
and zinc) and catalytic (zinc) Syndecan-1 108
metal ions. The 50–54 residues γ2-Subunit of laminin-5 109
at the carboxyl terminus of the
catalytic domain include a MMP15 MT2-MMP Fibrin 34,107
highly conserved MMP16 MT3-MMP Fibrin 34,107
HEXXHXXGXXH sequence Syndecan-1 108
(where X denotes any amino
acid), which includes a glutamic MMP17 MT4-MMP ND
acid residue (E) that provides MMP19# ND
the nucleophile that severs
peptide bonds and histidine MMP20 Enamelysin Amelogenin 110
residues that coordinate the MMP21 ND
zinc ions.
MMP22 ND
MET TURN MMP23 CA-MMP ND
On the carboxy side of the zinc
MMP24 MT5-MMP ND
active site, matrix
metalloproteinases have a MMP25 Leukolysin MT6-MMP ND
methionine residue that is
MMP26 Endometase Matrilysin-2 ND
always conserved. This residue
is part of a 1,4-β-turn that loops MMP27** ND
the polypeptide chain beneath MMP28 Epilysin ND
the catalytic zinc ion and forms
a hydrophobic base for the *Matrix metalloproteinase (MMP)-4, -5 and -6 were found to be identical to either MMP2 or MMP3 and, therefore, are not unique.
zinc-binding site. MMP18 has only been cloned from Xenopus laevis; a mammalian homologue has not been found. ‡This list of substrates is limited to
proteins that have been shown to be cleaved by an MMP by either a gain- or loss-of-function approach or both. For some substrates,
ADAM
only indirect evidence has been provided, but, importantly, the substrate is known to be cleaved. Excluded from this list are the many
proteins that have been shown to be cleaved by an MMP only in a defined in vitro setting. In addition, proteins that were identified only
A disintegrin and
by the treatment of cells with exogenous MMP or by overexpression of a particular MMP have also been omitted. §References are
metalloproteinase family
reports of identification of substrates only. ||Various assays and parameters, such as the formation of neoepitopes in vivo111–113, indicate
of proteases. They contain
that MMP1 and MMP13 probably act on fibrillar collagens (type I and type II). However, data that compare the collagenolytic activity of
disintegrin-like and these MMPs with that of MMP14 (REF. 114) question which enzymes created these neoepitopes. ¶|Collagenase-like protein A (ColA)
metalloproteinase-like and ColB are probably the murine homologues of MMP1 (REF. 115). On the basis of its chromosomal position and enzymatic activity,
domains and are involved McolA is a strong candidate to be the murine orthologue of human MMP1. #In the initial cloning paper116, human MMP19 was called
in the regulation of MMP18, but this designation was already assigned to X. laevis MMP18 (REF. 117). **In addition to being identified in a large scale
developmental processes, bioinformatic search of secreted proteins118, full-length cDNAs for MMP27 from humans, rats and northern tree shrews have been
cell–cell interactions and submitted to GenBank. CA, cysteine array; CCL, CC-chemokine ligand; CXCL, CXC-chemokine ligand; E-cadherin, epithelial cadherin;
protein processing, including ND, not determined; PUMP, putative metalloproteinase; TGF, transforming growth factor; TNF, tumour-necrosis factor; VEGF, vascular
ectodomain shedding. endothelial growth factor.

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CLAN
The superfamily of
metalloenzymes includes more
than 200 members. It has been
divided into eight clans based on
the similarity of protein folding
characteristics and ~40 families
according to evolutionary
relationships. The matrix-
metalloproteinase family belongs
to clan MB, the members of
which have three histidine
residues as zinc-binding ligands
in the consensus sequence
HEXXHXXGXXH (where X
denotes any amino acid).
HINGE REGION
A domain that is typically ~75
residues and links the catalytic
domain to the hemopexin-like
domain of most matrix
metalloproteinases.
HEMOPEXIN-LIKE DOMAIN
This matrix-metalloproteinase
domain comprises ~200
residues and contains four
repeats that resemble
hemopexin and vitronectin. It is
not essential for catalytic activity
but does modulate substrate
specificity and binding to tissue
inhibitors of metalloproteinases.
GLYCOSYLPHOSPHATIDYL-
INOSITOL (GPI)-ANCHORING
SIGNALS
A glycolipid modification that is
usually located at the carboxyl
terminus and anchors proteins
to the external surface of the
plasma membrane.
GELATIN-BINDING DOMAINS
These domains contain three
fibronectin-like modules (also
known as fibronectin type II
modules) and are present in the
catalytic domain of both matrix
metalloproteinase 2 and -9.
CYSTEINE-SWITCH MECHANISM
The pro-peptide maintains a
matrix metalloproteinase
(MMP) in an inactive state.
When the interaction between
the conserved cysteine residue in
the pro-domain and the active
site zinc ion is disrupted (for
example, by proteolytic removal
of the pro-peptide or by the
action of organomercurials and
chaotropic agents on the thiol of
the cysteine residue), the active
site becomes accessible, and the
MMP has been ‘activated’. The
pro-domain does not need to be
removed for a proMMP to
acquire activity; only disruption
of the zinc–thiol interaction is
absolutely required.
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Basic MMP1, -3, -8, SH Zn2+ Hinge
-10, -12, -13, -18, SP Pro Catalytic Hemopexin-like
-19, -20, -22, -27
ColA, ColB
Minimal MMP7, -26 SP Pro Catalytic
Furin- MMP11, -28 SP Pro Fr Catalytic Hemopexin-like
activated
TM Cs
Membrane- TM: MMP14, -15, -16, -24 SP Pro Fr Catalytic Hemopexin-like or
anchored GPI: MMP17, -25
GPI
Hemopexin-like
Gelatin- MMP2, -9 SP Pro Catalytic Fn Fn Fn or
binding
C5 Hemopexin-like
Type II MMP23 SA Pro Fr Catalytic Cys IgG-like
membrane
Figure 1 | Domain structure of the mammalian MMP family. The important features of matrix metalloproteinases (MMPs) are
illustrated, showing the minimal domain structures. Although MMPs are often subdivided into groups on the basis of differences in
domain composition (shown here), there is little consensus in the field about how such subdivisions should be assigned. Domain
structure alone does not predict function. One clear division is between MMPs that are secreted and those that are anchored to the
cell surface by an intrinsic motif: namely, a transmembrane (TM) domain (MMP14, -15, -16 and -24), a glycosylphosphatidylinositol
(GPI) anchor (MMP17 and MMP25) or an amino (N)-terminal signal anchor (SA) (MMP23). Both the TM domains and GPI anchors
are attached to the hemopexin-like domain by a short linker. As discussed in the text, the secreted MMPs might still be confined to
the cell surface through interactions with specific accessory macromolecules. Because the mechanisms that control activation (that
is, conversion of proMMP to active MMP) are key steps in the regulation of proteolysis, another grouping of the MMPs can be made
on the basis of intracellular activation by furin proteinases. Nine MMPs, including all of the membrane-anchored enzymes, have a
furin-recognition domain. C5, type-V-collagen-like domain; Col; collagenase-like protein; Cs, cytosolic; Cys, cysteine array; Fn,
fibronectin repeat; Fr, furin-cleavage site; Pro, pro-domain; SH, thiol group; SP, signal peptide; Zn, zinc.
(MMP2 and MMP9) have GELATIN-BINDING DOMAINS that a tissue or in culture, express MMPs. For the most part,
resemble a similar motif in fibronectin. This motif is the production of MMPs is regulated at the level of
involved in the binding of fibronectin to denatured colla- transcription by specific signals that are temporally
gen, and in MMP2 and MMP9, it probably enhances the limited and spatially confined.
interaction with gelatin or gelatin-like substrates. In addi- ProMMPs are kept in a catalytically inactive state by
tion to a common three-dimensional structure3, MMPs the interaction between the thiol group of a pro-domain
have a similar gene arrangement, indicating that they cysteine residue and the zinc ion of the catalytic site.
probably arose by duplications of an ancestor gene. At They are converted to active proteinases by disruption of
least eight of the known human MMP genes (MMP1, -3, this interaction (a process known as the CYSTEINE-SWITCH
5
-7, -8, -10, -12, -13 and -20) are clustered on chromosome MECHANISM ), which can be achieved by proteolysis of the
11 at 11q21–23, whereas other MMP genes are ‘scattered’ pro-domain or by modification of the cysteine thiol
along chromosomes 1, 8, 12, 14, 16, 20 and 22 (REF. 4). group. Several MMPs contain an RXKR or RRKR
sequence between the pro- and catalytic domains, which
Regulation of MMP activity functions as a target sequence for pro-protein convertases
Similar to all secreted proteinases, the catalytic activity or furins; this is known as a furin cleavage site (FIG. 1).
of MMPs is regulated at four points — gene expression, For the other MMPs, the mode of activation is more
compartmentalization (that is, the pericellular accu- presumed than proven. The best described non-furin
mulation of enzymes), pro-enzyme (or zymogen) proMMP activation mechanism is probably the cell-
activation and enzyme inactivation — and is further surface activation of proMMP2 by active MMP14
controlled by substrate availability and affinity. (REFS 6–9). After the pro-domain has been cleaved, the
Typically, MMPs are not expressed in normal healthy active MMPs can be inhibited by natural inhibitors (BOX 1)
tissues. By contrast, MMP expression can be detected in and internalization10,11. Furthermore, and of relevance to
all repair or remodelling processes, in all diseased or inflammation, oxidants that are generated by leukocytes
inflamed tissues and in all cell types grown in culture. or other cells can both activate MMPs (through oxida-
Although the qualitative patterns and quantitative levels tion of the pro-domain thiol group) and subsequently
of MMP expression vary among tissues, diseases, inactivate MMPs (through modification of amino acids
tumours, inflammatory conditions and cell types, a rea- that are crucial for catalysis), providing a mechanism to
sonable generalization is that activated cells, whether in control quantum bursts of proteolytic activity12–16.
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Box 1 | Blocking MMP activity
Natural inhibitors
Tissue inhibitors of metalloproteinases (TIMPs). Four TIMPs (TIMP1, -2, -3 and -4) inhibit
matrix-metalloproteinase (MMP) activity by binding to the catalytic site of MMPs91.
However, the TIMPs differ in their affinity for specific MMPs, and their interaction
does not always lead to inhibition. Indeed, binding of TIMP2 to the hemopexin-like
domain of MMP2 is required for activation of the enzyme (in a complex with active
MMP14). Some TIMPs also have activities that are independent of MMPs92,93.
RECK (reversion including cysteine-rich protein with kazal motifs). This membrane-
bound glycoprotein inhibits MMP2, -9 and -14. RECK probably has other functions,
because a null mutation in the gene is embryonic lethal in mice94.
α2-Macroglobulin. This broad-spectrum inhibitor traps several classes of active
enzymes in the circulation, thereby mediating their uptake by scavenger receptors.
Synthetic inhibitors
(For a discussion of the use and promise of MMP inhibitors in clinical trials for cancer
see Coussens et al.95 and Overall and López-Otin96.)
Peptidomimetics. These small hydroxamic-acid-based molecules mimic the substrate-
recognition site of collagen, chelate zinc ions and effectively inhibit MMP activity.
Examples are batimastat and marimastat. Similar to the other synthetic inhibitors, the
hydroxamic-acid-based inhibitors lack specificity. Not only do they block the activity
of MMPs, but they can act on most metalloenzymes.
Non-peptidomimetics. These are based on the conformation of the MMP active site
and include tanomastat and prinomastat. Tanomastat is particularly potent against
MMP2, -3 and -9.
Tetracycline and derivatives. Tetracycline and non-antibiotic derivatives reduce both
the synthesis and activity of MMPs. The only MMP inhibitor that has been approved by
the United States Food and Drug Administration is the doxycycline known as periostat,
which is used for treatment of periodontal and skin diseases.
Others (natural and synthetic). These include the biphosphonates, neovastat (an extract
of shark cartilage), green tea catechins, aspirin and CT-PCPE (a carboxy (C)-terminal
fragment of the pro-collagen C-terminal proteinase enhancer97).
In vitro studies have shown considerable overlap in
the substrates that MMPs can cleave, particularly among
the extracellular-matrix (ECM) substrates17. For exam-
ple, fibronectin, laminins, elastin and type IV collagen
can be degraded by various MMPs in vitro. In a setting
such as inflammation, in which essentially all MMPs are
present, the shared substrate potential would seemingly
allow biochemical redundancy. However, substrate
selectivity can be honed by two processes: enzyme affin-
ity and compartmentalization. Kinetic studies using
model substrates have shown that specific enzymes
degrade some substrates more efficiently than others.
For example, both MMP2 and MMP9 degrade cleaved
TIMPs
collagen more efficiently than other gelatinolytic
Tissue inhibitors of MMPs18, and MMP7 is a more potent proteoglycanase
metalloproteinases. A family than MMP3 or MMP9 (REF. 19). So, in tissues, which
of four (TIMP1, -2, -3 and -4) contain many potential substrates, the selectivity of
endogenous matrix-
MMP catalysis, in addition to being regulated by the
metalloproteinase (MMP)
inhibitors that bind the catalytic concentration of active enzyme, might be partly
site in activated enzymes. directed by the concentration of a preferred substrate
TIMP1 and TIMP3 also bind relative to that of other potential substrates that are in
the hemopexin-like domain proximity to a secreted MMP.
of the MMP9 and MMP13
zymogens, whereas TIMP2, -3
Compartmentalization (that is, where and how in
and -4 can bind this domain the pericellular environment an MMP is released and
in the MMP2 zymogen. held) is equally, if not more, important for regulating
620 | AUGUST 2004 | VOLUME 4
the specificity of proteolysis than the affinity of the
enzyme–substrate interaction. An important concept is
that cells do not indiscriminately release proteinases.
Instead, proteinases, such as MMPs, are typically
anchored to the cell membrane, thereby maintaining a
high enzyme concentration locally and targeting their
catalytic activity to specific substrates in the pericellular
space. In addition to the membrane-bound MMPs, sev-
eral examples of specific cell–MMP interactions have
been reported, such as the binding of MMP2 to the
αvβ3-integrin20, MMP1 to the α2β1-integrin21,22, MMP9
to CD44 (REF. 23) and MMP7 to surface proteoglycans24,25.
As has been suggested for CD44 (REF. 23) and the α2β1-
integrin21, these membrane anchors might function as
accessory factors that mediate both the activation of the
pro-enzyme and the binding of both substrate and pro-
teinase, thereby increasing the probability of proteolysis
(FIG. 2). So, as for most protein–protein interactions,
MMP specificity might be driven by an additional
component. It is probable that other MMPs are also
attached to cells or matrix through similar specific
interactions, and determining these anchors will be a
key advance towards identifying activation mechanisms
and substrates.
Do MMPs do more than degrade the ECM?
MMPs are thought to be responsible for the turnover
and degradation of connective-tissue proteins. In fact,
the first MMP, discovered by Gross and Lapiere in 1962
(REF. 26), was found in the regressing tadpole tail during
their search for an endogenous collagenase that func-
tions at neutral pH — the pH at which collagens turn
over in most tissues. Following this lead, essentially all
MMPs that have since been isolated have been shown to
be capable of degrading various protein components of
the ECM17. Consequently, as a family, MMPs are often
assigned the role of being the enzymes responsible for
the turnover, degradation, catabolism and destruction of
the ECM. This assumption has led to some unexpected
results in clinical trials using MMP inhibitors and,
eventually, led to a reconsideration of MMP function.
The role in ECM degradation attributed to MMPs
has mostly been based on findings in defined systems —
typically, using a purified MMP incubated under optimal
conditions with a purified ECM protein — showing
what a proteinase is capable of and not what it actually
does in tissues. Despite this caveat, a large body of com-
pelling biochemical and observational data, together
with recent genetic findings, indicate that some MMPs
do act on ECM proteins in vivo (TABLE 1). An important
concept is that in a complex setting, such as an inflamed
tissue or tumour, many diverse cell types are present and
carry out various processes, including angiogenesis,
remodelling and phagocytosis. As discussed earlier, cells
produce a spectrum of MMPs, and as a result of the dis-
tinct localization of MMPs, a specific MMP secreted by
one cell type (for example, a macrophage) would proba-
bly carry out a different function than the same enzyme
produced by another cell type (for example, an epithelial
cell). Furthermore, a particular MMP that is produced
by one cell type probably has more than one function.
www.nature.com/reviews/immunol

CHEMOKINES That is, particular MMPs can act on several protein sub-
A family of structurally related, strates in a given tissue, and recent evidence indicates
small glycoproteins (70–90 that these multiple functions are not always related to
amino acids) that have potent
ECM degradation. The key to understanding MMP
leukocyte activation and/or
chemotactic activity. They have function is to identify the physiological substrates.
pivotal roles in innate and
acquired immunity. These Finding physiological substrates
molecules, of which there are Many proteinases, particularly MMPs, are nonspecific
more than 50, are classified into
four subfamilies depending on
in vitro, so biologically relevant approaches have been
the arrangement of the amino- required to identify enzyme function: that is, to identify
terminal conserved cysteine the natural physiological substrates. Because MMPs are
residues: CC-, CXC-, C- and secreted or anchored to the cell surface, their potential
CX3C-chemokines (where X
substrates include all membrane proteins and proteins
denotes any amino acid). In
general, CC-chemokines attract in the secretory pathway and extracellular space, but it
monocytes, lymphocytes, is probable that only a small proportion of these are
basophils and eosinophils, actual substrates. A key question is how we find and,
whereas CXC-chemokines are importantly, verify substrates. The identification of
chemotactic for neutrophils.
substrates is not straightforward, but several approaches
have been used, including affinity-based approaches
using substrate-binding motifs27, proteomics28,29 and
deduction (BOX 2).
Undoubtedly, the development of protein-expression
technology and genetically defined mice has provided
extremely powerful investigative tools for modern biol-
ogists. The combination of a lack of protein cleavage in
a specific knockout mouse, together with the acquisition
of proteolysis upon ectopic expression of a specific
MMP, is a great tool for verifying substrates. Recently,
we have found evidence that MMPs can act on as many
non-matrix substrates as ECM substrates30. In TABLE 1,
we provide a list of MMP substrates that is limited to
those that have been identified and verified (at least in
part) by targeted mutagenesis and/or overexpression.
Of the MMPs that have been genetically targeted so
far, all show no phenotype or only a minor phenotype
in unchallenged mice, except MMP14-deficient mice,
which have severe bone deformations31,32. Although
these negative observations could be interpreted to indi-
cate that many MMPs do not have a direct role in the
turnover of ECM proteins, this function is known to be
carried out by some family members. For example,
Reaction product
(gain or loss of
Substrate substrate function)
Accessory/
adaptor
protein ProMMP Active MMP
(inactive enzyme)
Figure 2 | Minimal components of the proteolytic process. In vitro, proteolysis requires
only a proteinase and a substrate. By contrast, in vivo, at least one additional component is
typically included to augment, if not define, specificity, as well as perhaps catalytic rate. Various
types of these accessory factors or adaptors are used: they can be either proteins or
glycosaminoglycans, and either membrane-associated or extracellular-matrix components.
As indicated in this figure, a transmembrane accessory factor would simultaneously interact
with the proteinase and substrate, bringing both together at an effective concentration. In
addition, the activation of some pro-matrix metalloproteinases (proMMPs), whether autolytic
or non-autolytic, might be mediated by interaction with these factors. These factors could also
have other functions.
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MMP14 is a key physiological collagenase31,33,34, and
gain-of-function studies indicate that MMP7 expressed
by human macrophages is the relevant elastase pro-
duced by these cells35. However, matrix degradation is
neither the sole nor the main function of these enzymes.
An emerging view of MMPs is that they act primarily on
non-matrix proteins — such as cytokines, CHEMOKINES,
receptors and antimicrobial peptides — often potenti-
ating the activity of these proteins (TABLE 1). After chal-
lenge, such as by injury, cancer, inflammation or
infection, MMP-deficient mice reveal various pheno-
types (TABLE 2), indicating that these enzymes have
specific, and at times, essential roles in tissue repair,
angiogenesis, host defence, tumour progression and,
in particular, inflammation. So, it seems that MMPs (at
least those that have been studied using knockout mice)
have evolved to respond to the insults and pressures of
the extra-uterine environment. If any generalization
can be made about MMP function, it is that these pro-
teinases have a role in inflammation. This has been rec-
ognized for MMP9 for several years. However, as this
topic has been reviewed previously36, and because
recent data have provided new clues to the mechanisms
by which MMP7 influences inflammation, here we
often use MMP7 as an example.
MMPs in inflammation
Before discussing the role of MMPs as modulators of
inflammatory processes, we briefly describe the process
of inflammation and the possible roles MMPs might
have in the response of host tissues to environmental
insults. The inflammatory process (FIG. 3) comprises a
series of cellular responses that depend on integrating
information associated with the following: detection of
an injury and/or the presence of microorganisms, the
accumulation and intervention of cells that eliminate
invading microorganisms and infected host cells, and
the repair of tissues that are damaged by the initial
insult, trauma or the response of the host.
In a recent review of inflammation37, Nathan classi-
fied inflammatory disorders according to their possible
origin and the particular role of inflammation. In each
category there are diseases in which members of the
MMP family are upregulated. In fact, increased or mis-
regulated levels of many MMPs are observed in any
disease that is characterized by or associated with
inflammation. If only because matrix proteolysis is a
hallmark of the inflammatory process that is associated
with many conditions, the role of MMPs as matrix-
degrading proteinases justifies their inclusion as impor-
tant components of the host response to traumatic,
infectious, toxic or autoimmune insults, which we have
broadly defined as inflammation. Although MMP
inhibitors (BOX 1) are used as anti-inflammatory drugs in
periodontal disease38 and have been suggested as a ther-
apy to halt tissue destruction in inflammatory condi-
tions such as arthritis39 and vascular disease40, the exact
role of most MMPs in inflammatory conditions has not
yet been elucidated, even to the extent of understanding
whether they function to improve or worsen inflamma-
tion. Indeed, using arthritis as an example, mouse studies
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Box 2 | Identifying and verifying substrates
Identifying substrates
Proteomics. The basic strategy is to compare the pattern of pericellular proteins that is
produced when a specific proteinase is expressed with the pattern in the absence of the
proteinase28,29.
Affinity approaches. Proteinases bind substrates using regions outside the catalytic
domain (exosites), and these interactions can be exploited to design approaches to find
binding partners, such as direct binding in a yeast two-hybrid assay97.
Deduction. By far the most common approach to finding candidate substrates. Close
examination of the phenotypes of knockout mice might indicate candidates, such as
excess type I collagen deposition in matrix metalloproteinase 14 (MMP14)-deficient
mice31 and reduced apoptosis in the prostate glands of castrated MMP7-deficient mice53.
Verifying substrates
Location. A proteinase and substrate need to be in the same microenvironment during
proteolysis. This can be assessed by methods such as immunostaining, fractionation,
co-immunoprecipitation or ‘pull-down’. A caveat for immunoprecipitation and pull-
down approaches is that a proteinase might only interact transiently with its substrate.
However, a catalytically dead proteinase should bind stably to its substrate. The activity
of MMPs can be blocked by mutating the nucleophile-providing glutamatic acid residue
that is present in the conserved motif (HEXXHXXGXXH, where X denotes any amino
acid) of the catalytic domain.
Cleavage sites. If an MMP cleaves a protein in vivo, then it should make the same cut(s)
and only that cut (those cuts) in vitro using purified reagents.
Loss-of-function. At present, the favoured method is showing that ablation or mutation
of an MMP can prevent a specific proteolytic process. This can be carried out using
targeted mutagenesis, RNA interference, dominant-negative proteins or blocking
antibodies.
Gain-of-function. Overexpression or ectopic expression of an MMP or substrate can
be easily manipulated to examine questions about cleavage-site specificity, accessory
factors and more. As discussed in TABLE 1, overexpression studies need to be used with
caution in the identification of candidate substrates.
indicate that MMPs long thought to contribute to joint
destruction, such as MMP2 and MMP3, might actually
provide protection41,42 (TABLE 2). As we discuss later, there
are now several examples that show that these pro-
teinases are important effectors in various processes con-
trolling repair and leukocyte recruitment — processes
that are central to inflammation (FIG. 4).
MMPs in innate immunity and repair
Epithelial repair. INNATE IMMUNITY comprises various
ready-to-go mechanisms that defend against invading
microorganisms, contribute to tissue repair, and regu-
INNATE IMMUNITY late the activity and influx of cells that are involved in
The term generally refers to acquired immunity. By maintaining a tight barrier
innate pathogen-recognition
systems, as well as to
against the external environment, secreting antimicro-
antimicrobial peptides. Innate bial peptides, and producing chemokines and homing
immunity comprises receptors, the epithelium is the first line of innate
immediate responses that are defence. Injury disrupts the barrier function of the
generated without the
epithelium, creating an entry point for microorganisms
requirement for memory of, or
prior exposure to, the and toxins. However, injured epithelial cells respond
pathogen. It is mostly mediated rapidly to close wounds, a process that involves cell pro-
by receptors that have broad liferation, spreading and migration. Injury also induces
specificity (such as Toll-like the expression of several MMPs43, some of which are
receptors): that is, receptors
that recognize many related
crucial for wound closure. For example, the repair of
pathogen-associated molecular skin wounds requires the catalytic activity of MMP1.
patterns. MMP1 alters the migratory substratum, which consists
622 | AUGUST 2004 | VOLUME 4
of type I collagen, from a high-affinity ligand for the
α2β1-integrin to one of lower affinity. In this way, it
functions as the enzymatic machinery that drives the
forward movement of the repairing cells by allowing
them to attach, dislodge, then reattach to the wound-
bed matrix44. In mucosal tissues, such as lung and gut,
MMP7 is expressed by wound-edge epithelial cells and
is required for RE-EPITHELIALIZATION45. This presumably
occurs by the shedding of epithelial (E)-cadherin
ectodomains, which would loosen cell–cell contacts,
thereby facilitating cell migration46. Although MMP9
has been shown to be required for the migration of
airway epithelial cells over a collagen matrix in cell
culture47, no defect in epithelial closure is observed in
MMP9-deficient mice48.
Killing bacteria. Unlike the many MMPs that are
expressed in response to injury, inflammation or other
overt stimuli, MMP7 is expressed by non-injured, non-
inflamed mucosal epithelium in most, if not all, adult
tissues49. The expression of MMP7 in healthy epithe-
lium indicates that it functions in common homeostatic
processes, which seem to provide defence against
microorganisms and enable apoptosis. In mice, MMP7
activates intestinal pro-α-DEFENSINS (also known as
cryptdins)50, and because of the lack of mature, active
α-defensins, MMP7-deficient mice have an impaired
ability to battle enteric pathogens, such as pathogenic
Escherichia coli and Salmonella typhimurium51. In addi-
tion to killing bacteria through the action of defensins
and other antimicrobials, intestinal epithelial cells that
are infected with invasive bacteria participate in defence
by suicide: that is, by undergoing apoptosis and slough-
ing into the lumenal space (from which they are
excreted by the host), and thereby decreasing the bacter-
ial load52. Among its verified substrates (TABLE 1), MMP7
also sheds membrane-bound FAS ligand (FASL, also
known as CD95L). The binding of FASL to the death
receptor FAS controls programmed cell death53.
Although MMP7-mediated effects on apoptosis have
not yet been shown to be involved in defence mecha-
nisms, these effects might be another way in which
MMP7 is involved in innate immune responses (FIG. 4).
Whereas MMP7 has an indirect role in killing bacteria,
preliminary observations indicate that the hemopexin-
like domain of MMP12, which is readily released from
the intact enzyme, has bactericidal activity54.
The induction and activation of MMP7 in mucosal
epithelium is highly sensitive to the presence of virulent
bacteria, further extending a role for this MMP in innate
immunity55,56. All epithelial tissues in which MMP7 is
expressed are exposed to the external environment, so the
widespread production of MMP7 in epithelium might be
sustained by continual, low-level bacterial exposure.
Consistent with this idea, MMP7 is not detected in fetal
or perinatal tissues57 that have little exposure to micro-
organisms. In the small intestine of germ-free, adult mice,
MMP7 is present at almost undetectable levels, but its
production is induced in ex-germ-free mice that are colo-
nized with a single species of commensal bacteria55. So,
bacterial exposure seems to be the physiological signal
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 REVIEWS

that regulates MMP7 expression in intact epithelium. In
accordance with the suggested role of epithelial cells as
‘sensors’ for microbial infection58, MMP7 might be part
of a more general activation response to bacteria.
MMPs and chemokines
Chemokines are a group of chemotactic molecules that
specifically attract and recruit populations of immune
effector cells (including neutrophils, monocytes and
eosinophils) to sites of injury or infection and thereby
shape the evolution and outcome of the inflammatory
response59. Several studies have shown that specific
MMPs control chemokine activity. This control can be
direct, by MMP-mediated cleavage of these mole-
cules27,60–63, which results in enhancement, inactivation
or antagonism of chemokine activities, or it can be indi-
rect, by the proteinases acting on other substrates that
bind, retain or concentrate the chemotactic molecules in
particular locations64–66.
MMPs directly modulate chemokine activity. One
potential action of MMPs is to convert chemokines
from their true (or initial) nature as chemotactic mole-
cules into antagonistic derivatives, thereby disrupting
the further recruitment of cellular components and
contributors to sites of inflammation. This repressive
processing has been examined in detail by Overall and
colleagues27,60,61, who studied the monocyte-chemotactic
protein (MCP) group of CC-chemokines. Using the
C-terminal hemopexin-like domain of MMP2 as a bait
in a yeast two-hybrid system, they discovered, unexpect-
edly, that CC-chemokine ligand 7 (CCL7; also known as
MCP3) is a substrate of MMP2, which cleaves the four
amino (N)-terminal amino acids of the chemokine.
Truncated CCL7 still binds its cognate CC-chemokine
receptors (CCRs), but it no longer promotes chemo-
taxis; instead, it functions as a chemokine antagonist.
Similarly, MMP1, -3, -13 and -14 cleave the N-terminus
of CCL2 (also known as MCP1), CCL8 (also known as

Table 2 | Inflammatory and immune phenotypes of Mmp-null mice

Mouse Phenotypes that indicate a function in innate or acquired immunity, References
including repair-related processes
Mmp2–/– Decreased allergic inflammation 64
More-severe immune-mediated arthritis 41
Reduced tumour, corneal, retinal and choroidal neovascularization 119–122
Mmp3–/– More-severe immune-mediated arthritis 42
RE-EPITHELIALIZATION Reduced contact hypersensitivity 123
A mechanism of repair that Reduced neutrophil influx in immune-mediated lung injury 124
involves epithelial-cell Reduced number of macrophages in atherosclerotic plaques 125
proliferation and migration Reduced cartilage-derived macrophage-chemotactic activity 72
across a denuded surface to Mmp7–/– Lack of activation of cell-surface TNF displayed on macrophages 106
re-establish cell contact and Lack of active intestinal α-defensins and impaired ability to kill enteric pathogens 51
close a wound. During re- Spatially constrained chemokines and reduced neutrophil influx in injured lung 66
epithelialization, cells receive Markedly impaired re-epithelialization and reduced shedding of E-cadherin in injured lung 45,46
and process cues from a new Reduced shedding of active FAS ligand (CD95L) and epithelial-cell apoptosis 53
microenvironment (that is, Increased neovascularization in corneal wounds 126
the exposed wound) and Mmp8–/– Marked increased in chemically induced skin tumours* 68
coordinate various responses, –/–
including the induction of Mmp9 Prolonged contact hypersensitivity 123
matrix metalloproteinases and
Reduced experimental autoimmune encephalomyelitis 127
Protection against endotoxin-mediated shock 128
pro-inflammatory mediators,
Reduced dendritic (Langerhans)-cell migration 129
and the activation and
Reduced antigen-mediated blister formation 130
expression of integrins. Reduced angiogenesis in developing bone, tumours* and ischaemic tissues 131–133
Impaired bronchiolization after acute lung injury 48
DEFENSINS Altered chemokine gradients in models of allergen-induced airway inflammation 71
A class of antimicrobial peptides Altered leukocyte influx in models of allergen-induced airway inflammation 71,134,135
that have activity against Gram- Modulation of IL-13-induced lung inflammation 136
positive and Gram-negative Less-severe experimental arthritis 41
bacteria, fungi and viruses. Impaired macrophage infiltration in atherosclerosis 137
Defensins are classified into two Protection against macrophage-induced aneurysm formation* 138,139
main categories on the basis of
Reduced ischaemia-induced cerebral injury 140
the position of conserved Mmp2–/–Mmp9–/– Reduced choroidal neovascularization 141
cysteine and hydrophobic –/–
Mmp11 ND
residues and the linkages of
disulphide bonds: α-defensins Mmp12–/– Reduced macrophage migration and influx in smoke-induced emphysema 73,142
are produced by intestinal Reduced spinal cord injury 143
Paneth cells and neutrophils, Reduced release of active TNF from smoke-exposed macrophages 106
and β-defensins are expressed Reduced neutrophil influx and epithelial permeability in immune-mediated lung injury 144
by most epithelial cells. A third Modulation of IL-13-induced lung inflammation 136
category, the θ-defensins, arises Mmp14–/– ND
from the splicing of two α- –/–
Mmp20 ND
defensin-related peptides into a
circular molecule; at present, Mmp28–/– Markedly reduced inflammation in models of lung injury‡
these defensins have been *These phenotypes were reversed following transplantation of wild-type bone marrow, indicating that the effect observed in knockout
detected only in the neutrophils mice was caused by the lack of the matrix metalloproteinase (MMP) in an inflammatory cell or group of inflammatory cells. ‡W.C.P.,
of rhesus macaques. unpublished observations. E-cadherin, epithelial cadherin; IL, interleukin; ND, not determined; TNF, tumour-necrosis factor.

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REVIEWS
Lumen
β-defensins Cathelicidins
Epithelial cell
a Glycocalyx
Desmosomes and
adherens junctions
Blood vessel
Neutrophil
Injury and infection
Macrophage
Pathogen
c concentrations of processed mouse CXCL5 are recov-
b
ICAM1
b
Chemokines, cytokines
and lipid mediators
Neutrophil
Figure 3 | Mucosal immunity. The mechanisms that epithelia use to defend against
microorganisms encompass at least four processes. a | Barrier function. Epithelia form
specialized macromolecular structures and polymers that are diplayed on their lumenal face,
such as the cornified layer of the epidermis (which is mostly crosslinked intermediate filaments),
the highly ordered uroplakin layer that lines the bladder and the glycocalyx of mucosal epithelia
(shown). In addition to barring or partly restricting the passage of solutes and water, these
lumenal structures provide a physical barrier to invasive microorganisms. The barrier function of
epithelia is augmented by various junctional complexes, such as desmosomes and adherens
junctions, that weld epithelial cells together. Also, intact epithelia release a variety of
antimicrobial peptides and proteins, such as β-defensins, cathelicidins and lysozyme, that
directly kill microorganisms. b | Re-epithelialization. Epithelia are constantly subjected to injury
and trauma, which ranges from the denudation of a few cells to large wounds. A breached
epithelium provides an entry point for infection. In response to injury, the wound-edge
epithelium of all tissues responds rapidly and in a similar manner to close the tissue gap. That
is, the cells at the edge of the wound change from a stationary to a migratory phenotype,
spread out and cover the wound. This process of re-epithelialization uses the daughters of
hyperproliferative cells that are present just behind the wound front. c | Bacterial clearance.
There are several means that epithelia use to remove invading bacteria at a wound site: first,
enhanced production and release of antimicrobial compounds; second, (potentially) apoptosis
(not shown),which might be selective for cells that have been infected with invasive pathogens;
and third, various physical processes that drive bacteria from the body, such as the mucociliary
apparatus in the airways, fluid flow in the sweat glands and excretion. These mechanisms
would also be active in the defence of intact epithelium. d | Inflammation. By their production of
chemotactic molecules, such as chemokines and lipid mediators, and their expression of
adhesion receptors, such as intercellular adhesion molecule 1 (ICAM1), epithelia provide the
initial host-derived signals that mediate and direct the influx of inflammatory cells, such as
neutrophils, to sites of injury or infection.
624 | AUGUST 2004 | VOLUME 4
MCP2) and CCL13 (also known as MCP4) to produce
antagonist factors60. Although MMPs have long been
considered to augment inflammation and the associated
tissue damage, these examples indicate that they can
also dampen inflammatory processes (FIG. 4).
In addition to MCPs, the function of other CC- and
CXC-chemokines is altered by direct MMP proteolysis.
CXC-chemokine ligand 11 (CXCL11; also known as
SDF1) is a substrate of several MMPs (MMP1, -2, -3, -9,
-13 and -14) both in vitro using purified proteins and in
cell-culture models, and the processing of CXCL11 by
MMP2 yields a product that has potent neurotoxic
activity67. Similarly, processing by MMP9 leads to a loss
of chemotactic activity of various chemokines, such as
CXCL5 (also known as ENA78), CXCL6 (also known as
GCP2) and mouse CXCL5 (also known as GCP2 or
LIX). By contrast, the processing of CXCL8 (also known
as interleukin-8, IL-8) by MMP9 markedly increases its
chemotactic activity62,63. In vitro, MMP8 can also cleave
mouse CXCL5, generating peptides with enhanced
neutrophil-chemotactic activity; however, in vivo, high
ered from the inflamed lungs of MMP8-deficient
mice68, indicating that other proteinases can function to
regulate the activity of this CXC-chemokine. In addi-
tion, MMPs might regulate the expression of chemokine
receptors and signalling through these receptors, pro-
viding an indirect mechanism by which these proteinases
can regulate leukocyte influx69. Although the studies dis-
cussed here indicate that MMPs can act on chemokines
and alter their activity, a complete deficiency in the pro-
teolytic processing of a specific chemokine has not yet
been shown in any model of cellular recruitment in
MMP-deficient mice.
MMPs establish chemokine gradients. The bioavailability
of chemokines is regulated by the level of biosynthesis,
the expression of the cognate receptor(s) and the modi-
fication of chemokines by proteinases. In addition, it is
also regulated by immobilization of chemokines to the
ECM or to cell surfaces. In vivo, chemokines form
chemotactic gradients by binding to accessory macro-
molecules (typically the glycosaminoglycan side chains
of proteoglycans), thereby providing directional cues to
migrating leukocytes. By acting on these accessory mol-
ecules, MMPs can indirectly regulate chemokine activity
and, in turn, the influx of inflammatory cells. A good
demonstration of this mechanism is provided by
the ability of epithelial-derived MMP7 to shed the
ectodomain of syndecan-1, a transmembrane heparan
sulphate proteoglycan70. In this situation, three epithelial
components — a secreted proteinase (MMP7), a cell-
bound proteoglycan (syndecan-1), and a chemokine
(mouse CXCL1; also known as KC) — function coordi-
nately to control and confine acute inflammation,
specifically NEUTROPHIL TRANSEPITHELIAL MIGRATION, to sites of
injury (FIG. 4). In response to mucosal injury, epithelial
cells synthesize, secrete and deposit CXCL1 (or CXCL8
in humans) onto the heparan sulphate chains of pre-
existing syndecan-1 molecules. MMP7, which is also
induced by injury, cleaves the syndecan-1 core protein at
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 REVIEWS

a juxtamembrane site to release the ectodomain–
CXCL1 complex. The shed complex either actively or
passively creates a chemotactic gradient that guides neu-
trophils into the alveolar space. In bleomycin-injured
lungs of MMP7-deficient mice, neutrophils extravasate
easily from the vasculature, but they are either held at
the epithelial–interstitial interface or in markedly
expanded perivascular spaces, and do not enter the
lumen of the lung. Interestingly, the interaction between
MMP7 and heparan-sulphate-containing molecules
increases the catalytic activity of MMP7 (REF. 24). So, the
glycosaminoglycan chains on syndecan-1 might func-
tion as an accessory factor (FIG. 2) that links MMP7 to its
substrate, the core protein of the proteoglycan.

Lumen
MMP9

Soluble FASL d

FASL
Neutrophil
c elastase
Pathogen

Adherens junctions
Epithelial cell

b
a MMP12
e TNF-α
Syndecan-1 CXCL8/
CXCL1
MMP7
ADAM17
Macrophage

CCR3/CCR1

CXCR2 f
CCL7

Fibroblast
MMP2

Blood vessel

Neutrophil

NEUTROPHIL TRANSEPITHELIAL
MIGRATION
During bacterial infections at
mucosal sites, neutrophils
migrate from the vasculature
through the interstitial
compartment and across the
epithelial barrier. The activation
and migration of neutrophils
into lungs also contributes to
inflammatory tissue injury and
remodelling of tissue
architecture
Figure 4 | MMPs in inflammation in response to tissue injury. Injury initiates a programmed, coordinated series of responses to
both repair the damaged tissue and to defend against infection. Almost all resident cells, particularly epithelial cells, endothelial cells
and fibroblasts, participate in these processes and contribute to the regulation of inflammation. This occurs partly through the
specific activity of a variety of matrix metalloproteinases (MMPs) that are produced by these cells. a | Soon after injury, epithelial cells
at the wound edge produce a chemokine (in humans, CXC-chemokine ligand 8, also known as interleukin-8, and in mice CXCL1,
also known as KC) that accumulates on the heparan sulphate chains of syndecan-1, a transmembrane proteoglycan. At the same
time, these cells release MMP7, which sheds the ectodomains of syndecan-1, thereby establishing a local chemokine gradient that
controls the influx and activation of neutrophils. b | Later on in the repair process, epithelial-derived MMP7 cleaves the ectodomains
of epithelial (E)-cadherin, thereby disrupting adherens junctions and, in turn, facilitating cell migration. Re-epithelialization is also
facilitated by the action of other MMPs, such as MMP1 in skin and MMP9 in lung cells. c | MMP7 also sheds and activates FAS
ligand (FASL, also known as CD95L) that is produced by epithelial cells, thereby mediating apoptosis, which is a potential innate
defence mechanism (discussed in text). d | After activation, neutrophils release several proteases. Among them, neutrophil elastase,
a serine protease that is exclusively produced by neutrophils, which has direct antimicrobial activity. Mice deficient in this enzyme
have an impaired ability to defend against Gram-negative bacteria98. Activated neutrophils also release MMP9, which degrades and
neutralizes the serine protease inhibitor α1-antiproteinase99, a potent inhibitor of neutrophil elastase. In this setting, MMP9 provides
cover for the antimicrobial activity of neutrophil elastase, thereby assigning it an indirect role in innate immunity. e | The activation of
the latent form of tumour-necrosis factor (TNF) on the surface of cells such as macrophages is due to metalloproteinase-mediated
proteolysis. In addition to ADAM17 (a disintegrin and metalloproteinase; also known as TNF-converting enzyme, TACE), MMP7 and
MMP12 can activate latent TNF (TABLE 1). f | The influx of inflammatory cells is mainly directed by specific chemokines that are
released by resident cells. In addition to indirect effects on chemokine activity, as discussed in a, MMPs also directly act on
chemokines, either enhancing or abrogating their activity. For example, MMP2, which is typically produced by mesenchymal cells,
can cleave and inactivate CC-chemokine ligand 7 (also known as macrophage-chemotactic protein 3, MCP3). CCR, CC-chemokine
receptor; CXCR, CXC-chemokine receptor.

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REVIEWS
This inhibition of neutrophil recruitment in MMP7-
deficient mice that have been subjected to acute lung
injury results in reduced lethality compared with wild-
type animals66. However, if the neutrophils are forced
into the alveolar space, by administration of a bacterial
chemotactic peptide, nFNLP, then the MMP7-deficient
mice are more susceptible to the lethal effects of acute
lung injury than wild-type mice. These observations
indicate that neutrophil activation and its oxidative
burst, which probably cause the lethality, are held in
check until these leukocytes reach the lumenal space,
where they would first encounter any microorganism
attempting to enter the tissue through the breached
epithelium. Therefore, MMP7-mediated proteolysis
would be high in the hierarchy of events that control
neutrophil activation.
The observation that MMP7-deficient mice are pro-
tected against acute lung injury and the subsequent
lethal effects of a pro-inflammatory insult does not indi-
cate that in all cases MMP7 is a detrimental proteinase.
Although in the experimental model discussed, massive
neutrophil influx and oxidative burst cause indiscrimi-
nate, severe and potentially mortal damage, neutrophils
comprise an essential cellular component of innate host
defence; and in alternative situations, MMP7 has a ben-
eficial function in neutrophil influx and activation, as
well as in mucosal immunity and epithelial migration66.
Although MMP7 might not directly process chemo-
kines at these sites60, it has a leading role in regulating
the formation of a chemotactic gradient that controls
neutrophil influx and activation.
Other MMPs also regulate the formation of
chemokine gradients. Allergen-induced airway inflam-
mation is dampened in MMP2-deficient mice, an
observation associated with reduced levels of CCL11
(also known as eotaxin) in lavage fluid64. How MMP2
controls the bioavailability of CCL11 is not known, but
it is interesting that this pro-inflammatory function is
distinct from the anti-inflammatory action of MMP2
on MCP chemokines (CCL2, -7, -8 and -13). In the
same allergen model, MMP9 was also recently shown
to be required for formation of transepithelial gradients
of CCL11, as well as CCL7 and CCL17 (also known as
TARC). But again, the mechanism by which MMP9
facilitates the movement of these chemokines from one
tissue compartment to another is not known71. MMP9
also affects the ability of CXCL8 to stimulate the release
of leukocytes from bone marrow, but similar to the
allergen model, the mechanism by which this occurs
(that is, the identity of the target substrate) has not
been determined. In addition, MMP3 is known to
mediate the release of a macrophage-chemotactic activ-
ity from chondrocytes72, and MMP12 is required for
the influx of macrophages into smoke-exposed lungs73.
But the nature of these activities, and whether they
involve chemokines, is not known. Collectively, these
findings show that several MMPs can regulate an
inflammatory response by controlling the activity and
mobilization of chemokines, and further work to iden-
tify the key substrates in these processes will provide
insight into fundamental mechanisms of inflammation.
626 | AUGUST 2004 | VOLUME 4
MMPs and cytokines
Several studies have indicated that MMPs can directly or
indirectly affect the activity of various cytokines that
function in inflammation and repair processes, includ-
ing interferon-β74, vascular endothelial growth factor75,
epidermal growth factors76, fibroblast growth factors77
and transforming growth factor-β1 (TGF-β1). As shown
using TGF-β1-deficient mice, this cytokine functions to
restrain mononuclear inflammation78–80. TGF-β1 is
released by cells with its cleaved pro-domain still latently
associated, and several mechanisms, including MMP
proteolysis, have been proposed to release the active
cytokine from this complex. In both cells and tissue-
explant models, MMP3 (REF. 81), MMP9 (REF. 23) and
MMP14 (REF. 82) have been shown or suggested to acti-
vate a proportion of total TGF-β1. If these or other
MMPs can activate TGF-β1 in vivo, then this would be
another mechanism by which MMPs restrain, rather
than augment, inflammation.
Similarly, IL-1β, a potent pro-inflammatory
cytokine, requires proteolytic processing for activation, a
process attributed to the IL-1β-converting enzyme
(ICE, also known as caspase-1). Although the function
of ICE had been well established in vitro, studies using
ICE-deficient mice provided evidence of other mecha-
nisms of IL-1β activation83. At least three members of
the MMP family, MMP2, -3 and -9, can cleave and acti-
vate the IL-1β precursor84. Furthermore, after activating
IL-1β, MMP3 degrades the biologically active
cytokine84, which can also be inactivated in vitro by
MMP1, -2 and -9 (REF. 85). These data indicate a dual role
for MMPs in biphasic modulation of inflammatory-
mediator activity: they are involved in both activation
(MMP3 and MMP9, and more weakly MMP2) and
inactivation (MMP3).
Another essential pro-inflammatory mediator that
is regulated by metalloproteinase activity is tumour-
necrosis factor (TNF), which is produced as a 26-kDa
membrane-associated protein (proTNF) and is cleaved
by TNF-converting enzyme (TACE) into a soluble
17.5-kDa cytokine. Because synthetic metallo-
proteinase inhibitors block this cleavage, it was sug-
gested that TACE was an MMP86. However, when the
convertase activity was purified and cloned, TACE was
found to be identical to ADAM17 (REFS 87,88), a member
of the disintegrin family of metalloproteinases
(ADAMs). The cleavage of proTNF by ADAM17 is spe-
cific89, and because the release of active TNF is reduced
by 90% in cells derived from ADAM17-deficient mice,
ADAM17 does seem to be the principal physiological
TNF-converting enzyme in vivo. Even if ADAM17 is
the main modulator of the generation of TNF activity,
several MMPs (including MMP1, -2, -3, -9 and -17)
can process proTNF to its active form in vitro89,90.
Furthermore, as shown using cells from knockout
mice, MMP7 and MMP12 also activate proTNF on
macrophages (TABLE 1). So, whereas ADAM17 is seem-
ingly involved in the inducible, high-level release of
TNF in response to bacteria and toxic shock, MMP7
and MMP12 might elicit the constitutive release of
TNF from macrophages that is required for common
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 REVIEWS

functions, such as tissue resorption and resolution in
response to injury (FIG. 4).
The future
We have discussed many examples of how MMPs func-
tion in innate immunity and inflammation, but we have
not mentioned acquired immunity. Because many
MMPs influence macrophage behaviour (TABLE 2),
this might indicate that they also affect the antigen-
presentation function of these cells. In addition, func-
tions associated with lymphocyte influx and T helper 2
cell cytokines, such as IL-13 (TABLE 2), support the con-
tention that MMPs have diverse functions in immunity.
One challenge for the MMP field is to further uncover
the function of MMPs: that is, to identify authentic sub-
strates using physiologically relevant approaches and
systems, and to do this with an open mind. Importantly,
determining precise MMP–substrate interactions might
provide an alternative and more precise strategy to
block specific and potentially detrimental processes that
are associated with inflammation and immune-mediated
disease.

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Competing interests statement
The authors declare no competing financial interests.
Online links
DATABASES
The following terms in this article are linked online to:
Entrez Gene:
http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=gene
ADAM17 | CCL7 | CXCL1 | CXCL8 | FAS | FASL | IL-1β | MMPs |
syndecan-1 | TGF-β1 | TNF
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